Contents

Division of Bioconvergence Technology

3 BioNanotechnology Research Center

A-1 Synthesis and characterization of fluorescent fullerene-silica nanoparticles for bioimaging applications

A-2 Multifunctional polymer nanoparticles for the labeling and tracking of immunotherapeutic cells

A-3 High level expression and purification of thrombin-activatable procaspase-7(TAC) in E. coli

A-4 Detection of conformationally changed MBP using intramolecular FRET

A-5 Development of fluorescence probes specific to ß2-adrenergic receptor

A-6 Label-free array detection of miRNA by structure-specific RNA binder

A-7 Nanogap biosensor for label-free detection of prostate-specific antigen

A-8 Near-infrared emitting fluorescent nanocrystals-labeled natural killer cells as a platform technology for the

optical imaging of immunotherapeutic cells-based cancer therapy

6 Aging Research Center

A-9 A novel p21Cip1 phosphorylation/degradation pathway playing a role in the ERK-mediated cell proliferation

signal

A-10 Identification of age-related genes in mammalian muscle

A-11 HBx induced liver dysfunction related with IGF-II signaling pathway

A-12 The roles of peroxiredoxin II as a modulator of erythrocyte apoptosis

A-13 JNK/FOXO-mediated neuronal expression of fly homologue of peroxiredoxin II reduces oxidative stress

and extends lifespan

A-14 Processed short neuropeptide F peptides regulate growth through the ERK-insulin pathway in Drosophila

Melanogaster

8 Brain Research Center

A-15 The phosphorylation of PTPRT attenuated the synapse formation in the hippocampal neurons

A-16 Novel interaction between PTPRT and neuronal cell adhesion molecules

9 Integrative Omics Research Center

A-17 Identification and characterization of b-galactosidases of Mannheimia succiniciproducens

A-18 Genome-wide transcriptional responses of Escherichia coli to metagenomic DNA

A-19 Skn7 mediated oxidative stress response in the methylotrophic yeast Hansenula polymorpha

A-20 A novel surface-associated ß-galactosidase of Streptococcus pneumoniae with Galß1-3GlcNAc moiety

specific hydrolysis activity

A-21 High-throughput N-glycan analysis of recombinant antibodies derived from plant using a DNA

sequencing equipment

A-22 Osmosensing two-component signal transduction system of the methylotrophic yeast Hansenula

polymorpha

12 BioMonitoring Research Center

A-23 Expression of functional biotinylated scFv in cell-free protein synthesis system

A-24 Electrical detection of enzyme-catalyzed metal deposition on FIB-fabricated interdigital electrode

Division of Translational Research

3rd KRIBB Poster Festival Abstracts

15 Medical Genomics Research Center

B-1 Tristetraprolin regulates the stability of HIF-1 mRNA during prolonged hypoxia

B-2 Study of the action mechanism of anti-diabetic drugs by transcriptome profiling

B-3 Genome-wide methylation profiling with gastric cancer cells and embryonic stem cells

B-4 Peroxiredoxin I contributes to TRAIL resistance through suppression of redox-sensitive caspase activation

in human hepatoma cells

B-5 Development of fluorescent microsphere immunoassay for Cystatin B (CSTB) in human blood samples

B-6 MD 164, a novel migration inhibitor, inhibits PRL-3 induced migration activity in DLD-1 cells

B-7 YY22 induces apoptosis through down-regulation of heat shock proteins in human pancreatic cancer cells

B-8 PRL-3 promotes the tumor metastasis via up-regulation of MMP expression in human colorectal cancer cells

B-9 Action mechanism of a novel JAK/STAT pathway inhibitor ; KT-18618 suppresses interaction between

JAK3 and STAT3 via binding at the JAK3 molecule

19 Medical Proteomics Research Center

B-10 Screening of target genes of Yap2

B-11 Reduced formation of advanced glycation endproducts via interactions between glutathione peroxidase 3

and dihydroxyacetone kinase 1

B-12 The molecular interaction between HAX-1 and XIAP inhibits apoptosis

B-13 New opportunities for neutron protein crystallography

21 Development and Differentiation Research Center

B-14 Cytoskeleton-associated proteins are enriched in human embryonic-stem cell-derived neuroectodermal

spheres

B-15 Notch signaling is required for maintaining stem-cell features of neuroprogenitor cells derived from

human embryonic stem cells

B-16 Localization of oocyte-specific DNA methyltaansferase-1 during pocine early development

Division of Biosystems Research

25 Industrial Biotechnology & Bioenergy Research Center

C-1 Complete genome sequence of Escherichia coli BL21(DE3)

C-2 Biosynthesis of the peptide antibiotic polymyxin in the surrogate host Bacillus subtilis

C-3 Enhancing 1-butanol tolerance in Escherichia coli through repetitive proton beam irradiation

C-4 Autoinducible expression system in Bacillus subtilis

C-5 Comparative genome sequence analysis of Bifidobacterium bifidum BGN4

C-6 Genome analysis of the secondary metabolism in Streptomyces clavuligerus ATCC 27064

C-7 Functional analysis of the Type II secretion systems in Escherichia coli BL21(DE3)

C-8 The crystal structure of the human NDRG2 (N-Myc downstream-regulated gene 2) protein, a potential

tumor suppressor

C-9 Genetic encoding Escherichia coli to monitor catalytic activity In Vivo

C-10 Novel detection mode of multiple sugars using conformational relaxation of FRET sensors at high

temperature

C-11 One-step cloning and simultaneous expression in Escherichia coli mediated by homologous recombination

C-12 PESTAS: A web server for EST analysis and sequence mining

C-13 GarlicESTdb: an online database and mining tool for garlic EST

C-14 EKIS: An integrated database of EST information for research application

C-15 Evolutionary characterization of a highly repetitive sequence identified from the false killer whale

(Pseudorca crassidens)

C-16 Comparative analysis of expressed sequence tags from the white-rot fungi (Phanerochaete chrysosporium)

C-17 Airborne induction and priming of plant defenses against a bacterial pathogen

C-18 Induced systemic resistance elicited by bacterial genetic materials

C-19 Metabolic engineering for squalene production in Saccharomyces cerevisiae

32 Plant Systems Engineering Research Center

C-20 Silencing of the catalytic subunit of ferredoxin:thioredoxin reductase induces pathogenesis-related genes

and pathogen resistance in tomato

C-21 CaPIR1 is a RING-finger E3 ligase protein in pepper that negatively regulates plant basal resistance

C-22 Screening of tissue specific genes and functional analysis in tomato

C-23 Functional RNAi suppressor for Chlamydomoans reinhardtii

C-24 Classification of rice (Oryza sativaL. japonica Nipponbare) immunophilins (FKBPs, CYPs) and

expression patterns under water stress

35 Bioinformatics Research Center

C-25 Korean efforts on tomato chromosome 2 genome sequencing

C-26 SOLmiRNA : A comprehensive search tool for the identification of microRNAs and target genes in

Solanaceae.

C-27 Pepper EST database: comprehensive in silico tool for analyzing the chili pepper (Capsicum annuum)

transcriptome

C-28 CVD Finder: a web-based finder of integrated article resources for cerebrovascular disease

C-29 CVD Base: an integrated genetic information resource for cerebral stroke

C-30 CVD information site: web portal-site supplying essential information of cerebrovascular disease based

upon Oriental medicine

C-31 AsiDesigner: exon-based siRNA design server considering alternative splicing

38 Industrial Bio-materials Research Center

C-32 Anti-atherosclerotic effects of Redobovatol, analogue of obovatol on atherogenic lesion formation in

LDL receptor-deficient mice

C-33 Effects of ethanolic extracts of Artemisia princeps Pamp. cv. Sajabal in high-fat diet-induced obese mice

C-34 Ethanol extract of Torreya nucifera seed blocks hepatic steatosis and atherosclerosis in LDL receptor-

deficient mice

C-35 Novel GH10 xylanase, with a fibronectin type 3 domain, from Cellulosimicrobium sp. strain HY-13, a

bacterium in the gut of Eisenia fetida

C-36 Anti-inflammatory properties of Tellimagrandin I, isolated from Corylopsis coreana in mousemacrophages

C-37 Linoleic acid derivative (S)-glycerol-monolinoleate attenuate atherogenic lesion formation in LDL

receptor-deficient mice

41 Environmental Biotechnology Research Center

C-38 Optimization for the harvest of high-density Scenedesmus sp. culture using a biopolymer produced by

Paenibacillus polymyxa AM49

C-39 Anaerobic VC to ethene dechlorinating enrichment culture

C-40 Axenic Induction method for axenic culture of a cyanobacterium, Microcystis aeruginosa

C-41 Effect of light emitting diode wavelength for the lipid content and composition of Chlorella vulgaris

C-42 Vinaxanthone, a new type of FabI inhibitor from Penicillium sp.

C-43 New type of enoyl-ACP reductase inhibitor from Sporothrix sp.

C-44 New type of FabG inhibitor from Bacillus sp. AT28

C-45 Transgenic sweetpotato plants expressing a bacterial phytase gene

C-46 Enhanced tolerance to MV-mediated oxidative stress and high temperature in transgenic potato plants

overexpressing CuZnSOD, APX and NDPK2 genes

C-47 Stress-induced expression of choline oxidase in potato plant chloroplasts confers enhanced tolerance to

oxidative, salt and drought stresses

C-48 Overexpression of sweetpotato orange gene conferred enhanced carotenoid contents in sweetpotato and

Arabidopsis

C-49 Development of transgenic sweetpotato plants expressing swpa4 peroxidase gene

C-50 Transgenic crops with enhanced tolerance to multiple environmental stresses on marginal lands

C-51 Differential expression of starch biosynthesis-related genes by exogenous sucrose in sweetpotato cultivars

C-52 Green biotechnology of natural rubber in plants

Division of Bio R&D Infra

49 Korean Bioinformation Center

D-1 MitoInteractome: Mitochondrial protein interactome database, and its application in ‘aging network’ analysis

50 Biological Resource Center

D-2 Salt-tolerance genes as selectable marker for genetic plant transformation of duckweed

D-3 Arabidopsis mutants defective in anthocyanin biosynthesis are arranged in a cascade manner in the

multivariate analysis score plot of their FT-IR spectroscopic profiles

D-4 Salipiger sp. nov., a moderately halophilic bacterium isolated from a solar saltern

D-5 Roseivivax sp. nov., isolated from a solar saltern

D-6 Nocardioides panacisoli sp. nov. isolated from the soil of a ginseng field in South Korea

D-7 Homologous overexpression of omcZ, a gene encoding outer surface c-type cytochrome, critical for high-

dense electricity generation in Geobacter sulfurreducens by single-step gene replacement

D-8 Paenibacillus pini sp. nov., a cellulolytic isolate from the rhizosphere of a pine tree

D-9 Cohnella gyejoki sp. nov., xylanolytic bacteria isolated from the rhizosphere

D-10 Recognition of the ascomycetous yeast Candida acaciaensis sp. nov., as a distinct species based on

polyphasic taxonomic study

D-11 Candida rhododendronensis sp. nov., a novel member of the Metschnikowiaceae

D-12 Paenibacillus panacisegetis sp. nov., isolated from soil of a ginseng field

D-13 A novel bacterium, Patulibacter ginsengiterrae sp. nov., isolated from soil of ginseng

D-14 A novel bacterium, Solirhabdus panacisegetis gen. nov., sp. nov., isolated from soil of ginseng field

D-15 A polyphasic investigation on a novel halophilic archaeon isolated from sea salt

D-16 A tRNA based primer cocktail for mitochondrial COI barcoding of hexapoda

D-17 DNA barcoding the hemiptera

56 Biotechnology Process Engineering Center

D-18 Production of amino butyric acid by Lactobacillus sp. isolated from traditional Korean fermented foods

Others

59 Viral Infectious Disease Research Center

E-1 VDAC-2 regulates apoptosis in dexamethasone-induced human eosinophils

E-2 Application of poly-gamma-glutamic acid improves dermatitis in NC/Nga mice

E-3 Poly(gamma-glutamic acid)-chitosan nanoparticles induces antigen specific humoral and cellular immunity

60 International Biological Material Research Center

E-4 Inclusion of Luffa tuberosa Roxb. in Momordica L. (Cucurbitaceae): evidence from ITS sequences of

nrDNA

E-5 Relationships among subfamily cucurbitoideae (Family Cucurbitaceae) from India inferred from ITS

sequences of nrDNA

E-6 Cypsela morphology of Stephanomeriinae (Compositae) and its taxonomic significance

E-7 Studies on seedling morphology of some medicinal plants of Korea

E-8 A molecular systematic study of some species of Trichosanthes L. (Cucurbitaceae) using ITS sequences of

nrDNA and its taxonomic implication

E-9

63 Laboratory of Experimental Animals

E-10 Preventative effects of phellinus baumii on hepatic steatosis in high fat diet-fed C57BL/6 mice

64 DAEJEON-KRIBB-FHCRC Research Cooperation Center

Ochang Branch Institute

67 Therapeutic Antibody Research Center

F-1 The protease inhibitor, elafin, induces p53-dependent apoptosis in human melanoma cells

68 Stem Cell Research Center

F-2 The effects of tumor necrosis factor-alpha on in vitro differentiation of natural killer cells

69 Immune Modulator Research Center

F-3 Suppressive effects of methanol extract from Lilium lancifolium on inflammatory responses in raw264.7 cells

F-4 Anti-inflammatory and anti-asthmatic effects of Capsicum annuum L. on Ovalbumin-induced lung

inflammation in a mouse asthma model

F-5 Anti-inflammatory and anti-asthmatic effects of tiarellic acid (TA) in a mouse model of allergy

F-6 Constituents of Anthriscus sylvestris and suppressive effect on matrix metalloproteinase-9 in airway

remodeling

71 Molecular Cancer Research Center

F-7 Two positive regulation genes from the Geldanamycin biosynthetic cluster in Streptomyces hygroscopicus

JCM4427

72 Chemical Biology Research Center

F-8 Regulation of integrin alpha2 (ITGA2) expression by MAPK signal cascades in Ki-ras transformed

prostate cancer cell lines (267B1/Ki-ras)

F-9 Regulation of ARL4C (ARL7) expression by MAPK signal cascades in Ki-ras transformed prostate cancer

cell lines (267B1/Ki-ras)

F-10 ATM blocks tunicamycin-induced endoplasmic reticulum stress

F-11 Endoplasmic reticulum stress induction by antimycins isolated from actinomycete AN070088

F-12 Isolation and structure determination of PTP1B inhibitory peptide, enniatins from fungi isolate No. 332

F-13 Verrucarin A, a inhibitor ER-stress induced XBP-1 activation and GRP78 in FaO cells

F-14 ER stress-induced cervical cancer cell death by Pimaradienoic acid involving GRP78 regulated caspase-

12 activation and c-Jun N-terminal kinase

F-15 Clitocybins A-D of novel isoindolinone compounds from the culture broth of Clitocybe aurantiaca

F-16 5-pentyl-2-furaldehyde, a melanogenesis inhibitor from Clitocybe sp.

F-17 Isodeoxyhelicobasidin, a novel human neutrophil elastase inhibitor from the culture broth of Volvariella

bombycina

F-18 Anti-melanogenic effect of milk thistle extract

F-19 Reticulone, a novel free radical scavenger produced by Aspergillus sp.

76 Bio-Evaluation Center

F-20 Species differences in serum stability of hydroxamate-containing histone deacetylase inhibitors

F-21 Artemisinin inhibits lipopolysaccharide-induced nitric oxide production by blocking IFN-b/STAT-1

signaling

F-22 DBM1285 suppresses LPS-induced TNF-a production and attenuates development of rheumatoid arthritis

in adjuvant-induced arthritis model

F-23 CML-08-62 inhibits the growth of human lung cancer cell through the induction of apoptosis

F-24 YAL-1112, a novel NF-kB inhibitor, inhibits the growth of colon cancer cells through induction of

apoptosis and cell cycle arrest

F-25 Glabridin inhibits dendritic cell maturation and function by blocking the activation of NF-kB/Rel and

MAPKs

F-26 Hepatocellular carcinoma in genetically obese mice

F-27 Lack of vitamin D3 Up-regulated protein 1 (VDUP1) accelerates liver injury through AKT activity

F-28 Natural variations of genes controlling soybean seed coat and flower colors under domestication

F-29 Molecular genetic assessment and quantitative detassion of a CMV-tolerant transgenic pepper line

F-30 Seed longevity and dormancy of transgenic Capsicum annuum L. resistant to cucumber mosaic virus

80 Korea National Primate Research Center

F-31 Molecular characterization of outer membrane vesicles produced from shiga toxin mutants of Escherichia

coli O157:H7

F-32 Role of estradiol-17ß on porcine blastocysts produced by in vitro fertilization or somatic cell nuclear

transfer

F-33 Gain of new exons and promoters by lineage-specific transposable elements-integration and conservation

event on CHRM3 gene

F-34 Bioinformatic analysis of constitutive TE-containing exons and nonsense-mediated mRNA decay within

human genome

F-35 Functional analysis of VSIG1 in mammalian spermatogenesis

Jeonbuk Branch Institute

85 Molecular Bioprocess Research Center

G-1 Development new enzyme screening protocols for high throughput screening system

G-2 New screening strategy of Exo-type cellulase from metagenomic resources based on high-throughput

screening (HTS) system

G-3 Screening of various microbial hydrolases from bovine rumen

G-4 Elimination of by-products formation during production of 1,3-propanediol from glycerol in Klebsiella

pneumoniae by inactivation of the oxidative pathway for glycerol metabolism

G-5 Statistical optimization of two-step fermentation for 1,3-propanediol production by recombinant Klebsiella

pneumoniae

G-6 Identification and utilization of a 1,3-propanediol oxidoreductase isoenzyme for production of 1,3-propanediol

from glycerol in Klebsiella pneumoniae

G-7 Cloning and expression of Bacillus amyloliquefaciens type 1 levansucrase gene in Bacillus megateriumand

optimization of fermentation conditions for the production of levansucrase by recombinant strain

G-8 Expression of human papillomavirus Type 16, 18, 33, 58 L1 proteins and production of virus-like particles

for vaccination in Sf-9 cells

G-9 Influence of cellular fatty acids composition on ethanol tolerance in Escherichia coli

88 Bioindustry Research Center

G-10 Antiviral effect of isolated flavonols from Rhodiola rosea on influenza A virus

G-11 Characterization of bacteriocin produced by Lactococcus lactis ET-45 isolated from Kimchi

G-12 The pure ß-glucan production in Aureobasidium pullulans IMS-822 KCTC11179BP by Pullulan

synthetase gene disruption

Open Innovation

91 Cooperation Between Medicine and Biotechnology

H-1 Sentinel node detection with PET/MR/fluorescent multi-modal imaging using 68Ga-SCN-NOTAsilica

nanoparticle

H-2 HSP90 regulates angiotensin II-induced hypertrophy via STAT pathway and IL-6 secretion in vascular

smooth muscle cell

H-3 Clinical significance of L1 expression in breast and thyroid cancer

H-4 The function of mitochondrial in granular corneal dystrophy type 2

H-5 15-hydroxy prostagladin dehydrogenase (15-PGDH) is a possible early detection marker in coloncancer :

analysis of prostagladin pathway in human tissue

H-6 Identification of characteristic molecular signature of acute myocardial infarction from human peripheral

blood

H-7 Development of pH-sensitive MRI imaging carriers and evaluation the molecular imaging techniques

using ovarian cancer animal models

H-8 Gene polymorphism study for early detection of gastric precancerous lesions and search of gastric cancer

high-risk group

H-9

H-10 Proteome analysis for searching key marker associated with psoriasis vulgaris

H-11 Synthesis and characterization of fluorescent magneto polymeric nanoparticles for bimodal imaging

probes

H-12 VEGF gene therapy combined with mesenchymal stem cell improve neuronal survival

H-13 Therapeutic effects of hypoxia-inducible gene expression system in rat spinal cord injury model

96 BINT Convergence Technology

H-14 Design and evaluation of microfluidics based thermoelectric biosensor with split-flow microchannel

H-15 Novel method for coating the iron oxide nanoparticles with poly(amino acid) derivatives using

poly(succinimide)

H-16 Aqueous-processible photoresponsive polymers for addressable micro-patterning of multiple proteins

H-17 Novel ultra-sensitive immunoassay technology utilizing peroxidase-mimics from nano-biotechnology

H-18 pH responsive nanocomposites monitoring drug release

H-19 Fe3O4-Loaded pH-responsive polymer as a MRI probe for cancer diagnosis

H-20 Development of core technology for neuronal signal detection using Si nanowire-based nano-biochip

H-21 8-nitroguanine derivatives as potential candidates for selective bioconjugation

H-22 Dural stimuli-responsive dendritic-linear block copolymer

H-23 Liquid crystal-based monitoring of biomolecular interactions at aqueous-LC interfaces

H-24 Development of nanoplasmonics based biosensor for the real time, label-free detection of synaptic

network activity

H-25 Application of optofluidc sensor for highly sensitive bio/environmental detection

Division of Bioconvergence Technology

BioNanotechnology Research Center

Aging Research Center

Brain Research Center

Integrative Omics Research Center

BioMonitoring Research Center

3 r d K R I B B P o s t e r F e s t i v a l

3 Korea Research Institute of Bioscience and Biotechnology

A-1Synthesis and characterization of fluorescentfullerene-silica nanoparticles for bioimagingapplications

Jinyoung Jeong1, Miyoung Cho1, Yong Taik Lim1, NamWoong Song2, and Bong Hyun Chung1*

1BioNanotechnology Research Center, KRIBB2Nano-Bio Convergence Research Center, KRISS

Fluorescent nanomaterials are of great interest inbiotechnology for diverse applications such as cell imaging,targeting, and detecting agents. In this work, we prepared afluorescent nanoparticles based on fullerene-silicahybridization by a reverse-microemulsion method. Thefullerene-silica nanoparticles (FSNP) of 60 nm in diameterexhibited broad fluorescence with maximum peak at 600 nmby excitation 350 nm. Compared to conventional dye(Alexa488 fluorophor), the FSNP showed 10 times higherphotostability revealing by single dot fluoresecne analysis.XPS and FT-IR analysis revealed that fullerene werehybridized with silica network in form of C-O-Si which isoriginated in new fluorescence of FSNP. The fluorescencemicroscopic images of FSNP uptake various cell linedemonstrated that the FSNP were effective for cellpenetration and had potential in drug delivery. Besides, thecell viability result supported the utilization of FSNP as a safebioimaging agent. We believed that the new synthetic FSNPcan be a promising fluorescent probe for bioimaging analysis.(Spring Symposium of the Korean Institute of ChemicalEngineers : Gwangju, 2009)

Keywords : fullerene-silica hybridization, photoluminescence,bioimaing

A-2Multifunctional polymer nanoparticles for thelabeling and tracking of immunotherapeuticcells

Young-Woock Noh1, Yong Taik Lim2, and Bong HyunChung1

1BioNanotechnology Research Center, KRIBB 2ChungNam National University

In this study, we report on the development of biocompatiblepolymer-based nanoprobes for both magnetic resonanceimaging (MRI) and near-infrared (NIR) fluorescenceimaging. The biocompatible polymer nanoparticles-basedbimodal imaging contrast agents should be differentiatedfrom the superparamagnetic iron oxide nanoparticlesconjugated with fluorescent dyes in three points ; i) ICG isbiocompatible molecule which has been routinely used in

humans for over 40 years, while the in vivo toxicity ofmost fluorescent dyes used in conjugation with iron oxidenanoparticles is still less known. ii) the relaxivity (R2) ofPLGA nanoparticles encapsulating ICG and iron oxidenanoparticles, which is a measure of MR contrast, waslarger than that of superparamagnetic iron oxide nanoparticlesdue to the synergistic magnetism of multiple iron oxidenanoparticles satellites encapsulated in the PLGAnanoparticles matrix. iii) PLGA matrix is preferred for theencapsulation of therapeutic drugs in controlled releaseapplications. With the help of dual-functional PLGAnanoparticles, the migration of dendritic cells (DC) vialymphatic drainage was monitored through real-time NIRfluorescence imaging, and the homing of dendritic cellsinto lymph nodes through noninvasive MRI techniqueswas also accomplished. The biocompatible polymer-basedbimodal imaging contrast agents are expected to be usedfor the effective trafficking of DC to the lymph nodes ofpatients, which is highly required for the efficient DC-based immunotherapy of cancers. (First InternationalConference on Multifunctional Hybrid and Nanomaterials: Paris(France), 2009)

Keywords : NIR, molecular imaging, dendritic cells,PLGA

A-3High level expression and purification of thrombin-activatable procaspase-7(TAC) in E. coli

Young-Mi Lee1, Hyo Jin Kang1,2, and Sang J. Chung1,2

1BioNanotechnology Research Center, KRIBB 2University of Science and Technology (UST)

Caspase-7 is one of the executioner proteins of apoptosisand very similar to caspase-3 in the structure and substratespecificity. As caspase-3 and -7 are the final executionersof apoptosis, both inhibition and activation of their catalyticactivities are of significant interests as therapeutic targetsfor neurodegenerative diseases and cancers, therebyrequiring a large amount of the protein for drug discoveryresearches. Here we report a novel method for high-levelexpression and purification of caspase-7 precursors, whichwere strategically engineered to lack auto-activation duringexpression by mutating the auto-activation site to theamino acid sequences susceptible to thrombin hydrolysis.The engineered precursors were highly expressed as solubleproteins in E. coli, easily purified by affinity chromatography,to levels of 10-13 mg from 1 L of E. coli culture, and readilyactivated by thrombin digestion.

Seoul, 2009

Keywords : caspase-7, apoptosis, expression, precursor

BioNanotechnology Research Center

A-4Detection of conformationally changed MBPusing intramolecular FRET

Kyoungsook Park, So Yeon Yi, Yong-Beom Shin, BongHyun Chung, and Moonil Kim*

BioNanotechnology Research Center, KRIBB

Here we present the fluorescence resonance energy transfer(FRET) technology to explore maltose-induced conformationalchanges in MBP. For the FRET pair, ECFP and EYFPwere used as the donor and the acceptor, and were linkedgenetically to the C-terminal and N-terminal regions ofMBP (ECFP:MBP:EYFP), respectively. After the FRETreaction, maltose-treated MBP was shown to exhibit aconsiderable energy transfer (FRET efficiency (E) = ~0.11,Distance (D) = ~6.93 nm) at the ensemble level, which wasregarded as reflective of the increase in donor quenchingand the upshift in acceptor emission intensity, therebysuggesting that the donor and the acceptor had been broughtclose together as the result of structural alterations in MBP.However, upon glucose treatment, no FRET phenomenonwas detected, thereby implying the specificity of interactionbetween MBP and maltose. The in vitro FRET results werealso confirmed via the acceptor photobleaching method.Therefore, our data showed that maltose-stimulatedconformational changes of MBP could be measured byFRET, thereby providing biological information, includingthe FRET efficiency and the intramolecular distance.(Biotronics 2009 Conference : Seoul, 2009)

Keywords : FRET, ECFP, EYFP, MBP

A-5Development of fluorescence probes specific toß2-adrenergic receptor

Hyo Jin Kang and Sang J. Chung

BioNanotechnology Research Center, KRIBB

G-protein-coupled receptors (GPCRs) are the largest familyof integral membrane proteins coded by the human genomeand constitute significant medicinal targets. Among them,ß-2 adrenergic receptor (ß-2 AR) is structurally characterizedand serves as a well-studied prototype GPCR. Based onthe 3-dimensional structure of ß-2 AR complex with itsspecific ligand, fluorescence probes specific to ß-2 ARwere designed, synthesized, and tested for the specificrecognition of ß-2 AR overexpressed in a mammalian.When applied to ß-2 AR overexpressed cells over controlcells the probes showed a strong fluorescence intensity foronly ß-2 AR overexpressed cells but not the control cells.

Seoul, 2009

Keywords : protein, G-protein-coupled receptors, ß2-adrenergic receptor

A-6Label-free array detection of miRNA bystructure-specific RNA binder

Jeong Min Lee, Hyun Min Cho, Sun Hee Yeon, andYongwon Jung*

BioNanotechnology Research Center, KRIBB

MicroRNAs (miRNAs) are a class of small non-codingRNAs (19 - 25 nucleotides) that regulate gene expressionin viruses, plants, and animals. Accumulated studies havereported distinct miRNA expression patterns in varioushuman diseases, showing great potential of miRNAprofiling in clinical applications such as diagnosis anddrug efficacy evaluation. Microarray-based detectionoffers an efficient method for profiling large numbers ofmiRNA expressions simultaneously. Currently, mostmiRNA labeling methods utilize direct or indirectenzymatic labeling reactions against target miRNAs priorto (or sometimes after) hybridization to chip surfaceprobes. Developing an array detection method of miRNAsfree from labeling reactions will clearly simplify theprocess and greatly bolster the credibility of miRNAprofiling studies. Here we fabricated a novel structurespecific RNA binding protein, which acts like an antibodyand allows us to universally detect hybridized miRNAs onarray surfaces without the need of any enzymaticamplification or labeling reactions. (Biotronics 2009Conference : Seoul, 2009)

Keywords : microRNAs (miRNAs), microarray, novelstructure specific RNA binding protein

A-7Nanogap biosensor for label-free detection ofprostate-specific antigen

Sang Kyu Kim1,2, Hyunmin Cho1,2, and Bong HyunChung1,2

1BioNanotechnology Research Center, KRIBB2School of Engineering, University of Science and Technology(UST)

A simple method for fabrication of nanometer distancegap was developed using silicon anisotropic wet etchingtechnique on silicon-on-insulator (SOI) wafer with a finelycontrollable silicon device layer. Since the silicon anisotropic

Korea Research Institute of Bioscience and Biotechnology 4

wet etching resulted in the trapezoid-shaped structurewhose end side became narrower during the etching, thenanogap structure was simply fabricated by the siliconanisotropic wet etching on the SOI wafer. The size of thenanogap was controlled by adjusting the dimension of thewindow mask and finally the nanogap having a width lessthan 60 nm was obtained. Furthermore, we used the nanogapfor electrical biosensing of biomolecular interactionwithout labeling. The current was proportionally increased tothe concentrations of the charged molecules in the rangefrom 100 fg/ml to 100 ng/ml at 1 V bias. It is expected thatthe nanogap developed here can be used as a biosensorplatform for label-free detection of biomolecular interactions.(2009 Labautomation : Palm Springs (USA), 2009)

Keywords : nanogap, biosensor, PSA, Label-Free Detection

A-8Near-infrared emitting fluorescent nanocrystals-labeled natural killer cells as a platform technologyfor the optical imaging of immunotherapeuticcells-based cancer therapy

Mi Young Cho, Yong Taik Lim, Young-Woock Noh, JinWoong Chung, and Bong Hyun Chung

BioNanotechnology Research Center, KRIBB

This study describes the development of near-infrared opticalimaging technology for the monitoring of immunotherapeuticcell-based cancer therapy using natural killer (NK) cellslabeled with fluorescent nanocrystals. Although NK cell-based immunotherapeutic strategies have drawn interest aspotent preclinical or clinical methods of cancer therapy,there are few reports documenting the molecular imagingof NK cell-based cancer therapy, primarily due to the difficultyof labeling of NK cells with imaging probes. Human naturalkiller cells (NK92MI) were labeled with anti-human CD56antibody-coated quantum dots (QD705) for fluorescenceimaging. FACS analysis showed that the NK92MI cellslabeled with anti-human CD56 antibody-coated QD705have no effect on the cell viability. The effect of anti-humanCD56 antibody-coated QD705 labeling on the NK92MIcell function was investigated by measuring interferongamma (IFN- ) production and cytolytic activity. Finally,the NK92MI cells labeled with anti-human CD56 antibody-coated QD705 showed a therapeutic effect similar to thatof naive NK92MI cells. Images of intratumorally injectedNK92MI cells labeled with anti-human CD56 antibody-coatedcould be acquired using near-infrared optical imaging both in vivoand in vitro. This result demonstrates that the immunotherapeuticcells labeled with fluorescent nanocrystals can be a versatileplatform for the effective tracking of injected therapeuticcells using optical imaging technology, which is very importantin cell-based cancer therapies. ( )

Keywords : NIR, molecular imaging, natural killer cells,cancer therapy

Members

DirectorBong-Hyun Chung (chungbh@kribb.re.kr)

Principal Researcher Sang-Jeon Chung (sjchung@kribb.re.kr)Myung-Kyu Lee (mklee@kribb.re.kr)

Senior Researcher Chang-Su Lee (cslee@kribb.re.kr)Im-Sik Chung (cis123@kribb.re.kr)Yong-Won Jung (ywjung@kribb.re.kr)Moon-Il Kim (kimm@kribb.re.kr)Yoon-Kyung Kim (ykim@kribb.re.kr)

Post-DocDo-Hyung Kwon (kwondh@kribb.re.kr)Joong-Hyun Kim (joong@kribb.re.kr)Bea-Pan Kee (bpkee@kribb.re.kr)Hyo-Sook Chung (hyosookj@kribb.re.kr)

ResearcherJoo-Oak Keem (okbead4203@yahoo.co.kr)Young-Woock Noh (woock@kribb.re.kr)Kyoung-Mi Park (kyoungmi@kribb.re.kr)Kyoung-Sook Park (marsp@hanmail.net)Sun-Hee Yeon (lovesower@empal.com)So-Yeon Lee (yisoyeon@naver.com)Young-Mi Lee (utopia1981@hanmail.net)Ui-Jin Lee (uijin@kribb.re.kr)In-Hwan Lee (rodigy@kribb.re.kr)Jung-Min Lee (qnghkf@hanmail.net)Su-Jin Im (cocky35@kribb.re.kr)Do-Yun Cho (ds1bll@naver.com)Mi-Young Cho (fairylamp@naver.com)Hyang-Mi Cho (dkfk2782@hanmail.net)Hyun-Min Cho (hyunmin@kribb.re.kr)Yo Han-Choi (ryghkd@hanmail.net)Sun-Young Kim (kimsun@kribb.re.kr)Hyo-Jin Kang (jin0305@kribb.re.kr)Sang-Kyu Kim (tech81@hanmail.net)Ju-Hwan Kim (hiwnghks@kribb.re.kr)Hyun-Kyung Song (gkhs7519@kribb.re.kr)Eun-Ju Yun (salada@kribb.re.kr)Jin-Young Jeong (jin0305@kribb.re.kr)Hyun-Kyung Jeong (tousledhair@hanmail.net)Ji-Na Kwon (wlsk3170@nate.com)Renata Goo (tata1208@kribb.re.kr)Mi-Na Min (mina@kribb.re.kr)Il-Hwang Park (silver@kribb.re.kr)Hae-Yeon Jeong (seikotop@kribb.re.kr)Myung-Sun Jeong (jms0727@kribb.re.kr)Ae-Lee Jeong (aljung@kribb.re.kr)Eun-A Choi (euna83@kribb.re.kr)Jong-Min Choi (cjm2735@kribb.re.kr)

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A-9A novel p21Cip1 phosphorylation/degradationpathway playing a role in the ERK-mediatedcell proliferation signal

Chae Young Hwang1, Cheolju Lee2, and Ki-Sun Kwon1,*

1Aging Research Center, KRIBB2Life Sciences Division, Korea Institute of Science andTechnology

p21Cip1 is an inhibitor of cell cycle progression that promotesG1 phase arrest by direct binding to cyclin-dependentkinase and proliferating cell nuclear antigen. Here wedemonstrate that mitogenic stimuli, such as epidermalgrowth factor treatment and oncogenic Ras transformation,induce p21Cip1 downregulation at the post-translational level.We show that ERK2 interacts with and phosphorylatesp21Cip1, promoting p21Cip1 nucleocytoplasmic translocationand ubiquitin-dependent degradation. Phosphopeptideanalysis of in vitro ERK2-phosphorylated p21Cip1 revealedtwo phosphorylation sites, Thr57 and Ser130. Both thephosphorylation-deficient T57A/S130A p21Cip1mutant andthe NES mutant, that are resistant to ERK2-mediateddownregulation, retain the ability to potently inhibit theG1/S transition. These results indicate that ERK2 activationtransduces mitogenic signals, at least in part, by phosphorylation,translocation, and degradation of the cell cycle inhibitoryprotein, p21Cip1, suggesting an alternative mechanism ofRas-ERK signaling-mediated cell proliferation. (ASBMBAnnual Meeting : New Orleans (USA), 2009)

Keywords : p21Cip1, Ras-ERK signaling, ubiquitin,translocation

A-10Identification of age-related genes in mammalianmuscle

Seung-Min Lee1, Sung-Kyu Ju1. Seon-Young Kim2, andKi-Sun Kwon1

1Aging Research Center, KRIBB2Human Genome Research Center, KRIBB

In the previous study, we analysed three publicy availablegene expression data sets downloaded from Gene ExpressionOminibus (GSE5086, GSE80, GSE362) and screened genesshowing high correlation with aging. Filtering the data with|correlation coefficient|>0.2, p<0.001, and fold change>1.5,we identified 220 upregulated and 73 downregulated genes.In the present study, we examined age-related geneshereby identified are involved in aging of human dermalfibroblast and rat muscle by RT-PCR. Representatively,p21Cip1, p53, B-cell CLL/lymphoma 6 (BCL6) and myosin

heavy chain 11 (MYH11) were upregulated, whereas piwi-like 14 (PIWIL4) and pinin (PNN) were downregulated.Further analyses of these genes may provide insights intothe possible roles of aging. Suwon, 2009)

Keywords : gene expression, aging, human dermal fibroblast,muscle

A-11HBx induced liver dysfunction related withIGF-II signaling pathway

Hye-Lin Ha1, Hye-Jun Shin2, and Dae-Yeul Yu1,*

1University of Science and Technology(UST)2Aging Research Center, KRIBB

HCC is one of the most common cancers in the world. Chronicinfection with HBV is a major risk factor. Among the fourproteins from the HBV genome, HBx is a multifunctionalregulatory protein which has been reported to be linked withhepatocellular carcinogenesis. We reported HBx inducesliver cancer in transgenic mice. HBx mice showed dysplasiawithin 4 weeks, dysplastic nodules and HCA after 6 monthsand HCC after 9 months of age. IGF-II is the protein hormones,predominantly active in fetal liver. Higher expression ofIGF-II can be detected in HCC. HBx mice have an increasedliver/body weight ratio as well as cyclinD1 expressioncompared to wild mice after 2 weeks. At that point, highlyexpressed IGF-II in wild mice started to decrease but inHBx mice the IGF-II expression still remained high level.The HBx-HepG2 cell line also has a higher expressedIGF-II level than the Mock-HepG2 cell line. Like in vivodata, the HBx cell line showed fast growth and increasedcyclinD1 expression levels. In this study, we suggest IGF-II as an early diagnosis marker and it leads to liverdysfunction via over-proliferation of an HBx damagedhepatocyte. (The 21st Annual Meeting of the KoreanSociety for Molecular and Cellular Biology : Seoul, 2009)

Keywords : HBx, hepatocellular carcinoma, IGF-II, earlydiagnosis marker

A-12The roles of peroxiredoxin II as a modulator oferythrocyte apoptosis

Ying-Hao Han, Tae-Ho Kwon, Sun-Uk Kim, Hye-Lin Ha,and Dae-Yeul Yu*

Aging Research Center, KRIBB

Korea Research Institute of Bioscience and Biotechnology 6

Aging Research Center

Peroxiredoxin (Prx) II, a scavenger of hydrogen peroxide,has been reported to be essential for antioxidative defensein red blood cells (RBCs) [Lee et al., Blood 101(12), 2003].Prx II knockout mice showed higher levels of reactive oxygenspecies (ROS) and abnormal RBCs in their peripheral blood.To examine the physiological roles of Prx II in thesephenotypes, we checked apoptosis and state of erythrocytedifferentiation in Prx II knockout blood. CD71+ cells andannexin V binding were significantly increased. In addition,the RBC life span was significantly decreased in Prx IIknockout mice. To demonstrate the relationship amongROS, apoptosis and abnormal RBC cells, we treated withoxidative insults such as hydrogen peroxide and anilinehydrochloride in vitro and in vivo. By the treatment of theinsults, ROS level was significantly increased in Prx II-/-RBCs compared to wild-types, which resulted in moreadvanced phenotype of hemolytic anemia with increasedreticulocytes and reduced hematocrit level. These resultsare indicative of physiological roles of Prx II for protectionRBCs from oxidative stress-mediated apoptosis. (The 21stAnnual Meeting of the Korean Society for Molecularand Cellular Biology : Seoul, 2009)

Keywords : peroxiredoxin, RBCs, apoptosis, hemolyticanemia

A-13JNK/FOXO-mediated neuronal expression offly homologue of peroxiredoxin II reducesoxidative stress and extends lifespan

Kyu-Sun Lee1, Hye-Mi Ahn1, Jin-Yu Zhang1, Dong-SeokLee2,*, and Kweon Yu1,*

1Aging Research Center, KRIBB2College of Natural Sciences, Kyungpook NationalUniversity

Activation of c-Jun N-terminal kinase (JNK) signalingincreases stress resistance and extends lifespan throughFOXO-mediated transcription in Drosophila. However,the molecular identities of JNK/FOXO target genes are notfully understood. Here, we identified Jafrac1, a Drosophilahomolog of human Peroxiredoxin II (hPrxII), is adownstream effecter of JNK/FOXO signaling in neurons.We found that oxidative stress activated Jafrac1 expressionin flies. Neuronal overexpression of Jafrac1 or hPrxIIsuppressed oxidative stress-induced lethality in flies, whileinhibition of Jafrac1 expression in neurons aggravated thelethality. Neuronal expression of Jafrac1 also significantlyreduced the reactive oxygen species level, restoredmitochondrial function, and attenuated JNK activationcaused by oxidative stress. We demonstrated that activationof JNK/FOXO signaling increased the Jafrac1 expressionlevel, indicating that Jafrac1 is a target of JNK/FOXO-

mediated transcription. Moreover, neuronal over-expressionof Jafrac1 or hPrxII extended lifespan in flies while Jafrac1inhibition reduced lifespan. These findings indicate thatJNK/FOXO-mediated Jafrac1 expression in the nervoussystems play a protective role against oxidative stress andregulate lifespan in Drosophila. (The 21st AnnualMeeting of the Korean Society for Molecular andCellular Biology : Seoul, 2009)

Keywords : JNK/FOXO, Jafrac1, lifespan, Peroxiredoxin II

A-14Processed short neuropeptide F peptides regulategrowth through the ERK-insulin pathway inDrosophila Melanogaster

Kyu-Sun Lee, Seung-Hyun Hong, Ae-Kyeong Kim, andKweon Yu*

Aging Research Center, KRIBB

Drosophila short neuropeptide F (sNPF), an orthologue ofmammalian neuropeptide Y (NPY), and sNPF receptorsNPFR1 regulate food intake and body growth. Previouslywe report that sNPF/sNPFR1 signaling controls growth byextracellular signal-regulated kinase (ERK)-mediated insulinsignaling. sNPF. The sNPF gene encodes the precursorprotein for 4 active peptides including two RLRFamidepeptides. Here we have mapped the distribution of precursorprotein and two RLRFamide peptides in the central nervoussystem (CNS) with its receptor, sNPFR1. Localization ofthese peptides and sNPFR1 indicates that sNPF/sNPFR1proteins were co-expressed in some neurons of Drosophilalarval CNS. To analyze the tissue-specific regulation ofsNPF and sNPFR1 genes, we isolated the 2.5kb genomicfragment from 5' up-stream sequence of the sNPF andsNPFR1 genes and generated Gal4 lines, respectively.Overexpression of sNPF or sNPFR1 with own Gal4 linesshowed that sNPF/sNPFR1 signaling regulated food intakeand body growth. In cell-based assay, amidation in RFresidue of sNPF is necessary for its function. Theseobservations suggest that sNPF/sNPFR1 expressed innervous system may have an important role in feedingbehavior and growth regulation in Drosophila. (The 21stAnnual Meeting of the Korean Society for Molecularand Cellular Biology : Seoul, 2009)

Keywords : Drosophila short neuropeptide F, food intake,body growth,

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Members

DirectorKi-Sun Kwon (kwonks@kribb.re.kr)

Principal Researcher Sung-Sup Park (sspark@kribb.re.kr)Kweon Yu (kweonyu@kribb.re.kr)Dae-Yeul Yu (dyyu10@kribb.re.kr)

Senior Researcher Kyu-Sun Lee (ekuse74@kribb.re.kr)

Post-DocSoo-Jin Kwak (kwak@kribb.re.kr)Sun-Uk Kim (sunuk@kribb.re.kr)So- Hee Dho (shdho@kribb.re.kr)Sung-Kyu Ju (sannya@kribb.re.kr)Ying-Hao Han (hyh@kribb.re.kr)Seung-Hyun Hong (shhong77@kribb.re.kr)Chae-Young Hwang (cyhwang@kribb.re.kr)

ResearcherTae-Ho Kwon (taeho@kribb.re.kr)Ae-Kyeong Kim (iop234@kribb.re.kr)Young-Jae Bahn (yjbahb@kribb.re.kr)Keun-Woo Lee (zizigul@kribb.re.kr)Seung-Min Lee (lsjhm99@kribb.re.kr)Young-Lang Lee (winter79@kribb.re.kr)Mi-Ra Jung (jung@kribb.re.kr)Jeong-Yi Choi (cj673419@kribb.re.kr)Jin-Ok Jang (yeur@kribb.re.kr)Jong-Seong Ha (hadeng@kribb.re.kr)Hye-Lin Ha (stillbb@kribb.re.kr)Sang-Mi Cho (shosm@kribb.re.kr)Hye-Jun Shin (aromajun@kribb.re.kr)Kyung-Min Kim (79kkm@kribb.re.kr)Young-Ho Park (pyh2877@kribb.re.kr)Hye-Mi Ahn (cocohm5@kribb.re.kr)

A-15The phosphorylation of PTPRT attenuated thesynapse formation in the hippocampal neurons

So-Hee Lim, Eunha Park, and Jae-Ran Lee

Brain Research Center, KRIBB

PTPRT is the brain-specific expressed protein tyrosinephosphatase and was known to regulate the synapseformation of the hippocampal neurons. When PTPRT wasphosphorylated by neuronal tyrosine kinase Fyn, theinduction of synapse formation was attenuated. Thedimerization/multimerization of PTPRT was enhanced andthe catalytic activity was inhibited by phosphorylation.The key residue of phosphorylation was known to be 912tyrosine on the N-terminus of PTPRT catalytic domain.According to the structural modeling of PTPRT catalyticdomains the phosphorylation of 912 tyrosine residueseems to make the flexible wedge enter the active pocketof reciprocal catalytic domain and as a result the catalyticactivity was inhibited by the strengthened dimericinteraction. This novel regulation mechanism of PTPRTactivity by phosphorylation could be applied to thefunctions of many protein tyrosine phosphatases on thesynapse formation. (ISN/APSN 2009, 22nd BiennialMeeting of the International Society for Neurochemistry :Busan, 2009)

Keywords : PTPRT, Fyn, tyrosine phosphorylation,synapse formation

A-16Novel interaction between PTPRT and neuronalcell adhesion molecules

So-Hee Lim and Jae-Ran Lee

Brain Research Center, KRIBB

The receptor-type protein tyrosine phosphatases have beenlinked to signal transduction, cell adhesion, and neuriteextension. PTPRT/RPTP is exclusively expressed in thecentral nervous system and regulates synapse formation byinteracting with cell adhesion molecules and protein tyrosinekinase. Overexpression of PTPRT in cultured neuronsincreased the number of excitatory and inhibitory synapses.In contrast, knockdown of PTPRT inhibited synapseformation and withered dendrites. Endogenous interactionpartners of PTPRT were examined in the rat brainsynaptosome by immunoprecipitation using a PTPRT-specific monoclonal antibody and tandem mass spectrometryfor amino acid sequencing. As a result, neuroligin wasidentified as a candidate interaction partner of PTPRT.Both neuroligin and neurexin interacted with PTPRT

Korea Research Institute of Bioscience and Biotechnology 8

Brain Research Center

through the extracellular ecto-domains. Incubation ofcultured neurons with recombinant proteins containing theextracellular region of PTPRT reduced the number ofsynapses by inhibiting the interaction between ecto-domains. Neuroligin was recruited by overexpressedPTPRT and synapse formation by neuroligin was notinduced when PTPRT was knocked-down in culturedneurons. These results suggest that brain-specific PTPRTregulates synapse formation through interaction with celladhesion molecules. (The Fall International Conventionof the Parmaceutical Society of Korea : Seoul, 2009)

Keywords : PTPRT, cell adhesion molecules, neuroligin,neurexin

Members

Director Jae-Ran Lee (leejr@kribb.re.kr)

Senior Researcher Baek-Soo Han (bshan@kribb.re.kr)Kyung-Sim Kim (kskim@kribb.re.kr)

ResearcherEunha Park (emyth@hanmail.net)Areum Park (piggy225@hanmail.net)So-Hee Lim (im_sohee@hanmail.net)Hoe-Yune Jung (elijah98@kribb.re.kr)

A-17Identification and characterization of b-galactosidasesof Mannheimia succiniciproducens

Eun-Gyeong Lee1, Jong Moon Shin1, Doo-Byoung Oh1,Sang Yup Lee2, and Ohsuk Kwon1

1Omics and Integration Research Center, KRIBB2Department of Chemical and Biomolecular Engineering,Center for Ultramicrochemical Process Systems, andDepartment of BioSystems, BioProcess Engineering ResearchCenter and Bioinformatics Research Center, KAIST

Mannheimia succiniciproducens is a capnophilic Gram-negativebacterium isolated from bovine rumen and gainsincreasing attentions for its ability to produce a largequantity of succinic acid under anaerobic growthconditions. Various molecular biological and functionalgenomic tools such as shuttle vectors, gene knock-outsystem and DNA chip have been developed and employedfor metabolic engineering of organism. In this study, twoputative b-galctosidase genes of M. succiniciproducens,MS0806 and MS0749, were cloned and expressed inEscherichia coli. The recombinant proteins were used tostudy their biochemical propertied by using chromogenicsubstrates, o-nitrophenyl-b-D-galactopyranosid and p-nitrophenyl-b-D-galactopyranosi-de. To determine theirphysical roles, single and double disruption mutant strainswere constructed and their ability to use lactose as the solecarbon source was various growth conditions. Obtainedresults indicate that b-galactosidase reporter system will beuseful to evaluate the strength and behavior of a promoterin M. succiniciproducens. This work was supported by theGenome-based Integrated Bioprocess Project of theMinistry of Education, Science and Technology (MEST).(International Symposium & Annual Meeting of theKorean Society for Microbiology and Biotechnology :Daejeon, 2009)

Keywords : b-Galactosidase, operon fusion, Mannheimiasucciniciproducens

A-18Genome-wide transcriptional responses ofEscherichia coli to metagenomic DNA

Min Jee Kim1, Mun-Kyoung Min1, Jae Jun Song2, Doo-Byoung Oh1, and Ohsuk Kwon1,*

1Integrative Omics Research Center, KRIBB2Molecular Bioprocess Research Center, KRIBB

To explore the effect of metagenomic DNA on thegenome-wide expression in Escherichia coli, we comparedthe transcriptome profiles of E. coli transformed with the

9 3rd KRIBB Poster Festival

Integrative Omics Research Center

metagenomic DNA libraries to those of untransformed E.coli. When a fosmid library was transformed into E. coliEPI300 strain, 40 and 60 genes were up- and down-regulatedby at least twofold, respectively. Even though 60% of thedifferently expressed genes were found to encode proteinsof hypothetical or unknown functions, the majority of up-regulated genes upon fosmid transformation were functionallygrouped into those responsible for transport and bindingproteins. In contrast, the majority of down-regulated geneswere functionally categorized into energy metabolism. Further,when we compared changes in transcriptome profiles of E.coli upon transformation with different fosmid metagenomiclibraries, similar genes were shown to be differently expressedin both transformants. Thus, our transcriptome data willprovide useful information to understand the genome-wideresponse of E. coli to metagenome and to develop an efficienthost system for metagenome cloning and functional expressionfor activity-based screening. This work was supported byBio R&D Programs of the Korea Science and EngineeringFoundation (KOSEF). (International Symposium & AnnualMeeting of the Korean Society for Microbiology andBiotechnology : Daejeon, 2009)

Keywords : Escherichia coli, metagenomic, DNA

A-19Skn7 mediated oxidative stress response in themethylotrophic yeast Hansenula polymorpha

Eun Hye Kim1,2, Surisa Suwannarangsee1, Min Jee Kim1,Hyun Ah Kang3, Jeong-Yoon Kim2, Doo-Byoung Oh1,Warawut Chulalaksananukul4, and Ohsuk Kwon1,*

1Integrative Omics Research Center, KRIBB2Department of Microbiology, Chungnam NationalUniversity

3Department of Life Science, Chung-Ang University4Department of Botany, Chulalongkorn University,Thailand

The methylotrophic yeast Hansenula polymorpha is a modelorganism for various fundamental researches such asmethanol metabolism, peroxisome biogenesis and function,nitrate assimilation, and resistance to heavy metals, heatand oxidative stress. Here, we identified the H. polymorphaSkn7 response regulator which consists of 474 aminoacids with 3 domains: a receiver domain for phosphorelay,a DNA-binding domain, and coiled-coil region. The aminoacid sequence of Hpskn7 shows 32.5% of identity and41.6% of similarity to that of S. cerevisiae Skn7. Deletionof Hpskn7 gene (skn7 ) caused the cell sensitive to H2O2

and t-BOOH. To understand the physiological role of Skn7,transcriptome profiles of wild-type and skn7 mutant H.polymorpha strains cultured in the presence of H2O2 werecompared. Of 5,848 ORFs analyzed, 58 genes were up-

regulated more than two-folds in wild-type strain, whileonly 14 genes were up-regulated in skn7 mutant in thepresence of H2O2. Several genes known to be responsiblefor protection to oxidative damages were no longer inducedin skn7 mutant, suggesting that the HpSkn7 is involved inoxidative stress response In H. polymorpha. (InternationalSymposium & Annual Meeting of the Korean Society forMicrobiology and Biotechnology : Daejeon, 2009)

Keywords : Skn7, Oxidative stress, two component system

A-20A novel surface-associated ß-galactosidase ofStreptococcus pneumoniae with Galß1-3GlcNAcmoiety specific hydrolysis activity

Yun Mi Lee1, Jae Kap Jeong1, Jung Mi Lee1, Eun-HyeKim2, Tu Nhat Le2, Dong-Kwon Rhee2, Hyun-Ah Kang3,Doo-Byoung Oh1, and Ohsuk Kwon1

1Integrative Omics Research Center, KRIBB2College of Pharmacy, Sungkyunkwan University3Department of Life Science, Chung-Ang University

Streptococcus pneumoniae, the major causative agent forpneumonia, otitis media and meningitis, produces surfaceexoglycosidases such as neuraminidase (NanA), ß-galactosidase (BgaA) and N-acetylglucosaminidase(StrH). These enzymes play critical roles for colonizationand pathogenesis by sequentially hydrolyzing variousglycoconjugates to penetrate host defense molecules, toexpose binding sites on the surface of the epithelial cells orto obtain monosaccharides for growth. In this study, wecharacterized a novel ß-galactosidase encoded by the bgaCgene of S. pneumoniae. The recombinant BgaC proteinexhibited a highly regio- and sugar-specific hydrolysisactivity for Galß(1,3)GlcNAc moiety of oligosaccharides.Interestingly, the BgaC was shown to be expressed as asurface protein, even though it does not have a typicalsignal sequence or membrane anchorage motif. The bgaCmutant strains did not show detectable changes in growthor morphology compared to wild type stains whencultivated under normal laboratory conditions. However,the bgaC mutant showed higher colonization levels at 6and 12 h post-infection in vivo than the wild type. Our datastrongly indicate that S. pneumoniae BgaC is a surface-associated ß-galactosidase with a specific hydrolysisactivity that contributes significantly to the adherence andinvasion of pneumococci in vivo and in vitro. (

2009 , Koseong-aun, 2009)

Keywords : Streptococcus pneumoniae, ß-galactosidase(BgaC)

Korea Research Institute of Bioscience and Biotechnology 10

A-21High-throughput N-glycan analysis of recombinantantibodies derived from plant using a DNAsequencing equipment

Kyung Jin Lee1,2, Jin-Hee Jung1, Jung Mi Lee1, Yang KangSo2, Ohsuk Kwon1, Hyun Ah Kang3, Kisung Ko2,*, andDoo-Byoung Oh1,*

1Integratve Omics Research Center, KRIBB2Department of Biological Science, College of NaturalSciences, Wonkwang University

3Department of Life Science, College of Natural Science,Chung-Ang University

High-throughput quantitative analytical method for plantN-glycan has been developed. All steps, including peptideN-glycosidase (PNGase) A treatment, glycan preparationand exoglycosidase digestion, were optimized for high-throughput applications using 96-well format proceduresand automatic analysis on a DNA sequencer. The glycansof horseradish peroxidase with plant-specific core (1,3)-fucose can be distinguished by the comparison of theglycan profiles obtained via PNGase A and F treatments.The peaks of the glycans with (91%) and without (1.2%)

(1,3)-fucose could be readily quantified and shown toharbor bisecting ß(1,2)-xylose via simultaneous treatmentwith (1,3)-mannosidase and ß(1,2)-xylosidase. Thisoptimized method was successfully applied to analyze N-glycans of plant-expressed recombinant antibody, whichwas engineered to contain a minor amount of glycanharboring ß(1,2)-xylose. These results indicate that ourDNA sequencer-based method provides quantitativeinformation for plant-specific N-glycan analysis in a high-throughput manner, which has not previously beenachieved by glycan profiling based on mass spectrometry.( 2009 , Koseong-aun, 2009)

Keywords : DNA sequencer, plant N-glycan, Glycan anlysis,(1,3)-fucose, ß(1,2)-xylose

A-22Osmosensing two-component signal transductionsystem of the methylotrophic yeast Hansenulapolymorpha

Surisa Suwannarangsee1, Eun Hye Kim1, Eun Kyung Lee1,Min Jee Kim1, Won Hee Han1, Yun Mi Lee1, Hyun AhKang2, Doo-Byoung Oh1, Warawut Chulalaksananukul3,and Ohsuk Kwon1,*

1Integrative Omics Research Center, KRIBB2Department of Life Science, Chung-Ang University3Department of Botany, Chulalongkorn University,Thailand

The methylotrophic yeast Hansenula polymorpha is anattractive model organism to study resistance to variousstresses such as heavy metals, heat and oxidative stress. Inthis study, based on the recently determined whole genomesequence of H. polymorphaDL-1 strain we have identifiedgenes coding for protein homologous to SLN1-YPD1-SSK1 two-component signal transduction system of S.cerevisiae. Deduced amino acid sequences of HpSLN1,HpYPD1, and HpSSK1, respectively, showed 35%, 27%,and 20% of identities to corresponding proteins of S.cerevisiae. Each gene was expressed in E. coli and resultingrecombinant proteins were used to characterize theirphosphorylation and transphosphorylation properties invitro. Furthermore, to study the physiological role of thesystem, mutant strains deleted individually for sln1, ypd1,or ssk1 genes were constructed and their growth phenotypeunder various stress conditions were examined. Obtaineddata indicate that SLN1-YPD1-SSK1 system of H. polymorphaplays important role in osmoregulation. (InternationalSymposium & Annual Meeting of the Korean Society forMicrobiology and Biotechnology : Daejeon, 2009)

Keywords : Hansenula polymorpha, SLN1-YPD1-SSK1

Members

DirectorKwang-Lae Hoe (Kwanghoe@kribb.re.kr)

Principal Researcher Ohsuk Kwon (oskwon@kribb.re.kr)Young-Woo Park (ywpark@kribb.re.kr)

Senior Researcher Dongu Kim (kimdongu@kribb.re.kr)Doo-Byoung Oh (dboh@kribb.re.kr)

Post-DocSeong-Hun Kim (seonghun@kribb.re.kr)Ki-Won Jo (Kiwonjo@yahoo.co.kr)Dong-Hui Lee (dowon74@hanmail.net)Sun-Jeong Jo (csuns@hanmail.net)Si-Young Lee (lee1974@kribb.re.kr)

ResearcherMin-Jeong Sohn (picorna@kribb.re.kr)Jyung-Mi Lee (ladymi@hanmail.net)Eun-Mi Jung (99bilyu@hanmail.net)Yun-Mi Lee (ccoccalcon@nate.com)Eun-Gyeong Lee (ummny@hanmail.net)Kyung-Jin Lee (kyungjin@kribb.re.kr)Ji-Young Mun (moonji20@hanmail.net)Eun-Mi Kim (classic0311@naver.com)Mun-Kyeong Min (ximber@naver.com)Ji-Ae Shin (jiae0616@nate.com)Jin-Hee Choi (jinhee83@kribb.re.kr)Oh-Cheol Kim (kill572@kribb.re.kr)

11 3rd KRIBB Poster Festival

Ju-Young Kim (dabi5372@kribb.re,kr)Seul-Gi Yun (puppy-35@naver.com)Jae-Won Jeon (jeonjw74@naver.com)Ji-Hyun Park (a0148@hanmail.net)Yeong-Sun Jang (soonjy33@magicn.com)Jun-Gu Jeong (yulaman@hanmail.net)Hye-Won Lee (positivelyhw@naver.com)Jung Yu (purity0918@yahoo.co.kr)Eun-Jeong Song (potato0224@hanmail.net)Suk-Ho Yoo (mito0926@dreamwiz.com)Ji-Hyun Moon (mintjw@nate.com)Myung Ho Sohn (sohnho2@naver.com)Seon Ju Heo (cellhsj@hanmail.net)Hye-In Choi (hamadryades82@hanmail.net)Jeong-Eun Lee (biomodam@hotmail.com)Jin-Mi Oh (jinmi84@hanmail.net)Chan-Woong Park (bbara23@naver.com)Eun-Kyung Lee (eun0316@hanmail.net)Seon-Ha Yun (tozoo@hanmail.net)Kyu-Won Cho (hislove100@naver.com)Dong-Jin Kim (dongjin_k_@naver.com)Mi-Ju Park (lovelymiju@nate.com)A-Reum Lee (iwtbmd@kribb.re.kr)Won-Sun You (wsyou72@kribb.re.kr)Mi-Young Nam (nmy1219@kribb.re.kr)Hye-Mi Lee (hyemilee@kribb.re.kr)

A-23Expression of functional biotinylated scFv incell-free protein synthesis system

Ju-Young Byun, Dong-Myung Kim1,*, and Min-Gon Kim2,*

1Department of Fine Chemical Engineering and Chemistry,Chungnam National University

2Biomonitering Research Center, KRIBB

In this report, we described expression of functionalbiotinylated scFv in cell-free protein synthesis system. Anmethod, which can be used for site-specific biotinylationof recombinant antibodies, takes advantage of thecapability of the enzyme biotin ligase to catalyze theattachment of a biotin to a unique lysine residue in specificsequence. In order to biotinylate the scFv for detection, insitu biotinyltion of scFv was expressed in a cell-freeprotein synthesis reaction through the fusion of N-terminus and C-terminus of anti CRP-scFv with biotinligation site(called Avi-tag) by two-step PCR method. Andthen the scFv was isolated by affinity-tag purificationmethod. We have showed that recombinant scFv wassuccessfully prepared and we do not need to add the biotinligase(BirA) to express the biotinylated scFv. The resultsalso shown that the scFv fusion protein could be used fordirect detection of its antigen in ELISA and Western blotswhen stained with streptavidin conjugated horseradishperoxidase. Taken together, we are currently working toimmobilize the scFv using the capture antibody ofsandwich ELISA. (International Symposium & AnnualMeeting of the Korean Society for Microbiology andBiotechnology : Daejeon, 2009)

Keywords : cell-free protein synthesis, biotinylation, scFv

A-24Electrical detection of enzyme-catalyzed metaldeposition on FIB-fabricated interdigitalelectrode

Junhyoung Ahn1,2, Tae Han Lee1,2, Kwang Heo3, SeunghunHong3, Yong-Beom Shin1,2, and Min-Gon Kim1,2,*

1School of Engineering, University of Science andTechnology (UST)

2BioMonitoring Research Center, KRIBB3Department of Physics and Astronomy, Seoul NationalUniversity

Electrical detection method has been used widely onbiosensor technology, such as cyclic voltammetry, andimpedommetry. For increasing sensitivity, there has beenstudied to enhance electrical signal [1]. Enzyme-catalyzedmetal deposition is useful in biosensor system for detecting

Korea Research Institute of Bioscience and Biotechnology 12

BioMonitoring Research Center

low concentration targets as enhancing electrical signal.Silver ion is easily reduced and deposited on metal particle,i.e. gold nanoparticle. However there are not severalmethods with enzyme-catalysis [2]. Therefore, we needsilver-enhancement by enzyme catalysis for further studyof electrical detection with ELISA. Horse-radish peroxidase(HRP) has been used for enzyme-linked immunosorbentassay (ELISA). We established the electrical detection oninterdigital electrode filled with HRP-catalyzed silver-precipitator. (Nano Korea 2009 : Koyang, 2009)

Keywords : interdigitated electrode, FIB, silver precipitation

Members

DirectorMin-Gon Kim (mgkim@kribb.re.kr)

Principal Researcher Yong-Beom Shin (ybshin@kribb.re.kr)

Senior Researcher Tae-Hwan Ha (taihwan@kribb.re.kr)Sang-Jick Kim (sjick@kribb.re.kr)

Post-DocSeung-Woo Lee (swlee7607@gmail.com)Dong-Hwan Choi (dhchoi1@hanmail.net)

ResearcherJun-hyoung Ahn (cultenigma@empal.com)Ju-Young Byun (bjy8349@nate.com)Tae-Han Lee (mars02_20@hanmail.net)Hwa-Young Sung (shy8338@hanmail.net)Young-Kyoung Oh (hggnei@hanmail.net)Yun-Ju Sung (sungv7@naver.com)Min-Su Kang (mskang4u@gmail.com)Song-I Yang (lovely-nest@hanmail.net)

13 3rd KRIBB Poster Festival

Division of Translational Research

Medical Genomics Research Center

Medical Proteomics Research Center

Development and Differentiation Research Center

3 r d K R I B B P o s t e r F e s t i v a l

B-1Tristetraprolin regulates the stability of HIF-1mRNA during prolonged hypoxia

Tae Woo Kim, Sujin Yim, Byung Joong Choi, Yejin Jang,Jung Ju Lee, Bo Hwa Sohn, Hyang-Sook Yoo, Young IlYeom*, and Kyung Chan Park*

Medical Genomics Research Center, KRIBB

Hypoxia inducible factor-1 (HIF-1) is a transcriptionfactor involved in the cancer cell adaptation to hypoxia, aleading cause of tumor malignancy. Thus, control of HIF-1 expression may assist in treatment of cancer. Theexpression of HIF-1 is finely regulated via alterations intranslation rate, and mRNA stability as well as proteinstability, which occurs in response to changes in O2 levels.Within the 3'-untranslated region (UTR) of HIF-1mRNA, there are seven pentamer (AUUUA) and onenonamer (UUAUUUAUUUAUU) AU-rich elements,which are putative motifs for RNA binding factors such astristetraprolin (TTP). Here, we show that TTP mRNAexpression was reduced and HIF-1 mRNA expressionelevated in hepatocellular carcinoma tissues. We alsofound that the TTP protein bound directly to the 3'-UTR ofHIF-1 mRNA to downregulate stability. Furthermore, TTPexpression was induced in hypoxic cells and overexpressionof this protein repressed the hypoxic induction of HIF-1protein. Taken together, these data suggest that TTP is amodulator of HIF-1 expression during prolonged hypoxiaand may play a physiological role in cellular adaptation tohypoxia. In addition, cancer cells may benefit from thedownregulation of TTP, which subsequently increasesHIF-1 expression and assists with the adaptation ofcancer cells to hypoxia. (The 21st Annual Meeting of theKorean Society for Molecular and Cellular Biology : Seoul,2009)

Keywords : TTP, HIF-1, ARE, mRNA stability

B-2Study of the action mechanism of anti-diabeticdrugs by transcriptome profiling

Suk-Jin Yang1, Keum-Jin Yang2, Young-Hoon Kim2,Seomg-Min Park1, Ye-Jin Jang1, Young-Seong Kim2,Chul-Ho Lee2, and Young Il Yeom1*

1Medical Genomics Research Center, KRIBB2Laboratory of Experimental Animals, KRIBB

The present study aimed at identifying candidate markergenes useful for the classification of anti-diabeticsaccording to their pharmacological efficacy by applyingthe DNA chip technology (Illumina 24K Mouse DNA

chip) and statistical methods for large-scale transcriptomeanalyses. We treated db/db and db/+ mice with 5 differentanti-diabetic agents having distinct action mechanisms.We found that, except for the insulin secretagogue, the-glucosidase inhibitor and insulin sensitizers could exert

therapeutically valid anti-diabetic efficacy in our animal model,that the TZD family drugs show significant hepatotoxicityand induce hypertrophy in fat tissue, weight gain and reductionof blood lipids, and that metformin has hepatoprotectiveeffects and induce significant weight loss. Transcriptomeprofiles for the total RNA of 4-5 target organs (liver, fat,muscle, pancreas and intestine) identified statisticallyvalidated predictor gene sets that can classify anti-diabeticdrugs according to their pharmacological action mode infat and liver tissues (p<0.01-0.02, 33 and 103 genes for fatand liver, respectively). Finally, we performed geneontology and pathway analyses and literature mining forthe predictor gene sets, and collected bioinformatic datathat are relevant with the pharmacological action of eachanti-diabetic drug. (The Fall International Conventionof the Parmaceutical Society of Korea : Seoul, 2009)

Keywords : transcriptomics, anti-diabetics, biomarker, DNAchip, mouse model

B-3Genome-wide methylation profiling with gastriccancer cells and embryonic stem cells

Han-Chul Lee, Mirang Kim, Jong-Lyul Park, Seon-YoungKim, and Yong Sung Kim

Medical Genomics Research Center, KRIBB

The epigenetic regulation may play an important role indevelopment of both embryonic stem cell and cancer cell.DNA methylation, one of epigenetic components, hasexperienced a large increase in interest in the last yearsand the analysis of DNA methylation either on a genome-wide or gene-specific scale has come to centre stage formany biological and medical questions. Here we appliedIllumina’s infinium methylation array to the discovery ofmethylation signatures that are specific in gastric cancercells (GCCs) or human embryonic stem cells (hESCs). Wefound a significant difference between gastric 11 GCCsand three hESCs, suggesting that genome-wide methylationpattern depend on cell lineage. Cell type-specifichypermethylation was found in 82 CpG sites from 72 genesfor GCCs or in 65 CpG sites from 61 genes for in hESCs,respectively. We addressed these cell type-specific hyper-or hypomethylated genes to the functional enrichment usingthe GOstat tool. Hypermethylated genes in GCCs wereenriched with function annotations such as DNA bindingand hypermethylated genes in hESCs were enriched withfunction annotations such as olfactory receptor activity,

15 3rd KRIBB Poster Festival

Medical Genomics Research Center

chemokine activity. We need functional study of specificmethylated genes but our results may help to understand ofgastric cancer development and hES cell development. (The100th Annual Meeting of American Association forCancer Research : Denver (USA), 2009)

Keywords : gastric cancer, human embryonic stem cells,methylation

B-4Peroxiredoxin I contributes to TRAIL resistancethrough suppression of redox-sensitive caspaseactivation in human hepatoma cells

In-Sung Song, Nang-Su Oh, Ga-Hee Ha, So-Young Jung,and Nam-Soon Kim

Medical Genomics Research Center, KRIBB

We evaluated the role of peroxiredoxin (Prx) I in TRAILresistance governed by coupling of nicotinamide adenosinedinucleotide phosphate oxidase (Nox)-derived ROSsignaling with the p38 mitogen-activated protein kinase(MAPK)/caspase-signaling cascade in liver cancer cells.Upregulated Prx I expression was found in neoplastic regionsof human patient liver, and Prx I knockdown resulted inaccelerated TRAIL induced cell death in SK-Hep-1 humanhepatoma cells. The TRAIL cytotoxicity by Prx I knockdownwas dependent on activation of caspase-8/3 cascades, whichwas ablated by addition of inhibitors for p38 MAPK, ROSor Nox, suggesting the association with Nox-driven redoxsignaling. Furthermore, we found that Nox4 was constitutivelyexpressed in both SK-Hep-1 cells and tumor regions ofpatient livers, knockdown of Nox4 expression couldalleviate ROS generation and TRAIL-mediated cytotoxicity.In accordance with previous findings, increased activationof both p38 MAPK and caspase cascades by Prx I knockdownwas inhibited byeither Nox4 knockdown or SB203580addition. Collectively, these data suggest that Prx Ifunctions to block propagation of Nox-derived ROSsignaling to the p38 MAPK/caspase/cell death cascadeduring TRAIL treatment and also provides a molecularmechanism by which Prx I contributes to TRAIL resistancein liver cancers. (2009 Meeting on Cell Death, Cold SpringHarbor Laboratory : New York (USA), 2009)

Keywords : ROS, TRAIL, p38, cell death

B-5Development of fluorescent microsphereimmunoassay for Cystatin B (CSTB) in humanblood samples

Na Young Ji1,4, Chung IL Lee1, Mi-YoungPark1, Hee GuLee1, Jae Wha Kim1, DaeGhon Kim2, Jong Wan Kim3, andEun Young Song1

1Medical Genomics Research Center, KRIBB2Department of Internal Medicine, Chonbuk National University,Medical School and Hospital

3Department of Laboratory Medicine, Dankook UniversitySchool of Medicine

4University of Science & Technologh (UST)

Serum Cystatin B (CSTB) concentrations have been reportedto be elevated in patients with hepatocellular carcinoma (HCC)as compared with that of normal subjects. In this study, wehave developed a “Fluorescent Microsphere Immunoassay”(FMI) which is capable of specifically detecting CSTB inserum specimens. We have produced polyclonal antibody(pAb) and monoclonal antibody (mAb) to CSTB. We haveselected four hybridoma clones which generated mAbs thatrecognize CSTB based al ELISA and Western blot analysis.The FMI was composed with fluorescent microsphereconjugated pAb to CSTB and biotinylated mAb, as captureprotein and probe protein, respectively. The results wereobtained using the Luminex200system. The dose-responserelationship betweel aSTB and fluorescent intensity showedlinearity in a range 0~eaaa pg/ml and a sensitivity was 7 pg/ml.The FMI was more sensitive, and more reproducible thanthe ELISA. We have determined the aSTB concentrationsof serum specimens with the FMI assay system. The resultswere similar to those measured with ELISA. The nbilydeveloped FMI method can be used as a multiplexedimmunoassay tool for the detection of CSTB with otherbiomarkers in human serum, particularly for cancer diagnostics.This work was supported by a Frontier Function HumanGenomic Project and a KRIBB Research Initiative fundfrom the Korean Ministry of Education, Science andTechnology (MEST) provided by the Korean Government.(BIEN 2009 : INWES Asian Network : Busan, 2009)

Keywords : CSTB, fluorescent microsphere immunoassay,ELISA

B-6MD 164, a novel migration inhibitor, inhibitsPRL-3 induced migration activity in DLD-1 cells

Jina Kim, Hye-Nan Kim, Yujin Lee, Dong Cho Han, andByoung-Mog Kwon

Medical Genomics Research Center, KRIBB

PRL-3 is a metastasis-associated phosphatase. PRL-3 isexpressed at higher levels and at a greater frequency incolorectal cancer metastases compared with primary colorectaltumors and normal colon tissue and knockdown of PRL-3

Korea Research Institute of Bioscience and Biotechnology 16

expression in DLD-1 cells by RNA interference effectivelyabrogated the activities of cancer cell motility in vitro andhepatic colonization in vivo. Potent and selective PRLinhibitors may ultimately constitute a novel and effectivefamily of anti-cancer agents. Thus we are attempts searchingof PRL-3 inhibitors. In the course of the search for PRL-3inhibitors through migration assay in PRL-3-overexpressingDLD-1cells using 783 clinically used drugs and 57 herbalmedicines, MD164 was discovered as a hit. This drug iswidely used as a cough suppressant in non-productive cough.In our results, treatment of MD164 effectively suppressedmigration and invasion of PRL-3-overexpressing DLD-1cells using trans-well migration assay, trans-well invasionassay, and tube formation assay. These results suggest thatMD164 inhibits the migration and invasion of PRL-3-overexpressing DLD-1 cells and can be good candidate forthe therapeutic control of cancer cell invasion or metastasis.(The 66th Annual Meeting of the Korean Society forBiochemistry and Molecular Biology : Seoul, 2009)

Keywords : PRL-3, migration

B-7YY22 induces apoptosis through down-regulation of heat shock proteins in humanpancreatic cancer cells

Young Ju Yoon, Ki-Deok Shin, Byung-Mog Kwon, andDong Cho Han

Medical Genomics Research Center, KRIBB

Heat shock proteins (HSPs) are highly expressed in a widerange of tumors and are correlated with a poor prognosisand resistance to therapy. HSPs help unfolded protein toadequately fold and protect cells from external stressors.Knockdown of HSPs using siRNA decreased proliferationof cancer cells. Pancreatic cancer is a malignant diseaseand has the lowest survival rate among the human cancers.It has been reported that numerous pancreatic carcinomahas a great dependency on HSPs to their growth and survival.Therefore, targeting HSPs using small molecules is anattractive for human cancer therapy. Heat shock factor 1(HSF1) is the master switch of HSPs expression in eukaryotes.To discover selective modulators of HSF1 activity, wedeveloped cell-based reporter assay. An initial screeningof synthetic and natural compounds library identified YY22as a HSF1 inhibitor. YY22 inhibited HSF1-dependentexpression of HSP70 in a concentration-dependent manner,causing cell cycle arrest and apoptosis in pancreatic cancercell. Collectively, these data indicate that YY22 inducesapoptosis through inhibition of HSF1 and can be a promisinglead compound for anti-cancer drug. (The 66th AnnualMeeting of the Korean Society for Biochemistry andMolecular Biology : Seoul, 2009)

Keywords : heat shock factor 1 (HSF1), heat shock protein70 (Hsp70), apoptosis, anti-cancer activity

B-8PRL-3 promotes the tumor metastasis via up-regulation of MMP expression in humancolorectal cancer cells

Dae-Seop Shin1, Young Ran Ha1, Su-Kyung Lee1, Dongcho Han1, and Byoung-Mog Kwon1,2

1Medical Genomics Research Center, KRIBB2Biomolecular Sciences, University of Science and Technology(UST)

Cancer metastasis to secondary organs is the major causeof death among cancer patients. However, molecular changesin a tumor cell that promote the metastatic process arelargely unclear. Matrix metalloproteinases (MMPs) are afamily of zinc endopeptidases that degrade extracellularmatix (ECM) components. MMPs have been proved to bea main process in local invasion and migration. PRL-3 issubgroup of protein tyrosine phosphatases (PTPs) whichwould also degrade the ECM and overexpressed in colorectalcancer metastasis compared with primary tumors. To determinewhether migration enhanced by PRL-3 was dependent onMMPs expression, we engineered DLD-1 human colorectalcancer cell overexpressing PRL-3. When expressions ofMMPs (2, 7, 8, 9, 13 and 14) were tested with real-timequantitative PCR, mRNA level of MMP-2, 13 and 14 wereincreased in PRL-3 overexpressed cells. We confirmedthat expression of PRL-3 could promote human colorectalcancer cell migration and invasion. Knockdown of MMP-2, 13 and 14 with siRNA completely inhibited the migrationand invasion of the PRL-3 overexpressed cells. Theseresults suggest that MMPs are target genes of PRL-3 andare directly or indirectly associated with PRL-3 signalingpathway to promote cell migration and invasion of tumorcells. (The 66th Annual Meeting of the Korean Society forBiochemistry and Molecular Biology : Seoul, 2009)

Keywords : PRL-3, MMP, migration, colon

B-9Action mechanism of a novel JAK/STAT pathwayinhibitor ; KT-18618 suppresses interactionbetween JAK3 and STAT3 via binding at theJAK3 molecule

Dae-Seop Shin1, Young Min Han1, Young Ju Yoon1, KiYong Min3, Dong Cho Han1, and Byoung Mog Kwon1,2

1Medical Genomics Research Center, KRIBB

17 3rd KRIBB Poster Festival

2Biomolecular Sciences, University of Science and Technology(UST)

3KRICT

We screened small-molecules inhibiting STAT3 activity inchemical library containing 1,333 synthetic compoundsobtained from “Korea Chemical Bank” through a cell-basedluciferase assay system. KT-18618 was selected as a novelinhibitor of JAK/STAT3 pathway. KT-18618 inhibitsSTAT3 phosphorylation at 1 hour and the expressfer of theSTAT3-regulated genes at the same time point. We postulatedthat inhibitier of JAK family proteins or c-Src result in inhibitierof STAT3 phosphorylcifer. Interestingly, phosphorylationof these kinases was mildly inhibited at 1 hour. This resultimplies that inhibitier of STAT3 phosphorylation by KT-18618 is independent event with phosphorylc-tier of up.Team kinases. Co-immunoprecipitation shows that KT-18618 inhibits the JAK3-STAT3 interaction. Moreover,JAK3 molecules were captured by biotylcied KT-18618implying that KT-18619 bind to JAK3 molecules. Finally,KT-18618 inhibits kinase activity of JAK3 about 30% at10 µM in vitro kinase assay. From these results, we suggestthat KT-18618 binds to JAK3 molecules and disturbsJAK3-STAT3 interaction which leads to inhibition ofSTAT3 phosphorylation. (The 21st Annual Meeting of theKorean Society for Molecular and Cellular Biology :Seoul, 2009)

Keywords : JAK3, STAT3, breast, cancer

Members

DirectorYoung-Il Yeom (yeomyi@krib.re.kr)

Principal Researcher Yong-Sung Kim (yongsung@kribb.re.kr)Seon-Young Kim (kimsy@kribb.re.kr)Byoung-Mog Kwon (kwonbm@kribb.re.kr)Dong-Cho Han (dchan@kribb.re.kr)Nam-Soon Kim (nskim37@kribb.re.kr)Hee-Gu Lee (hglee@kribb.re.kr)Jae-Wha Kim (wjkim@kribb.re.kr)Eun-Young Song (eysong@kribb.re.kr)Mi-Sun Won (mswon@kribb.re.kr)Kyung-Sook Chung (kschung@kribb.re.kr)

Senior Researcher Kyung-Chan Park (kpark@kribb.re.kr)Cho-Rok Jung (crjung@kribb.re.kr)

Post-DocDong-Chul Lee (dclee@kribb.re.kr)In-Young Park (iypark@kribb.re.kr)Ye-Jin Jang (erythro1@kribb.re.kr)

Su-Jin Chae (sjchae@kribb.re.kr)Mi-Rang Kim (mirang@kribb.re.kr)Han-Chul Lee (nan217@kribb.re.kr)Hyoung-Seok Ju (hsju@kribb.re.kr)Dae-Seop Shin (parady5@kribb.re.kr)Ki-Deok Shin (gdshin@kribb.re.kr)In-Sung Song (microsis@kribb.re.kr)Ho-Beom Kang (hbkang@kribb.re.kr)Heon-Sook Kwon (sook4269@kribb.re.kr)Jong-Tae Kim (jongtae@kribb.re.kr)Bo-Kyeung Kim (kimbk@kribb.re.kr)

ResearcherYoung-Soo Park (mnbz76@kribb.re.kr)Seok-Jin Yang (sukjini@kribb.re.kr)Hyun-Ahm Sohn (imcm1109@kribb.re.kr)Seong-Min Park (lastmhc@kribb.re.kr)Tae-Woo Kim (cavalry9@kribb.re.kr)Byung-Joong Choi (z8t8@kribb.re.kr)Jeong-Hwan Kim (alternar@kribb.re.kr)Tae-Wook Kang (twkang76@kribb.re.kr)Hee-Jin Kim (heejin1@kribb.re.kr)Yeo-Jin Jeon (jyj0242@kribb.re.kr)Sung-Joon Park (gotodr@kribb.re.kr)Jong-Lyul Park (nlcguard@kribb.re.kr)Kee-Ok Haam (totalkee@kribb.re.kr)Oh-Hyung Kwon (ohkwon95@kribb.re.kr)Kwun-Il Kim (ceed@kribb.re.kr)Ji-na Kim (jina@kribb.re.kr)Young-Ju Yoon (yjuru11@kribb.re.kr)Young-Min Han (hanring2@kribb.re.kr)Hong-Mei Lee (lhm0403@kribb.re.kr)Joo-Ae Kim (jooae@kribb.re.kr)Yu-Jin Lee (yujini@kribb.re.kr)Nang-Soo Oh (ds5wdm@kribb.re.kr)Ga-Hee Ha (hghee@kribb.re.kr)Yun-Hee Kang (@kribb.re.kr)Chung-Il Lee (@kribb.re.kr)Ji-Heon Kang (@kribb.re.kr)Yoo-Jin Lee (@kribb.re.kr)Min-A Kang (@kribb.re.kr)Hyo-Ran Yun (nep1015@kribb.re.kr)Na-Young Ji (charry07@kribb.re.kr)Mi-Kyeong Kim (@kribb.re.kr)Eun-Wha Choi (@kribb.re.kr)Jin-Ju Kim(@kribb.re.kr)Hyang-Ran Yun (yhr1205@kribb.re.kr)Hae-Kyeung Hong (hhk0719@kribb.re.kr)Jung-Eun Kang (flykje@kribb.re.kr)Chung-Hae Choi (monday27@kribb.re.kr)Kyung-Jae Won (wkj0423@kribb.re.kr)Seung-Yeob Kim (@kribb.re.kr)Ji-Won Ahn (jiwon@kribb.re.kr)Ji-Hee Seo (@kribb.re.kr)Woo-Jung Lee (@kribb.re.kr)Jung-Hwa Lim (jhwa@kribb.re.kr)Yoon-Jung Choi (ojungsis@kribb.re.kr)Kyeung Su Park (kyeungsu@kribb.re.kr)

Korea Research Institute of Bioscience and Biotechnology 18

B-10Screening of target genes of Yap2

Seo Young Bang1, Byoung Chul Park1, Kwang Hee Bae1,Sung Hyun Kang1, Seung Wook Chi1, Pyung KeunMyung2, Sayeon Cho3, Gwan-Su Yi4, and Sung Goo Park*

1Medical Proteomics Research Center, KRIBB2College of Pharmacy, Chungnam National University3College of Pharmacy, Chung-Ang University4Department of Bio and Brain Engineering, KAIST

Glutathione peroxidase (GPX) is one of the antioxidantenzyme and a ubiquitously expressed isoform that scavengesthe intracellular reactive oxygen species (ROS). In otherto search for the novel interaction partners of Gpx3,immunoprecipitation/two dimensional gel electrophoresis(IP-2DE) after formation of the mixed-disulfide intermediatesof the target proteins with Gpx3. The results revealed thatcadmium resistance protein 1 (CAD1), which also namedAP-1 like factor, Yap2, interacts with Gpx3. To furtherunderstand the role of Yap2, we performed proteomicanalysis to search Yap2-specific targets. For the screeningof Yap2 target genes, we used a strain overexpressingYap2. Under this condition, we found 15 proteins that isinduced by Yap2. To validate these genes as a target geneof Yap2, real-time PCR and binding motif analysis wereperformed. Furthermore, we will carry out quantitativeanalysis with target genes in the presence/absence ofinducers. (The 62th Annual Meeting of the KoreanAssociation of Biological Sciences : Daejeon, 2009)

Keywords : glutathione peroxidase 3, cadmium resistanceprotein 1, target genes

B-11Reduced formation of advanced glycationendproducts via interactions between glutathioneperoxidase 3 and dihydroxyacetone kinase 1

Hana Lee1, Seung Wook Chi1, Phil Young Lee1, SunghyunKang1, Sayeon Cho2, Chong-Kil Lee3, Kwang-Hee Bae1,Byoung Chul Park1, and Sung Goo Park1

1Medical Proteomics Research Center, KRIBB2College of Pharmacy, Chung-Ang University3Department of Pharmacy, Chungbuk National University

Dihydroxyacetone (DHA) induces the formation of advancedglycation endproducts (AGEs), a heterogeneous group ofnon-enzymatic glycation products of proteins, play a criticalrole in protein deterioration during chronic diabeticcomplications, atherosclerosis, hypertension, Alzheimer’sdisease and aging. Previously, we identified dihydroxyacetone

kinase 1 (Dak1) as a candidate glutathione peroxidase 3(Gpx3)-interacting protein in Saccharomyces cerevisiae.This is an unexpected finding, as no apparent evidence hasbeen found supporting the involvement of the oxidativestress system in DHA-induced AGE formation. Here, wedemonstrate that Gpx3 interacts with Dak1, alleviatesDHA-mediated stress by upregulating Dak activity, andconsequently suppresses AGE formation. Based on theseresults, we propose that defense systems against oxidativestress and DHA-induced AGE formation are related viainteractions between Gpx3 and Dak1. (The 21st AnnualMeeting of the Korean Society for Molecular and CellularBiology : Seoul, 2009)

Keywords : advanced glycation endproduct, dihydroxyacetone,dihydroxyacetone kinase 1, glutathione peroxidase3, oxidative stress

B-12The molecular interaction between HAX-1 andXIAP inhibits apoptosis

Young Ji Kang1,2, Mi Jang1, Seung-Wook Chi1, Chong KilLee2, Byoung Chul Park1, Sunghyun Kang1, Sayeon Cho3,Do Hee Lee4, Kwang-Hee Bae1, and Sung Goo Park1

1Medical Proteomics Research Center, KRIBB 2College of Pharmacy, Chungbuk National University3College of Pharmacy, Chung-Ang University4College of Natural Sciences, Seoul Women's University

Apoptosis is a fundamental and complex biological processthat kills and removes unwanted cells during animaldevelopment, normal homeostasis, and diseases. Regulatorsof apoptosis are thought to act in concert, but the molecularinteractions of this process are not well understood.Previously, HS-1 binding protein (HAX-1) was identifiedas a new substrate of caspase-3. HAX-1 is an anti-apoptoticprotein that shares homology to the BH1 and BH2domains from the Bcl-2 family of proteins. Our previousimmunoprecipitation and 2D electrophoresis studiesidentified X-linked inhibitor of apoptosis protein (XIAP)as a new HAX-1 interacting protein. In this study, wedemonstrated the direct interaction between HAX-1 andXIAP at the molecular and cellular level. Also, we showedthat the C-terminal domain of HAX- binds to BIR2 andBIR3 domains of XIAP by GST-pull down assay. Weshowed that HAX-1 inhibits polyubiquitination of XIAPand the interaction between HAX-1 and XIAP increasescell viability in cardiac myoblast H9c2. (The 21st AnnualMeeting of the Korean Society for Molecular and CellularBiology : Seoul, 2009)

Keywords : apoptosis, HAX-1, XIAP, ubiquitination

19 3rd KRIBB Poster Festival

Medical Proteomics Research Center

B-13New opportunities for neutron proteincrystallography

Simranjeet Singh Sekhon1, Elena Magay1, Dae Gwin Jeong1,2,*,and Tae-Sung Yoon1,2,*

1Medical Proteomics Research Center, KRIBB2Bio-Analytical Science, University of Science & Technology(UST)

Neutron crystallography can readily visualize hydrogenatoms in protein crystal structures as compared to X-raywhere these atoms are rarely seen. Neutrons penetratedeeply in biological materials and can easily differentiatebetween hydrogen and deuterium isotopes due to thedifference in magnitude and phases between them.1,2

Neutron beams are unique non-destructive probe andstructures can be determined at room temperature.1,2 Thestructures solved by neutron crystallography provide uniqueinsights into hydrogen-bonding interactions, protonationstates, catalytic mechanisms and hydration states of biologicalstructures that are not available from X-ray analysis alone.3

The enhanced visibility of hydrogen atoms from water,substrates and proteins enables direct determination ofprotonation state, and helps to provide a more completepicture of atomic and electronic structure.3 This is beneficialfor determining enzyme mechanism, for studies of ligand-binding interactions.2,3 The new generation of spallationneutron source enhances the 10-50 fold gain in performance.2

The number of structures deposited yearly at PDB usingneutron crystallographic techniques have increased fromtwo or three in 2005~2007 to seven in 2008. We hope thatsignificant advances in instrumentation and more ‘candidate’protein crystals available from structural genomics/proteomicsprojects will help us to better understand the details ofprotein structure by neutron diffraction method. (Structurefestival “Recent trends in structual biology” : Daejeon,2009)

Keywords : SGM, Sasang constitution, SNP markers

Members

DirectorByoung-Chul Park (parkbc@kribb.re.kr)

Principal Researcher Sang-Chul Lee (esach@kribb.re.kr)Sung-Goo Park (sgpark@kribb.re.kr)Seung-Jun Kim (ksj@kribb.re.kr)Eui-Jeon Woo (ejwoo@kribb.re.kr)Tae-Sung Yoon (yoonts@kribb.re.kr)Kwang-Hee Bae (khbae@kribb.re.kr)Seung-Wook Chi (swchi@kribb.re.kr)

Senior Researcher Sung-Hyun Kang (skang@kribb.re.kr)Dae-Gwin Jeong (dgjeong@kribb.re.kr)Jeong-Hee Moon (jhdae@kribb.re.kr)

Post-DocElena Magay (elena@kribb.re.kr)Jae-Eun Park (jepack@hanmail,net)

ResearcherJin-Sun Choi (sm9603@hanmail.net)Joo-Hyun Ryu (jhryu80@naver.com)Ji-Young Choi (sonatine82@naver.com)Hye-Yeon Shin (hy9781@nate.com)Ha-na Lee (ghetto21@kribb.re.kr)Young-Ji Kang (wow-kyj-peak@hanmail.net)Ki-Beom Heo (kbheo@kribb.re.kr)Min_Beom Heo (hmbgo@nate.com)Seo-Young Bang (vbbangv@kribb.re.kr)Ju- Hui Kim (cjdfnaldks@naver.com)Se-Mi Yun (sei_mee@naver.com)Hye-Yun Jung (hyjung@kribb.re.kr)Sun-Young Kim (khh2809@kribb.re.kr)Won-Kok Kim (wkkim@kribb.re.kr)Eun-Young Kim (issuesky@kribb.re.kr)Hye-Ryung Choi (heyryung779@hanmail.net)Eon-Ryung Kim (ksullove@hanmail.net)Suk Kyung Jeong (jsk0818@kribb.re.kr)Geum-ran Yoo (kiwisoda5@nate.com)Tae-Yang Jeong (sunny@kribb.re.kr)Hyeong-Nam Song (hotdog707@hanmail.net)Arti (artidumbre@gmail.com)Ji-Hyang Ha (hjh82@hanmail.net)Eun-Young Won (matrix302@naver.com)Dong-hwa Lee (ddong1879@nate.com)Ju-Hui Kim (wngml0989@naver.com)Song-Yi Kim (syi86@hanmail.net)Simranjeet Singh Sekhon (simran@kribb.re.kr)

Korea Research Institute of Bioscience and Biotechnology 20

B-14Cytoskeleton-associated proteins are enrichedin human embryonic-stem cell-derivedneuroectodermal spheres

Jung-Il Chae1,3, Janghwan Kim1, Sun-Mi Woo1, Hyo-WonHan1, Young Keun Cho1, Keon-Bong Oh1, Ki-Hoan Nam2,and Yong-Kook Kang1

1Development and Differentiation Research Center, KRIBB2Bio-Evaluation Center, KRIBB3Graduate School of Life Science, CHA Stem Cell Institute,Pochon CHA University

The ability to generate neural lineages from human embryonicstem cells(hESCs) in a controlled manner would furtherinvestigation of human neurogenesis and development.However, generating such neural stem cells(NSCs) remainsa challenge. In an attempt to characterize the cellularmechanisms involved in hESC differentiation intoneuroprogenitor cells, we performed 2-DE using proteinextracts from hESC-derived embryoid bodies(EBs) andneuroectodermal spheres(NESs). Of 47 differentiallyexpressed protein spots, 28 nonredundant spots were shownto be upregulated in the NESs; these protein spots includedneurogenesis-related proteins. Interestingly, 6 of these 28protein spots were cytoskeleton-associated proteins(CAPs).Western-blot analyses confirmed the increased levels ofthese proteins in the NESs. Furthermore, immunostaininganalysis showed that CAPs were expressed in the innerrims of neural rosettes. Our results suggest that, in additionto the induction of those genes involved in neural development,hESC differentiation into the NES is associated with amarked reorganization of the cellular cytoskeleton. (The21st Annual Meeting of the Korean Society for Molecularand Cellular Biology : Seoul, 2009)

Keywords : human embryonic stem cell(hESCs), cytoskeleton-associated proteins(CAPs), neuroectodermalspheres(NESs)

B-15Notch signaling is required for maintainingstem-cell features of neuroprogenitor cellsderived from human embryonic stem cells

Sun-Mi Woo1, Janghwan Kim1, Hyo-Won Han1, Jung-IlChae1,2, Mi-Young Son1, Sunwha Cho1, Hyung-MinChung2, Yong-Mahn Han3, and Yong-Kook Kang1,*

1Development and Differentiation Research Center, KRIBB2Stem Cell Research Laboratory, CHA Stem Cell Institute,Pochon Cha University

3Department of Biological Sciences, KAIST

Studies have provided important findings about the roles

of Notch signaling in neural development. However, mostof these studies have investigated the neural stem cells(NSCs) of mice or other laboratory animals rather thanhumans. It prompted us to focus on neuroectodermal spheres(NESs) which are derived from human embryonic stemcell (hESC) and inhabited by NSCs. We here investigatedthe role of Notch signaling with the hESC-derived NESs.We derived NESs from hESCs. NES formation was confirmedby various NSC marker genes and the emergence of rosettestructures. We found that Notch signaling is active in thesehESC-derived NESs. Expression levels of Notch signalingmolecules were increased in the NESs. Inhibition of theNotch signaling reduced rosette structures, expressionlevels of NSC marker genes and proliferation potential inthe NESs, and, if combined with withdrawal of growthfactors, triggered differentiation toward neurons. Our resultsindicate that the hESC-derived NESs maintain stem cellcharacteristics mainly through Notch signaling, whichsuggests that the hESC-derived NESs could be an in-vitromodel for in-vivo neurogenesis. (The 21st Annual Meetingof the Korean Society for Molecular and Cellular Biology: Seoul, 2009)

Keywords : human embryonic stem cell(hESCs),neuroectodermal spheres(NESs)

B-16Localization of oocyte-specific DNAmethyltaansferase-1 during pocine earlydevelopment

Young Sun Jeong1, Keon Bong Oh2, Jung Sun Park1, Ji-SuKim3, and Yong-Kook Kang1

1Development and Differentiation Research Center, KRIBB2Animal Biotechnology Division, National LivestockResearch Institute, RDA

3National Primate Research Center, KRIBB

DNA methyltransferase-1 (Dnmt1) is involved in themaintenance of genomic methylation patterns. Rather thanfull-length Dnmt1, mouse oocytes have a truncated variantcalled Dnmt1o. Immunofluorescence data showed thatDnmt1o localized to the cytoplasm, but this has not beenconfirmed using more direct methods. The cytoplasmiclocalization of Dnmt1o has been assigned to the main causeof global DNA demethylation in early mouse embryos. Westudied localization of Dnmt1o in mouse and pig embryos.We identified pig Dnmt1o protein and its transcript withunique 5'-end sequence. Physically separating mouse andpig 2-cell embryos into their nuclear and cytoplasmiccomponents demonstrated that Dnmt1o of both specieslocalized to the cytoplasm. Cloned pig embryos had Dnmt1oas the main form, with no indication of somatic Dnmt1.These findings indicate that Dnmt1o is cytoplasmic duringearly development; its presence in both pig and mouse

21 3rd KRIBB Poster Festival

Development and Differentiation Research Center

embryos further suggests that Dnmt1o is conserved inmammals. This work was supported by grant from KRIBBResearch Initiative Program and Biogreen 21(RDA). (The21st Annual Meeting of the Korean Society for Molecularand Cellular Biology : Seoul, 2009)

Keywords : DNA methyltransferase-1, Dnmt1

Members

DirectorYee-Sook Cho (june@kribb.re.kr)

Principal Researcher Yong-Kook Kang (ykkang@kribb.re.kr)

Senior Researcher Jeong-Woong Lee (jwlee@kribb.re.kr)Mi-Young Son (myson@kribb.re.kr)

Post-DocHwa-Jin Jung (junghj@khu.ac.kr)

ResearcherJung-Sun Park (jspark@kribb.re.kr)Min-Joung Kim (dandekmj@kribb.re.kr)Bin-Na Seol (nana8610@kribb.re.kr)Hyun-Jin Kim (invu115@nate.com)Jong-Jin Park (mrpro@kribb.re.kr)Young-Sun Jung (ysjeong@kribb.re.kr)Ji-Hyun Kim (bananadrops@naver.com)Teselkin Oleksiy (teselkin.oleksiy@gmail.com)Lin Tao (ailuomansi@sina.com)Mi-Ra Jang (doobe@kribb.re.kr)Eun-Jeong Jeong (lovelove0314@hanmail.net)Jeong-Hee Hwang (tkfkdgo0629@nate.com)Min-Hwa Do (flower841204@hanmail.net)Sun-Ok Oh (633zzang@kribb.re.kr)

Korea Research Institute of Bioscience and Biotechnology 22

Division of Biosystems Research

Industrial Biotechnology & Bioenergy Research Center

Plant Systems Engineering Research Center

Bioinformatics Research Center

Industrial Bio-materials Research Center

Environmental Biotechnology Research Center

3 r d K R I B B P o s t e r F e s t i v a l

C-1Complete genome sequence of Escherichia coliBL21(DE3)

Haeyoung Jeong1, Ji-Hoon Shim1, F. William Studier2, TaeKwang Oh3, and Jihyun F. Kim1

1Industrial Biotechnology & Bioenergy Research Center,KRIBB

2Biology Department, Brookhaven National Laboratory, USA321C Frontier Microbial Genomics & Applications Center

Escherichia coli BL21(DE3) is a derivative of strain B that wasspecially designed for high-level expression of recombinantproteins using the T7 expression system. It has severaladditional features that are ascribable to B strain-specificcharacteristics such as protease deficiency, little acetateproduction at high glucose levels, simpler cell surfacestructure, etc. We determined the complete genome sequenceof BL21(DE3) through NimbleGen’s comparative genomesequencing, 454 pyrosequencing, and Sanger pair-endedsequencing. BL21(DE3) has a single circular chromosomeof 4,557,508 bp without plasmid, which makes it the smallestgenome among the sequenced E. coli strains. Comparisonbetween BL21(DE3) and K-12 MG1655 reveals that twogenomes are closely related and ~92% of them could bealigned in blocks with >99% sequence identity. Though noinversions or arrangements were observed, interruptionsby mobile genetic elements, deletions and horizontal genetransfer were apparent, which are relevant to variousgenome-specific characteristics. (2009 CSHL Meeting onGenome Informatics : Cold Spring Harbor (USA), 2009)

Keywords : genome sequencing, E. coli B strain, comparativegenomics

C-2Biosynthesis of the peptide antibiotic polymyxinin the surrogate host Bacillus subtilis

Soo-Young Park1, Soo-Keun Choi1, Seong-Bin Kim1,Choong-Hwan Lee2, Jihyun F. Kim1, and Seung-Hwan Park1

1Industrial Biotechnology & Bioenergy Research Center, KRIBB2Division of Bioscience and Biotechnology, Konkuk University

Polymyxin is a cyclic-lipopeptide antibiotic which producedby Paenibacillus polymyxa, synthesized by non-ribosomalpeptide synthetase (NRPS). It has been reintroduced inclinical practice for the treatment of multidrug-resistantGram-negative pathogenic bacteria. Here, we present theinformation of polymyxin synthetase gene cluster identifiedfrom P. polymyxa E681. The gene cluster consists of fiveopen reading frames, designated pmxA, pmxB, pmxC, pmxD,and pmxE. The pmxC and pmxD genes are similar to genes

encoding transport proteins, and pmxA, pmxB, and pmxEencode polymyxin synthetases. The pmxE disrupted mutantstrain could not produce polymyxin completely. Introductionof the pmx gene cluster into the amyE locus of the Bacillussubtilis chromosome resulted in the production of polymyxinin the presence of extracellularly added L-2,4-diaminobutyricacid (DAB) which is a major amino acid in polymyxin,absent in B. subtilis 168. Finally the biosynthetic gene ofDAB (ectB) from P. polymyxa E681 was introduced intosurfactin biosynthetic gene (srfC). The resulting strain couldproduce polymyxin successfully without the addition ofexogenous DAB. (International Symposium & AnnualMeeting of the Korean Society for Microbiology andBiotechnology : Daejeon, 2009)

Keywords : polymyxin, Paenibacillus polymyxa, Bacillussubtilis, surrogate host

C-3Enhancing 1-butanol tolerance in Escherichiacoli through repetitive proton beam irradiation

Haeyoung Jeong and Jihee Han

Industrial Biotechnology & Bioenergy Research Center,KRIBB

Low butanol tolerance of most microorganisms severelylimits the production of 1-butanol at an economical scaleusing alternative hosts other than the natural butanol producerClostridium acetobutylicum, which is not amenable forgenetic manipulation and requires demanding cultureconditions. To generate butanol-tolerant E. coli, we devised acyclic selection strategy that consisted of iterative applicationof proton irradiation at a dose of ~250 Gy using 45 MeVprotons and selection by daily serial transfer in a minimalmedium containing increasing amounts of 1-butanol.Application of five rounds of the cyclic selection of E. coliATCC 8739 (C strain) over 61 days resulted in a mutantpopulation that could tolerate 1.2% 1-butanol (v/v).However, without proton irradiation, the cells were unableto grow at 0.8% 1-butanol in a control experiment.Seven different mutations were identified in one clonefrom the evolved population through 454 pyrosequencingof the genome. Tracing each mutation in terms of theprevalence in the population during the period of evolutionsuggested that proton beam irradiation-induced mutationswere rapidly fixed during the early phase of the selectionprocedure. This approach, which is still being applied toincrease butanol tolerance beyond 1.3%, is considered tobe useful for improving traits whose corresponding genesare not known. (13th International Conference onAccelerator & Beam Utilization : Gyeongju, 2009)

Keywords : 1-butanol, proton beam, irradiation, E. coli

25 3rd KRIBB Poster Festival

Industrial Biotechnology & Bioenergy Research Center

C-4Autoinducible expression system in Bacillussubtilis

Su-Jin Lee, Jae-Gu Pan, Seung-Hwan Park, and Soo-KeunChoi

Industrial Biotechnology & Bioenergy Research Center,KRIBB

The industrial production of heterologous proteins in Bacillussubtilis was not much successful to date because of manypitfalls including the lack of strong promoter, instability ofrecombinant proteins by extracellular proteases, and autotoxiceffect on the constitute expression of the foreign geneexpression. To overcome such problems, we developed thenovel Bacillus expression systems using strong andautoinducible phoB and cry promoters. The phoB promoterthat is completely repressed in the vegetative growth phase issharply induced when the bacteria face phosphate starvationcondition. The other inducible promoter, crystal protein gene(cry) promoter from Bacillus thuringiensis express adownstream gene only in the stationary phase. To improvethe two autoinducible promoters, the modification of thecry promoter sequences to the consensus sequence of sA-dependent promoter of B. subtilis and the directed evolutionof phoB promoter were carried out. The improved phoB andcry promoters significantly expressed downstream genes,suggesting that the systems can be useful to produce targetprotein in B. subtilis. (International Symposium & AnnualMeeting of the Korean Society for Microbiology andBiotechnology : Daejeon, 2009)

Keywords : Bacillus subtilis, heterologous expression,autoinduction

C-5Comparative genome sequence analysis ofBifidobacterium bifidum BGN4

Dong Su Yu1, Haeyoung Jeong1, Myeong-Soo Park2, GeunEog Ji3, Tae Kwang Oh1,4, and Jihyun F. Kim1

1Industrial Biotechnology and Bioenergy Research Center,KRIBB

2Anyang Technical College3Department of Food and Nutrition, Research Institute ofHuman Ecology, Seoul National University

421C Frontier Microbial Genomics and Applications Center

Bifidobacteria are high-G+C gram-positive anaerobes thrivingin the intestine of infants. As a probiotic stimulating gastro-intestinal health, Bifidobacterium bifidum BGN4 was isolatedas a ß-glucosidase-negative strain that does not releasecarcinogenic or mutagenic aglycons from some dietary

glycosides. Other features of B. bifidum BGN4 include aprominent adhesion activity to intestinal epithelial cells andthe production of polysaccharides (BB-pol) that can inhibitthe growth of a colon cancer cell line. The genome sequenceof BGN4 was determined by 454 pyrosequencing at ca. 25Xcoverage and capillary sequencing for gap closing. From thesequence of 2,223,664 Mbps and 62.65% G+C, we predicted1,835 coding sequences (CDSs) using CRITICA, GLIMMERand AutoFACT. Through comparative genome analysis ofBGN4 with those of four Bifidobacterium species in GenBank,1,064 common CDSs and 363 CDSs unique to BGN4 wereidentified. Through sequence analysis, we demonstrated thepresence of the bifid shunt pathway and metabolic pathwayof chiro-inositol that is the key ingredient of BB-pol. [Financialsupport from the 21C Frontier Microbial Genomics andApplication Center Program, MEST, Korea] (MSK's 50thAnniversary International Symposium on Microbiology: Jeju, 2009)

Keywords : Bifidobacterium bifidum BGN4, comparativegenome analysis, probiotic

C-6Genome analysis of the secondary metabolismin Streptomyces clavuligerus ATCC 27064

Ju Yeon Song1, Haeyeong Jeong1, Sang-Haeng Choi1,Hong-Seog Park1, Jae Jong Kim3, Tae Kwang Oh1, KyeJoon Lee2, and Jihyun F. Kim1

1Industrial Biotechnology and Bioenergy Research Center,KRIBB

2School of Biological Sciences, Seoul National University3GenoTech Co., Ltd.

Streptomyces clavuligerus ATCC 27064 is a high G+C,Gram-positive soil bacterium that produces antibiotics suchas clavulanic acid, cephamycin C and clavams. To explorethe biosynthesis of secondary metabolites in S. clavuligerusand to identify novel bioactive compounds for the developmentof strains better producing these molecules, a whole-genome-based approach was applied. We employed a Sanger/pyrosequencing hybrid approach using the end reads ofplasmid/fosmid libraries produced from ABI 3730xl andshotgun reads generated from GS systems. All reads ofapprox. 82xgenome coverage were assembled into 330contigs by PCAP assembler and the genome size wasestimated to be ca. 9.2 Mb as calculated from the total contiglength. More than 30 gene clusters for the secondarymetabolite biosynthesis of non-ribosomal peptides,polyketides, siderophores, terpenes and pigments wereidentified by homology searches of 8,103 protein-codinggenes predicted by Glimmer. This work was supported bythe 21C Frontier Microbial Genomics and ApplicationCenter Program, MEST, Republic of Korea. (International

Korea Research Institute of Bioscience and Biotechnology 26

Symposium & Annual Meeting of the Korean Societyfor Microbiology and Biotechnology : Daejeon, 2009)

Keywords : Streptomyces clavuligerus, genome analysis,secondary metabolites

C-7Functional analysis of the Type II secretionsystems in Escherichia coli BL21(DE3)

Ji Hoon Shim, Seong Keun Kim, Sung Ho Yoon, ChoongHoon Lee, Min Jung Kwak, and Jihyun F. Kim

Industrial Biotechnology & Bioenergy Research Center,KRIBB

Gram negative bacteria have inner and outer membranesto compartmentalize the cell from environment. To secreteproteins, bacteria employ several secretary machinery topass through the membrane barrier. Especially, the Type IIsecretion system (T2SS) known as the general secretorypathway exports periplasmic proteins into the extracellularmilieu. This system also has important roles in pathogenesisand outer membrane biogenesis. From the analysis of twoE. coli B strains, REL606 and BL21(DE3), we found thatthe B genomes have an additional gene cluster for T2SS ascompared to K-12 strains. Phylogenetic analysis showedthat clear distinctions could be made between the homologsof the T2SS conserved in both B and K-12 and those ofthe additional T2SS in B strains. To find out the roles ofthe two T2SSs, the gene clusters were deleted and we foundthat the complete loss of the two T2SSs in BL21(DE3)results in pleiotropic effects including the growth defect inrich media, increased quantities of extracellular proteins.[Supported by grants from the 21C Frontier MicrobialGenomics and Applications Center Program, MEST,Republic of Korea] (International Symposium & AnnualMeeting of the Korean Society for Microbiology andBiotechnology : Daejeon, 2009)

Keywords : BL21(DE3), Type II secretion system.

C-8The crystal structure of the human NDRG2(N-Myc downstream-regulated gene 2) protein,a potential tumor suppressor

Yoonjeong Kim, Jungwon Hwang, Hwiseop Lee, Won-Chan Choi, Tae-Kwang Oh, and Myung Hee Kim

Industrial Biotechnology & Bioenergy Research Center,KRIBB

Accumulated data have implied that the members of thehuman N-Myc downstream-regulated gene (NDRG)family, namely NDRG1-4, are tumor suppressors, and arealso involved in the development of other diseases.Primary amino acid sequence in formation suggests thatthe NDRG family proteins belong to an alpha /betahydrolase fold superfamily; however, detailed informationrelated to its function from the sequence has not beenobtained. Here, we present the crystal structure of theNDRG2 protein determined at 2.0Å resolution. NDRG2shows remarkable structural similarity to the alpha/ betahydrolase family of proteins, despite the limited sequencesimilarity. Functional assignments reveal that NDRG2 is anon-enzyme homolog of the alpha/ beta hydrolase familyof proteins, which lacks the catalytic signature residuesand has a disrupted substrate-binding site. Such structuralaspects are conserved in the NDRG family. This work wassupported by funding from the 21C Frontier MicrobialGenomics and Applications Center Program, MEST,Republic of Korea. (Structure festival "Recent trends instructual biology" : Daejeon, 2009)

Keywords : NDRG2, tumer suppression, Myc, alpha/betahydrolase

C-9Genetic encoding Escherichia coli to monitorcatalytic activity In Vivo

Eugene Rha1, Su-Lim Choi1, Jae Jun Song2, and Seung-Goo Lee1

1Industrial Biotechnology & Bioenergy Research Center,KRIBB

2Molecular Bioprocess Research Center, KRIBB

The underlying goal of synthetic biology is to make theprocess of engineering biological systems easier. Thisstudy is about a genetic enzyme screening system (GESS)for a hyper sensitive detection of target enzyme usingbiologically redesigned genetic circuitry. The redesignedgenetic circuitry is composed of three major parts, (i) thegene encoding transcriptional regulating factor (ii) theregulatory part for the expression of the circuitry, and (iii)the reporter proteins. The redesigned genetic circuitry wasnot detectable only phenol but also very broad rangephenolic compounds like as o-nitrophenol, 2-aminophenol,2-methoxyphenol, catechol, and so on. Here, we confirmedthat the activities of ß-galactosidase, tyrosine-phenol lyase,phenyl phosphatase, and methyl parathion hydrolase couldbe analyzed by the GESS. The present system enables thedetection of target enzyme from various gene sourcesincluding genomic and metagenomic libraries as well as asingle cloned gene. Furthermore, this method may enablethroughput quantification of enzyme activity, which is

27 3rd KRIBB Poster Festival

usefully applicable to protein engineering (directedevolution) of target enzyme. This project was supported byBio R&D Programs of the Korea Science and EngineeringFoundation (KOSEF). (International Symposium &Annual Meeting of the Korean Society for Microbiologyand Biotechnology : Daejeon, 2009)

Keywords : Synthetic biology, genetic circuitry, highthroughput screening, enzyme engineering

C-10Novel detection mode of multiple sugars usingconformational relaxation of FRET sensors athigh temperature

Jae-Seok Ha, Yu Jung Kim, Su Jin Kim, and Seung-Goo Lee

Industrial Biotechnology and Bioenergy Research Center,KRIBB

Monitoring sugar concentration during the saccharificationor alcohol fermentation process is becoming more importantfor the production of cellulosic ethanol. As a reliable andrapid method for quantifying multiple sugars, we developedfluorescence-based sugar sensors which constitute 3 domainfusion proteins in the order of ECFP-periplasmic-bindingprotein-EYFP. Various cellulosic sugars could be determinedquantitatively by reading the FRET levels. In this study, wedeveloped a novel detection mode of various sugars basedon conformational relaxation of periplasmic-binding proteinmoiety at critical high temperature, expanding the dynamicrange and allow quantification of individual sugars undermixed carbohydrate conditions. The sugar sensors withimproved signal intensity were based on the stabilizationof conformationally relaxed PBPs in the presence of lignadsat high temperature, thereby increasing the sugar-dependentFRET signals from 4 to 15 folds under optimal conditions.This study provides more sensitive sugar detection methodsusing fluorescence-based sugar sensors, thus facilitatingrapid and accurate detection of fermentable sugars.(International Symposium & Annual Meeting of theKorean Society for Microbiology and Biotechnology :Daejeon, 2009)

Keywords : FRET sensors, conformational relaxation,sugar detection

C-11One-step cloning and simultaneous expressionin Escherichia coli mediated by homologousrecombination

Jeongmin Lee1, Sun-Hwa Kim1, Eugene Rha1, Jae-SeokHa1, Eui-Sung Choi1, Jae Jun Song2, and Seung-Goo Lee1

1Industrial Biotechnology and Bioenergy Research Center,KRIBB

2Molecular Bioprocess Research Center, KRIBB

A new genetic tools based on homologous recombinationin Escherichia coli was developed. In this method, we usea special recipient plasmid (pRMT) consisting of an inactiveselection marker missing ribosome binding site (RBS) andthe start codon of selection marker gene. Whena specificform of amplified insert (iTGR) was introduced into thecells, the inactive reporter on the pRMT can switch to theactive one via the reconstitution of RBS and start codon,which are needed for the translation of the reporter gene.To demonstrate this concept, we tried to clone GFPuv(0.7kb), MBP-EGFP fusion protein (2kb), and carotene-biosynthesis operon (4kb), an astaxanthin-biosynthesisgene cluster (6kb) and the 6kb plus mevalonate pathwaygenes (10kb). As a result, the target DNAs could be clonedefficiently and the expression of each pathway genes wasshown by the development of characteristic colors. Thenon-homogeneity problem of recovered plasmids from theE. coli clones could be overcome by introducing a suicideselection maker (sacB) in the pRMT. The selection on 7%sucrose media resulted in mostly homogeneous clones,based on the analyses of recovered plasmids. Consequently,this recombination cloning method could be applied formassive parallel gene expression, and also for the synthesisof new metabolic pathway. (International Symposium &Annual Meeting of the Korean Society for Microbiologyand Biotechnology : Daejeon, 2009)

Keywords : homologous recombination, red-recombinase,synthetic biology, in vivo cloning

C-12PESTAS: A web server for EST analysis andsequence mining

Seong-Hyeuk Nam1,2, Dae-Won Kim1,2, Tae-Sung Jung1,Young-Sang Choi3, Dong-Wook Kim3, Han-Suk Choi3,Sang-Haeng Choi1, and Hong-Seog Park1,2

1Industrial Biotechnology & Bioenergy Research Center,KRIBB

2University of Science and Technology(UST)3Mokpo National University

We have developed a web server for the high-throughputannotation of expressed sequence tags (ESTs) calledpipeline for EST analysis service (PESTAS). PESTASprocesses entire datasets with an automated pipeline of 13analytic services, then deposits the data into the MySQL

Korea Research Institute of Bioscience and Biotechnology 28

database and transforms it into three kinds of reports:preprocessing, assembling and annotation. All annotatedinformation is provided to the scientist and can bedownloaded through a web browser. To get more relevantfunctional annotation results, a curation function wasintroduced with which biologists can easily change thebest-hit annotation information. We included a gene chipmodule that detects gene expression differences betweenlibraries by comparing accession number counts fromBLAST search results. PESTAS also provides access tothe pathway information of KEGG, which is useful formapping the relationships among networks of annotatedenzymes, and is especially valuable for those researchersinterested in biological pathways. (The 64th AnnualMeeting of the Korean Association of Biological Sciences :Daejeon, 2009)

Keywords : EST annotation, web services

C-13GarlicESTdb: an online database and miningtool for garlic EST

Dae-Won Kim1,2, Tae-Sung Jung1, Seong-Hyeuk Nam1,2,Hyuk-Ryul Kwon1, Aeri Kim1,2, Sung-Hwa Chae3, Sang-Haeng Choi1, Dong-Wook Kim4, Ryong Nam Kim1,2, andHong-Seog Park1,2

1Industrial Biotechnology & Bioenergy Research Center,KRIBB

2University of Science and Technology(UST)3Gnc Bio Co Ltd, Daejeon 4Mokpo National University

GarlicESTdb is an integrated database and mining tool forlarge-scale garlic (Allium sativum) EST sequencing. A totalof 21,595 ESTs collected from an in-house cDNA librarywere used to construct the database. The analysis pipelineis an automated system written in JAVA and consists ofthe following components: automatic preprocessing of ESTreads, assembly of raw sequences, annotation of theassembled sequences, storage of the analyzed informationinto MySQL databases, and graphic display of all processeddata. A web application was implemented with the latestJ2EE (Java 2 Platform Enterprise Edition) softwaretechnology (JSP/EJB/JavaServlet) for browsing andquerying the database, for creation of dynamic web pageson the client side, and for mapping annotated enzymes toKEGG pathways, the AJAX framework was also usedpartially. The online resources, such as putative annotation,single nucleotide polymorphisms (SNP) and tandem repeatdata sets, can be searched by text, explored on the website,searched using BLAST, and downloaded. To archive moresignificant BLAST results, a curation system was introducedwith which biologists can easily edit best-hit annotation

information for others to view. (The 64th Annual Meetingof the Korean Association of Biological Sciences : Daejeon,2009)

Keywords : Garlic EST, EST annotation

C-14EKIS: An integrated database of EST informationfor research application

Dae-Won Kim1,2, Tae-Sung Jung1, Young-Sang Choi3,Seong-Hyeuk Nam1,2, Hyuk-Ryul Kwon1, Dong-WookKim3, Han-Suk Choi3, Sang-Heang Choi1, and Hong-SeogPark1,2

1Industrial Biotechnology & Bioenergy Research Center,KRIBB

2University of Science and Technology(UST)3Mokpo National University

The EST Knowledge Integrated System, EKIS (http://ekis.kribb.re.kr), was established as a part of Korea‘sMinistry of Education, Science and Technology initiativefor genome sequencing and application research of thebiological model organisms (GEAR) project. The goals ofthe EKIS are to collect EST information from GEAR projectsand make an integrated database to provide transcriptomicand metabolomic information for biological scientists. TheEKIS constitutes five independent categories and severalretrieval systems in each category for incorporating massiveEST data from high-throughput sequencing of 65 differentspecies. Through the EKIS database, scientists can freelyaccess information including BLAST functional annotationas well as Genechip and pathway information for KEGG.By integrating complex data into a framework of existingEST knowledge information, the EKIS provides newinsights into specialized metabolic pathway informationfor an applied industrial material. (The 64th AnnualMeeting of the Korean Association of Biological Sciences :Daejeon, 2009)

Keywords : EST knowledgebase, EST annotation

C-15Evolutionary characterization of a highly repetitivesequence identified from the false killer whale(Pseudorca crassidens)

Dae-Won Kim1, Aram Kang1,2, Sang-Haeng Choi1, ZangGeun Kim3, Woo-Jin Kim4, Hyung-Cheol Kim1, andHong-Seog Park1,2

1Industrial Biotechnology & Bioenergy Research Center,

29 3rd KRIBB Poster Festival

KRIBB2University of Science and Technology(UST)3Gnc Bio Co Ltd, Daejeon 4Biotechnology Research Institute, National FisheriesResearch and Development Institute

We report here that a novel 1,869 bp repetitive sequenceidentified from the false killer whale (Pseudorca crassidens)could be a new molecular phylogenetic marker in cetaceans.Results of PCR amplification and southern blot hybridizationusing 16 species’ genomic DNAs from five different familiesrevealed that the repetitive sequence is highly conservedwithin all Delphinidae species. Notably, specific primersdesigned for this repetitive sequence effectively amplifiedthe targeted repetitive units, which were critically dependentupon the genetic phylogenies in the members of the Delphinidaecetaceans. Therefore, the novel sequences can be used as auseful phylogenetic marker for understanding the molecularevolutional studies in members of the Delphinidae familyof cetaceans. (The 64th Annual Meeting of the KoreanAssociation of Biological Sciences : Daejeon, 2009)

Keywords : Pseudorca crassidens, repetitive sequence,phylogenetic analysis

C-16Comparative analysis of expressed sequencetags from the white-rot fungi (Phanerochaetechrysosporium)

Dae-Won Kim1, Aeri Kim1,2, Ryong Nam Kim1, Seong-Hyeuk Nam1,2, Aram Kang1,2, Wan-Tae Chung3, Sang-Haeng Choi1, and Hong-Seog Park1,2

1Industrial Biotechnology & Bioenergy Research Center,KRIBB

2University of Science and Technology(UST)3Technology Services Division, National Institute ofAnimal Science, Rural Development Administration

Comprehensive analysis of the transcriptome of the P.chrysosporium is a useful approach to improve ourunderstanding of its special and unique enzyme systemand fungal evolution in molecular and industrial aspects.In order to unveil the functional diversity of this white-rotfungus in gene level and the expression patterns of its genes,in this study we carried out sequencing and annotation of4,917 P. chrysosporium expressed sequence tags (ESTs).Through our bioinformatic ESTs analysis, we elucidatedthat 1,751 genes were derived from the present dataset of4,917 ESTs, based on clustering and comparative genomicanalyses of the ESTs. Of the 1,751 unique ESTs, 1,006(57.5%) had homologues and orthologues in similaritysearches. Our P. chrysosporium ESTs showed many genesfor encoding 23 secreted proteins, many proteins for the

degradation of cellulose and hemicelluloses, and heat shockproteins for stress resistance, which explain the reason whyP. chrysosporium is very important and unique white-rotfungus in dealing with contaminated resources and indegrading lignin and in applying this organism to severalindustrial aspects. In addition, comparative analysis hasshed the fresh light on the mystery about how its uniqueenzyme system and stress resistance have been evolveddifferently from its closest relatives. (The 64th AnnualMeeting of the Korean Association of Biological Sciences :Daejeon, 2009)

Keywords : white-rot fungi EST, transcriptome analysis,bioinformatics

C-17Airborne induction and priming of plantdefenses against a bacterial pathogen

Hwe-Su Yi1,2, Martin Heil3, Rosa M. Adame-Alvarez3,Daniel J. Ballhorn4, and Choong-Min Ryu1,5

1Industrial Biotechnology & Bioenergy Research Center,KRIBB

2School of Life Science, Kyungpook National University3Departamento de Ingenieria Genetica, CINVESTAV,Mexico

4Department of General Botany-Plant Ecology, Universityof Duisburg-Essen, Germany

5Field of Functional Genomics, School of Science, Universityof Science and Technology (UST)

Herbivore-induced plant volatiles affect the systemic responseof plants to local damage and hence represent a kind of planthormone. These signals are transported outside the plants andcan, therewith, also lead to ‘plant-plant communication’, adefense induction in yet undamaged plants that growcloseto damaged neighbors. We observed a similar phenomenonin the context of disease resistance. Lima bean (Phaseoluslunatus) plants became more resistant against a bacterialpathogen, Pseudomonas syringae pv. syringae, when theywere located close to conspecific neighbors in which systemicacquired resistance (SAR) to pathogens had been chemicallyinduced with benzothiadiazole (BTH) or biologically byinfection with avirulent P. syringae. Challenge inoculationafter exposure to induced and non-induced plants revealedthat the air coming from induced plants mainly primedresistance, since expression of pathogenesis-related protein2 (PR-2) was significantly stronger in exposed than in non-exposed individuals when the plants were subsequentlychallenged by P. syringae. Among others, the plant-derivedvolatile, nonanal, was present in the headspace of BTH-treated plants and significantly enhanced PR-2 expression inthe exposed plants. Negative effects on growth of BTH-treated plants, which usually occur as a consequence of high

Korea Research Institute of Bioscience and Biotechnology 30

costs of direct resistance induction, were not observed in VOC-exposed plants. Volatile mediated priming appears to be ahighly attractive means for the tailoring of SAR against plantpathogens. (The 2009 KSPP Fall Meeting and the 1stJapan-Korea Joint Symposium : Jeju, 2009)

Keywords : Lima bean, VOCs, BTH, Nonanal

C-18Induced systemic resistance elicited by bacterialgenetic materials

Boyoung Lee1,2, Soohyun Lee2, and Choong-Min Ryu1,2

1Field of Functional Genomics, University of Science andTechnology (UST)

2Industrial Biochemistry & Bioenergy Research Center,KRIBB

Plant need to protect themselves against multiple pathogenson the infected site as well as at the distal part. Recent reportpresented that plant percept general cues from microbesreferred to as microbe-associated molecular patterns(MAMPs). The plant innate immunity elicited by recognitionof MAMPs such as fllagellin and elongation factor andstructural materials has intensively studied. Here, we providean evidence that bacterial genetic materials can be a MAMPsto increase plant defense responses against a bacterialpathogen. Infiltration of total RNA from a rhizobacteriumPaenibacillus polymyxa and a pathogenic bacteriaumPseudomonas syringae pv. tomato resulted in enhancingresistant to subsequent infection by P. syringae pv. tomatocompared to water control on Arabidopsis thaliana. 50KArabidopsis microarray was employed to analyze genomewide transcriptional level of Arabidopsis plants followingtreatment of bacterial RNA. The transcriptional expressionof plant defense related genes and its transcription factorwas up-regulated. Our results indicate that bacterialgenetic material can be a potential MAMP and bacterialdeterminant for induced resistance. (8th InternationalPGPR Workshop : Portland (USA), 2009)

Keywords : MAMP, ISR, Pseudomonas syrinage pv.tomato DC3000, bacterial RNA

C-19Metabolic engineering for squalene productionin Saccharomyces cerevisiae

Hee-Kyoung Ryu, Mi-Ran Jo, Jake Whang, Seon-Won Kim,and Eui-Sung Choi

Industrial Biotechnology & Bioenergy Research Center, KRIBB

Squalene is a triterpenoic hydrocarbon (C30H50). Dietarysqualene is able to stimulate nonspecific immune functionsand reduce the levels of low-density lipoprotein as well astriglyceride. The shark liver oil, the current major naturalresource, is limiting and its future supply is uncertain. In thiswork, Saccharomyces cerevisiae, some strains of whichareknown to accumulate a large amount of ergosterol, wasmetabolically engineered to overproduce squalene. Ineukaryotes, isoprenoids are synthesized from acetyl CoAthrough mevalonate. The catalytic domain of the gene encodinghydroxymethylglutaryl CoA reductase (HMGR), a keyregulatory step in mevalonate pathway, was over-expressedunder the GAL10 promoter in S. cerevisiae 2805 strain. Over-expression of HMGR resulted in the intracellular accumulationof squalene (0.49 g/L) in a shake flask experiment. Othergenes involved in isoprenoid synthesis such as ispA andErg20(bifunctional synthase for geranyl diphosphate andfarnesyl diphosphate), GPP (geranyl diphosphate) synthase orMVAE (acetyl-CoA acetyltransferase) were also overexpressedin S. cerevisiae 2805 strain. Over-expression of these genesalone did not show any improvements in squaleneaccumulation. Coexpression of ispA or Erg20 with HMGR,however, dramatically increased the level of squalene (upto 1.8 g/L). The level of squalene accumulation variedamong different strains of S. cerevisiae. (Fall Meeting &International Symposium of the Korean Society forBiotechnology and Bioengineering : Daejeon, 2009)

Keywords : squalene, Saccharomyces cerevisiae,hydroxymethylglutaryl CoA reductase,farnesyl pyrophosphate synthase

Members

DirectorJihyun F. Kim (jfk@kribb.re.kr)

Principal Researcher Jae-Goo Pan (jgpan@kribb.re.kr)Eui-Sung Choi (choi4162@kribb.re.kr) Seung-Hwan Park (shpark@kribb.re.kr)Hong-Seog Park (hspark@kribb.re.kr)Seung-Goo Lee (hspark@kribb.re.kr)Jung Hoon Sohn (sohn4090@kribb.re.kr) Choong-Min Ryu (cmryu@kribb.re.kr)Jung-Hoon Yoon (jhyoon@kribb.re.kr)

Senior Researcher Sang-Haeng Choi (shchoi@kribb.re.kr)Soo-Keun Choi (sookeun@kribb.re.kr)Hae-Young Jeong (hyjeong@kribb.re.kr)Sung-Ho Yoon (moncher@kribb.re.kr)

Post-DocSeong-Bin Kim (sbkim17@kribb.re.kr)Eugene-Rha (josephin@kribb.re.kr)Soo-Young Park (psy4114@kribb.re.kr)

31 3rd KRIBB Poster Festival

Ju-Yeon Song (songjy@kribb.re.kr)Jae-Seok Ha (hayashi@kribb.re.kr)Dae-Won Kim (todaewon@kribb.re.kr)Ryong-Nam Kim (jkrn1968@kribb.re.kr)

ResearcherDong-Su Yu (axxa76@kribb.re.kr)Ji-Hee Han (jiheehan@kribb.re.kr)Su-Jin Lee (sjlee@kribb.re.kr)Seong-Keun Kim (draman97@kribb.re.kr)Kun-Hyang Park (kun-hyang@hanmail.net)Min-Young Kim (jouner@kribb.re.kr)Sun-Hong Kim (bb0070@nate.com)Jong-Hoon Lee (genesisseven@nate.com)Choon-Hoon Lee (chlee@kribb.re.kr)Jung-Wom Hwang (jwhwang@kribb.re.kr)Sun-Hwa Kim (coco78@kribb.re.kr)Min-Jung Kwak (slayersrina@naver.com)Hwe-Su Yi (yihwesu@kribb.re.kr)Bo-Young Lee (boyoung@kribb.re.kr)Hyo-Bee Park (dolly0804@naver.com)Jung-Wook Yang (yjw787@kribb.re.kr)Soo-Hyun Lee (ysh76@kribb.re.kr)Seong-Hyeuk Nam (odysseus@kribb.re.kr)Ae-Ri Kim (ksang45@nate.com)Aram Kim (rami@ust.ac.kr)Dong-Wook Kim (sanctus@nate.com)

C-20Silencing of the catalytic subunit of ferredoxin:thioredoxin reductase induces pathogenesis-relatedgenes and pathogen resistance in tomato

Chan Ju Lim, Woong Bum Kim, Bok Sim Lee, and Suk-Yoon Kwon*

Plant Systems Engineering Research Center, KRIBB

Ferredoxin:thioredoxin reductase (FTR) catalyses the light-dependant activation of several photosynthetic enzymes. Asa heterodimer protein, FTR is composed of variable subunitand catalytic subunit. Among these, catalytic subunitcontains a redox-active disulfide and a [4Fe-4S] center asactive site. However, detailed physiological functions ofcatalytic subunit are still unclear. To elucidate these, weisolated catalytic subunit gene of FTR, designated SlFTR-c, from tomato (Solanum lycopersicum L.) by using RT-PCR. The tanscripts of SlFTR-c were detected all testedtissues such as root, leaf, flower, fruit, and seed. Under theseveral stressed and phytohormone treated condition, geneexpression levels were not affected, but ethylene treatmentscould change the expression of SlFTR-c. Interestingly,Virus-Induced Gene Silencing (VIGS) of SlFTR-c producednecrotic lesions with typical cell death symptoms in tomatoleaves. Moreover, these silenced plants of SlFTR-c resultedin enhanced disease resistance against bacterial phathogens(Pseudomonas syringae pv. tomato DC3000) by inductionof several defense-related genes (SlPR-1, SlPR-2, SlPR-5,SlGlucA, SlChi3, and SlChi9). These results indicate thatSlFTR-c play as a regulator of PCD and pathogen resistancein tomato. (2009 Crop Functional Genomics Workshop: Gangneung, 2009)

Keywords : VIGS, tomato, SlFTR-c, pathogen resistance

C-21CaPIR1 is a RING-finger E3 ligase protein inpepper that negatively regulates plant basalresistance

Sora Choi and Jeong Mee Park*

Plant Systems Engineering Research Center, KRIBB

The RING finger family of protein is known to possessubiquitin ligase activity and play pivotal role in proteindegradation. Here, we show that the CaPIR1 gene, whichencodes a pepper RING-finger E3 ligase, is involved ininnate immunity. Expression of CaPIR1 was specificallyinduced during the hypersensitive response (HR) inresponse to both non-host and host pathogens andtreatment of plant hormones, methyl jasmonate andethylene. Plants expressing the GUS under the control of

Korea Research Institute of Bioscience and Biotechnology 32

Plant Systems Engineering Research Center

CaPIR1 promoter confirmed specfic induction of GUSexpression in leaf mesophyll cells and vascular tissueunder pathogen infection. We studied the function ofCaPIR1 gene in hypersensitive response (HR) and basalresistance using an Agrobacterim-mediated transientexpression system in Nicotiana benthamiana leaves.Overexpression of CaPIR1 led to a significantly enhancedsusceptibility to Pseudomonase singae pv. tabaci. and alsomarkerd promotion of HR cell death induced by variousR/avr interactions and Bcl2-associated X (Bax) protein.We further demonstrate that expression levels of a numberof defense-related genes are reduced and delayed inCaPIR1 expressed leaves during pathogen infection. Theseresults suggest that a pepper CaPIR may have a role as anegative regulator in plant innate immunity. (XIVInternational Cogress on Molecular Plant-MicrobeInteractions : Quebec (Canada), 2009)

Keywords : E3 ligase, pepper, plant basal resistance

C-22Screening of tissue specific genes and functionalanalysis in tomato

Bok-Sim Lee1, Chan ju Lim1, Woong Bom Kim1, Ha-YounLee1, youn-il Park2, and Suk-Yoon Kwon1,*

1Plant Systems Engineering Research Center, KRIBB2Chungnam National University

Tomato (Solanum lycopersicum) is an important fruit cropin the world. To develop transgenic Solanaceae (includingtomato) plants with special purposes, tissue specificpromoters are required because its specificity limitingseveral negative effects on growth and developments intransgenic plants. Therefore, we were trying to developtissue specific promoters in tomato. Through statisticalmethod by comparing ESTs from several libraries oftomato, hot pepper, and Arabidopsis, 650 ESTs werescreened as putative tissue specific genes. To determinetheir specificities in each tissue, quantitative RT-RCRanalysis were performed. Among them, we focused onSlPOD, SlWRKY, SlSDR-1, and SlSDR-2 to investes te thedetail expression profiles, and generated transgenicArabidopsis expressing GUS reporter gene under the controlof each promoter of selected genes. Histochemical assayof these transgenic plants revealed that the expression ofPSlPOD::GUS and PSlWRKY::GUS were specific in Arabidopsisroot. Furthermore, the expression of PSlSDR-1::GUS andPSlSDR-2::GUS were detected seed in silique and flower,respectively, but small expressions in other tissues werealso detected. Virus induced gene silencing (VIGS) intomato plant was also performed using these selected fourgenes to investigate function of each gene. Interestingly,silenced plants of SlSDR-1 contained alsised seed number

comparing to control plants. However, it was hard to findany differences comparing to control plants, though genesilencing was confirmed in each silenced plant. Since oneof the major challenges in plant genetic engineering is todesign a transformation-cassette that would enable theprecise control of transgene activity by choosing adequatepromoters and genes, these characterized tissue specificpromoters and genes may be available to use in plantbiotechnological applications.

Seoul, 2009

Keywords : tissue specific genes, VIGS, tomato, SlSDR-1

C-23Functional RNAi suppressor for Chlamydomoansreinhardtii

Chun ji Yin, Yaw Joo Kim, and Won Joong Jeong*

Plant Systems Engineering Research Center, KRIBB

Codon-optimized RNAi suppressor (CrCMV2b andCrCMV2b2) genes, synthesized for expression inChlamydomonas reinhardtii with high GC content incodong region, was active for suppressing gene silencingin Nicotiana benthamiana and C. reinhardtii. Strongsuppressor activity of CrCMV2b against gene silencingmachinery was confirmed with transient expression in N.benthamiana using agroinfiltration experiment. Theaccumulation of miRNAs, siRNAs, and their putativetarget mRNA were altered in C. reinhardtii transformedwith CrCMV2b gene. These results suggest that codonoptimized RNAi suppressor interferes with RNAi pathwayin C. reinhardtii as in higher plants.

Seoul, 2009

Keywords : Chlamydomonas, RNAi suppressor, genesilencing

C-24Classification of rice (Oryza sativaL. japonicaNipponbare) immunophilins (FKBPs, CYPs)and expression patterns under water stress

Jun Cheul Ahn2, Dae-Won Kim1, Young Nim You1, MinSook Suk1, Hyun Sik Hwang3, Beom Gi Kim3, Hong SukPark1,*, and Hye Sun Cho1,*

1Plant Systems Engineering Research Center, KRIBB2Seonam Uinversity, 3National Academy of Agricultural Science

The FK506 binding proteins (FKBPs) and cyclophilins

33 3rd KRIBB Poster Festival

(CYPs) are abundant and ubiquitous proteins belonging topeptidyl-prolyl cis/transisomerase (PPIase) superfamily,which have been known to regulate a lot of biological activitythrough the function as a chaperone or an isomerization ofproline residues in protein folding. These are collectivelyreferred to immunophilin and are present in almost all cellularorgans. Particularly, a number of immunophilins have beenknown to relate to environmental stresses. Thus, theidentification and classification of putative rice FKBPs andCYPs could be a help to the rice breeding study for stresstolerance against global warming. We have identified andclassified FKBP and CYP proteins in rice (Oryza sativacv. Japonica), and gave the proper name for each PPIaseconsidering the ortholog-relation with Arabidopsis andChlamydomonasPPIases or molecular weight of the proteins.In addition, to select rice immunophilins related in the waterstress, we analyzed the expression of rice FKBPs and CYPsunder salt and drought stress. 29 genes for FKBPs and 27genes for CYPs are putatively identified, and among them,a number of genes could be putatively classified as orthologswith Arabidopsis immunophilins. However, some geneswere identified as novel genes which have not beenreported in other organisms containing Arabidopsis andChlamydomonas, and several genes were identified asparalogs by genetic replication. The significant number ofrice immunophilin genes found to be regulated by saltand/or drought stress. (The 21st Annual Meeting of theKorean Society for Molecular and Cellular Biology :Seoul, 2009)

Keywords : immunophilin, FKBP, CYP, Oryza sativa,water stress

Members

DirectorSuk-Yoon Kwon (sykwon@kribb.re.kr)

Principal Researcher Jang-Ryol Liu (jrliu@kribb.re.kr)Jae-Heung Jeon (jeonjh@kribb.re.kr)Jeong-Mee Park (jmpark@kribb.re.kr)

Senior Researcher Hyun-Soon Kim (hyuns@kribb.re.kr)Jae-Sun Moon (jsmoon@kribb.re.kr)Hye-Sun Cho (hscho@kribb.re.kr)Won-Joong Jeong (wonjoong@kribb.re.kr)Sung-Ran Min (srmin@kribb.re.kr)

Post-DocChan-Ju Lim (cjlim@kribb.re.kr)Serry-Koh (skohtn@kribb.re.kr)Jong-Hyun Kim (kjh1052@kribb.re.kr)Jung-Won Youm (yjw0116@kribb.re.kr)Joon-Woo Ahn (joon194@empal.com)Jeong-Hee Lee (jjonga7282@hanmail.net)

ResearcherWoong-Bom Kim (wbkim@kribb.re.kr)Ha-Yeon Lee (tulip1229@hanmail.net)Sora Choi (csora0321@naver.com)Bok-Sim Lee (alsgur04@kribb.re.kr)You Yopung Nim (dudsla83@hanmail.net)Min-Sook Suk (tnrl2104@empal.com)

Korea Research Institute of Bioscience and Biotechnology 34

C-25Korean efforts on tomato chromosome 2 genomesequencing

Sung-Hwan Jo1, Bo-Ra Kang1, Sang-Mi Kim1, JungeunKim1, Doil Choi2,*, and Cheol-Goo Hur1,*

1Bioinformatics Research Center, KRIBB2Department of Plant Science, Seoul National University

The genome of tomato (Solanum lycopersicum L.) is beingsequenced by an international consortium of 10 countriesas part of the project "International Solanaceae GenomeProject (SOL): Systems Approach to Diversity andAdaptation". The goal of the consortiumis to sequence theapproximately 220 Mb of euchromatin that may containthe majority of gene coding sequences and to use it as areference genome for the other solanaceous species. TheKorean SOL is aiming to complete sequencing of about 22Mb euchromatic region of chromosome 2 using BAC byBAC approach. From the initial sequencing stage, 60 BACclones, which are anchored on genetic markers, wereselected as "seed BACs". Further BAC extension from the"seed BACs" has been performed by utilization of BAC-end sequence and fosmid-end sequence database validatingtheir location with FISH and IL mapping. Currently, 23Mb of unique sequence was determined from 194 BACsusing conventional Sanger method and pool of 85 BACsand 95 fosmids using 454 FLX titanium platform. Amongthem, 184 BACs (14.7Mb) were mapped on chromosome2 and assembled in 29 super-contigs in order. An annotationeffort is also underway by collaboration with InternationalTomato Annotation Group. To completethe chromosome 2sequencing project, we are applying next generationsequencing platforms and comparative genomic analysis.(SOL2009 The 6th Solanaceae Genome Workshop :Delhi (India), 2009)

Keywords : tomato, Solanum lycopersicum L., chromosome 2

C-26SOLmiRNA : A comprehensive search tool forthe identification of microRNAs and targetgenes in Solanaceae.

Hyun-Jin Kim1, Kwang-Hyun Baek2, BongWoo Lee1, DoilChoi3,*, and Cheol-Goo Hur1,*

1Bioinformatics Research Center, KRIBB2School of Biotechnology, Yeungnam University3Department of Plant Science, Seoul National University

Mature microRNAs (miRNAs) are a class of small single-stranded and non-coding RNAs that range in length from

19 nucleotides(nt) to 25nt. Evolutionally conservedmiRNAs regulate negatively gene expression at theposttranscriptional levels by binding the target mRNAs formRNA cleavage or at the mRNA translation. In plants,miRNAs function to control tissue differentiation anddevelopment, signal transduction, vegetative growth orreproductive growth, and the response to biotic and abioticstresses. The family Solanaceae has important values innutrition, health, and economy, however, the research ofSolanacea miRNAs has been very limited. Therefore, wehave built the SOLmiRNA database, which holds theinformation of miRNAs and the target genes in the familyof Solanaceae. SOLmiRNA was constructed based oncomprehensively analyzed of EST data of Solanaceaespecies, especially of pepper, tomato, potato, tobacco, andNicotiana benthamiana. As the first web-based database ofmiRNAs in the family of Solanacea, SOLmiRNA isexpected to be a workbench for identifying miRNAs andthe target genes in Solanacea, and for comparing thefunctions of miRNAs with those in other plant species.SOLmiRNA is freely available and can be accessed athttp://genepool.kribb.re.kr/SOLmiRNA. (2009 CropFunctional Genomics Workshop : Gangneung, 2009)

Keywords : SOLmiRNA database, microRNAs, Solanaceae.

C-27Pepper EST database: comprehensive in silicotool for analyzing the chili pepper (Capsicumannuum) transcriptome

Hyun-Jin Kim1, Kwang-Hyun Baek2, Seung-Won Lee1,JungEun Kim1, BongWoo Lee1, Doil Choi3,*, and Cheol-Goo Hur1,*

1Bioinformatics Research Center, KRIBB2School of Biotechnology, Yeungnam University, Gyeongsan3Department of Plant Science, Seoul National University

Recent advances in the high-throughput sequencing projectfor the Expressed Sequence Tags (ESTs) of chili pepperplants (Capsicum annuum cv. Bukang) have producedhundreds of thousands of complementary DNA (cDNA)sequences. We have built the Pepper EST database, whichis the first available database for mining the complexity ofESTs of chili pepper. The database was built on 122,582sequenced ESTs and 116,412 refined ESTs from the 21EST libraries. The ESTs were clustered and assembled intofull-length cDNA, and assigned metabolic pathway, geneontology, and MIPS functional catalog. The Pepper ESTdatabase is designed to provide a workbench for (i) findingunigenes in pepper plants, (ii) studying the expressionpatterns in different developmental tissues and in stressfulconditions, and (iii) comparing the ESTs with those of otherSolanaceae family. The Pepper EST database is expected

35 3rd KRIBB Poster Festival

Bioinformatics Research Center

to provide a high-quality resource and potentially help toaddress numerous topics such as systemic understanding ofplant diseases and genetic-based population studies. Thedatabase is also anticipated to contribute to analyzing genesynteny on the chili pepper sequencing project through theEST mapping to the genome. The Pepper EST database isfreely available at http://genepool.kribb.re.kr/pepper. (2009Crop Functional Genomics Workshop : Gangneung,2009)

Keywords : chili pepper (Capsicum annuum), pepper ESTdatabase

C-28CVD Finder: a web-based finder of integratedarticle resources for cerebrovascular disease

Young-Uk Kim, Young-Kyu Park, and Young-Joo Kim

Bioinformatics Research Center, KRIBB

Complex diseases such as cerebrovascular disease, diabetes,and cancers have two or more genetic inheritance and are aswell affected by environmental factors, which complicatedthe diseases even worse. Due to the complex characteristics ofthe diseases, it is important to search and study comprehensiveliterature-based article resources. Some disease relatedliterature databases have been introduced through specialjournal issues or major websites. However, most of themare remained scattered throughout the internet and literaturesand users have difficulties to find accurate and comprehensiveinformation easily and timely manner. We developed CVDFinder, a web-based finder of integrated article resourcesfor cerebrovascular disease, which is freely available. Thesystem allows users to explore PubMed search resultscategorized by the MeSH (Medical Subject Headings) andthe symptom classification of Oriental medicine. CVDFinder collects data from important sites such as PubMed,Scirus, and Scopus automatically to maintain higher qualityand updated contents. Currently, the system indexes morethan 2,000,000 PubMed abstracts that are related to stroke,stroke etiology, and Oriental medical classification. Thesystem will contribute valuable literature information to ascientific and medicine society in cerebrovascular disease.(BioInfo2009: CBI-KSBSB Joint International SymposiumThe 62th Annual Meeting of the Korean Association ofBiological Sciences : Busan, 2009)

Keywords : stroke, literature, text-mining, etiology,bioinformatics

C-29 CVD Base: an integrated genetic information

resource for cerebral stroke

Young-Kyu Park, My-Young Cheong, and Young-Joo Kim

Bioinformatics Research Center, KRIBB

Stroke is the second most common cause of death in Koreaand worldwide. Also, it is a major cause of disability evenif the patients are survived. The interaction between lots ofgenes, environmental factors, and many etiologies includinghypertension, atherosclerosis, diabetes, hyperlipidemia, andcardiovascular disease, etc. are responsible for the disease.Up to date, various studies on the pathology and mechanismin terms of genetic experiments have been conducted andapproximately a hundred of genes were reported as strokeassociated genes. To support the studies on the cause andmechanism of stroke, an efficient database system whichprovides genetic variant information related to stroke andits etiologies is required. CVD Base is an integrated web-based genetic information resource for the cerebral strokedesigned to service researchers the genomic variants, genes,and secondary information. It provides numerous effectivesearch interfaces including browse function based on geneentries, search function based on keywords, compare functionbased on population, and showing the statistics of data stored.Raw stroke genomic variant data and its etiologies wereretrieved and extracted from journal papers and the dataentries of PubMed, OMIM, and UniProt databases. Werefined them based on gene information and stored into therelational database. CVD Base will be an effectiveinformation resource for the study on the cause andmechanism of cerebral strokes by providing one-stop servicefor the genomic variants and derived secondary informationassociated with stroke and its etiologies. (BioInfo2009:CBI-KSBSB Joint International Symposium The 62thAnnual Meeting of the Korean Association of BiologicalSciences : Busan, 2009)

Keywords : stroke, pathway, genomics, etiology,bioinformatics

C-30CVD information site: web portal-site supplyingessential information of cerebrovascular diseasebased upon Oriental medicine

Jinho Kim, Young-Kyu Park, and Young-Joo Kim

Bioinformatics Research Center, KRIBB

Cerebrovascular disease (CVD) is the second most frequentcause of death, followed by cancer, in Korea. CVD is acomplex disease resulting from the interaction of manygenetic and environmental factors. A series of research hasbeen concerted regarding causes and mechanisms of CVDand the information is provided through many web services.

Korea Research Institute of Bioscience and Biotechnology 36

Traditionally, in Korea, a good portion of CVD is diagnosedand treated based upon Oriental medicine such as acupuncture,moxibustion, and herbal medicine. Especially in CVDtreatment, conventional medicine and Oriental medicinecoexist. CVD information site integrated essentialcerebrovascular disease information for web serviceregarding stroke pathology, diagnostics, and treatments basedupon Oriental medicine. Raw information was collectedfrom various sources, such as journal articles, books, webs,and news stories and it was refined, classified, and storedinto a relational database system through automaticclassification as well as manual curation. To provide theinformation effectively to users, specialized retrieval systembased on web interfaces was implemented. CVD informationsite services the information to not only experts but alsopatients and/or ordinary people. CVD information site willserve investigators by providing expert information, patientsby providing knowledge of effective treatments, and ordinarypeople by providing many preventive tools. (BioInfo2009:CBI-KSBSB Joint International Symposium The 62thAnnual Meeting of the Korean Association of BiologicalSciences : Busan, 2009)

Keywords : stroke, pathway, portal, etiology, bioinformatics

C-31AsiDesigner: exon-based siRNA design serverconsidering alternative splicing

Young-Joo Kim1, Misun Won2, and Young-Kyu Park1

1Bioinformatics Research Center, KRIBB2Medical Genomics Research Center, KRIBB

RNA interference (RNAi) with small interfering RNA (siRNA)has become a powerful tool in functional and medicalgenomic research through directed post-transcriptional genesilencing. In order to apply RNAi technique for eukaryoticorganisms, where frequent alternative splicing results indiversification of mRNAs and finally of proteins, we needspliced mRNA isoform silencing to study the function ofindividual proteins. AsiDesigner is a web based siRNAdesign software system which provides siRNA designcapability to account for alternative splicing for mRNAlevel gene silencing. It provides numerous novel functionsincluding the designing of common siRNAs for the silencingof more than two mRNAs simultaneously, a scoring schemeto evaluate the performance of designed siRNAs by adoptingstate-of-the-art design factors, a stepwise off-target searchingwith BLAST and FASTA algorithms and checking thefolding secondary structure energy of siRNAs. To do this,we developed a novel algorithm to evaluate the commontarget region where siRNAs can be designed to knock down aspecific mRNA isoform or more than two mRNA isoformsfrom a target gene simultaneously. The developed algorithm

and the AsiDesigner were tested and validated as veryeffective throughout widely performed gene silencingexperiments. It is expected that AsiDesigner will play animportant role in functional genomics, drug discovery, andother molecular biological research. AsiDesigner is freelyaccessible at http://sysbio.kribb.re.kr:8080/AsiDesigner/.(2008 ASHG Conference 58th Annual Meeting :Philadelphia (USA), 2008)

Keywords : siRNA, alternative splicing, design,bioinformatics

Members

DirectorCheol-Goo Hur (hurlee@kribb.re.kr)

Principal Researcher Kyou-Hoon Han (khhan600@kribb.re.kr)Young-Ju Kim (yjkim8@kribb.re.kr)In-Sun Chu (chu@kribb.re.kr)

Senior Researcher Nam-Shin Kim (deepreds@kribb.re.kr)Seung-Won Lee (swlee@kribb.re.kr)

Post-DocSung-Hwan Jo (shjo@kribb.re.kr)Hyun-Seo Jang (jhsc79@kribb.re.kr)Hea- Young Oh (helloaz@kribb.re.kr)

ResearcherJung-Eun Kim (jekim@kribb.re.kr)Sang-Mi Kim (pepepoint@hanmail.net)Hyun-Jin Kim (hjkim@kribb.re.kr)Bong_Woo Lee (huk5432@kribb.re.kr)Young-Kyu Park (ykpark@kribb.re.kr)Jin-Ho Kim (jinho76@kribb.re.kr)My-Young Cheong (chongmy@kribb.re.kr)Young-Uk Kim (kimuks@kribb.re.kr)

37 3rd KRIBB Poster Festival

C-32Anti-atherosclerotic effects of Redobovatol,analogue of obovatol on atherogenic lesionformation in LDL receptor-deficient mice

Jong-Min Han1, Seung-Hwa Baek1, Hua Li1, Ji-Seon Park1,Moon-Hee Cho1, Young-Min Han2, Byoung-Mog Kwon2,and Tae-Sook Jeong1,*

1Industrial Bio-materials Resource Research Center,KRIBB

2Medical Genomic Research Center, KRIBB

Atherogenesis is a complex process involving mechanical,chemical and biological factors. Damage to endothelialcells and activation, attachment and transmigration ofmonocytes through the endothelium are early events inatherosclerosis. Biphenolic components in Magnoliaobovata have shown several pharmacological activitiessuch as antitumor, anti-oxidant and anti-inflammatoryeffects. Among them, obovatol has anti-inflammatoryactivity through inhibition of NF- B in LPS-treatedmacrophage RAW264.7 cells and in UVB-treatedfibroblast. Redobovatol (Rd-obo) was synthesized fromobovatol, and we found that Rd-obo inhibited activation ofNF- B and AP-1 which have been known to be a significanttranscriptional factor to control of endothelial cellinflammation. The present study was designed to examinewhether the Rd-obo could prevent atherosclerosis in LDLreceptor deficient (LDLR-/-) mice fed with a Western dietfor 12 weeks. The mice were fed with a chow diet alone(ND group), a Western diet containing 0.15% cholesterolalone (control group) or a Western diet supplemented with0.2% Rd-obo (wt/wt diet, Rd-obo group). A Western diet hasbeen shown to cause hyperlipidemia and atherosclerosis inLDLR-/- mice. Rd-obo group significantly lowered plasmatotal cholesterol and triglyceride levels as compared to thecontrol group. In aortic arch, atherosclerotic plaque wassignificantly induced in the control group (129.6 ± 45.7 m2x103), however, Rd-obo group showed lower plaqueabout 62.6 ± 53.3 m2x103. We found that this reductionwas associated with a reduced expression of markers formacrophages and NF- B mediated factors, CCL2,CX3CL1, TNF- , iNOS, and MMP9 in the vascular wallby real-time quantitative polymerase chain reaction. Theseresults suggest that redobovatol is involved with adecreased atherosclerosis and with the suppression of pro-inflammatory mediators in the LDLR-/- mice.

Seoul, 2009

Keywords : redobovatol, atherosclerosis, cholesterolmetabolism, proinflammatory mediators

C-33Effects of ethanolic extracts of Artemisia princepsPamp. cv. Sajabal in high-fat diet-induced obesemice

Ji-Seon Park1, Seung-Hwa Baek1, Hua Li1, Moon-Hee Cho1,Jong-Min Han1, Hae-Gon Chung2, Nam-In Baek3, Myung-Sook Choi4, Kyung-Tae Lee5, and Tae-Sook Jeong1,*

1Industrial Bio-materials Research Center, KRIBB2Gangwha Agricultural R&D Center3,5Kyung Hee University4Kyungpook National University

Obesity is the most common nutritional disorder worldwideand it is characterized by an increase in fat mass. Anti-obesity targets on suppression of food intake, anti-adipogenesis and increased lipolysis, fat oxidation andenergy expenditure. Artemisia princeps Pamp. is acommonly known herbaceous plant in Korea, Japan, andChina. A. princeps Pamp. cv. Sajabal is specially cultivated inGanghwa County, a place located in the west coast ofKorea. It contains a high content of flavonoids such aseupatilin and jaceosidin as compared with Artemisia herbsfrom other regions. The anti-obese effect of ethanolicextracts of A. princeps Pamp. cv. Sajabal (ESJ) wasevaluated in diet-induced obese mice. Male C57BL/6Jmice were used and the mice were divided into control andESJ groups. The control group was fed a high-fat (45kcal% fat) diet, while the ESJ group was fed a high-fatdiet with the ESJ (1% wt/wt diet) for 12 weeks, while theoverall amount of food intake was not affected. ESJsupplementation significantly decreased body weight gain,food efficiency ratio and adipose tissue weight and size.ESJ lower the plasma levels of triglyceride, glucose andglutamate oxaloacetate transaminase. The activities ofsuperoxide dismutase and catalase in erythrocytes werehigher in the ESJ group compared with the control group.ESJ treatment induced the expression in white adiposetissues and liver of lipolysis- and energy expenditure-relatedgenes including hormone sensitive lipase, peroxisomeproliferator-activated receptor (PPAR ), PPAR ,peroxisome proliferator-activated receptor gamma coactivator-1 and uncoupling protein. ESJ also reversed the high-fatdiet-induced change in the expression pattern of genes suchadiponectin, PPAR , acetyl-CoA carboxylase 1 (ACC1),ACC2, fatty acid synthase and diacylglycerol acyltransferase.These results suggest that ESJ has potential as an effectiveanti-obese agent via the inhibition of adipocyte lipidaccumulation and synthesis and the stimulation of energyexpenditure. Seoul, 2009

Keywords : Artemisia princeps Pamp. cv. Sajabal, anti-obese effect, proliferator-activated receptor,lipid metabolism, white adipose tissue

Korea Research Institute of Bioscience and Biotechnology 38

Industrial Bio-materials Research Center

C-34Ethanol extract of Torreya nucifera seed blockshepatic steatosis and atherosclerosis in LDLreceptor-deficient mice

Jong-Min Han1, Jang-IL Han1, Seung-Hwa Baek1, Hua Li1,Ji-Seon Park1, Moon-Hee Cho1, Yong-Hoon Kim2, Chul-Ho Lee2, and Tae-Sook Jeong1,*

1Industrial Bio-materials Research Center, KRIBB2Laboratory of Experimental Animals, KRIBB

Atherosclerosis may be the result of genetic susceptibilitycombined with environmental factors such as diet, lifestyle,or possibly microbial infections. Dietary cholesterol playsan important role in determining the plasma levels ofcholesterol and the risk of cardiovascular diseases. Torreyanucifera seed oil was known as useful hypolipidemic oilbecause it can reduce the plasma and liver triacylglycerollevels by suppressing hepatic fatty acid synthesis. Theobjective of this study was to investigate whether ethanolextract of Torreya nucifera seed (ETNS) blocks highcholesterol diet-induced liver steatosis and atherosclerosisin LDL receptor deficient (LDLR-/-) mice. The mice werefed with a chow diet alone (ND group), a Western dietcontaining 0.15% cholesterol alone (control group) or aWestern diet supplemented with 1.0% ETNS (wt/wt diet,ETNS group) for 12 weeks. A Western diet has been shownto cause obesity, fatty liver, and increased Hyperlipidemiaand atherosclerosis in LDLR-/- mice. ETNS group significantlylowered plasma total cholesterol and triglyceride levels ascompared to the control group. Liver sections were stainedwith oil Red O, and ETNS groups showed lower hepaticsteatosis compared to the control group. Further analysisusing RT-PCR, ETNS inhibits expression of enzymesrelated to the fatty acid synthesis, such as FAS and ACC-1via blocked SREBP-1 and PPAR . In aortic arch,atherosclerotic plaque was significantly induced in thecontrol group (129.6 45.7 m2x103), however, ETNSgroup showed lower plaque about 65.5 36.1 m2x103.Moreover, expression of proinflammatory mediators suchas, CCL2, CX3CL1, IL-1ß, and TNF- was suppressed inaorta of ETNS group. These results suggest that the anti-atherogenic effect of the ETNS is involved with a decreasedhepatic steatosis and with the suppression of pro-inflammatory mediators in the LDLR-/- mice. (The 66thAnnual Meeting of the Korean Society for Biochemistryand Molecular Biology : Seoul, 2009)

Keywords : Torreya nucifera, ETNS, atherosclerosis,liver steatosis, pro-inflammatory mediators

C-35Novel GH10 xylanase, with a fibronectin type 3domain, from Cellulosimicrobium sp. strainHY-13, a bacterium in the gut of Eisenia fetida

Do Young Kim1, Mi Kyiung Han1, Doo-Sang Park2, JongSuk Lee3, Hyun-Woo Oh1, Dong-Ha Shin4, Tae-Sook Jeong1,Sung Uk Kim1, Kyung Sook Bae2, Kwang-Hee Son1, andHo-Yong Park1

1Industrial Bio-materials Research Center, KRIBB2Biological Resources Center, KRIBB3Industrial Biotechnolgy & Bioenergy Research Center,KRIBB

4Insect Biotech Co. Ltd.

The gene encoding a novel modular xylanase from anearthworm-symbiotic bacterium, Cellulosimicrobium sp.HY-13 was identified and expressed in E. coli and itstruncated gene product was characterized. The isolatedXylK1 gene contained a 1,671-bp open reading frame thatencodes a protein of 556 amino acids with a deduced molecularmass of 58,296 Da and a calculated pI of 4.59. The enzymeconsisted of three distinct functional domains, an N-terminalcatalytic GH10 domain (Leu38 to Asp330), a fibronectintype 3 (Fn3) domain (Pro359 to Gly430), and a C-terminalcarbohydrate-binding module 2 (CBM2) (Cys454 to Cys553)and no xylanase with domain architecture identical to that ofXylK1 with an Fn3 domain has been reported to date. Thecatalytic GH10 domain of XylK1 showed the highestsequence identity (67%) with that of the Cellulomonasfimi xylanase (AAA56792). However, its CBM 2 and Fn3domain were most identical to that (64%) of C. fimi GH6cellulase (AAC36898) and that (70%) of the Acidothermuscellulolyticus 11B GH48 enzyme (ABK52390), respectively.The binding ability of the C-terminal CBM 2-lacking rXylK1to avicel and insoluble oat spelt xylan clearly indicatedthat the Fn3 domain plays an important role in the enzyme-substrate binding because rXylK1∆Fn3 was not bound toavicel. A significant decrease in the catalytic activity ofrXylK1∆Fn3 induced by deletion of the Fn3 domain alsosuggested that the Fn3 domain may participate in promotionof the catalytic hydrolysis of xylosic substrates by modifyingtheir surfaces. The relatively high cleavage activity of rXylK1toward PNP-cellobioside and its transxylosylation activitythat enables it to produce longer xylooligosaccharidesfrom X3 or X4 differentiate it from other known GH10xylanases. (1st International Symposium on InsectBiotechnology and 2nd Korea-China Joint Symposiumon Insect Biotechnology : Busan, 2009)

Keywords : Cellulosimicrobium sp. HY-13, Eiseniafetida, xylanase, Fn3 domain

C-36Anti-inflammatory properties of TellimagrandinI, isolated from Corylopsis coreana inmousemacrophages

Jong-Min Han, Tae-Hoon Kang, Sung-Wook Kim, andTae-Sook Jeong*

39 3rd KRIBB Poster Festival

Industrial Bio-materials Research Center, KRIBB

Nitric oxide (NO) is a free radical produced from L-arginineby a catalytic reaction of two types of constitutive or inducibleNOS and an important intercellular signal messenger.Although the large amount of NO produced by iNOS helpsto kill pathogens, excessive NO production contributes tonot only harmful cellular responses but also initiation andprogression of inflammation, sepsis, atherosclerosis, andcancer. In general, it has been known that NO and reactiveoxygen species (ROS) are responsible for regulation on thetranscriptional pathways of nuclear factor- B (NF- B).Therefore, NF- B is a good therapeutic target forcardiovascular disease and numerous efforts are beingmade to develop safe NF- B inhibitors. Tannins have beenshowed to possess a variety of pharmacological activitiessuch as antiviral, antimicrobial, antioxidant, antimutagenic,and anti-tumor activities. Tellimagrandin I is a hydrolysabletannin compound widely present in plants. TellimagrandinI (Tel I) had inhibitory effects on erythroid and megakaryocyticdifferentiation, and tannins like Tel I could influence theanti-tumor efficiency of some drugs and the hematopoiesisprocesses. The goal of the present study was to investigatethe role of Tel I in LPS-induced expression of proinflammatorymediators and chemokines in RAW264.7 macrophages.Tel I inhibited accumulation of ROS, such as hydrogenperoxide, and was able to dose-dependently reduce theLPS-induced NO production as well as iNOS proteinexpression and the expression of various inflammation-associated genes (iNOS, IL-1ß, IL-6, and CX3CL1). We alsoexamined the ability of Tel I to modulate the transcriptionfactors, NF- B DNA binding activity in a dose-dependentmanner by western blotting and EMSA. In conclusion,Tellimagrandin I appears to selectively prevent the nuclearentry of activated NF- B, and this may be the basis of itsbeneficial effect in inflammation-related diseases.

Seoul, 2009

Keywords : tellimagrandin I, nuclear factor- B,proinflammatory mediators, chemokines

C-37Linoleic acid derivative (S)-glycerol-monolinoleateattenuate atherogenic lesion formation in LDLreceptor-deficient mice

Jong-Min Han1, Jang-IL Han1, Seung-Hwa Baek1, Hua Li1,Ji-Seon Park1, Moon-Hee Cho1, Hyung-Jae Jeong2, WooSong Lee2, and Tae-Sook Jeong1,*

1Industrial Bio-materials Research Center, KRIBB2Bioindustry Technology Research Center, KRIBB

The atherosclerotic lesion begins with the entry of low-density lipoprotein (LDL) cholesterol into the vascularintima. The subsequent oxidation of this LDL-cholesterol

leads to the expression of adhesion and chemotactic molecules.At present, the two main conceptual approaches to therapyfor atherosclerosis are manipulation of plasma lipoproteinmetabolism or cellular cholesterol metabolism, andmanipulation of inflammatory processes. In this study, weinvestigated whether Fatty acid glycerol (FAG), (S)-glycerol-monolinoleate, blocks inflammation-related atherosclerosis inhigh cholesterol diet-fed LDL receptor deficient mice. Themice were fed with a chow diet alone (ND group), a Westerndiet containing high cholesterol alone (control group) or adiet supplemented with 0.02% FAG (wt/wt diet, FAG group)for 12 weeks. Surprisingly, FAG group significantly loweredplasma total cholesterol and triglyceride levels as comparedto the control group. Furthermore, atherosclerotic plaquein the aortic sinus was significantly reduced in FAG group(oil red O-stained lesions was 129.6 ± 45.7 m2x103 incontrol group versus 51.0 ± 24.5 m2x103 in FAG group)and in the ascending aorta. Moreover, expression of mRNAand protein which regulated by NF- B such as, CCL2,CX3CL1, and TNF- were suppressed in aorta. In conclusion,our results suggest that FAG could be useful in the preventionand management of hyperlipidemia and atherosclerosis.(The 66th Annual Meeting of the Korean Society forBiochemistry and Molecular Biology : Seoul, 2009)

Keywords : (S)-glycerol-monolinoleate, atherosclerosis,inflammation, hyperlipidemia

Members

DirectorTae-Sook Jeong (tsjeong@kribb.re.kr)

Principal Researcher Young-Ik Lee (yilee@kribb.re.kr)Sung-Uk Kim (kimsu@kribb.re.kr)Ho-Yong Park (hypark@kribb.re.kr)Kwang-Hee Son (sonkh@kribb.re.kr)

Senior Researcher Hyun-Woo Oh (hwoh@kribb.re.kr)

Post-DocJong-Min Han (malius@kribb.re.kr)Do-Young Kim (kdy119@kribb.re.kr)Jong-Seok Kim (jskim901@kribb.re.kr)Hye-Suk Han (sion5081@hanmail.net)

ResearcherJi-Hyun Kang (bsloaucl@kribb.re.kr)Sung-Eun Kim (kse9456@hanmail.net)Yi-Joon Kim (kimiyu@kribb.re.kr)Ji-Ae Kim (jiaekim@kribb.re.kr)Jin-Sung Mok (mokujin@kribb.re.kr)Sun-A Park (seona@kribb.re.kr)Sung-Ho Park (psh9787@naver.com)Min-Kyu Song (ecoli@kribb.re.kr)

Korea Research Institute of Bioscience and Biotechnology 40

Song-I Yoo (anyonestar@naver.com)Lan-Hee Lee (wendy83@kribb.re.kr)Eun-Na Lee (saysayna@nate.com)Hwa Li (leehua@kribb.re.kr)Hoon Jang (janghoon@kribb.re.kr)Young-Eun Jeon (jye3533@hanmail.net)Moon-Hee Cho (munhui@kribb.re.kr)Ji-Hwa Ju (qhtitizzz@nate.com)Won-Sik Choi (highelfu@hanmail.net)Mi-Kyung Han (rosy22@kribb.re.kr)Su-Jin Ham (sjcom86@kribb.re.kr)Pung-Mi Hyun (pungmi@hanmail.net)

C-38Optimization for the harvest of high-densityScenedesmus sp. culture using a biopolymerproduced by Paenibacillus polymyxa AM49

Kim, Dong-Geol1, Chi-Yong Ahn1, Yong-Ha Park2, andHee-Mock Oh1,*

1Environmental Biotechnology Research Center, KRIBB2Yeungnam University, Gyeongsan

Biodiesel from microalgae seems to be desirable renewablebiofuel that has a potential to replace the petroleum-derivedtransport fuels. The harvesting of microalgal culture is oneof major processes in microalfal mass-production. Therefore,the development of efficient harvesting method is stillneeded for the cost-effective production of biodiesel frommicroalgal biomass. The optimum conditions for the harvestby flocculation of Scenedesmus sp. with culture broth ofPaenibacillus polymyxa AM49 were investigated. Theflocculating activities of 5 mM FeCl3 and 10 mM CaCl2 asa co-flocculant for 1.0 g DCW/L of Scenedesmus sp. were83.8% and 49.8% which were higher than those of NH4Cl,KCl, MgSO4 and Al2(SO4)3. As the density of Scenedesmussp.culture increase, the flocculating activity was declinedfrom 47.2% (at 0.832 g DCW/L) to 4.3% (at 2.329 g DCW/L)with only CaCl2. That was also same with a co-flocculantFeCl3. With the consecutive treatment of 10 mM CaCl2followed by 1 mM FeCl3, the flocculting activityincreased up to 99.7% at a relatively high density ofScenedesmus sp. (2.329 g DCW/L). The flocculatingactivity of a biopolymer produced by P.polymyxa AM49was higher than a chemical flocculant, alum AlNH4(SO4)2.The consecutive treatment of 2 co-flocculants also reducedsettling time in the process of Scenedesmus sp. flocculation.With a recycled culture broth after biopolymer flocculation,the growth of Scenedesmus sp. was 55% of that with a freshBG11 medium and which was higher than 34% with arecycled culture broth after alum flocculation. (2009 AlgaeBiomass Summit : San Diego (USA), 2009)

Keywords : bioflocculant, microalgal, Paenibacilluspolymyxa

C-39Anaerobic VC to ethene dechlorinatingenrichment culture

Byung-Hyuk Kim1,3, Kyung-Hwa Baek1, Dae-Hyun Cho1,Youlboong Sung2, Chi-Yong Aha1, Hee-Mock Oh1, Sung-Cheol Koh3, and Hee-Sik Kim1,*

1Environmental Biotechnology Research Center, KRIBB2Environmental and Energy Research Center, RIST3Division of Construction and Environmental Engineering,Korea Maritime University

41 3rd KRIBB Poster Festival

Environmental Biotechnology Research Center

Vinyl chloride (VC) is a known human carcinogen. It is thehighest ranked organic compound on the 2007 CERCLAPriority List of Hazardous Substances. VC is found in thesubsurface, due to incomplete degradation of other prioritypollutants such as PCE, TCE, and cis-DCE. Other sourcesof VC in groundwater include waste from polyvinyl chlorideplastic production. Under anaerobic conditions, thesecompounds can be reductively dehalogenated to innocuousethene by microorganisms through dehalorespiration. Wehave developed the anaerobic enrichment culture, calledFMC-12T which completely converted 1.4 mM VC to ethenewithin 5 weeks. The bacterial diversity of the establisheddechlorinating enrichment culture, FMC-12T was analyzedbysequencing of 16S rRNA gene clones and PCR-DGGE.The results of sequencing, FMC-12T consisted ofSulfurospirillum sp. (75.0 %), Denitrovibrio acetophilus (8.3%), and Dehalococcoides sp. (8.3 %). The well-studiedorganisms, Sulfurospirillum sp. dechlorinated PCE to cis-DCE. In conclusion, we established the VC dechlorinatingenrichment culture and confirmed the existence ofSulfurospirillum sp. in the FMC-12T. (MSK's 50thAnniversary International Symposium on Microbiology: Jeju, 2009)

Key words : anaerobic enrichment culture, dechlorination,microbial diversity, Sulfurospirillum sp.,vinyl chloride.

C-40Axenic Induction method for axenic culture ofa cyanobacterium, Microcystis aeruginosa

Eun-Kyung Kim, Chi-Young Ahn, Gang-Guk Choi, Young-Ki Lee, and Hee-Mock Oh

Environmental Biotechnology Research Center, KRIBB

µmol photons/m2/s MA

KW

µg/ml

µg/ml

µg/ml KW

ug/ml

96 well plat well KW

KW

Geoje, 2009

Keywords : antibiotic, axenic, Microcystis, high throughput

C-41Effect of light emitting diode wavelength forthe lipid content and composition of Chlorellavulgaris

Jae-Yon Lee, Young-Ki Lee, Chi-Yong Ahn, and Hee-Mock Oh

Environmental Biotechnology Research Center, KRIBB

Light Emitting Diode (LEDs)

LED

Chlorella vulgaris N2170

LED (White, Red,

Green, Blue) (fatty

acid) C. vulgaris

N2170 Blue LED White, Red,

Green LED

: Geoje, 2009

Keywords : lipid content, lipid composition, cell growth,LEDs, chlorella vulgaris

Korea Research Institute of Bioscience and Biotechnology 42

C-42Vinaxanthone, a new type of FabI inhibitorfrom Penicillium sp.

Yun-Ju Kwon, Mi-Jin Sohn, Chang-Ji Zheng, and Won-Gon Kim

Environmental Biotechnology Research Center, KRIBB

Bacterial enoyl-ACP reductase that catalyzes the final andrate-limiting step in the type II fatty acid synthesis has beenvalidated as a novel target for antibacterial drug development.There are three isoforms, FabI, FabK and FabL, of enoyl-ACP reductase. FabI is highly conserved among mostbacteria. Our screening program for new FabI inhibitorsled to the selection of Penicillium sp. producing a stronginhibitory metabolite. The active compound was purifiedas a single compound through bioassay-guided fractionation.Its chemical structure was elucidated to be vinaxanthone.It selectively inhibited Staphylococcusaureus FabI with anIC50 of 0.9 mM, while didn’t effect FabK.Consistent withits inhibition of FabI, the inhibitor prevented intracellularfatty acid synthesis as well as the growth of S. aureus,MRSA, and QRSA with MIC of 32µg/ml. Importantly,FabI-overexpressing S. aureus showed reduced susceptibilityto the inhibitor compared to wild strain, confirming that itsantibacterial action is mediated by the inhibition of FabI.In conclusion, vinaxanthone is a new type of FabI-directedantibacterial of microbial origin. This work was supportedby the 21C Frontier Microbial Genomics and ApplicationCenter Program. (International Symposium & AnnualMeeting of the Korean Society for Microbiology andBiotechnology : Daejeon, 2009)

Keywords: enoyl-ACP reductase, antibacterial,Staphylococcus aureus, microbial metabolite,inhibitor, penicillium

C-43New type of enoyl-ACP reductase inhibitorfrom Sporothrix sp.

Yun-Ju Kwon, Yi Fang, and Won-Gon Kim

Environmental Biotechnology Research Center, KRIBB

Bacterial enoyl-ACP reductase has been validated as anovel target for antibacterial drug development. There arethree isoforms, FabI, FabK and FabL, of enoyl-ACPreductase. FabI is highly conserved among most bacteria,while Streptococcus pneumoniae conatin only FabK,Enterococcus faecalis and Pseudomonas aeruginosacontain both FabI and FabK, and Bacillus subtilis containboth FabI and FabL. Our continued screening of FabIinhibitors led to the selection of a fungal strain, Sporothrix sp.

FN611, producing a strong inhibitory metabolite. The activecompound was purified as a single compound throughbioassay-guided fractionation using ethylacetate extraction,silica gel column chromatography and preparative TLC.Chemical structure of the purified compound namedcompound 611 was elucidated by MS and NMR spectraldata. Compound 611 inhibited Staphylococcus aureus FabIwith an IC50 of 3.2 uM. The compound also inhibited S.pneumoniae FabK with an IC50 of 9.2 uM, which was 3-times weaker than that aganist FabI. The degalactosylationof aquastatin A did not affect the FabI and FabK-inhibitory orantibacterial activities, thereby suggesting that the sugarmoiety within its molecular structure was not involved inthese activities. The compound showed mixed-type inhibitionagainst FabI with respective to both the substrate and thecofactor NADPH. Consistent with its inhibition of FabIand FabK, the compound also exhibited antibacterial activityon S.aureus and methicillin-resistant Staphylococcus aureus(MRSA) with MIC of 16 - 32 µg/ml, while showed a weakantibacterial activity on S. pneumoniae with a MIC of 128µg/ml. This work was supported by the 21C FrontierMicrobial Genomics and Application Center Program.(International Symposium & Annual Meeting of theKorean Society for Microbiology and Biotechnology :Daejeon, 2009)

Keywords: FabI, FabK, antibacterial, MRSA, Staphylococcusaureus, microbial metabolite, inhibitor

C-44New type of FabG inhibitor from Bacillus sp.AT28

Yun-Ju Kwon, Chang Ji Zheng, Mi-Jin Sohn, and Won-Gon Kim

Environmental Biotechnology Research Center, KRIBB

Bacterial fatty acid synthase (FAS), which is essential forbacterial growth, consists of multiple individual enzymes,each encoded by a separategene, in contrast to the mammalianfatty-acid synthase. There is low protein sequence homologyand that there are different arrangements of enzymatic-activesites between bacterial and mammalian FAS. Thesedifferences have been exploited to establish bacterial FASas a target for antibacterial drug discovery. b-Ketoacyl-acyl carrier protein (ACP) reductase (FabG) is an essentialenzyme which catalyzes the reduction of b-ketoacyl-ACPto b-hydroxyacyl-ACP in the bacterial fatty acid synthesispathway. Since the FabG gene is well conserved in mostpathogenic bacteria, it is considered to be a new potentialtarget for broad-spectrum antibacterial agents. In thecourse of screening for FabG inhibitors from microbialsources, a new 24-membered ring lactone named macrolactinS,along with the known compound macrolactin B, has been

43 3rd KRIBB Poster Festival

isolated from the mycelium of liquid fermentation cultures ofBacillus sp. AT28. The structure of macrolactin S wasdetermined on the basis of MS and NMR data. Macrolactin Sshowed a dose-dependent inhibition of Staphylococcusaureus FabG, not inhibiting S. aureus FabI. Also macrolactinS inhibited the growth of S. aureus, Bacillus subtilis, andEscherichia coli. This work was supported by the 21CFrontier Microbial Genomics and Application CenterProgram. (International Symposium & Annual Meetingof the Korean Society for Microbiology and Biotechnology: Daejeon, 2009)

Keywords: bacterial fatty acid synthase, FabG,antibacterial, Staphylococcus aureus,microbial metabolite, inhibitor

C-45Transgenic sweetpotato plants expressing abacterial phytase gene

Kyoung-Sil Yang1, Tae-Kwang Oh2, Il-Gin Mok1, Haeng-Soon Lee1, and Sang-Soo Kwak1*

1Environmental Biotechnology Research Center, KRIBB2Microbial Genomics & Applications Center, KRIBB

Phytic acid (myo-inositol hexakisphosphate, phytate) is notavailable to monogastric animals and phosphatesupplementation is required for optimal animal growth.Undigested phytate in animal manure is considered a majorsource of phosphorus pollution to the environment fromagricultural production. Phytase (myo-inositol hexakisphosphatephosphohydrolase) catalyze the hydrolysis of the phosphatemoieties from phytic acid, thereby, resulting in the loss ofability of phytic acid to chelate metal ions. Sweetpotato[Ipomoea batatas (L.) Lam.] is one of the most importantcrops to produce functional food and feed materials onmarginal lands. In this study, to develop the transgenicsweetpotato (cv. Yulmi) with a functional feed material,we constructed the transformation vectors using phytasegene under the control of enhanced CaMV 35S promoteror sporamin promoter with high expression in the storageroots of sweetpotato (referred to as E35Sp::Phytase andSPOp::Phytase, respectively). Transgenic sweetpotatoplants were successfully generated by Agrobacterium-mediated transformation. Kanamycin-resistant embryogeniccalli were selected on MS medium containing 400 mg/Lclaforan and 100 mg/L kanamycin. Embryogenic callitransferred to hormone-free MS medium with kanamycingave rise to somatic embryos and then converted intoplantlets in the same medium. The putative transgenicplants were selected by PCR with nptII or phytase-specificprimer. Further characterization of transgenic sweetpotatoplants is under study. (Joint Meeting of The BotanicalSociety of Korea and The Korean Society for Plant

Biotechnology : Gwangju, 2009)

Keywards : Sweetpotato, Phytic acid, Phytase, Functionalfeed, phosphorous pollution

C-46Enhanced tolerance to MV-mediated oxidativestress and high temperature in transgenicpotato plants overexpressing CuZnSOD, APXand NDPK2 genes

Myoung Duck Kim1, Yun-Hee Kim, Suk-Yoon Kwon2,Sang-Soo Kwak1, and Haeng-Soon Lee1*

1Environmental Biotechnology Research Center, KRIBB2Plant Genomic Reserch Center, KRIBB

Oxidative stress is one of the major factors causing injuryto plants exposed to environmental stress. To developtransgenic plants with enhanced tolerance to multipleenvironmental stresses, we are trying to manipulate theantioxidative mechanism under the control of an oxidativestress-inducible SWPA2 promoter. In a previous study, wedeveloped SSA potato plants expressing genes of bothsuperoxide dismutase (CuZnSOD) and ascorbate peroxidase(APX) in chloroplasts (referred to as SSA plants) ornucleoside diphosphate kinase 2 (NDPK2) in cytosols (SNplants) under the control of SWPA2 promoter. Both SSAand SN plants showed a strong tolerance to methyl viologen(MV)-mediated oxidative stresses and high temperature. Inthis study, NDPK2 gene was further introduced into SSAplants (SSAN plants) under the control of SWPA2promoter by an Agrobacterium tumefaciens-mediated transSWmation to expect the synergistic effect of SSA plantsand SN plants. SSAN potato plants showed enhancedtolerance to MV treatment in leaf and whole plant levelscompare to SN, SSA or non-transgenic plants, respectively.In addition, SSAN plants showed enhanced tolerance tohigh temperature stress. These results ind inte that thesimultaneous expression of antioxidant enzymes and ROSsignal pathway-regulating upstream enzyme are moreeffective than overexpression of double antioxidantenzyme (SSA) or single signal pathway regulating enzyme(SN) for developing transgenic plants with enhancedtolerance to oxidative stress. We anticipate that SSANpotato plants would be useful for commercial cultivationin unfavorable growth conditions. (Joint Meeting of TheBotanical Society of Korea and The Korean Society forPlant Biotechnology : Gwangju, 2009)

Keywords : ascorbate peroxidase (APX), nucleosidediphosphate kinase 2 (NDPK2), oxidativestress-inducible SWPA2 promoter, superoxidedismutase (CuZnSOD)

Korea Research Institute of Bioscience and Biotechnology 44

C-47Stress-induced expression of choline oxidase inpotato plant chloroplasts confers enhancedtolerance to oxidative, salt and drought stresses

Myoung Duck Kim, Raza Ahmad, Haeng-Soon Lee, andSang-Soo Kwak*

Environmental Biotechnology Research Center, KRIBB

Transgenic potato plants (Solanum tuberosum L. cv. Superior)with the ability to synthesize glycinebetaine (GB) inchloroplasts (referred to as SC plants) were developed viathe introduction of the bacterial choline oxidase (codA) geneunder the control of an oxidative stress-inducible SWPA2promoter. SC1 and SC2 plants were selected via theevaluation of methyl viologen (MV)-mediated oxidativestress tolerance, using leaf discs for further characterization.The GB contents in the leaves of SC1 and SC2 plantsfollowing MV treatment were found to be 0.9 and 1.43mmol/g fresh weight by HPLC analysis, respectively. Inaddition to reduced membrane damage after oxidative stress,the SC plants evidenced enhanced tolerance to NaCl anddrought stress on the whole plant level. When the SC plantswere subjected to two weeks of 150 mM NaCl stress, thephotosynthetic activity of the SC1 and SC2 plants wasattenuated by 38% and 27%, respectively, whereas that ofnon-transgenic (NT) plants was decreased by 58%. Underdrought stress conditions, the SC plants maintained higherwater contents and accumulated higher levels of vegetativebiomass than was observed in the NT plants. In addition,A codA gene was further introduced into transgenic potatoplants expressing both genes of CuZnSOD and APX inchloroplasts (SSAC potato plants). SSAC potato plantsshowed synergistically tolerant to various abiotic stresses.

Seoul, 2009

Keywords : abiotic stresses, bacterial choline oxidase(codA), glycinebetaine (GB), oxidative stress-inducible SWPA2 promoter

C-48Overexpression of sweetpotato orange geneconferred enhanced carotenoid contents insweetpotato and Arabidopsis

Sun Ha Kim1, Young Ock Ahn1, Joon-Seol Lee2, Il-GinMok1, Haeng-Soon Lee1, and Sang-Soo Kwak1*

1Environmental Biotechnology Research Center, KRIBB2Bioenergy Crop Research Center, NICS, RDA

Sweetpotato (Ipomoea batatas L.) is an important food andindustrial crop to produce useful components includingvarious antioxidants such as carotenoids and anthocyanins.

We are trying to develop industrial sweetpotato withincreased carotenoid contents by metabolic engineering ofcarotenoids biosynthesis. Orange gene (Or) was firstreported from cauliflower (Brassica oleracea cv. botrytis)regarding carotenoid accumulation in chromoplasts.Previously, we have reported the cloning and characterizationof Or gene from the storage roots of yellow-fleshedsweetpotato (cv. Shinhwangmi). In this study, we haveconstructed two splicing variants of Or gene by overlappingPCR. These constructs were transformed into sweetpotatocallus and Arabidopsis for constitutive expression underthe 35S promoter control. IbOr transgenic callus waseasily selected from its fine orange color while controlcallus showed faint yellow. We also analyzed carotenoidcontents in transgenic sweetpotato callus and compared withthat of control. Transgenic callus showed higher ß-carotenecontents than control. Furthermore, transgenic Arabidopsisseed coats (T2) were bright yellow color showing highercarotenoid production in transgenic plants. This resultindicates that IbOr would be applicable to the developmentof transgenic plants with high carotenoid contents bymetabolic engineering. (Joint Meeting of The BotanicalSociety of Korea and The Korean Society for PlantBiotechnology : Gwangju, 2009)

Keywards : sweetpotato, IbOr, carotenoid, -carotene,transgenic plants, metabolic engineering

C-49Development of transgenic sweetpotato plantsexpressing swpa4 peroxidase gene

Yun-Hee Kim, Il-Gin Mok, Haeng-Soon Lee, and Sang-Soo Kwak

Environmental Biotechnology Research Center, KRIBB

The secretory class III peroxidases (EC 1.11.1.7) have beenimplicated in a broad range of physiological processes,including defense against pathogenic attack and a varietyof abiotic stress tolerances (Cosio and Dunand 2009). Wepreviously characterized the 10 peroxidase cDNA clonesfrom cell cultures of sweetpotato (Ipomoea batatas) (Parket al. 2003; Jang et al. 2004; Kim et al. 2007). Among them,the expression of the swpa4 gene was profoundly inducedby a variety of abiotic stresses and pathogenic infections insweetpotato plant. Additionally we demonstrated that theoverexpression of apoplastic swpa4 showed tolerance to avariety of abiotic and biotic stresses (Kim et al. 2008). Inthis study, for a better understanding of the roles of swpa4,we are trying to generate swpa4 transgenic sweetpotato ina sense or antisense gene expression manner under the controlof CaMV 35S promoter. Kanamycin-resistant embryogeniccalli were selected on selection medium and normallyconverted into plantlets. Molecular characterization of

45 3rd KRIBB Poster Festival

transgenic plants are under investigation. The transgenicplants will be also characterized in terms of environmentalstress and agronomical characters including starch contents.(Joint Meeting of The Botanical Society of Korea andThe Korean Society for Plant Biotechnology : Gwangju,2009)

Keywords : environmental stress, peroxidase, sweetpotato

C-50Transgenic crops with enhanced tolerance tomultiple environmental stresses on marginallands

Yun-Hee Kim1, Wen-Bin Wang2, Xi-Ping Deng2, DaifuMa3, Il-Gin Mok1, Hang-Soon Lee1, and Sang-Soo Kwak1

1Environmental Biotechnology Research Center, KRIBB 2Institure of Soil and Water Conservation, CAS, Yangling,Shannxi, China

3Institute of Sweetpotato, CAAS, Xuzhou, Jiangsu, China

The crop with global crises over food and energy suppliesas well as environmental problems, it is urgently requiredto develop new crop varieties to be grown on marginallands including desertification areas for sustainableagriculture. In this respect, to generate transgenic cropssuch as potato (Solanum tuberasum L), sweetpotato(Ipomoea batatas) and alfalfa (Medicago sativaL.) plantswith enhanced tolerance to environmental stress, multiplestress-tolerant genes were introduced into plant genomeunder the control of an oxidative stress-inducible SWPA2promoter. Transgenic potato (cv. Superior) plants with theability to synthesize glycinebetaine (GB) in chloroplastsby introduction of the bacterial choline oxidase (codA)gene (referred to as SC potato plants) were characterized.The GB contents in the leaves of SC potato plants followingmethyl viologen (MV) treatment were found to be 1.43mmol/g fr wt by HPLC analysis. In addition to reducedmembrane damage after MV-mediated oxidative stress, theSC plants showed enhanced tolerance to NaCl and droughsstress on the whole plant level. Transgenic plants (potato,sweetpotato and alfalfa) expressing Arabidopsis nucleosidediphosphate kinase 2 (AtNDPK2) gene under the control ofSWPA2 promoter (SN plants) were generated. SN plantsshowed enhanced tolerance to multiple stress includingdroughs, high salt and extreme temperature. In addition,the strategies for the development of industrial transgeniccrops to combat desertification in China will be introducedin terms of global collaboration. (INTERDROUGHT-III: Shanghai (China), 2009)

Keywords : alfalfa, drought, oxidative stress, potato, saltstress, sweetpotato

C-51Differential expression of starch biosynthesis-related genes by exogenous sucrose in sweetpotatocultivars

Young Ock Ahn1, Sun Ha Kim1, Cha Young Kim1, Joon-Seol Lee2, Il-Gin Mok1, Sang-Soo Kwak1, and Haeng-SoonLee1*

1Environmental Biotechnology Research Center, KRIBB 2Bioenergy Crop Research Center, NICS, RDA

Three sweetpotato cultivars (Ipomoea batatas L. cv. Yulmi,Ym; cv. Sinhwangmi, Hm; cv. Sinzami, Zm) were investigatedfor their starch contents and amylose :amylopectin ratio.We also analyzed the expression levels of starch and sucrosebiosynthesis-related genes. All genes tested in thisexperiment were detected only in Ym, while several genesshowed very faint or no expression in Zm and Hm. Wealso measured tissue-specific expression of those genes inwhole plants of Ym. Most of the genes are expressed inthe stem and roots of the plants. Expression profiles ofstarch and/or sucrose synthesis-related genes of sweetpotatoleaves were investigated after supplementing the mediumwith different concentrations of sucrose solution. All genesin Ym were clearly induced by sucrose but the expressionlevels of some of those genes did not change in Zm andHm. Total starch contents of Ym, Zm, and Hm graduallyincreased over time on addition of 3%, 6%, and 9% sucroseconcentrations. The greatest accumulation was observed inYm at 48 h and it was almost 2.24 times higher than that of(0%) control, while Zm and Hm showed 1.76 and 1.91 timeshigher levels of starch, respectively. These results indicatethat cooperative expression of all related genes is essentialfor starch biosynthesis from sucrose. (Joint Meeting ofThe Botanical Society of Korea and The Korean Societyfor Plant Biotechnology : Gwangju, 2009)

Keywards : sweetpotato, starch, starch biosynthesis,sucrose feeding, sucrose

C-52Green biotechnology of natural rubber inplants

Eun-Ju Lee1, Hae Jin Kim1, Jeong Sheop Shin2, Jun YoungChoi2, Hunseung Kang3, Je Hwan Park4, Tae Kwon Jung4,and Stephen Beungtae Ryu1

1Environmental Biotechnology Research Center, KRIBB2Graduate School of Biotechnology, Korea University,3Department of Biotechnology, Chonnam NationalUniversity,

4Kumho Tire Co.

The Heveabrasiliensis tree is currently the only commercial

Korea Research Institute of Bioscience and Biotechnology 46

rubber producing Crop. Because of the poor geneticvariations, there is always a potential danger of crop failurewith diseases. Hevea also has another fundamental problemof life-threatening allergy caused by the proteins in the latex.It is, therefore, highly desirable to develop alternative rubbercrops that produce a high quality rubber without allergy.Most candidates for the alternative rubber crops have somedrawbacks such as relatively shorter length of rubber polymerand/or lower rubber production compared to the Hevea. Inorder to make them commercially viable rubber crops, it isrequired to either increase the size of rubber polymer andimprove rubber production. We propose to geneticallymanipulate the pathway of rubber biosynthesis by higroducingfunctional genes and/or by blocking the expression of certaingenes. We are investing the gene(s) that determine therubber polymer size or rubber production by transformingplants such as Russian dandelion and sunflower with potentialgenes. Ultimately we aim to apply the basic technology toan alternative target crop and produce high quality naturalrubber without allergy. (The 21st Annual Meeting of theKorean Society for Molecular and Cellular Biology :Seoul, 2009)

Keywords : natural rubber, green biotech, bio-materials

Members

DirectorHee-Mock Oh (heemock@kribb.re.kr)

Principal Researcher Sang-Soo Kwak (sskwak@kribb.re.kr)Won-Gon Kim (wgkim@kribb.re.kr)Hee-Sik Kim (hkim@kribb.re.kr)Beung-Tae Ryu (sbryu@kribb.re.kr)Haeng-Soon Lee (hslee@kribb.re.kr)

Senior Researcher Chi-Yong Ahn (cyahn@kribb.re.kr)Jae-Cheol Jeong (jcjeong@kribb.re.kr)

Post-DocMyoung-Duck Kim (dorrf@kribb.re.kr)Yun-Hee Kim (cefle@kribb.re.kr)Young Ock Ahn (yoahn@kribb.re.kr)Young-Ki Lee (8896lyk@hanmail.net)Eun-Ju Lee (zoor@hanmail.net)So-Young Jun (sy6921@hanmail.net)

ResearcherDong-Geol Kim(kdg09@kribb.re.kr)Byung-Hyuk Kim (tiger417@kribb.re.kr)Dae-Hyun Cho (ryuwoon@kribb.re.kr)Chan Yoo (brightchan@naver.com)Eun-Kyung Kim (ekkim@kribb.re.kr)Jae-Yon Lee (jylee@kribb.re.kr)Yun-Ju Kwon (mars005@daum.net)Jun-Young Cho (jjyoung3542@nate.com)

Nyung Kim (rlanyoung@naver.com)Jung-Ahn Kim Jung-Mae Liu (liujung43@naver.com)Sun-Ha Kim (sunha82@kribb.re.kr)Sung-Chul Park (heypsc@empal.com)Yoo-Sun Seo (sunny3646@hanmail.net)Hae-Jin Kim (bioen97@korea.ac.kr)Krishna Kumar (krishnakr.agri@gmail.com)

47 3rd KRIBB Poster Festival

Division of Bio R&D Infra

Korean Bioinformation Center

Biological Resource Center

Biotechnology Process Engineering Center

3 r d K R I B B P o s t e r F e s t i v a l

49 3rd KRIBB Poster Festival

D-1MitoInteractome : Mitochondrial proteininteractome database, and its application in‘aging network’ analysis

Rohit Reja1,2,*, Venkatakrishnan A.J.1,*, Jungwoo Lee3,Byoung-Chul Kim1, Jea-Woon Ryu1, Sungsam Gong4,Jong Bhak1,2, and Daeui Park1

1Korean Bioinformation Center, KRIBB2Bioinformatics Department, University of Science andTechnology (UST)

3Scotch College, Australia4Department of Biochemistry, University of Cambridge, UK.

Mitochondria play a vital role in the energy productionand apoptotic process of eukaryotic cells. Proteins in themitochondria are encoded by nuclear and mitochondrialgenes. Owing to a large increase in the number of identifiedmitochondrial protein sequences and completed mitochondrialgenomes, it has become necessary to provide a web-baseddatabase of mitochondrial protein information. We present‘MitoInteractome’, a consolidated web-based portalcontaining a wealth of information on predicted protein-protein interactions, physico-chemical properties,polymorphism, and diseases related to the mitochondrialproteome. MitoInteractome contains 6,549 protein sequenceswhich were extracted from the following databases: SwissProt,MitoP, MitoProteome, HPRD and Gene Ontology database.The first general mitochondrial interactome has beenconstructed based on the concept of ‘homologous interaction’using PSIMAP (Protein Structural Interactome MAP) andPEIMAP (Protein Experimental Interactome MAP). Usingthe above mentioned methods, protein-protein interactionswere predicted for 74 species. The mitochondrial proteininteraction data of humans was used to construct a networkfor the aging process. Analysis of the ‘aging network’ gave usvital insights into the interactions among proteins thatinfluence the aging process. MitoInteractome is a comprehensivedatabase that would (1) aid in increasing our understandingof the molecular functions and interaction networks ofmitochondrial proteins, (2) help in identifying new targetproteins for experimental research using predicted protein-protein interaction information, and (3) help in identifyingbiomarkers for diagnosis and new molecular targets fordrug development related to mitochondria. MitoInteractomeis available at http://mitointeractome.kobic.kr/. (BioInfo2009:CBI-KSBSB Joint International Symposium The 62thAnnual Meeting of the Korean Association of BiologicalSciences : Busan, 2009)

Keywords : protein-protein interaction, database,mitochondria, aging

Members

Director Joon-Ki Jung (jkjung@kribb.re.kr)

Senior Researcher Byung-Uk Lee (bulee@kribb.re.kr)Sung-Hoon Lee (ishoon@kribb.re.kr)Bo-Kyung Hou (bkher71@kribb.re.kr)Sung-Woo Hwang (swhwang@kribb.re.kr)Sung-Soo Kang (suskang@kribb.re.kr)Jin-Ok Yang (joy@kribb.re.kr)

Post-Doc Chul-Hong Kim (chulhong@kribb.re.kr)Phan-Kyu Kim (pgkim@kribb.re.kr)Young-Seok Lee (dolsemtl@kribb.re.kr)Jung-Hee Lim (jhlim@kribb.re.kr)

ResearcherHo-Young Kang (kangho@kribb.re.kr)Gun-Hwan Ko (koguns@kribb.re.kr)Yu-Jin Kwak (who321@kribb.re.kr)Je-Keun Kwon (kwon@kribb.re.kr)Sung-Jun Kim (sjkim84@kribb.re.kr)Hank-Min Kim (howmany2@kribb.re.kr)Rohit Reja (rohit@kribb.re.kr)Je-Un Ryu (jwryu@kribb.re.kr)Sung-Jin Park (psj420@kribb.re.kr)Hyun-Chul Park (pgkim@kribb.re.kr)Ik-Soo Byun (isbyeon@kribb.re.kr)Hana Byun (habyun@kribb.re.kr)Sung-Woo Yang (josh617@kribb.re.kr)Sang-Ho Oh (sangho@kribb.re.kr)Nam-Hee Yu (namhee@kribb.re.kr)Hye-Sook Yoo (hsyoo@kribb.re.kr)Kyung-A Lee (kalee@kribb.re.kr)Lang-Ho Lee (lhlee@kribb.re.kr)Jae-Kyung Jun (roachq@kribb.re.kr)Dong-Soo Jung (dsjung@kribb.re.kr)Sung-Wung Cho (sucho@kribb.re.kr)Su-An Cho (chosuan@kribb.re.kr)Kyung-Suk Choi (choi@kribb.re.kr)Tae-Hui Hong (taehui@kribb.re.kr)Ji-Min Hwang (jhwang@kribb.re.kr)Muhammad Ilyas Zulfiqur Ali Mir

Korean Bioinformation Center

Korea Research Institute of Bioscience and Biotechnology 50

D-2Salt-tolerance genes as selectable marker forgenetic plant transformation of duckweed

Myung Jin Oh2, Jong Hyun Kim2, Myung Suk Ahn2, JongMi Park1, Jang Ryol Liu2, and Suk Weon Kim1

1Biological Resource Center, KRIBB2Plant System Research Center, KRIBB

An antibiotic resistance gene is a commonly used selectionmarker. However, the spread of antibiotic resistant pathogensis a serious worldwide problem. Therefore the antibioticresistance genes cannot have extensive use in thecommercialization. Alternatively, antibiotic-free selectionsystems have been suggested. Salt-tolerance gene sll-0170,Slr-0813, STL-slr0746, and STL-slr1894 were identifiedfrom chip data of Synechocystis PCC6803. For Salt-tolerancetest, these genes were introduced into DH5a harboring thevector TIA-Rclp GAH. Cell were cultured in LB brothcontaining of 1%, 5%, 6% NaCl. Growth rate was determinateby measuring the optical density of suspended cells at600nm. Growth rate of sll0170 was 68%, 29% in LB brothcontaining 5%, 6% NaCl, respectively. Growth rate ofsll0746, sll0813 gene were observed above 75%, 30% inLB broth containing 5%, 6% NaCl, respectively. Growthof control cell was not apparent in 6% concentration ofNaCl. It was determined a suitable selective pressure ofNaCl concentrations with fronds of duckweed. Growth ofduckweed fronds did not occur in the presence of 150 mMconcentration of NaCl. Fronds of duckweed were transformedwith Agrobacterium tumefecience(GV3101) harbored withsalt-tolerance gene sll-0170, Slr-0813, STL-slr0746, andSTL-slr1894 in independent vectors. Putative transgeniclines were selected by 1/2 MS medium supplemented with1mg/L BA and 150mM NaCl. Transgenic lines were growthin medium containing 150mM NaCl, but non-transgeniclines were chlorosis and necritized. Transformation wasconfirmed by PCR analysis. Salt-tolerance gene could useas the effective, and safe tool for transgenic plant selection.(Joint Meeting of The Botanical Society of Korea andThe Korean Society for Plant Biotechnology : Gwangju,2009)

Keywords : antibiotics selection, duckweed, salt tolerence,Synechocystis PCC6803, transformation.

D-3Arabidopsis mutants defective in anthocyaninbiosynthesis are arranged in a cascade mannerin the multivariate analysis score plot of theirFT-IR spectroscopic profiles

Jong Hyun Kim1, Suk Weon Kim2, Myung Suk Ahn1, andJang Ryol Liu1

1Plant System Research Center, KRIBB2Biological Resource Center, KRIBB

To develop a system for screening out mutants with genesor with deleted genes that affect a common biosyntheticpathway by metabolic profiling, we analyzed FT-IR profilesof knockout mutants for genes found in anthocyaninbiosynthesis. When the profiles were analyzed usingmultivariate analysis, these mutants were grouped in thesame cluster. Furthermore, these mutants were arranged ina cascade manner corresponded to the order of enzymescatalyzing from early intermediates to late intermediateswithin the cluster on the linear discriminant analysis scoreplot. Therefore, it was expected that mutants for unknowngenes located within the cluster could be with genes orwith deleted genes that affect anthocyanin biosyntheticpathway. When FT-IR profiles of approximately 1,050Arabidopsis activation-tagged mutants were analyzed usingmultivariate analysis, some mutant lines were screened outthose were obviously located within the cluster of anthocyaninbiosynthetic knock-out mutants. We finally choose threeactivation-tagged mutants based on the analyzing 1H-NMRand anthocyanin contents. Those mutants showed adramatically reduced gene expression level of regulatorygenes for anthocyanin biosynthesis than wild type usingqRT-PCR. Overall results suggest that dispersed mutantson the score plot represent a networking relationship amongmutants at the metabolome level. (Joint Meeting of TheBotanical Society of Korea and The Korean Society forPlant Biotechnology : Gwangju, 2009)

Keywords : anthocyanin biosynthesis, Arabidopsis,discriminant analysis, FT-IR spectroscopy

D-4Salipiger sp. nov., a moderately halophilicbacterium isolated from a solar saltern

Jae-Chan Lee1, Yu-Qin Zhang2, Dong-Jin Park1, andChang-Jin Kim1

1Biological Resource Center, KRIBB2Peking Union Medical College, Beijing, China

Salipiger sp. nov. is a moderately halophilic, Gram-negativerod isolated from a solar saltern in south Korea. The bacteriumis strictly aerobic and motile. It is catalase negative andphosphatase positive. It does not produce acids fromcarbohydrates. It cannot grow with carbohydrates or aminoacids as sole sources of carbon and energy. It grows best at5–12 % w/v NaCl and requires the presence of Na+. Itsmajor fatty-acid component is C18:1 w7c (59.9%). Thepredominant respiratory lipoquinone found in the strain isubiquinone with ten isoprene units. The G+C content is 60mol%. Phylogenetic analyses strongly indicate that this

Biological Resource Center

strain forms a distinct line within a clade containing thegenus Salipiger. The similarity value with Salipiger mucosusis 95.6 %. On the basis of the polyphasic evidence gatheredin this study it is proposed that the isolate a new species,Salipiger sp. nov. The proposed type strain is strain BH028T

(=KCTC 22651). (International Symposium & AnnualMeeting of the Korean Society for Microbiology andBiotechnology : Daejeon, 2009)

Keywords : Salipiger sp. nov. phylogeny, taxonomy,moderate halophile

D-5Roseivivax sp. nov., isolated from a solar saltern

Jae-Chan Lee1, Yu-Qin Zhang2, Dong-Jin Park1, andChang-Jin Kim1

1Biological Resource Center, KRIBB2Peking Union Medical College, Beijing, China

Phenotypic and phylogenetic studies were performed withstrain (BH21) of aerobic bacteria isolated from a solarsaltern located on the west coast of Korea. The strain wasGram-negative and motile rods with subpolar flagella.Catalase and oxidase were produced. ONPG reaction waspositive. The strain could grow in 0-15% (w/v) NaCl and5-8% NaCI was optimum. The major cellular fatty acidswere C18:1 w7c and C19:0 cyclo w8c. The results of 16SrRNA sequence comparisons revealed that strain BH21formed a new phyletic line in the genus Roseivivax of theclass Proteobactera. The similarity value of the 16S rRNAsequences between strain BH21 and Roseivivax halotoleransand Roseivivax halodurans were 94.7% and 94.3%,respectively. Therefore, it was concluded that the strainshould be placed in a genus Roseivivax as a new species.The type strain is BH21 (=KCTC 22650). (InternationalSymposium & Annual Meeting of the Korean Societyfor Microbiology and Biotechnology : Daejeon, 2009)

Keywords : Roseivivax sp. nov. phylogeny, taxonomy,identification

D-6Nocardioides panacisoli sp. nov. isolated fromthe soil of a ginseng field in South Korea

Song-gun, Kim, Dong-Shan An, and Tae Woong Whon

Biological Resource Center, KRIBB

A Gram-positive, rod-shaped, non-spore-forming bacterium(GSoil 346T) was isolated from the soil of a ginseng field

in South Korea. GSoil 346T was shown to belong to thegenus Nocardioides in the family Nocardioidaceae, withthe most closely related species being Nocardioides aquiterrae(96.6%); however, it comprised a distinct branch of thephylogenetic tree together with N. kongjuensis (96.2%),N.aromaticivorans (96.1%), N. nitrophenolicus (96.1%),and N. simplex (95.9%). GSoil 346T was characterizedchemotaxonomically as having LL-2,6-diaminopimelicacid in the cell wall peptidoglycan, phosphatidylinositoland phosphatidylglycerol as the major polar lipids, MK-8(H4) as the predominant menaquinone, and iso-C16:0,C17:1 8c, and C18:1 9c as the major fatty acids. TheG+C content of its genomic DNA was 73.0 mol%. Theresults of physiological and biochemical tests along withthe phylogenetic analysis allowed us to differentiate strainGSoil 346T genotypically and phenotypically from previouslypublished Nocardioides species. Therefore, strain GSoil346T represents a novel species, for which the nameNocardioides panacisoli sp. nov. is proposed, with GSoil346T as the type strain (= KCTC 19470T = DSM 21348T).(MSK’s 50th Anniversary International Symposium onMicrobiology : Jeju, 2009)

Keywords : Nocardioides panacisoli sp. nov., taxonomy

D-7Homologous overexpression of omcZ, a geneencoding outer surface c-type cytochrome,critical for high-dense electricity generation inGeobacter sulfurreducens by single-step genereplacement

Byoung-Chan Kim1,2* and Jung-Sook Lee2

1Department of Microbiology, University of Massachusetts,USA

2Biological Resource Center, KRIBB

Dissimilatory Fe(III)-reducing Geobacter sulfurreducenscan obtain energy by coupling the oxidation of organiccompounds to the reduction of several soluble and insolubleelectron acceptors including Fe(III) oxide, uranium, andelectrode. Therefore, G. sulfurreducnes can play animportant role both in the bioremediation of radionuclidesand toxic metals and in biological electricity production.G. sulfurreducens has been one of the most intensivelystudied microorganisms to date because it produces highcurrent densities, the complete genome sequence of thisorganism is available, and a genetic manipulation systemis available. OmcZ is one of the major outer surface c-typecytochromes of G. sulfurreducens critical for electricitygeneration in the poised flow-through microbial fuel cells.In order to make stable overexpression system for OmcZ,recombinant PCR product containing ompJ promoter of G.sulfurreducens and omcZ coding sequence was combined

51 3rd KRIBB Poster Festival

Korea Research Institute of Bioscience and Biotechnology 52

and integrated into the chromosome of wild type G.sulfurreducens by homologous DNA recombination. Thequantitative RT-PCR analysis showed that the amount ofomcZ transcript in the knock-in strain was 6-fold moreabundant than in wild type. The stable expression of OmcZwithout selective pressure by antibiotics was observed.Notably, higher rate of electricity generation on initial stepwas observed, indicating genetic engineering might aid inthe optimization and improvement of electricity productionby G. sulfurreducens. (International Meeting of theFederation of Korean Microbiological Societies : Seoul,2009)

Keywords : Geobacter sulfurreducens, microbial fuel cell,outer surface cytochrome, electricity generation

D-8Paenibacillus pini sp. nov., a cellulolytic isolatefrom the rhizosphere of a pine tree

Byung-Chun Kim, Kang Hyun Lee, Mi Na Kim, Eun-MiKim, Sun Beom Kwon, and Kee-Sun Shin*

Biological Resource Center, KRIBB

A novel cellulolytic bacterium, strain S22T, was isolatedfrom the rhizosphere of a pine tree in Korea. This isolatewas characterized taxonomically by using phenotypic andphylogenetic study. S22T was Gram-positive, endospore-forming, and motile rods that grew at 10-30ºC, optimallyat 25ºC, and pH 5.5-7.5, optimally at pH 7.2. This isolaterepresented positive activity for catalase, oxidase, esteraselipase, leucine arylamidase, ß-galactosidase, N-acetyl-ß-glucosaminidase, urease, and hydrolysis of esculin. Themajor cellular fatty acid profiles were anteiso-C15:0 (58.4%),iso-C15:0 (12.3%), and iso-C16:0 (8.2%). The predominantisoprenoid quinone was menaquinone 7 (MK-7), andmeso- diaminopimelic acid was contained in the cell-wallpeptidoglycan. Phylogenetic analysis based on 16S rRNAin the sequences showed that this isolate belong to thePaenibacillaceae. Strain S22T represented less than 97.0%16S rRNA in the similarity with all relative type strains inthe genus Paenibacillus, and most closely related strainwas Paenibacillus anaericanus MH21T and Paenibacillusginsengisoli Gsoil 1638T, with a similarity of 95.79%. Basedon the phylogenetic, chemotaxonomic and phenotypiccharacteristics, strain S22T is considered to represent anovel species in the genus Paenibacillus, for which thename Paenibacillus pini sp. nov. is suggested. The typespecies is strain S22T (= KCTC 13694T = KACC 14198T).(International Meeting of the Federation of KoreanMicrobiological Societies : Seoul, 2009)

Keywords : Paenibacillus pini sp. nov. novel cellulolyticbacterium, taxonomy

D-9Cohnella gyejoki sp. nov., xylanolytic bacteriaisolated from the rhizosphere

Byung-Chun Kim, Kang Hyun Lee, Mi Na Kim, Eun-MiKim, Sun Beom Kwon, and Kee-Sun Shin*

Biological Resource Center, KRIBB

Two novel xylanolytic bacteria, strain S1 and S15T, wereisolated during the study of culturable heterotrophic bacteriain rhizosphere. Strain S1 and S15T were Gram-positive,motile, rod-shaped bacteria and have a terminal or subterminalellipsoidal spore. These isolates grew at 15-40ºC, optimumat 30ºC, at pH 5.5-8.0, optimum at pH 7.5, and in thepresence of 1% (w/v) NaCl. These isolates were positivefor oxidase, acid phosphatase, alkalin phosphatase, esterase,esterase lipase, -galactosidase, ß-galactosidase, and ß-glucosidase. MK-7 was the predorminat isoprenoid quinone.Anteiso-C15:0 and iso-C15:0 were the major cellular fattyacids. meso-Diaminopimelic acid was contained in the cell-wall peptidoglycan. 16S rRNA gene sequences analysisrepresented that the S1 and S15T shared 99.46% sequencessimilarity and were affiliated with a cluster within thefamily Paenibacillaceae. The type strains that werer mostclosely related to S1 and S15T was Cohnella laevire osiRI-39T (95.67-95.74%) and then Cohnella thermotoleransCCUG 47242T (95.21-95.34%) and Cohnella hongkongensisHKU3T (94.48-94.78%). The polyphasic characteristics ofthe isolates allowed these two isolates to be clearlydistinguished from other Cohnella species. On basis ofthese data, the isolates are considered to represent a noveltaxon, for which the name Cohnella gyejoki sp. nov. isproposed. The type strain S15T. (International Meeting ofthe Federation of Korean Microbiological Societies :Seoul, 2009)

Keywords : Cohnella gyejoki sp. nov. novel xylanolyticbacteria, taxonomy

D-10Recognition of the ascomycetous yeast Candidaacaciaensis sp. nov., as a distinct species basedon polyphasic taxonomic study

Kee-Sun Shin1, Soon Gyu Hong2, Kyung Sook Bae1, KangHyun Lee1, Sun Beom Kwon1, Mina Kim1, and Moon-SooRhee1

1Biological Resource Center, KRIBB2Korea Polar Research Institute (KOPRI)

An undescribed anamorphic yeast strain of ascomycetousaffinity was isolated from Rhizosphere soil of Acacia sp.The taxonomic position of the yeast strain SG02-69T was

examined. Sequence analysis of D1/D2 variable domain ofthe large subunit rRNA cording gene as well as physiologicaland chemosystematic studies on strain SG02-69T placed itunder the genus Candida, including Candida valdivianaNRRL Y-7791T, Candida sp. NRRL YB-1835T and Candidasp. NRRL YB-3827T. However, there were several differencesin some physiological characteristics and 26S partialsequence, which are making the strain recognizable fromother Candida species. The results of the present studyindicated that strain SG02-69T should be placed under thegenus Candida as a new species. The type strain of the newspecies is strain SG02-69T (= KCTC 17837T). (MSK’s 50thAnniversary International Symposium on Microbiology: Jeju, 2009)

Keywords : Candida acaciaensis sp. nov. novel yeastspecies, taxonomy

D-11Candida rhododendronensis sp. nov., a novelmember of the Metschnikowiaceae

Kee-Sun Shin, Kang Hyun Lee, Sun Beom Kwon, and Mina Kim

Biological Resource Center, KRIBB

A yeast strain isolated from Rhododendronsp. was studiedto determine its taxonomic position. The strain wascharacterized by budding cells, negative diazonium blue B(DBB) and urease reactions and the presence of Q-9 as themajor ubiquinone isoprenologue. A phylogenetic analysisof the D1/D2 region of the 26S rRNA gene placed strain07-Y5T within the Metschnikowiaceae. The results of apolyphasic taxonomic study and phylogenetic analysisindicated that the strain belonged to an described speciesof the genus Candida. As such, the strain was designated anovel species with the name Candida rhododendronensissp. nov. and type strain 07-Y5T(= KCTC17838T). (MSK’s50th Anniversary International Symposium onMicrobiology : Jeju, 2009)

Keywords : Candida rhododendronensis sp. nov. novelyeast species, taxonomy

D-12Paenibacillus panacisegetis sp. nob., isolatedfrom soil of a ginseng field

Jung-Sook Lee, Keun Chul Lee, and Kwang Kyu Kim

Biological Resource Center, KRIBB

Ginseng is very famous herb in the medicinal plants.

Especially, Korean ginseng is known for the excellent propertiesfor helping good health. We investigated the microbialcommunity diversity of the soil in the ginseng field, isolatednovel bacterial species, and characterized it by polyphasicapproach. We carried out the studies of the 16S rRNA genesequences comparison and DNA-DNA hybridization,analyses of chemotaxonomic characteristics including fattyacid profiles, G+C contents, isoprenoid quinines, and polarlipids, and phenotypic characteristics. A novel Gram-positive,spore-forming rod-shaped bacterial strain, P11-6T,wasisolated from soil of a ginseng field, Korea. The straincontained MK-7 as the dominant menaquinone and anteiso-C15:0 and C16:0 as the major fatty acids. Phylogeneticanalysis, based on 16S rRNA gene sequencing, showedthat the isolate 11-6T was most closely related to the genusPaenibacillus, Paenibacillus ginsengisoli (with 96%sequence similarity). On the basis of combined phenotypic,chemotaxonomic and phylogenetic data, it is proposed thatstrain P11-6T represents a novel species, Paenibacilluspanacisegetis sp. nov. The type strain is P11-6T (=KCTC13564T).(FEMS Congresses of European Microbiologists :Gothenburg (Sweden), 2009)

Keywords : Paenibacillus ginsengisoli, taxonomy,ginseng

D-13A novel bacterium, Patulibacter ginsengiterraesp. nov., isolated from soil of ginseng

Jung-Sook Lee, Keun Chul Lee, Kwang Kyu Kim, MiJeong Kim, and Mi Kyung Eom

Biological Resource Center, KRIBB

A novel Gram-positive, short-rod bacterial strain, P4-5T,was isolated from soil of a ginseng field, Korea. The strainhad glucose, galactose, mannose, arabinose and rhamnoseas whole-cell sugar, and meso-diamino pimelic acid as thediagnostic diamino acid in the peptidoglycan. It containedC18:1 w9c, summed feature7 (C19:0 cyclow 10c/ 19w6)and C18:0 as the predominant cellular fatty acids and DNAG+C content of 75.0 mol%. Phylogenetic analysis, basedon 16S rRNA gene sequencing, showed that the isolateP4-7T was most closely related to the genus Patulibacter,Patulibacter minatonensis (with 97.2 % sequencesimilarity). On the basis of combined phenotypic,chemotaxonomic and phylogenetic data, it is proposed thatstrain P4-7T represents a new genus and a novels pecies,Patulibacter ginsengiterrae sp. nov. The type strain is P4-5T(=KCTC19427T). (BIEN 2009 : INWES AsianNetwork : Busan, 2009)

Keywords : Patulibacter ginsengiterrae, taxonomy,ginseng

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Korea Research Institute of Bioscience and Biotechnology 54

D-14A novel bacterium, Solirhabdus panacisegetisgen. nov., sp. nov., isolated from soil of ginsengfield

Jung-Sook Lee, Keun Chul Lee, Kwang Kyu Kim, MiJeong Kim, and Mi Kyung Eom

Biological Resource Center, KRIBB

A novel Gram-positive, short-rod bacterial strain, P4-7T,was isolated from soil of a ginseng field, Korea. The strainhad MK-8(H4) as the major menaquinone, glucose,mannose, xylose and rhamnose as whole-cell sugar,diphosphatidylglycerol as major polar lipid and meso-diaminopimelic acid as the diagnostic diamino acid in thepeptidoglycan. It contained iso-C16:0 and anteiso-C15:0as the predominant cellular fatty acids and DNA G+Ccontent of 70.2 mol%. Phylogenetic analysis, based on16S rRNA gene sequencing, showed that the isolate P4-7T

was most closely related to the genus Humicoccus,Humicoccus flavidus (with 97.0 % sequence similarity).However, strain P4-7T showed distinct differences fromHumicoccus flavidus in content of the major menaquinone,cell-wall sugar, major polar lipid and fatty acid profile. Onthe basis of combined phenotypic, chemotaxonomic andphylogenetic data, it is proposed that strain P4-7T representsa new genus and a novel species, Solirhabdus panacisegetisgen. nov., sp. nov. The type strain is P4-7T (= KCTC 19426T).(International Symposium & Annual Meeting of theKorean Society for Microbiology and Biotechnology :Daejeon, 2009)

Keywords : Patulibacter ginsengiterrae, taxonomy,ginseng

D-15A polyphasic investigation on a novel halophilicarchaeon isolated from sea salt

Kwang Kyu Kim, Keun Chul Lee, and Jung-Sook Lee

Biological Resource Center, KRIBB

Three halophilic isolates, strains B1T, B3 and B4, wereisolated from an evaporitic salt crystal from Namhae, Korea.Cells of the strains were Gram-negative, non-motile andpleomorphic (mainly oval-shaped), and formed circular,slightly convex, semi-translucent, red-pigmented colonies.The three strains grew at NaCl concentrations of 1.7-5.1 M(optimum at 3.4 M NaCl) and required at least 10 mMMg2+ for growth. They were able to grow over a pH rangeof 5.0-8.0 and a temperature range of 15-50ºC, with optimalpH of 7.0 and optimal temperature of 42ºC. The majorpolar lipids of the strains were phosphatidylglycerol,

phosphatidylglycerophosphate methyl ester and twoglycolipids (S-DGD-1 and DGD-1). On the basis of 16SrRNA gene sequence comparisons, strains B1T, B3 and B4,which shared 100 % similarity, showed the highest similarityto Halogranum rubrum RO2-11T (99.3 %). However, DNA-DNA relatedness data and their phenotypic propertiessupported that they represent a novel species of the genusHalogranum, for which the name Halogranum namhaensesp. nov. is proposed; the type strain is B1T (= KCTC 4066T).(International Meeting of the Federation of KoreanMicrobiological Societies : Seoul, 2009)

Keywords : halophilic archaeon, halogranum

D-16A tRNA based primer cocktail for mitochondrialCOI barcoding of hexapoda

Park, D.-S.1,2, Hebert, P.D.N.2

1Biological Resource Center, KRIBB2Biodiversity Institute of Ontario, University of Guelph,Canada

DNA barcoding uses a 650 bp segment of the mitochondrialcytochrome c oxidase I (COI) gene as the basis for anidentification system for members of the animal kingdomand some other groups of eukaryotes. PCR amplificationof the barcode region is a key step in the analytical chain,but it sometimes fails because of a lack of homologybetween standard primer sets and target DNA. In thisstudy we describe a primer design strategy which enabledresolution of this problem for the most diverse group ofanimals - the Hexapoda. We began by analyzing the genearrangements for all arthropod mitochondrial genomes inthe Mitom database. This analysis revealed that the tRNA-W gene was invariably located within 200 bp upstream ofCO1 gene, while sequence alignment showed two distinctgroups of tRNA-W with high internal homogeneity. Twoforward PCR primers were developed to best representthese two groups and were combined with a standardreverse primer (LepR1) to produce a cocktail which wastested against 141 specbes representing 126 differentfamilies of Hexapoda, including groups such as scale insectsthat invariably fail to amplify with standard primers. Thiscocktail generated 111 amplicons, allowing characterizationof the usual primer binding region in COI 5' as well as thebarcode segment. These cases of success included groupssuch as the scale insects whose unusual nucleotide sequenceat the regular forward primer binding site provide anexplanation for past failures in amplification. Laterperformance tests on more taxa revealed that the newcocktail amplifies CO1 for nearly all hexapods which failto amplify with regular barcode primers. We suggest thatthe design of primers that bind in highly conserved gene

regions upstream of COI will be a useful strategy inresolving problems in amplification. (Third InternationalBarcode of Life Conference : Mexico City (Mexico),2009)

Keywords : COI barcoding, PCR primers, hexapoda,tRNATrp

D-17DNA barcoding the hemiptera

Park, D.-S.1,3, Foottit, R.2, Maw, E.2, Hebert, P.D.N.3

1Biological Resource Center, KRIBB2CNC of Insects, Arachnids and Nematodes, Agricultureand Agri-Food Canada

3Biodiversity Institute of Ontario, University of Guelph,Canada

Although DNA barcode coverage has grown rapidly forsome insect orders, prior work on the Hemiptera hasfocused on just two families (Aphididae, Adelgidae) of theapproximately 160 families belonging to this order. Thisstudy sought to broaden coverage, largely through theanalysis of material in museum collections. DNA wasextracted from 1700 specimens with an average age of12.2 years. Barcode records over 500 bp were recoveredfrom 1173 of these specimens providing coverage for 374species representing 33 families and 190 genera. Sequencesdivergences (K2P distance) between congeneric speciesaveraged 9.99%, a value which was 11-fold higher thanthe mean intra-specific variation. Moreover, the intra-specific divergence value was likely inflated by thepresence of species overlooked by current taxonomictreatments because several ‘species’ showed divergencesgreater than 2.0%. Other clusters included individualsassigned to several species, suggesting the need for moredetailed morphological and molecular studies. Takencollectively, the present results suggest both the feasibilityof creating a comprehensive barcode library for theHemiptera and its value in both revealing taxonomicsituations worthy of deeper analysis and in creating aneffective system for identifying species in this group.(Third International Barcode of Life Conference :Mexico City (Mexico), 2009)

Keywords : COI barcoding, hemiptera

Members

DirectorJung-Sook Lee (jslee@kribb.re.kr)

Principal Researcher Chang-Jin Kim(changjin@kribb.re.kr)Kyung-Sook Bae (ksbae@kribb.re.kr)

Senior Researcher Suk-Weon Kim (kimsw@kribb.re.kr)Song-Gun Kim (sgkim@kribb.re.kr)Doo-Sang Park (dspark@kribb.re.kr)Kee-Sun Shin (ksshin@kribb.re.kr)Byoung-Chan Kim (bckim@kribb.re.kr)Young-Hyo Chang (yhchang@kribb.re.kr)

Post-DocJin-Hee Choi (jhchoi77@kribb.re.kr)Jae-Chan Lee (jcle777@kribb.re.kr)Dong-Shan An (dsan1122@kribb.re.kr)Kwang Kyu Kim (kkkim@kribb.re.kr)Byung-Chun Kim (kimbc@kribb.re.kr)

Researcher Dong-Jin Park (dongjin@kribb.re.kr)In-Soon Park (ispark@kribb.re.kr)Kang-Hyun Lee (khlee@kribb.re.kr)Keun-Chul Lee (kclee@kribb.re.kr)Moon-Soo Lee (mslee@kribb.re.kr)Yong-Jae Lee (tmx@kribb.re.kr)Jong-Ok Jang, (olliber6@kribb.re.kr)Hyeon-Seong Yoon (hgyoon@kribb.re.kr)Pan-Kyung Kim (pkjin@kribb.re.kr)Young-Hee Kim (kyhee@kribb.re.kr)Geun-Hye Lee (leegh78@kribb.re.kr)Jong-Mi Park (pjm335@kribb.re.kr)Chang-Hee Jung (tanksboy@kribb.re.kr)Hyang-Mi Kim (hyang0706@nate.com)Mi-Jeong Kim (midali424@nate.com)Mi-Kyung Eom (alrodrl@kribb.re.kr)Mi-Na Kim (z8k4@kribb.re.kr)Kyung-Im Kang (kikang78@kribb.re.kr)Tae-Woong Whon (twwhon@kribb.re.kr)Min-Young Jung (myjung@kribb.re.kr)Yong-Kook Kwon (ykkwon@kribb.re.kr)Hye-Young Kang (khy5460@kribb.re.kr)Jeong-Ran Han (hanjrjr@kribb.re.kr)Keun-Ae Park, (keunae6@kribb.re.kr)Ji-Hye Kim (kim0424@kribb.re.kr)Eun-Mi Kim (kem82@kribb.re.kr)Dan-Oh Gu (danoh@kribb.re.kr)

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Korea Research Institute of Bioscience and Biotechnology 56

D-18Production of amino butyric acid by Lactobacillussp. isolated from traditional Korean fermentedfoods

Dong-Soo Kim2, Hae-Dong Jang2, Young-In Kwon2, EunGyo Lee1, Hong-Won Lee1, and Dae-Kun Choi3

1Biotechnology Process Engineering Center, KRIBB2Department of Food and Nutrition, Hannam University3R&D Center, CTC Bio

-Aminobutyric acid (GABA) known as a non-proteinamino acid, acts in the central nervous system as a post-synaptic inhibitory neurotransmitter in the brain, increasesthe flow of blood to the brain in human/animals, andenhances the metabolism of cells in brain by stimulatingthe oxygen supply. GABA is produced primarily by aglutamate decarboxylase in the cell, which catalyses theirreversible decarboxylation of L-glutamate to GABA. Inthis study, we isolated 690 strains of lactic acid bacteria(LAB) from kimchi, yoghurt, soy pastes and fish pastes.Selected 10 strains were cultured in a MRS mediumcontaining 5% and 10% (w/v) monosodium glutamate(MSG) by incubating at 30ºC for 48 hours. The strainsgrown in the medium with high concentration of MSGwere cultured in the liquid medium containing MSG andmeasured the productivity of GABA using both thin layerchromatography (TLC) and HPLC methods. The GABAyield reached maximum after 12 h. Furthermore, thebiotransformation conditions of MSG to GABA wereoptimized with the cheaper industrial medium instead ofMRS. Under the optimized culture conditions, the yield ofGABA reached over 20 g/L with MSG precursor and2.282 mg/L without MSG, respectively. (InternationalSymposium & Annual Meeting of the Korean Societyfor Microbiology and Biotechnology : Daejeon, 2009)

Keywords : aminobutyric acid, lactobacillus bacteria,glutamate decarboxylase

Members

DirectorHong-Weon Lee (hwlee@kribb.re.kr)

Principal Researcher Joon-Ki Jung (jkjung@kribb.re.kr)Eun-Gyo Lee (eglee@kribb.re.kr)

Senior Researcher Jung-Oh Ahn (ahnjo@kribb.re.kr)

Post-DocMin-Hee Hong (hee1023@kribb.re.kr)Yeon-Gu Kim (ygkim@kribb.re.kr)

ResearcherChun-Sug Kim (chskim@kribb.re.kr)Hyeok-Won Lee (tntn7616@kribb.re.kr)Joo-Hwan Lee (jhlee@kribb.re.kr)Jin-Ho Lee (jhcholgo@kribb.re.kr)Jong-Woon Jeon (confess@kribb.re.kr)Ji-Myung Ryu (jmryu@kribb.re.kr)Seung-Hui Lee (leesh@kribb.re.kr)Ji-Yeon Hong (jyhong79@kribb.re.kr)Seung-Hoon Cho (chosh@kribb.re.kr)Hye-Min Park (gohworld@kribb.re.kr)Eun-Young Han (hiy10844@kribb.re.kr)Soon-Han Kwon (soonhan@kribb.re.kr)So-Yeon Shin (syshin@kribb.re.kr)Hye-Yeong Kim (bvcadmin@kribb.re.kr)Eun-Kyung Jeon (ekjeon@kribb.re.kr)

Biotechnology Process Engineering Center

Others

Viral Infectious Disease Research Center

International Biological Material Research Center

Laboratory of Experimental Animals

DAEJEON-KRIBB-FHCRC Research Cooperation Center

3 r d K R I B B P o s t e r F e s t i v a l

59 3rd KRIBB Poster Festival

E-1VDAC-2 regulates apoptosis in dexamethasone-induced human eosinophils

Sun-Woo Yoon1 and Haryoung Poo1

Viral Infectious Disease Research Center, KRIBB

Eosinophilia is clinically manifested in a number ofinflammatory diseases, particularly in allergic diseasessuch as atopic dermatis (AD), asthma. In these diseases,the accumulation of eosinophils occurs in an anti-apoptoticmechanism. Previously, we used a proteomics approach toanalyze eosinophil proteins from AD patients witheosinophilia and healthy donors. Among the identifiedproteins, we observed increased expression of voltage-dependent anion channel protein 2 (VDAC2) in eosinophilsfrom AD patients in comparison to healthy donors and itsanti-apoptosis function has been studied in eosinophils.Interleukin-5 (IL-5) is a potent eosinophils viability-enhancingfactor and known to extend eosinophils survival throughinhibition of programmed cell death. Glucocorticosteroidssuch as dexamethasone have among allergic diseasestherapeutic effects the inhibition of inflammatory cytokineproduction induction of eosinophils apoptosis. In order tostudy the mechanism of the anti-apoptosis activity ofVDAC2 to regulate in dexamethasone-induced humaneosinophil, eosinophils isolated from human peripheralblood by a negative immunomagnetic procedure weretreated with IL-5 for 24 h and then exposed to dexamethasonefor 24 h. Dexamethasone induced DNA fragmentation,cytochrome c release and its increased expression of pro-apoptotic molecule such as Bak. During apoptosis,VDAC2 expression was reduced in dexamethasone-induced eosinophils and VDAC2 interaction with Bak wasdetected by immunoprecipitation. Additionally, the role ofVDAC2 in regulating cell survival and death wasinvestigated by silencing endogenous VDAC2 expressionby using a short hairpin RNA (shRNA)-expressing vector.VDAC2 knockdown cells showed the reducedproliferation and increased apoptosis after dexamethasonetreatment compared with dexamethasone treated controlcells. These results suggest that VDAC2 may play animportant role in anti-apoptosis mechanism ofeosinophilia. (BIEN 2009 : INWES Asian Network :Busan, 2009)

Keywords : VDAC2, apoptosis, atopic dermatis

E-2Application of poly-gamma-glutamic acidimproves dermatitis in NC/Nga mice

Tae-young Lee1, Jai-Chul Choi2, Moon-Hee Sung2,3, andHaryoung Poo1

1Viral Infectious Disease Research Center, KRIBB2Bioleaders Cooperation3Department of Bio & Nanochemistry, Kookmin University

Atopic dermatitis (AD) is a chronic inflammatory skindisease characterized by pruritic and eczematous lesions,along with increased total IgE level in plasma,inflammatory cell infiltration, and the expression of Thelper 2 (Th2) cytokines. NC/Nga mice kept inconventional conditions are known to develop skin lesionresembling human AD. Poly-gamma-glutamic acid (g-PGA) is an edible biomaterial naturally synthesized byBacillus subtilis (chungookjang). Previously, we reportedthat g-PGA induced TLR4-dependent activation of innateimmunity, such as the production of IL-12 to shift theimmune response from type Th2 to Th1. We examinedwhether g-PGA could prevent the development of the skinlesions in NC/Nga mice. To examine the effect of g-PGAon NC/Nga, the g-PGA was applied once a day for 7weeks, starting at 14 weeks old on to the shaved skin. g-PGA remarkably changed the immune response from typeTh2 and Th1 as determined from cytokine level. Theserum IgE and IgG1 level was decreased and theexpression of IgG2a was upregulated. Also, the spleniclevel of IL-10 was downregulated, whereas that of IFN-gand IL-12 was increased. Furthermore, histologicalanalysis of the skin revealed that application of g-PGAsignificantly inhibited the thickening of epidermiscompared to control mice. These results suggest that g-PGA is effective for immunotherapy agents for thetreatment of AD. (BIEN 2009 : INWES Asian Network :Busan, 2009)

Keywords : poly-gamma-glutamic acid, atopic dermatitis

E-3Poly(gamma-glutamic acid)-chitosan nanoparticlesinduces antigen specific humoral and cellularimmunity

Kyong soon Lee1, Moon-Hee Sung2,3, and Haryoung Poo1

1Viral Infectious Disease Research Center, KRIBB2BioLeaders Corporation3Department of Bio & Nano Chemistry, Kookmin University

Nanoparticles are considered to be efficient tools forinducing potent immune responses by an Ag carrier. Inthis study, we examined the effect of Ag-carringbiodegradable poly(gamma-glutamic acid)(gamma-PGA)chitosan nanopartles on the induction of immune responsesin mice. gamma-PGA chitosan nanoparticles stronglyinduced up-regulation of costimulatory molecules, and theenhancement of T cell stimulatory capacity in DCs. Theimmunization of mice with OVA-carrying nanoparticles

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Korea Research Institute of Bioscience and Biotechnology 60

induced Ag-specific CTL activity and Ag-specific productionof IFN-gamma in splenocytes as well as potent productionof Ag-specific IgG Abs in serum. These results suggestthat Ag-carrying gamma-PGA chitosan nanoparticles arecapable of inducing strong cellular and humoral immuneresponses and might be potentially useful as effectivevaccine adjuvants for the therapy of infectious diseases.(International Symposium & Annual Meeting of theKorean Society for Microbiology and Biotechnology :Daejeon, 2009)

Keywords : nanoparticle, adjuvant

Members

DirectorHa-Ryoung Poo (haryoung@kribb.re.kr)

Post-DocSun-Woo Yoon (swyoon@kribb.re.kr)

ResearcherTae-Young Lee (taeyoung@kribb.re.kr)Kyong-Soon Lee (ghoshow@kribb.re.kr)

E-4Inclusion of Luffa tuberosa Roxb. in MomordicaL. (Cucurbitaceae): evidence from ITS sequencesof nrDNA

M. Ajmal Ali1, Soo-Yong Kim1, Arun K. Pandey2, S.Karuppusamy3, and Joongku Lee1

1International Biological Material Research Center, KRIBB2Botany Department,University of Delhi, Delhi, India3Botany Department, Madurai College, Madurai, TN, India

The phylogenetic position of long been systematicallydebatable species Luffa tuberosa is evaluated in thepresent study using ITS sequence of nuclear ribosomalDNA data. The study sampled a total number of 16accessions which include five accessions of Luffa (underfour species i.e. Luffa acutangula, L. cylindrica, L. aegyptiacaand L. tuberosa), nine accessions of Momordica (undereight species i.e. M. angustisepala, M. balsamina, M.cabraei, M. charantia, M. charantia subsp. macroloba, M.cissoides, M. cochinchinensis, M. dioica and M. foetida)and two accessions of Trichosanthes under two species(i.e. T. lepiniana and T. tricuspidata). The bootstrap strictconsensus tree clearly reveals two major group i.e. Luffagroup (100% bootstrap support) and Momordica groupwith L. tuberosa (94% bootstrap support). Both the groupsshow strong relationship to each other (99% bootstrapsupport). We herein support inclusion of Luffa tuberosainto the genus Momordica as M. tuberosa (Roxb.) Cogn.based on its nesting as a polytomy within the base ofMomordica group. (The 64th Annual Meeting of theKorean Association of Biological Sciences : Daejeon,2009)

Keywords : Momordica, Cucurbitaceae, phylogeneticposition, ITS

E-5Relationships among subfamily cucurbitoideae(Family Cucurbitaceae) from India inferredfrom ITS sequences of nrDNA

M. Ajmal Ali1, Arun K. Pandey2, and Joongku Lee1

1International Biological Material Research Center, KRIBB2Department of Botany, University of Delhi, India

The ITS sequences of nrDNA were analysed from 14species of the subfamily Cucurbitoideae to assess phylogeneticrelationships. The analysis resolves four major groups i.e.(1) Edgaria darjeelingensis; (2) Momordica dioica - Luffatuberosa; (3) Luffa - Trichosanthes and (4) Benincasa -Coccinia - Cucumis - Diplocyclos - Lagenaria - Melothria.Edgaria darjeelingensis (tribe Herpetospermeae) is well

International Biological Material Research Center

distinct from the other tribes i.e Joliffieae, Luffeae,Trichosantheae and Benincaseae. Luffa tuberosa forms aclade with Momordica dioica (95% bs). Trichosanthes andLuffa are grouped in a single clade (71% bs). The membersof the tribe Benincaseae are grouped into a single clade(84% bs). Lagenaria siceraria is very closely related toDiplocyclos palmats - Coccinia grandis subgroup. Diplocyclospalmatus clade together (88% bs) with Coccinia grandis.Benincasa hispida clade with Cucumis sp. (85% bs).Melothria heterophylla does not clade with Cucumis. Weherein support the retention of tribe Melothrieae underCucurbitoideae instead of merging it within tribe Benincaseae.(The 64th Annual Meeting of the Korean Associationof Biological Sciences : Daejeon, 2009)

Keywords : Cucurbitoideae, India, phylogenetic relationships,ITS

E-6Cypsela morphology of Stephanomeriinae(Compositae) and its taxonomic significance

Sang-Hong Park1, Joo-Hwan Kim2, and Joongku Lee1

1International Biological Material Research Center, KRIBB2Department of Biology, Daejeon University, Daejeon

Cypsela of 15 species (under 5 genera) of subtribeStephanomeriinae were scanned using SEM and analysedunder PAUP in order to evaluate its systematicsignificance. Cypsela morphological characteristic clearlyreveals its potential significance in classifying subtribeStephanomeriinae. The results reveal three major groups,i.e. (i) Rafinesquia, (ii) Prenanthella-Munzothamnus-Pleiacanthus, and (iii) Stephanomeria. Prenanthelladiffers from Munzothamnus and Pleiacanthus in cypselasurface characteristic and shape in transverse section.Within the genus Stephanomeria, S. exigua, S. diegensis,S. paniculata, and S. elata grouped together. The genusPrenanthella, Munzothamnus, Pleiacanthus showsplumose type of pappus bristles which differs fromStephanomeria in having barbellte type of pappus bristles.Pleiacanthus differs from Stephanomeria in havingrectangular pattern of cypsela surface in Pleiacanthuswhile striate in Stephanomeria. Rectangular surfacepattern of cypsela can be easily distinguished Prenanthellafrom Munzothamnus, Pleiacanthus, Rafinesquia andStephanomeria. Cypsela morphology was beyondcontroversy about disposal among Munzothamnus,Pleiacanthus and Stephanomeria. (The 64th AnnualMeeting of the Korean Association of BiologicalSciences : Daejeon, 2009)

Keywords : Stephanomeriinae, Cypsela, SEM, taxonomicsignificance

E-7Studies on seedling morphology of some medicinalplants of Korea

Jinki Kim, Geonrae Kim, Changyoung Lee, M. Ajmal Ali,Sang-Hong Park, and Joongku Lee

International Biological Material Research Center, KRIBB

The knowledge of seedling morphology is significant insystematic delimitation of taxa and as such can be utilizedas one of the character states for a comparative study oftaxa at specific and generic level as well. Being quiteconservative in nature, the seedling morphology is beingutilized to establish systematic relationships among thetaxa. Out of 3034 species taxonomically treated in thegenera of vascular plants of Korea (Park et al. 2007), thereare over 1,000 species of medicinal plants that have beenimportant as a means of treating and preventing diseasetraditionally in Korea (www.wpro.who.int). The presentcommunication document a detailed account of seedlingcharacteristic of medicinal plants (70 species) of Korea,under 62 genera and 39 familiy for identification of taxonin seedling stage which will be useful to botanist,conservation biologist, and other interested in medicinalplants of Korea. (The 64th Annual Meeting of theKorean Association of Biological Sciences : Daejeon,2009)

Keywords : seedling morphology, medicinal plants, korea

E-8A molecular systematic study of some speciesof Trichosanthes L. (Cucurbitaceae) using ITSsequences of nrDNA and its taxonomicimplication

M. Ajmal Ali1, Arun K.Pandey2, Sang-Hong Park1, andJoongku Lee1

1International Biological Material Research Center, KRIBB2Department of Botany, University of Delhi, India

Principally an Asiatic genus Trichosanthes L. of tribeTrichosantheae, subtribe Trichosanthinae, familyCucurbitaceae is the largest genus in the family with aboutapproximately 100 accepted species which is either anIndo-Malayan or Chinese Centre of origin. Majority ofTrichosanthes species grow in wild. The genus is ofimmense economic importance. Phylogenetic relationshipamong some species of Trichosanthes sampled from Indiaand Korea was assessed using ITS sequence of nuclearribosomal DNA. All bootstrap strict consensus treesresulting from the analysis resolves two major clades i.e(1) T. lepiniana-T. tricuspidata (100 bootstrap support)

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Korea Research Institute of Bioscience and Biotechnology 62

and (2) T. diocia-T. cucumerina (74% bootstrap support).T. kirilowii sampled from South Korea show polytomy atbase, however clearly nested with T. lepiniana-T.tricuspidata group within NJ tree. We herein based on thepresent study strongly support the treatment of T.cucumerina var. anguina (L.) Haines as cultivated variantof T. cucumerina. (The 64th Annual Meeting of theKorean Association of Biological Sciences : Daejeon,2009)

Keywords : Trichosanthes, Cucurbitaceae, ITS, taxonomicimplication

E-9

1 1 2

1

2

Salsola collina

Galeola septentrionalis

(The 64th Annual Meeting of theKorean Association of Biological Sciences : Daejeon, 2009)

Keywords :

Members

Director Hyouk Joung (joungh@kribb.re.kr)

Senior Researcher Joongku Lee(joongku@kribb.re.kr)Sang-Ho Choi (decoy0@kribb.re.kr)

Post-DocSang Hong Park (shpark7@kribb.re.kr)M. Ajmal Ali (majmalali@rediffmail.com)

ResearcherJin-Ki Kim (nemesis9@kribb.re.kr)Geonrae Kim (mtnwind@kribb.re.kr)Chang-Young Lee (changags@kribb.re.kr)Sung-Rae Oh (osr520@kribb.re.kr)Hye-Seon Yim (zone753@kribb.re.kr)Jeong-Uk Eom (zxzxv82@kribb.re.kr)Tae-Yang Song (ddansun@kribb.re.kr)Bum-Jung Kim (kbjman@kribb.re.kr)

E-10Preventative effects of phellinus baumii onhepatic steatosis in high f f diet-fed C57BL/6mice

Jung-Ran Noh1,, Bong-Sik Yun2, In-Kyoung Lee2, Yong-Hoon Kim1, Gil-Tae Gang1, Keum-Jin Yang1, and Chul-Ho Lee1

1Laboratory of Experimental Animals, KRIBB2College of Environmental & Bioresource Science, ChonbukNational University

This study was performed to evaluate the effects of(phellinus baumii, PB) on development of hepatic steatosisin high fat diet (HFD)-fed C57BL/6 mice. Eight week-oldmale C57BL/6 mice were randomly divided into threegroups (n=6 per group); HFD (21% lard and 0.15%cholesterol, w/w) plus daily vehicle (1% DMSO, 0.1%Tween 80) as a control group, HFD plus xenical (50mg/kg), HFD plus the extract of PB (PB, 500 mg/kg), andthe PB was administered daily by oral gavage for 12weeks. Although there were no significant changes inbody weight and fat mass, hepatic total lipid, totalcholesterol and triglyceride (TG) concentrations wereremarkably decreased in xenical- and PB-treated groupscompared with control group. Especially, hepatic TGcontent appeared to be approximately 34.2% less inxenical group and 36.9% less in PB group as comparedwith the control group (p<0.05). On the basis ofmorphometric of liver tissue, the score index, which wascalculated as graded numbers from 1 to 4 for fatty liverchanges, was significantly decreased in the xenical-(2.3±0.5) or PB-treated groups (2.5±0.5) as compared withthe control group (4.0±0.0). Furthermore, to evaluate theeffect of PB on TG absorption, TG inhibition experimentwas performed and a marked reduction was observed inxenical- and PB-treated groups as compared with controlgroup (P<0.05). Based on these results, we speculate thatPB has shown preventative effects of fatty liver viasuppressing hepatic lipid accumulation and inhibiting ofTG absorption. (Annual Meeting of the KoreanAssociation for Laboratory Animal Science : Cheonan,2009)

Keywords : Phellinus baumii, hepatic lipid, TG absorption

Members

DirectorChul-Ho Lee (chullee@kribb.re.kr)

Principle Research Byung-Hwa Hyun (hyunbh@kribb.re.kr)

Senior Researcher Hwan-Jung Hwang (coccs99@kribb.re.kr)

Post-DocKeum-Jin Yang (nadia@kribb.re.kr)

ResearcherDong-Hee Choi (heedc@kribb.re.kr)Yong-Hoon Kim (milknut@kribb.re.kr)Gil-Tae Gang (kkt0303@kribb.re.kr)Jung-Ran Noh (sspera@kribb.re.kr)Ji-Sun Moon (mmjs@kribb.re.kr)Hyo-Jeong Kim (hhjung88@kribb.re.kr)In-Bok Lee (inbok4526@kribb.re.kr)

63 3rd KRIBB Poster Festival

Laboratory of Experimental Animals

Korea Research Institute of Bioscience and Biotechnology 64

Members

DirectorHyang-Sook Yoo (yoohyang@kribb.re.kr)

Principal Researcher Jeoung-Heun Ko (jhko@kribb.re.kr)Eun-Wie Cho (ewcho@kribb.re.kr)

Senior Researcher Yong-Sam Kim (omsys1@kribb.re.kr)

ResearcherSun-Hee Kim (griguri@hanmail.net)Yun-Mi Seo (yunmi0714@lycos.co.kr)Su-Jin Jung (sujin4088@hanmail.net)Mi-Kyung Woo (mllove81@nate.com)Hai-Min Hwang (purity-32@hanmail.net)Dong-Il Kim (1111kdi@hanmail.net)Chang-Kyu Heo (hck40@kribb.re.kr)Jee-Ae Jung (najiae86@hanmail.net)Chang-Hee Cho (arztcho@naver.com)

DAEJEON-KRIBB-FHCRC Research Cooperation Center

Ochang Branch Institute

Therapeutic Antibody Research Center

Stem Cell Research Center

Immune Modulator Research Center

Molecular Cancer Research Center

Chemical Biology Research Center

Bio-Evaluation Center

Korea National Primate Research Center

3 r d K R I B B P o s t e r F e s t i v a l

67 3rd KRIBB Poster Festival

F-1The protease inhibitor, elafin, induces p53-dependent apoptosis in human melanoma cells

Kyung Sook Yu1, Yangsoon Lee1,2, Chung Mi Kim1, Eui-Chul Park3, Juhyun Choi2, Dae-Sik Lim2, Young-HwaChung4, and Sang Seok Koh1*

1Therapeutic Antibody Research Center, KRIBB2National Research Laboratory, Department of BiologicalScience, KAIST,

3LG Life Sciences, Ltd./R&D Park4Department of Nanomedical Engineering, BK21 NationalTechnology Team, Pusan National University

Expression of the protease inhibitor elafin is deregulated inseveral human cancers. However, functions of the proteinin cancer are yet to be established. Here we show thatelafin elicits pro-apoptotic effects in melanoma cells.Elafin triggered the intrinsic apoptotic pathway asevidenced by the increased caspase-9 activity andunaltered caspase-8 activity. Caspase-9-specific siRNA,dramatically abrogated elafin-induced apoptosis. Elevatedlevel of p53 was observed, resulting in increasedtranscriptional activation and consequent expression ofdownstream effector molecules. Moreover, the apoptoticeffect of elafin was inhibited by p53-specific siRNA andthe p53 inhibitor pifithrin- . Elafin treatment of xenograftmice of melanoma cells led to significantly smaller tumorsizes compared with those of untreated control mice.Western blot and reverse transcription analyses indicatedtranscriptional repression of the elafin gene in melanomacells. Our results collectively indicate that elafin inducesapoptosis in melanoma cells through a p53-dependentintrinsic apoptotic pathway, and that repression of elafinexpression in melanoma may contribute to diseaseprogression. (The 21st Annual Meeting of the KoreanSociety for Molecular and Cellular Biology : Seoul,2009)

Keywords : elafin, apoptosis, p53, melanoma, therapeutics

Members

Director Hyo-Jeong Hong (hjhong@kribb.re.kr)

Principal Researcher Sang-Seok Koh (sskoh@kribb.re.kr)Se-Mi Kim (semikim@kribb.re.kr)

Senior Researcher Jeong-Ki Min (jekmin@kribb.re.kr)Ju-Yeon Jung (jjung@kribb.re.kr)

Post-DocIn-Soo Park (ipark@kribb.re.kr)Mi-Jin Shon (mijin@kribb.re.kr)

ResearcherYon-Sung Son (sonysung@kribb.re.kr)Eung-Seuk Lee (les91@kribb.re.kr)Jin-Hong Kim (toveless@kribb.re.kr)Moon-Sik Jeong (15mpjms@kribb.re.kr)Hyun-Ho Yoon (swat95@kribb.re.kr)Tae-Woo Park (ptw44@kribb.re.kr)Ji-Hyun Moon (jhMoon@kribb.re.kr)Seul-Ki Cho (Seul8502@kribb.re.kr)Dong-Gwang Lee (creator@kribb.re.kr)Tae-Hyoung Kwon (biokth@kribb.re.kr)Rohot Sing (rohits@kribb.re.kr)Seong-Kuk Jeon (heaven78@kribb.re.kr)Su-Jin Kim (doppijin@gmail.com)Eun-Hee Nam (enjjng@kribb.re.kr)Min-Hye Jin (tophj012@kribb.re.kr)Eun-Hye Park (dms1127@kribb.re.kr)Kyung-Sook Yu (redcamel@kribb.re.kr)Yang-Soon Lee (jyslee@kribb.re.kr)

Therapeutic Antibody Research Center

Korea Research Institute of Bioscience and Biotechnology 68

F-2The effects of tumor necrosis factor-alpha on invitro differentiation of natural killer cells

Jiwon Lee1,2, Suk Hyung Lee1,2, Mi Sun Kim1, and InpyoChoi1,2

1Cell Therapy Research Center, KRIBB2Functional Genomics, University of Science and Technology(UST)

Natural killer (NK) cells are differentiated fromhematopoietic stem cells (HSCs) in bone marrow. Thedifferentiation of NK cells is regulated by various factorsincluding soluble growth factors and transcription factors.Here, we demonstrated that tumor necrosis factor- (TNF-) is a positive regulator of NK cell differentiation. HSC-

derived precursor NK (pNK) cells were furtherdifferentiated into mature NK (mNK) cells in the presenceof IL-15 invitro. The potential role of TNF- on NK cellmaturation was evaluated in the presence or absence of IL-15. TNF- itself induced the expression of NK1.1 andCD122 of mature NK cells and its effects weresignificantly augmented in the presence of IL-15. IFN-production was maximal when NK cells were matured inthe combination of IL-15 and TNF- . Moreover mRNAexpression of several transcription factors including T-betand GATA-3 was increased during TNF- -mediatedinvitro NK cell differentiation. Overall, these data indicatethat TNF- regulates NK differentiation by increasingtranscription factors which are crucial for NK maturation.(96th Annual Meeting of the American Association ofImmunologists : Seattle (USA), 2009)

Keywords : NK cells, TNF- , differentiation, T-bet, Gata-3, NF- B

Members

DirectorIn-Pyo Choi (ipchoi@kribb.re.kr)

Senior Researcher Suk-Ran Yoon (srtoon@kribb.re.kr)Tae-Don Kim (tdkim@kribb.re.kr)Hai-Young Jung (haiyoung@kribb.re.kr)

Post-DocHyun-Woo Suh (hhyunw@empal.com)Hu-Nan Sun (sunmkbb@hanmail.net)Ai-Guo Wang (wangaiguotl@sohu.com)Lee Suui (iamsuui@kribb.re.kr)Mi-Sun Kim (mskim@kribb.re.kr)

ResearcherSo-Hyun Yun (thgusdldi@hanmail.net)Seung-Hee Yang Dong-Hee Shin (dalcy@naver.com)Mi-Jung Kim (tito006@kribb.re.kr)Dong Oh Kim (jesusdd@kribb.re.kr)Ji-Won Lee (pasff200@hanmail.net)Soo-Yeon Park (mspsy82@kribb.re.kr)JI-Hye Park (jhpark@kribb.re.kr)Swok-Hyung Lee (daddy4u@kribb.re.kr)Ju-yeong Park (teajonboy@hotmail.com)Chan-Mi Park Hye Min Lee Seong-Jin Cho (chonnom001@kaist.ac.kr)

Stem Cell Research Center

F-3Suppressive effects of methanol extract fromLilium lancifolium on inflammatory responsesin raw264.7 cells

Ok-Kyoung Kwon1, Mee-Young Lee2, Ji-EunYuk1, Sei-Ryang Oh1, Hyeong-Kyu Lee1, and Kyung-Seop Ahn1,*

1Immune Modulator Research Center, KRIBB 2Herbal medicine EBM Research Center, KIOM

Lilium lancifolium is a species of lily native to northernand eastern Asia, including Korea, Japan, China andSakhalin. It was commonly used in the bronchitis, thepneumonia, nourishment, a tonicity and the strongstomache. In this study, anti-inflammatory effects of themethanol extract from the root of Lilium lancifolium wereinvestigated in LPS-stimulated mouse macrophage cells,Raw264.7 cells. Levels of NO, PGE2 and proinflammatorycytokines (IL-6 and TNF-a) in supernatant were determinatedby sandwich ELISA. The expression of COX-2 and iNOS,the phosphorylation of MAPK subgroups (ERK and JNK)and the activation of NF-kB in extract were detected byWestern blot analysis and immunocytochemistry assayrespectively. The methanol extract significantly inhibitedthe production of NO, PGE2, IL-6 and TNF-a in LPS-stimulated cells. Furthermore, the extract also suppressedthe expression of iNOS and COX-2, the phosphorylationof ERK1/2 and JNK and the translocation of NF-kB p65subunit into nuclear. The extract also inhibited interleukin-4 and interleukin-13 production in Con A-inducedsplenocytes. These results indicated that the anti-inflammatory effects of methanol extract from L.lancifolium might be due to the down-regulation of NO,COX-2, IL-6 and TNF-a via the suppression of NF-kBactivation and conversation of ERK and JNK productionin LPS-stimulated Raw264.7 cells. (The Fall InternationalConvention of the Parmaceutical Society of Korea :Seoul, 2009)

Keywords : Lilium lancifolium; Inflammation; iNOS;COX-2; NF-kB

F-4Anti-inflammatory and anti-asthmatic effectsof Capsicum annuum L. on Ovalbumin-induced lung inflammation in a mouse asthmamodel

Ji-Eun Yuk1, Mi-Young Lee2, Ha-Young Jang1, Se-miKim1, Ok-Kyoung Kwon1, Sei-Ryang Oh1, Hyeong-KyuLee1,*, and Kyung-Seop Ahn1,*

1Immune Modulator Research Center, KRIBB2EBM Research Center, Korea Institute of Oriental Medicine

Introduction: Bronchial asthma represents the chroniclung inflammation, airway hyperresponsiveness, andairway remodeling. It has been reported that pepper fruitCapsicum annuum L. has anti-oxidative activity. However,the effect of Capsicum annuum L. on asthma has notreported yet. Based on this study, Capsicum annuum L.could be developed as an anti-asthmatic drug. Methods:We used a mouse asthma model, produced by ovalbuminsensitization and challenge. Mice were sensitized byinjection of 20 µg OVA, which was emulsified in 2 mgaluminum hydroxide, in a total volume of 200 µl on days 1and 14. The mice were exposed to a 1 % ovalbuminsolution aerosolized using an ultrasonic nebulizer for 20min per day form days 28 to 30 after the secondsensitization. Capsicum annuum L. (30 mg/kg, PO) wastreated 1 hr before the ovalbumin challenge. Results:Capsicum annuum L. significantly reduced the T-helper-2-type cytokines and chemokines such as IL (interleukin)-4,IL-13, and eotaxin. Eosinophils and leukocytes, therecruited cells in BALF, were decreased after Capsicumannuum L. treatment. Histological analysis showed thatCapsicum annuum L. reduced the goblet cell hyperplasiaand mucus production and attenuated eosinophil-richleukocyte infiltration compared with OVA-challengedmice. This extract significantly inhibited mRNAexpression of IL-4, IL-5, IL-13 following asthmainduction in lung tissue. Also it effectively suppressedimmunoglobulin E (IgE) level and OVA specific IgE levelin both BALF and serum. Conclusions: These resultssuggest that Capsicum annuum L. could be a herbalmedicine for treat asthma. (The Fall InternationalConvention of the Parmaceutical Society of Korea :Seoul, 2009)

Keywords : asthma inflammation, anti oxidant, Capsicumannuum

F-5Anti-inflammatory and anti-asthmatic effectsof tiarellic acid (TA) in a mouse model ofallergy

Mi-Young Lee1, Ji-Eun Yuk1, Ok-Kyoung Kwon1, Kim HS1,2,Sei-Ryang Oh1, Hyeong-Kyu Lee1, and Kyung-Seop Ahn1

1Immune Modulator Research Center, KRIBB 2Biomolecular Science, University of Science & Technology(UST)

Asthma is an inflammatory disease of the airways, and thecurrent focus in managing asthma is the control ofinflammation. We investigated anti-inflammatory andanti-asthmatic effects of tiarellic acid isolated fromTiarella polyphylla on asthmatic parameters–such asimmunoglobulinE (IgE) level, cytokine release,

69 3rd KRIBB Poster Festival

Immune Modulator Research Center

Korea Research Institute of Bioscience and Biotechnology 70

eosinophilia, airway hyperresponsiveness (AHR) and mucushypersecretion–in an ovalbumin (OVA)-sensitized/challengedmouse model. Tiarellic acid significantly inhibited increasesin total immunoglobulinE (IgE), T-helper-2-type cytokinessuch as interleukin (IL) -4 and IL-5 in bronchoalveolarlavage fluid (BALF), and also effectively suppressedairway hyperresponsiveness, eosinophilia, and mucushypersecretion, in the asthmatic mouse model. The efficacyof tiarellic acid was comparable to montelukast, an anti-asthmatic drug that is currently available. These resultssuggest that tiarellic acid could be a major marker in herbalmedicines that are used for asthma treatment, and couldalso act as a lead for the development of anti-inflammatoryand anti-asthmatic drugs. (57th International Congress &Annual Meeting of the Society for Medicinal Plant andNatural Product Research : Geneva (Switzerland),2009)

Keywords : tiarellic acid; asthma; AHR; IgE

F-6Constituents of Anthriscus sylvestris andsuppressive effect on matrix metalloproteinase-9 in airway remodeling

Hye-Sun Lim1, Sei-Ryang Oh1*, Ho-Jae Lee2, Xing FuCai1, Soo-Hyun Kim1, Kyung-Seop Ahn1, Dai-Eun Sok3,and Hyeong-Kyu Lee1

1Immune Modulator Research Center, KRIBB2Laboratory of Chemoprevention, Gachon University ofMedicine and Science Lee Gil Ya Cancer and DiabetesInstitute

3College of Pharmacy, Chungnam National University

Airway remodeling is a major change responsible forirreversible asthmatic airflow restriction. The Th-2 cytokines-dominant eosinophilic inflammatory mechanism cannotfully explain the progressive subepithelial fibrosis andstructural changes in the extracellularmatrix(ECM). Matrixmetalloproteinases (MMPs) are endopeptidases belongingto the metzincin superfamily which are known as the keyenzymes responsible for ECM degradation. Among theMMPs, MMP-9 may play a pivotal role in chronic airwayinflammation and remodeling in asthma. Therefore, themodulators of MMP-9 expression and activity may be apromising therapeutic target of airway remodeling. In thecourse of screening for suppressors of MMP-9 expression/production from natural products, the methanolic extractof Anthriscus sylvestris showed a significant inhibitoryeffects on MMP-9 gene expression (reporter assay) andproduction (zymographic assay) in vitro. Activity-guidedseparation was carried out to isolate the active constituentsof A. sylvestris, and the structural elucidation of the activesagainst MMP-9 expression/production is in process. The

active compounds isolated from A. sylvestris will be suggestedas candidates for the treatment of airway remodeling inasthma. Gwangju, 2009)

Keywords: airway remodeling, anthricus sylvestris, MMP-9, luciferase assay, zymography

Members

DirectorSei-Ryang Oh (seiryang@kribb.re.kr)

Principal Researcher Hyeong-Kyu Lee (hykylee@kribb.re.kr)Kyung-Seop Ahn (ksahn@kribb.re.kr)Young-Kook Kim (kimyk@kribb.re.kr)Sang-Ku Lee (sangku@kribb.re.kr)Hyun-Jun Lee (hjlee7@kribb.re.kr)

Senior ResearcherDur-Han Kwon (dhkwon@kribb.re.kr)Young-Won Jin (f2744@kribb.re.kr)

Post-DocXing Fu Cai (cxf2007@kribb.re.kr)

ResearcherOk-Kyoung Kwon (dooli@kribb.re.kr)Ji-Eun Yuk (lenayuk@naver.com)Hye-Sun Lim (qp1015@kribb.re.kr)Soo-Hyun Kim (beluga@kribb.re.kr)Hui-Seong Kim (fungdori@naver.com)Se-Mi Kim (elfdeer@nate.com)

F-7Two positive regulation genes from theGeldanamycin biosynthetic cluster in Streptomyceshygroscopicus JCM4427

Oksik Choi, Woncheol Kim, Cheng-Zhu Wu, Su HeunRoh, Kyeong Lee, Jung Joon Lee, and Young-Soo Hong*

Molecular Cancer Research Center, KRIBB

Streptomyces cells have very complex regulatory networksof secondary metabolism, in which many regulators thatgovern production of secondary metabolites showfunctional interactions. Pathway-specific regulators onlyaffect a single antibiotic biosynthetic pathway, the gene ofwhich is clustered, and regulation of antibiotic biosyntheticgene expression is exerted at the transcriptional level. Thegel14 and gel17 genes from the geldanamycin biosyntheticcluster in Streptomyces hygroscopicus JCM4427 wereanalyzed, and each deduced products was found to haveamino acid sequence homology with the LuxR transcriptionalregulatory proteins, which is included an N-terminal ATP-binding domain and a C-terminal DNA-binding domain.Inactivation by gene disruption of gel14 or gel17 in genomeresulted in a complete loss of geldanamycin production. Inthis report we provide a detailed the transcriptionalregulation of the geldanamycin biosynthetic gene clusterby RT-PCR studies on each gene disruption mutant.(International Symposium & Annual Meeting of theKorean Society for Microbiology and Biotechnology :Daejeon, 2009)

Keywords : streptomyces, geldanamycin, biosynthesis,regulation

Members

DirectorHyun-Sun Lee (leehs@kribb.re.kr)

Principal Researcher Jung-Joon Lee (jjlee@kribb.re.kr)Young-Soo Hong (hongsoo@kribb.re.kr)Kyeong Lee (kaylee@kribb.re.kr)

Post-DocJin Xuejun (xjjin@kribb.re.kr)Byung-Jun Ryu (bjryu93@kribb.re.kr)

ResearcherOk-Sik Choi (dhrtlrl79@hanmail.net)Cheng-Zhu Wu (wcz0611@kribb.re.kr)Su-Heun Roh (nsh1052@nate.com)Jee-Hee Seo (jhseo@kribb.re.kr)Il-Soon Kim (kiss79@kribb.re.kr)Nam-Ye Kim (whiteday@kribb.re.kr)Sun-Hwa Lee (enddlsh@kribb.re.kr)Yinglan Jin (jinyl@kribb.re.kr)Yan Xia (xiayan1980@kribb.re.kr)Jung-Jin Kwon (kjjzzang1@naver.com)Hong Ri Jin (hrjinooo@kribb.re.kr)

71 3rd KRIBB Poster Festival

Molecular Cancer Research Center

Korea Research Institute of Bioscience and Biotechnology 72

F-8Regulation of integrin alpha2 (ITGA2) expressionby MAPK signal cascades in Ki-ras transformedprostate cancer cell lines (267B1/Ki-ras)

Kyoung-A Kim, Sook Jung Jeong, Osong Kwon, Bo YeonKim, and Jong Seog Ahn

Chemical Biology Research Center, KRIBB

The integrin alpha2beta1 heterodimer, present on a variableof cell types including fibroblasts and platelets, serves as acell surface receptor for collagen. And integrin alpha2’snormal expression fordifferentiation is altered duringcarcinogenesis, but the regulatory mechanism is unclear.We investigated the regulatory mechanism of ITGA2 up-regulation in 267B1 cells stably or transientyl expressingKi-ras. We performed microarray and realtime PCRanalysis with 267B1 human prostate epithelial cellstransformed viral Ki-ras transformed cells (267B1/Ki-ras), and found that Ki-ras is associated with ITGA2 up-regulation in mRNA and protein levels. The MAPK signalpathways, ERK, JNK, and p38 were activated, and wereexpected to be involved in ITGA2 up-regulation, it couldnot repressed in Ki-ras stably knock down cell line (KD#16) even though inhibited ERK and JNK activationexcept p38. The ERK inhibitor, PD98059 decreasedITGA2 expression in mRNA and protein level. Our datashow that Ki-ras overexpressed in 267B1 cells constitutivelyactivate the ERK signaling, ultimately leading to theincreased expression of ITGA2. (The 21st AnnualMeeting of the Korean Society for Molecular andCellular Biology : Seoul, 2009)

Keywords : ITGA2, prostate cancer, Ki-ras, Ets-1

F-9 Regulation of ARL4C (ARL7) expression byMAPK signal cascades in Ki-ras transformedprostate cancer cell lines (267B1/Ki-ras)

Sook Jung Jeong, Kyoung-A Kim, Osong Kwon, JongSeog Ahn, and Bo Yeon Kim

Chemical Biology Research Center, KRIBB

ADP-ribosylation factor-like 4C (ARL4C, previouslydesignated ARL7), a 22 kDa GTP-binding proteinbelonging to the family of ADP-ARFs, seems to functionas a GTP-operated molecular switch that regulatesintracellular vesicle and/or protein transport. In this study,using microarray and realtime PCR analysis, we foundARL4C expression was up-regulated in transformedhuman epithelial prostate cancer cells with mutated viralKi-ras (267B1/Ki-ras). Stable or transient transfection of

the normal 267B1 cells with Ki-ras, activated MAPKsignaling pathways, ERK, JNK and p38 as well as ARL4Coverexpress, but ARL4C could not repressed in Ki-rasstably knock down cell line (KD #16) even thoughinhibited ERK and JNK activation except p38. Also, theERK inhibitor, PD98059, decreased ARL4C expression.Our data show that Ki-ras overexpression in 267B1 cellsconstitutively activates the ERK pathway and induces theactivation of the Ets-1 transcription factor, ultimatelyleading to the increased expression of ARL4C andactivation of JNK and p38 pathway. (The 21st AnnualMeeting of the Korean Society for Molecular andCellular Biology : Seoul, 2009)

Keywords : ARL4C, prostate cancer, Ki-ras, Ets-1

F-10ATM blocks tunicamycin-induced endoplasmicreticulum stress

Long He, Kim Kyoung A, Osong Kwon, Sun Ok Kim,Sook Jung Jeong, Jong Seog Ahn, and Bo Yeon Kim*

Chemical Biology Research Center, KRIBB

Endoplasmic reticulum stress (ER-stress) is associatedwith ataxia telangiectasia mutated (ATM) gene. Wepresent here conclusive data showing that ATM blocksER-stress induced by tunicamycin or ionizing radiation(IR). X-box protein-1 (XBP-1) splicing, GRP78expression and caspase-12 activation were increased bytunicamycin or IR in Atm-deficient AT5BIVA fibroblasts.Activation of caspase- 12 and caspase-3 by tunicamycinwas significantly reduced in cells transfected with wild-type Atm (AT5BIVA/wtATM). Atm knockdown bysiRNA, however, noticeably elevated ER-stress andchemosensitivity to tunicamycin. In summary, we presentsubstantial data demonstrating that ATM blocks the ERstress signaling associated with cancer cell proliferation.(The 21st Annual Meeting of the Korean Society forMolecular and Cellular Biology : Seoul, 2009)

Keywords : ATM, ER stress, UPR, IR

F-11Endoplasmic reticulum stress induction byantimycins isolated from actinomycete AN070088

JunPil Jang*, Sun-Ok Kim, Bo Yeon Kim, and Jong SeogAhn

Chemical Biology Research Center, KRIBB

Chemical Biology Research Center

Endoplasmic reticulum stress (ER stress) is key mediatorfor various type of diseases, including cancer, diabetes andimmune system. Thus, development of ER stress regulatorwould be a promising drug for the therapy of those diseases.In the course of searching for ER stress inducer frommicrobial sources, antimycins has been isolated from theculture broth if actinomycete AN070088. Antimycinsinduced XBP-1 mRNA splicing and accordingly upregulatedthe expression of GRP78/Bip chanperone in HeLa cell.(International Symposium & Annual Meeting of theKorean Society for Microbiology and Biotechnology :Daejeon, 2009)

Keywords : endoplasmic reticulum (ER) stress, XBP-1,antimycins, GRP78/Bip

F-12Isolation and structure determination ofPTP1B inhibitory peptide, enniatins from fungiisolate No. 332

Duc Manh Hoang, In-Ja Ryoo, KiHwan Bae, Bo YeonKim, and Jong Seog Ahn*

Chemical Biology Research Center, KRIBB

Protein tyrosine phosphatase 1B (PTP1B) is considered asa potential therapeutic target for the treatment of diabetesand obesity. In an effort to search for PTP1B inhibitorsfrom natural sources and microorganisms, an ethylacetate-soluble extract of fungi broth was found to inhibitPTP1B activity (75% inhibition at 30µg/mL). Bioassay-guided fractionation resulted in the isolation of Enniatin B(1), Enniatin D (2), Enniatin F(4), Enniatin G (3), andAntibiotic MK1688 (5) as PTP1B inhibitors. Compounds1-5 inhibited PTP1B with IC50 value ranging from 12±0.5µM to 31±0.5 µM. (International Symposium & AnnualMeeting of the Korean Society for Microbiology andBiotechnology : Daejeon, 2009)

Keywords : protein tyrosine phosphatase 1B (PTP1B)

F-13Verrucarin A, a inhibitor ER-stress inducedXBP-1 activation and GRP78 in FaO cells

Eun Young Bae*, Kim Sun Ok, Hyuncheol OH, Hyun GilChoi, Bo Yeon Kim, and Jong Seog Ahn

Chemical Biology Research Center, KRIBB

The endoplasmic reticulum (ER) is a perinuclear, cytoplasmic

compartment where proteins and lipids are synthesized.The 78-kDa glucose protein GRP78 is a central regulatorof endoplasmic reticulum (ER) homeostasis, functioningin protein folding, ER calcium binding and modulation oftransmembrane ER stress sensor activity. ER stressuncouples the interaction between GRP78 and ER stresssensors, leading to activation of the unfolded proteinresponse (UPR), including upregulation of ER chaperoneproteins. In the course of screening for an inhibitor of ERstress-induced XBP-1 activation and regulators ofGRP78/Bip molecular chaperone expression, a fungalisolate CRM00190, was selected on the basis of its pontentinhibitory effect. To identify ER stress inhibitors fromCRM00190, the organic extract form the fungus wasfractionated and purified by using the solvent extraction,RP-C18 column chromatography, and HPLC. The structureanalysis of the active compounds revealed the presence ofseveral highly substituted trichothecenes type metabolites.(International Symposium & Annual Meeting of theKorean Society for Microbiology and Biotechnology :Daejeon, 2009)

Keywords : endoplasmic reticulum (ER) stress, verrucarinA, GRP78, XBP-1

F-14ER stress-induced cervical cancer cell death byPimaradienoic acid involving GRP78 regulatedcaspase-12 activation and c-Jun N-terminalkinase

Sun-Ok Kim, Kyoung-A Kim, Long He, Jong Seog Ahn,and Bo Yeon Kim

Chemical Biology Research Center, KRIBB

Endoplasmic reticulum stress (ER stress) is a key mediatorfor various types of diseases, including cancer, obesity,diabetes and immune system. Thus, development of ERstress regulator would be a promising drug for the therapyof those diseases. Recently, pimaradienoic acid (PA)having a strong induction of endoplasmic reticulum stresswas isolated from a plant Aralia continentalis. PA inducedthe XBP-1 mRNA splicing and accordingly up-regulatedthe expression of Grp78/Bip chaperone in HeLa cells. AndPA induced the activation of several ER stress-relevantregulater, including CHOP/GADD153, caspase-12 and c-Jun N-terminal kinase (JNK). CHOP/GADD153expression was also increased by PA treatment,presumably through both IRE1 phosphorylation and p-eIF 2 phosphorylation. Moreover, using Grp78 siRNA, itwas found that GRP78-regulated caspase-12 activationand JNK increase responsible for HeLa cervical cancercells death. In this study, the mechanism of pimaradienoicacid compound for induction of ER stress and cancer cell

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death is presented. (The 21st Annual Meeting of theKorean Society for Molecular and Cellular Biology :Seoul, 2009)

Keywords : ER stress, Grp78, JNK, Caspase-12

F-15Clitocybins A-D of novel isoindolinonecompounds from the culture broth of Clitocybeaurantiaca

Young-Hee Kim, In-Ja Ryoo, Soo-Jin Choo, Guang-HuaXu, and Ick-Dong Yoo*

Chemical Biology Research Center, KRIBB

Clitocybin A and D, novel antioxidative compoundswereisolated from the culture broth of Clitocybeaurantiaca. These compounds were purified by solventextraction, silica gel, Sephadex LH-20 columnchromatography, and finally by preparative HPLC. Thesestructures were determined as 4,6-dihydroxy-2-(4-hydroxyphenyl)-isoindolin-1-one and 4-(4,6-dihydroxy-3-methoxy-3H-isoindol-1-yl)-benzoic acid on the basis ofthe NMR and MS spectroscopic analysis. Clitocybin Asynthesized for biological activity together with twocompounds in synthetic procedure,and two compoundsnamed clitocybin B and C. The antioxidant activity ofclitocybins A-D were evaluated by measuring free radicalscavenging effects using three different assays. ClitocybinA and B exhibited potently superoxide radical scavengingactivity with IC50 value of 10.3 and 11.2 M (Catechin16.0M), and displayed significant ABTS radicalscavenging activity with IC50 value of 6.4 and 7.2 M(Catechin 7.1M), while exhibited no DPPH radicalscavenging activity. The comet assay showed that theexposure of the cells to H2O2 increased their tail length to61.1, whereas their treatment with clitocybins A-Cresulted in a decrease of their tail length to 35.4, 38.3 and32.3 as shown in comet image. (InternationalSymposium & Annual Meeting of the Korean Societyfor Microbiology and Biotechnology : Daejeon, 2009)

Keywords : clitocybins A-D, Clitocybe aurantiaca,antioxidant activity, comet assay.

F-165-pentyl-2-furaldehyde, a melanogenesisinhibitor from Clitocybe sp.

Young-Hee Kim1, Soo-Jin Choo1, In-Ja Ryoo1, Guang-Hua

Xu1, Soon-Ja Seok2, and Ick-Dong Yoo1,*

1Chemical Biology Research Center, KRIBB2National Institute of Agricultural Science and Technology,RDA

In the continued search for melanogenesis inhibitors frommicrobial metabolites, we found that the culture broth ofClitocybe sp. MKACC 53267 inhibited melanogenesis inB16F10 melanoma. The active compound was purified bysolvent extraction, silica gel chromatography, SephadexLH-20 column chromatography, and finally by preparativeHPLC. Its structure was determined as 5-pentyl-2-furaldehydeon the basis of the UV, NMR, and MS spectroscopicanalysis.5-pentyl-2-furaldehyde potently inhibitedmelanogenesis in B16F10 cells with an IC50 value of 28.4µg/ml without cytotoxicity. (The Spring InternationalConvention of the Parmaceutical Society of Korea :Daejeon, 2009)

Keywords : melanogenesis, 5-pentyl-2-furaldehyde,Clitocybe sp.

F-17Isodeoxyhelicobasidin, a novel human neutrophilelastase inhibitor from the culture broth ofVolvariella bombycina

Guang-Hua Xu1, Young-Hee Kim1, Soo-Jin Choo1, In-JaRyoo1, Chang-Ji Zheng1,2, Soon-Ja Seok3, Won-Gon Kim1,and Ick-Dong Yoo1,*

1Chemical Biology Research Center, KRIBB2Key Laboratoy of Natural Resources and FunctionalMolecules of Changbai Mountain, Affiliated Ministry ofEducation, Yanbian University College of Pharmacy, PRChina.

3National Institute of Agricultural Science andTechnology, RDA

Elastin, an important structural protein of extracellularmatrix (ECM), is the main component of elastic fiber whichprovides resilience and elasticity to many tissues such asskin, lungs, ligaments, and arterial walls. Human neutrophilelastase(HNE), a serine protease primarily located in theazurophil granules of polymorphonuclear leukocytes, isthe only enzyme capable of degrading extracellular matrixproteins such as elastin, collagen, fibronectin, laminin andproteoglycan. Biologically, elastase activity significantlyincreases with age and results in a reduced skin elasticiticproperty. In the course of our screening program for HNEinhibitors, we isolated a novel compound, isodeoxyhelicobasidin(1), from the culture broth of Volvariella bombycina. Thechemical structure of 1 was elucidated on the basis ofspectroscopic analysis. Compound 1 showed significant

HNE inhibitory and antibacterial activites. (InternationalSymposium & Annual Meeting of the Korean Societyfor Microbiology and Biotechnology : Daejeon, 2009)

Keywords: human neutrophil elastase, isodeoxyhelicobasidin,volariella bombycina

F-18Anti-melanogenic effect of milk thistle extract

Soo-Jin Choo, In-Ja Ryoo, Young-Hee Kim, Guang-HuaXu, and Ick-Dong Yoo*

Chemical Biology Research Center, KRIBB

In our search for the melanogenesis inhibitors from naturalresources, we found that silymarin exhibited the inhibitoryeffect on melanogenesis in a spontaneously immortalizedmouse melanocyte cell line, Mel-Ab. Silymarin is astandardized extract obtained from the dried seeds of milkthistle (Silybum marianum Gaertn.). Silymarin significantlyprevents melanin production in a dose-dependent mannerwith an IC50 value of 28.2 µg/ml without effects on cellviability. Also, silymarin inhibited tyrosinase activity inmelanocyte, while it did not affect the catalytic activity ofcell-free tyrosinase. Furthermore, Western blot analysisindicated that silymarin decreased the expression oftyrosinase protein, the rate limiting melanogenic enzyme.Silybin A/B and isosilybin A/B were also able to inhibitmelanin production and tyrosinase expression in proteinlevel. In clinical test using double-blind methods, weobserved treatment with 2% silymarin-containing creamresulted in significant effect on skin whitening. Therefore,this study suggests that silymarin may be useful as anatural skin whitening agent. (International Symposium& Annual Meeting of the Korean Society for Microbiologyand Biotechnology : Daejeon, 2009)

Keywords: melanogenesis, silymarin, silybin

F-19Reticulone, a novel free radical scavengerproduced by Aspergillus sp.

In-Ja Ryoo1, Guang-Hua Xu1, Young-Hee Kim1, Soo-JinChoo1, Jong- Seog Ahn1, Ki-Hwan Bae2, and Ick-Dong Yoo1,*

1Chemical Biology Research Center, KRIBB2College of Pharmacy, Chungnam National University

Free radicals induce oxidative damage of cellular lipids,nucleic acids and proteins, are thought to be one of the

major risks for various diseases caused by oxidative stress.Thus, free radical-scavenging antioxidants have the potentialas protective agents against various diseases caused byoxidative damage. In the course of our screening programfor the free radical scavengers, we isolated a new compound(1) along with a known compound (2, reticulol) from theculture broth of Aspergillus sp.. Their chemical structureswere elucidated on the basis of UV, IR, NMR, and MSspectroscopic analysis. The antioxidative acitvity of 1 wasevaluated by DPPH and ABTS assay with minor modification.Compound 1 exibited potent radical scavenging activitywith IC50 value of 6.91µM for ABTS radical and 3.05 µMfor DPPH radical. (International Symposium & AnnualMeeting of the Korean Society for Microbiology andBiotechnology : Daejeon, 2009)

Keywords : Aspergillus sp. reticulone, DPPH, ABTS

Members

DirectorJong-Seog Ahn (jsahn@kribb.re.kr)

Principal Researcher Ick-Dong Yoo (idyoo@kribb.re.kr)Bo-Yeon Kim (bykim@kribb.re.kr)Jong-Pyung Kim (kimjp@kribb.re.kr)

Post-DocJae-Hyuk Jang (jangjh@kribb.re.kr)Asami Yukihiro (yasami@kribb.re.kr)Sook-Jung Jeong (sjjeong@kribb.re.kr)Kyoung-A Kim (lskka@kribb.re.kr)Eun-Young Bae (eunjs39@kribb.re.kr)Young-Hee Kim (kimyh93@kribb.re.kr)Soo-Jin Choo (joosch@hanmail.net)Guang-Hua Xu (Xgh0125@kribb.re.kr)

ResearcherIn-ja Ryoo (ijryoo@kribb.re.kr)O-Song Kwon (pine4102@kribb.re.kr)Duc-Manh Hoang (manhhd@kribb.re.kr)Jun-Pil Jang (jpjang@kribb.re.kr)Sun-Ok Kim (cuteok@kribb.re.kr)Long He (aaron001@kribb.re.kr)Phamthi Thu Huong (pthuong@kribb.re.kr )Hyun-Gil Choi (chg21@kribb.re.kr)

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F-20Species differences in serum stability ofhydroxamate-containing histone deacetylaseinhibitors

Soo Jin Oh1, Kye Sook Lee1, Song-Kyu Park1, GyoonheeHan2, Mihee Nam2, Jong Soon Kang1, Hwan Mook Kim1,and Kiho Lee1

1Bio-Evaluation Center, KRIBB2Department of Biotechnology, Yonsei University

Histone deacetylase inhibitors (HDACIs) are being developedas a new class of therapeutic agents with potent antitumoractivities in a broad spectrum of human cancers. To evaluateinter species differences in hydrolysis of hydroxamic acid-based HDACIs, the in vitrostability of KBH-A40 (N-hydroxy-3-(2-oxo-1-phenethyl-1,2,5,6-tetrahydropyridin-3-yl)propanamide), suberoylanilide hydroxamic acid(SAHA) and trichostatin A (TSA) was investigated in rat,dog, monkey and human serum up to 24 hours. KBH-A40was not degraded significantly when incubated with humanand monkey serum for 24hr. In contrast, it was rapidlyhydrolyzed to its carboxylic acid form in rat and dogserum (t1/2 = 1.24 hr and 0.66 hr, respectively). Hydrolysisrate of KBH-A40 was observed in the rank order ofdog>rat>monkey> human. SAHA was hydrolyzed in dogand monkey (t1/2 = 5.96 hr and 7.94 hr, respectively) whilethe hydrolysis was relatively slow in rat and human (t1/2 =19.35 hr and >24 hr, respectively). The rank order ofhydrolysis of SAHA was dog>monkey>rat>human. AlthoughTSA was relatively stable in the serum of all species(t1/2>24 hr), it was more stable in human and monkeyserum than in dog and rat serum. Taken collectively, ourresults show that hydroxamic acid-containing HDACIs aregenerally stable in human serum and less stable in theserum of experimental animals with varying degreesdepending on species. Also, these results suggest that it isnecessary to select the most relevant animal species interms of serum stability for evaluation of pharmacokineticsof HDACIs. Further studies are warranted, to elucidate theenzyme(s) involved in hydrolysisof hydroxamic acids. (TheSpring International Convention of the ParmaceuticalSociety of Korea : Daejeon, 2009)

Keywords: KBH-A40, SAHA, TSA, histone deacetylase,hydrolysis, hydroxamic acid hydrolysis

F-21Artemisinin inhibits lipopolysaccharide-induced nitric oxide production by blockingIFN-b/STAT-1 signaling

Ki Hwan Park1, Yeo Dae Yoon1, Sang-Bae Han2, Hyunju

Lee1, Kiho Lee1, Song-Kyu Park1, Hwan Mook Kim1, andJong Soon Kang1

1Bio-Evaluation Center, KRIBB2College of Pharmacy, Chungbuk National University

Nitric oxide (NO) produced by macrophages has beenimplicated in a host defense against microbial pathogensand tumor cells. Artemisinin is a well-known anti-malarialdrug and has been shown to inhibit NO production. In thisstudy, we investigated the effect of artemisinin onlipopolysaccharide (LPS)-induced NO production inmacrophages and the molecular mechanisms responsiblefor its effects. Artemisinin significantly suppressed NOproduction and inducible nitric oxide synthase (iNOS)mRNA expression in LPS-stimulated macrophages. NF-B activity, which is known to be important for NOproduction and iNOS expression, was minimally inhibitedonly by highest concentration of artemisinin used andMAPKs were not affected by artemisinin treatment. Incontrast, STAT-1 activation was suppressed by artemisinintreatment in a concentration-dependent manner. Furtherstudies demonstrated that inhibition of STAT-1 activationby artemisinin might be mediated by blocking interferon-ß(IFN-ß) expression. These results suggest that artemisininsuppresses NO production and iNOS expression, at least inpart, by blocking IFN-ß production and concomitantdown-regulation of STAT-1 signaling. (The 21st AnnualMeeting of the Korean Society for Molecular andCellular Biology : Seoul, 2009)

Keywords: NO (nitric oxide), LPS, iNOS, IFN-ß, STAT-1

F-22DBM1285 suppresses LPS-induced TNF-aproduction and attenuates development ofrheumatoid arthritis in adjuvant-inducedarthritis model

Hyunju Lee1, Jong Soon Kang1, In Young Choi2, Sang-BaeHan3, Yeo Dae Yoon1, Kiho Lee1, Hwan Mook Kim1, andSong-Kyu Park1

1Bio-Evaluation Center, KRIBB2Dongbu HiTek Co., Ltd.3College of Pharmacy, Chungbuk National University

TNF- is a major inflammatory cytokine that plays animportant role in the development of various inflammatorydiseases. In this study, we report DBM1285 as a novelinhibitor of TNF- production and a therapeutic candidatefor RA treatment. DBM1285 concentration-dependentlyinhibited lipopolysaccharide (LPS)-induced TNF-secretion in various cells of macrophage/monocytelineage, including bone marrow macrophages, THP-1 cells

Bio-Evaluation Center

and RAW 264.7 cells. However, LPS-induced mRNAexpression of TNF- was not affected by DBM1285 inthese cells. Further studies demonstrated that the inhibitoryeffect of DBM1285 on TNF- production might bemediated by post-transcriptional regulation through themodulation of the p38 mitogen activated protein kinase(MAPK)/MAPKAP kinase 2 (MK2) signaling pathway.We also confirmed that DBM1285 directly inhibits p38MAPK enzymatic activity. In vivo administration ofDBM1285 inhibited LPS-induced increase in serum levelof TNF- in mice. DBM1285 also suppressed progressionof RA in an adjuvant-induced arthritis model. Theseresults suggest that DBM1285 might be a promisingtherapeutic candidate for the treatment of RA. (The 21stAnnual Meeting of the Korean Society for Molecularand Cellular Biology : Seoul, 2009)

Keywords: TNF- , LPS, DBM1285, MAPK (mitogenactivated protein kinase)

F-23CML-08-62 inhibits the growth of human lungcancer cell through the induction of apoptosis

Jang Hyun Kim1, Hee Soon Lee2, Moo Rim Kang1, JongSoon Kang1, Jeong Wook Yang1, Sang-Bae Han2, HwanMook Kim1, and Song-Kyu Park1

1Bio-Evaluation Center, KRIBB2Chungbuk National University

We investigated the anti-cancer activity of CML-08-62, anovel NF- B inhibitor. This compound significantlysuppressed the proliferation of 17 human cancer cell linestested. Among them, the human lung cancer cell line NCI-H23 was the most sensitive to CML-08-62. Apoptosisinduction was measured using annexin V staining andcaspase activity assay. Flow cytometry analysis showedthat the apoptotic death of NCI-H23 cells by CML-08-62increased by 50% compared to the control group of cells.In addition, the activities of caspases, such as caspase 3/7,caspase 8 and caspase 9, of NCI-H23 cells treated byCML-08-62 increased 1.5 to 3 times those of the controlgroup of cells. Caspase 3, cleaved caspase 3 and caspase 7and cleaved caspase 7 and caspase 8 and cleaved caspase 8and PARP and cleaved PARP, which were active formsinvolved in apoptotic signaling pathway, were alsodetected in NCI-H23 cells treated with CML-08-62. Theseresults suggest that CML-08-62 is a promising anticancerlead compound. (The 21st Annual Meeting of theKorean Society for Molecular and Cellular Biology :Seoul, 2009)

Keywords: CML-08-62, NF- B, cell cycle arrest,apoptosis

F-24YAL-1112, a novel NF-kB inhibitor, inhibits thegrowth of colon cancer cells through inductionof apoptosis and cell cycle arrest

Moo Rim Kang1, Heesoon Lee2, Ki Hoon Lee1, Kiho Lee1,Jong Soon Kang1, Chang Woo Lee1, Jang Hyun Kim1,Jeong Wook Yang1, Bo Geun Kim1, Hwanmook Kim1, andSong-Kyu Park1

1Bio-Evaluation Center, KRIBB2Chungbuk National University

Aberrant activation of NF- B is frequently observed inmany cancers. It has been reported that suppression of NF-B limits the proliferation of cancer cells. Hence, methods

of inhibiting NF- B signaling have potential therapeuticapplication in cancer. In this study, we investigated theanti-tumor activity of YAL-1112, a novel synthetic NF- Binhibitor. YAL-1112 suppressed growth of 9 cancer celllines. Among the cell lines examined, HCT116 wa one ofthe cell lines most sensitive to YAL-1112. YAL-1112induced cell cycle arrest at the G2/M phase. In addition,YAL-1112 also induced apoptosis in these cells, which wasaccompanied by the activation of caspases, includingcaspase-9, caspase-8 and caspase-3. The pan-caspasesinhibitor,Z-VAD-fmk, partially blocked the cell death induced byYAL-1112. Our results indicate that YAL-1112 exerts ananti-proliferation activity against colon cancers byperturbing the cell cycle and activating the caspasespathways. (The 21st Annual Meeting of the KoreanSociety for Molecular and Cellular Biology : Seoul,2009)

Keywords: YAL-1112, NF- B, cell cycle arrest, apoptosis,Z-VAD-fmk

F-25Glabridin inhibits dendritic cell maturationand function by blocking the activation of NF-kB/Rel and MAPKs

Jong Soon Kang1, Jee Youn Kim2, Sang-Bae Han2, KihoLee1, Song-Kyu Park1, and Hwan Mook Kim1

1Bio-Evaluation Center, KRIBB2College of Pharmacy, Chungbuk National University

Glabridin is known to have anti-inflammatory,antimicrobial and cardiovascular protective activities. Theobjective of this study was to investigate the effect ofglabridin on dendritic cell maturation and function and themolecular mechanisms responsible for this effect.Glabridin dose-dependently suppressed lipopolysaccharide(LPS)-induced expression of maturation markers of

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dendritic cells, such as CD80, CD86 and MHC-class IImolecules, without affecting cell viability. The inhibitoryeffect of glabridin on functional maturation of dendriticcells was confirmed by enhancement of endocytosisdecreased by LPS treatment and suppression of LPS-induced IL-12 production and allogeneic T cellstimulation. The production of IFN- and IL-4 inallogeneic T cells was also suppressed by glabridintreatment. Further study demonstrated that LPS-inducedactivation of NF- B/Rel and MAPKs, which are known tobe important for dendritic cell maturation and functionalactivation. Collectively, these results demonstrate thatglabridin inhibits LPS-induced maturation and functionalactivation of dendritic cells, at least in part, by blockingNF- B/Rel and MAPK pathway. (96th Annual Meetingof the American Association of Immunologists : Seattle(USA), 2009)

Keywords: glabridin, LPS, NF- B, MAPK

F-26Hepatocellular carcinoma in genetically obesemice

Hyo-Jung Kwon1,2,Young-Suk Won1, Dae-Yong Kim2, andHyoung-Chin Kim1,*

1Bio-Evaluation Center, KRIBB2Department of Veterinary Pathology, College of VeterinaryMedicine, Seoul National University

Various epidemiological studies have shown that obesitymay be risk factor for hepatocellular carcinoma. However,underlying mechanisms of how obesity promotes hepaticcarcinogenesis remain unknown. In this study, weinvestigated the susceptibility of genetically obese mice todiethylnitrosamine (DEN) induced hepatic carcinogenesis.The ob/ob, db/db and wild type (WT) mice were injectedwith a single dose of DEN at 12 days of age and killed atappropriate time points. Visible tumors developed as earlyas 16 weeks of age in genetically obese mice. Histologicalexamination revealed that 100% of ob/ob and db/db micedeveloped hepatic adenoma and 100% of ob/ob and 30%of db/db mice developed hepatocellular carcinoma by 26weeks, while no hepatic tumor was observed in WT mice.Phenotypically, body and relative liver weights, serumcholesterol, glucose and triglyceride levels of ob/ob anddb/db mice were significantly higher than WT mice.Microarray analysis showed approximately 200 of theknown genes was commonly expressed in both hepatictumors of ob/ob and db/db mice, which were mostlyrelated to the regulation of cell proliferation and translation.In conclusion, our data show that development of hepatictumor is enhanced in genetically obese ob/ob and db/dbmice. Futuremore, gene expression profiling of these mice

models may provide novel insights into obesity-associatedhepatic carcinogenesis. (Society of Toxicologic Pathology28th Annual Symposium : Washington DC (USA), 2009)

Keywords : hepatocellular carcinoma, obesity, ob/ob, db/db

F-27 Lack of vitamin D3 Up-regulated protein 1(VDUP1) accelerates liver injury through AKTactivity

Sae-Bhom Lee1, Jong-Tak Han1, Sang-Woon Kim1, Hyo-Jung Kwon1, Won-Kee Yoon1, Ki-Hoan Nam1, Hyoung-Chin Kim1, Soo-Young Choe2, and Young-Suk Won1,*

1Bio-Evaluation Center, KRIBB2Department of Biology Graduate School, ChungbukNational University

Vitamin D3 up-regulated protein 1 (VDUP1) is a stress-response gene that mediates oxidative stress via suppressingthioredoxin function. Although there is accumulatingevidence for the involvement of VDUP1 in apoptosis, theroles of VDUP1 in apoptosis regulation have not fullybeen understood. In this study, we examined the role ofVDUP1 on hepatocyte apoptosis using lipopolysaccharide(LPS)/D-galactosamine (DGA)-induced liver injurymodel. After LPS/DGA injection, VDUP1 KO miceshowed significantly less survival rate, higher serum ALTand AST levels, more prominent hepatic necrosis, andhigher hepatocyte apoptosis. Caspase-3, caspase-8 andcaspase-9 activities were significantly increased in VDUP1KO mice compared with those in WT mice. AlthoughTNF-a level after LPS/DGA injection was highly expressedin VDUP1 KO mice, its downstream targets, JNK, andp38 MAPK, showed no significant difference between twogroups. In VDUP1 KO mice, AKT activity and its downstreamtarget, NF-kB p50, were dramatically reduced in KO micethan in WT mice. Taken together, these results indicatethat VDUP1 prevents hepatocyte apoptosis by regulatingAKT induced NF-kB activation. (The 21st Annual Meetingof the Korean Society for Molecular and Cellular Biology: Seoul, 2009)

Keywords : VDUP1, apoptosis, AKT, NF-KB

F-28Natural variations of genes controlling soybeanseed coat and flower colors under domestication

Kiwoung Yang1, Namhee Jeong1, Jung-Kyung Moon2,Yeong-Ho Lee2, Suk-Ha Lee4, Hwan Mook Kim1, Cheol

Ho Hwang5, Kyoungwhan Back3, and Soon-Chun Jeong1

1Bio-Evaluation Center, KRIBB2National Institute of Crop Science, RDA3Molecular Biotechnology Major, Chonnam NationalUniversity

4Department of Crop Science, Seoul National University5Department of Crop Science and Biotechnology,Dankook University

Proanthocyanidins and anthoyanins derived from thephenylpropanoid pathway most likely play a protectivefunction from pathogens and UV light exposure within theplant and act as signal molecules in plant-microbeinteractions. This study aimed to elucidate geneticcorrelation between genetic loci regulating pigmentationand polymorphisms of the genes involved in thebiosynthesis of anthocyanins. Association studies betweenthe polymorphisms in the anthocyanin biosynthetic genesand seed coat(I, R, and T) and flower(W1, W2, W3, W4,Wm and Wp) colors suggested that markers generated fromsites presumed to cause functional defects were alwayscompletely associated with the color trait variations butmarkers generated from the sites in the intron or distantlyflanking to casual genes were weakly associated with thevariation. The results provide an important empirical datafor the design of association studies for soybean in thefuture. (The 64th Annual Meeting of the KoreanAssociation of Biological Sciences : Daejeon, 2009)

Keywords : proanthocyanidin, anthoyanin, seed coatcolor, flower color

F-29Molecular genetic assessment and quantitativedetassion of a CMV-tolerant transgenic pepperline

Pack, IS1, YJ Kim1, ES Youk1, KH Lee1, WK Yoon1, C-GKim1, CH Harn2, HM Kim1, and SC Jeong1

1Bio-evaluation Center, KRIBB2Nongwoo Bio Co.

Transgenic plants that overexpress virus coat protein geneshave attracted particular interest from researchers, byvirtue of their tolerance to virus infection. The Cucumbermosaic virus (CMV-CP)-tolerant chili pepper contains theCMV coat protein under the control of CaMV 35Spromoter and NOS. In our study, the CMVP0-CP chilipepper event 7 was selected by screening. SouthernBlotting, Genomewalker and Inverse PCR analysisrevealed that the selected CMVP0-CP chili pepper event 7contained a single copy number of the inserted genecassette that was inserted into a noncoding, intergenic

region. We searched for pepper-specific DNA sequencecandidates for endogenous reference gene of GM-pepperdetection. We found that only one copy of CaSIG4 andLipocalin genes are present in pepper genome andcharacterized pepper-specific DNA sequences of CaSIG4and Lipocalin genes. The information of the integrated siteand the internal positive control sequence were used toestablish a CMVP0-CP event-specific PCR-baseddetection method, including GMO analysis. (2009 CropFunctional Genomics Workshop : Gangneung, 2009)

Keywords : transgenic plants, CMV

F-30Seed longevity and dormancy of transgenicCapsicum annuum L. resistant to cucumbermosaic virus

T-S Cha, E-M Ko, DY Kim, K-H Choi, C-G Kim, S-CJeong, WK Yoon, HM Kim, and KW Park

Bio-Evaluation Center, KRIBB

A new trait of transgenic plants may alter seed longevityand consequently increase the possibility of weediness ofthe plants. Studies to estimate seed longevity and dormancyof a cucumber misaic virus (CMV) resistant transgenicchili pepper (Capsicum annuum L.) line, CMV-CP, werecinducted from 2007 to 2009 at the Korea ResearchInstitute of Bioscience and Biotechnology, Cheongwon-gun, Chungcheongbuk-do, Korea. Seeds from the transgenicchili pepper and its non-transgenic parent line, P915 wereused. Chili pepper seeds were buried at 5, 15, and 30cm inthe upland fields in November 2007. Three differentenvironmental conditions were treated in the fields; blackplastic mulching, weed management, and no weedmanagement. The fields were managed and removed 5, 7,9, 11, and 18 months later. Based on the ANOVA, differencesin germination were not detected among environmentalconditions. However, seed germination rates of CMV-CPand P915 depended on the seed burial depth. Seeds of theP915 deteriorated faster than the CMV-CP. While 6.3% ofP915 seeds germinated five month after seed burial, 15.1%of CMV-CP seeds were germinated. Though the germinationrates were very low, seeds of both CMV-CP and P915germinated after two winter season (18 month after seedburial). If buried deeply, it could be expected that viablechili pepper seeds will persist at least 18 month after theseeds are introduced to a site. (2009 Crop FunctionalGenomics Workshop : Gangneung, 2009)

Keywords : CMV-CP, transgenic chili pepper, seedlongevity, seed dormancy

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Members

DirectorHwan-Mook Kim (hwanmook@kribb.re.kr)

Principal Researcher Hyoung-Chin Kim (hckim@kribb.re.kr)Ki-Hoan Nam (namk@kribb.re.kr)Soon-Chun Jeong (scjeong@kribb.re.kr)Won-Kee Yoon (wkyoon@kribb.re.kr)

Senior Researcher Chang-Gi Kim (cgkim@kribb.re.kr)Jong-Soon Kang (kanjon@kribb.re.kr)Kee-Woong Park (park@kribb.re.kr)Young-Suk Won (yswon@kribb.re.kr)

Post-DocHyo-Jung Kwon (hjkwon9@kribb.re.kr)Moo-Rim Kang (mrkang@kribb.re.kr)Kyung-Hwa Choi (choi978@kribb.re.kr)

ResearcherIn-Soon Pack (bis74@kribb.re.kr)Tae-Sik Cha (last-1004@hanmail.net)Do-Young Kim (nemodol2@kribb.re.kr)Eun-Soo Youk (gomi6@kribb.re.kr)Hyun-Ju Lee (hunju3575@hanmail.net)Jong-Tak Han (majong82@kribb.re.kr)Ki-Hwan Park (pkhw@kribb.re.kr)Jang-Hyun Kim(fck108@naver.com)Nam-Hee Jeong (Z699@kribb.re.kr)Soo-Jin Oh (diatree@kribb.re.kr)Sae-Bhom Lee (dameya5053@kribb.re.kr)Sang-Woon Kim (z5r5@kribb.re.kr)Yu-Jin Kim (liu3k@kribb.re.kr)

F-31Molecular characterization of outer membranevesicles produced from shiga toxin mutants ofEscherichia coli O157:H7

Sang-Hyun Kim, Keun-Su Kim, Ara Ko, Sang-Rae Lee,Ekyune Kim, and Kyu-Tae Chang

National Primate Research Center, KRIBB

Shiga toxins (STx) elaborated by an Escherichia coliO157:H7 are believed to be localized in the periplasm andsecreted into extracellular milieu via export mechanism(s)that still remains to be fully elucidated. In this study, wehave shown that STx toxins are secreted extracellularlythrough the released outer membrane vesicles (OMVs) inwhich STx are entrapped within the vesicle, in a normalgrowth condition. In an attempt to produce STx-negativeOMVs from E. coli O157:H7, series of stx-specificmutations were introduced into the chromosomal loci inwhich two types of STx-encoding operon exist. By site-specific deletions of the stx1A and stx2A subunit genes, weengineered the mutant to produce OMVs showing theSTx-negative phenotype when assessed by Vero cellcytotoxicity assay. Western blot analyses with the STx-negative OMVs showed that the non-toxic B subunitswere present without the A subunit counterpart.Furthermore, the B subunits were in its homo-pentamericcomplexes, as revealed by immunoprecipitation analysiswith anti-STxB monoclonal antibody. These resultssuggest that the B subunit alone can be oligomerized intothe B-pentamer in the periplasm without thetoxic Asubunit, and subsequently incorporated into the OMVs.Thus, considering that STx-B subunit has been proposedto have antigen carrier inducing CTL response, the OMVscontaining STx-B subunit fused with a particulate antigenwould be utilized for development of safer combinedvaccine delivery vehicles. (International Meeting of theFederation of Korean Microbiological Societies : Seoul,2009)

Keywords : OMV toxicity, shiga toxin, O157:H7, LD50,vaccine vehicles

F-32Role of estradiol-17ß on porcine blastocystsproduced by in vitro fertilization or somatic cellnuclear transfer

Ji-Su Kim1, Bong-Seok Song1, Deog-Bon Koo2, and Kyu-Tae Chang1

1National Primate Research Center, KRIBB2Department of Biotechology, College of Engineering,Daegu University

Korea National Primate Research Center

We evaluated the developmental competence of in vitrofertilization (IVF) and somatic cell nuclear transfer(SCNT) embryos using in vitro embryo culture systems.The major role of estradiol-17ß (E2) is to prepare thereproductive system for breeding and pregnancy, and peakE2 levels are reached prior to ovulation. In this study, weevaluated the effect of E2 in porcine embryos duringpreimplantation stages. Porcine oocytes were matured inthe NCSU-23 medium supplemented with 10% (v/v)porcine follicular fluid, 10 ng/ml EGF, 25 µM ß-mercaptoethanol, 0.57 mM cysteine, 10 IU/ml PMSG, 10IU/ml hCG, 1 µg/ml E2 for 22 h and further culturedwithout hormones for an additional 22 h. After in vitrofertilization, porcine embryos were cultured in NCSU23medium with 4 % BSA and 200ug/ml at 39ºC, 5 % CO2 inair for 6d. In present result, E2 treated (85.5 ± 3.8%)oocytes showed a higher proportion of the metaphase IIstage than non-treated (72.8 ± 2.6%) ones (P<0.05). In theE2 treated group, polyspermic rate (38.3 ± 1.0%) wasreduced as compared to that of non-treated group (47.4 ±0.9%). Furthermore, the rate (22.0 ± 5.4%) of blastocystformation in E2 treated group was higher than non-treatedgroup (17.2 ± 5.6%; P<0.05). Also, culturing IVF andSCNT embryos with E2 significantly increased the numberof total cells in blastocysts and decreased TUNELpositive-apoptotic nuclei (P<0.05). Our results indicatethat maturation condition of porcine oocytes may affectapoptosis and embryonic quality during preimplantationdevelopment. (The 21st Annual Meeting of the KoreanSociety for Molecular and Cellular Biology : Seoul,2009)

Keywords : estradiol-17ß (E2), SCNT, pocine

F-33Gain of new exons and promoters by lineage-specific transposable elements-integration andconservation event on CHRM3 gene

Young-Hyun Kim, Jae-Won Huh, Dae-Soo Kim, Sang-JePark, and Kyu-Tae Chang

National Primate Research Center, KRIBB

The CHRM3 is a member of the muscarinic acetylcholinereceptor family that plays important roles in the regulationof fundamental physiological functions. The evolutionarymechanism of exon-acquisition and alternative splicing ofthe CHRM3 in relation to transposable elements (TEs)were analyzed. Five different transcripts (T1, T2, T3, T3-1, and T4) derived from three distinct promoter regions(T1: L1HS, T2, T4: original, T3, T3-1:THE1C) were identified.A placenta (T1) and testis (T3 and T3-1)-dominatedexpression pattern appeared to be controlled by differentTEs that were integrated into the common ancestor or

genome during primate evolution. Remarkably, the T1transcript was formed by the integrationevent of the humanspecific L1HS element. Among the 12 different brain regions,the brain stem, olfactory region, and cerebellum showeddecreased expression patterns. Evolutionary analysis ofsplicing sites and alternative splicing suggested that theexon-acquisition event was determined by a selection andconservation mechanism. Furthermore, continuousintegration events of TEs could produce lineage specificalternative transcripts by providing novel promoters andsplicing sites. (The 21st Annual Meeting of the KoreanSociety for Molecular and Cellular Biology : Seoul,2009)

Keywords : CHRM3 gene, transposable elements, alternativesplicing, exon-acquisition, L1HS element

F-34Bioinformatic analysis of constitutive TE-containing exons and nonsense-mediated mRNAdecay within human genome

Dae-Soo Kim, Jae-Won Huh, Young-Hyun Kim, Sang-JePark, and Kyu-Tae Chang

National Primate Research Center, KRIBB

About half of the human genome derives from discernibletransposed elements. Recently, the existence of Alu whichcan trigger nonsense-mediated mRNA decay wasexperimentally proved. In this study, we systematicallyidentified retrotransposable elements which can lead NMDfrom human EST involving genes. The result shows that139 retrotransposable element can make prematuretermination codon (PTC) to nonsense-mediated decaywhen they are analyzed with human expressed sequencetags and mRNA dataset. We found that NMD-triggeringretrotransposable elements are enriched in an Alu family.We believe this discovery can provide biologists insightinto a function of retrotransposable elements as sources ofnonsense-mediated decay, which are able to affect phenotypesincluding diseases. We suggest that retrotransposableelements to NMD process are a mechanism contributing tothe evolution of gene regulation in human genome as wellas having implications to diseases. (The 21st AnnualMeeting of the Korean Society for Molecular andCellular Biology : Seoul, 2009)

Keywords : retrotransposable element, nonsense-mediatedmRNA decay, Alu

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Korea Research Institute of Bioscience and Biotechnology 82

F-35Functional analysis of VSIG1 in mammalianspermatogenesis

Ekyune Kim1, Youngjeon Lee1, Sang-Hyun Kim1, Sang-Rae Lee1, Yonggeun Hong2, and Kyu-Tae Chang1

1National Primate Research Center, KRIBB2Department of Rehabilitative Science, Graduate School ofInje University

VSIG1 has been characterized as a member of theadhesion molecules, which consists of two extracellularimmunoglobulin(Ig)-like domains, a transmembranedomain, and a cytoplasmic domain. The expression ofVSIG1 was reported predominantly in the stomach andtestis. However, the function has not been fully characterized.We investigated the role of VSIG1 in spermatogenesis.We generated an antibody against mouse VSIG1 andexamined the localization. RT-RCR and Western blotanalysis of the mouse tissues indicated that VSIG1 wasexpressed specifically in testis. Trypsinization andbiotinylation assays support VSIG1 is localized on thegerm cell surface. In order to determine whether VSIG1participate in homotypic interactions, we producedglutathione S-transferase(GST) fusion proteins againsteach domain. Western blot analysis demonstrated thateach Ig-like domain could form homophilic complex,while the cytoplasmic domain could not. Pull-down assayalso demonstrated that each GST-fusion Ig-like domainhas homotypic binding. These results show VSIG1 is anew component of testicular specific members, andpotentially interacts with the sertoli cell surface proteinsduring spermatogenesis. (The 21st Annual Meeting ofthe Korean Society for Molecular and Cellular Biology: Seoul, 2009)

Keywords : VSIG1, spermtogenesis, testis, Ig-like domain

Members

DirectorKyu-Tae Chang (changkt@kribb.re.kr)

Principal Researcher Myeong-Su Kim (myeongsu@kribb.re.kr)

Senior Researcher Ekyune Kim (kimek@kribb.re.kr)Sang-Hyun Kim (skim@kribb.re.kr)Sang-Lae Lee (srlee@kribb.re.kr)Jae-Won Huh (huhjw@kribb.re.kr)

Post-DocDae-Soo Kim (kds2465@kribb.re.kr)Ji-Su Kim (vj-man@hanmail.net)Bong-Seok Song (sbs6401@hanmail.net)

ResearcherYoung-Hyun Kim (kyh@@kribb.re.kr)Young-Jeon Lee (neurosci@kribb.re.kr)Sang-Je Park (sangje82@kribb.re.kr)Sung-Woo Kim (adamsw123@hanmail.net)Kang-Jin Jeong (nemo9426@kribb.re.kr)Sang-Il Lee (qq2qq4@kribb.re.kr)Bong-Sang Cho (finalfz@kribb.re.kr)Cheon-Kyu Park (pcg2813@kribb.re.kr)Sang-Yong Lee (active@kribb.re.kr)Pil-Yong Kang (gt1300@kribb.re.kr)

Jeonbuk Branch Institute

Molecular Bioprocess Research Center

Bioindustry Research Center

3 r d K R I B B P o s t e r F e s t i v a l

85 3rd KRIBB Poster Festival

G-1Development new enzyme screening protocolsfor high throughput screening system

YunJon Han, YongMo Kim, Hyun Jung, Cho, Jong HyunChoi, Joong Su Kim, and Jae Jun Song

Molecular Bioprocess Research Center, KRIBB

Glycosyltransferases(GTs) constitute a superfamily ofenzymes involved in synthesizing the carbohydrate moietiesof several biological compounds including proteins, lipids,steroids, and other small molecules. The GT superfamilyis categorized into 78 families. Of these 78 families, theFamily 1 GT contains the UDP-glycosyltransferases(UGTs)that have been found in plants, animals, fungi, andbacteria. The UGTs can convert many small compoundssuch as flavonoids and alkaloids. Aldolases catalyze thereversible addition of a donor onto an acceptor and arehighly attractive tool in biosynthesis of rare sugar or itsderivatives. Among them, deoxyribose 5-phosphatealdolase(DERA) is important and useful in industrialbiosynthesis of biological active compounds such asstatins and epothilone. Both of above enzymes are difficultto determine activity by the plate assay method. Thus itwas difficult for mass screening. In this study, wedeveloped mass screening methods for UGT and DERA.UGT activity was measured fluorescence disappear usingthe 4-methylumbelliferone as substrate. DERA activitywas measured fluorescence increase using synthesizedsubstrate(5’-(4-methylumbelliferyl)-2’-deoxy-D-ribose).Finally optimized screening methods for high throughputsystem led to 8,000 clone active verification per day wereavailable. (International Meeting of the Federation ofKorean Microbiological Societies : Seoul, 2009)

Keywords : HTS system, glycosyltransferases, aldolase,4-methylumbelliferone

G-2New screening strategy of Exo-type cellulasefrom metagenomic resources based on high-throughput screening (HTS) system

Kyong-Cheol Ko, Jong Hyun Choi, Yun-Jon Han, JoongSu Kim, and Jae Jun Song

Molecular Bioprocess Research Center, KRIBB

In general, various endo-type cellulases can be easily screenedon agar plates by staining degraded residual polysaccharideswith various dyes. However, it is difficult to screen outexo-type cellulases from various bacterial resources due tolimited assay protocol, expensive substrate and low sensitivity.In the present study, we describe new strategy fol screening

of novel exo-type cellulases from various metagenomicresources by using high-throughput screening (HTS)system based on robot control. This new strategy combinedwith HTS system could screen exo-type cellulases more fast,sensitive and easy than previously reported screeningmethods. This approach would be powerfully applied forother useful enzyme screening from metagenomic resources.(International Meeting of the Federation of KoreanMicrobiological Societies : Seoul, 2009)

Keywords : high-throughput screening (HTS), netagenome,Exo-type cellulase, enzyme assay

G-3Screening of various microbial hydrolases frombovine rumen

Kyong-Cheol Ko, Jong Hyun Choi, Yun-Jon Han, JoongSu Kim, and Jae Jun Song

Molecular Bioprocess Research Center, KRIBB

The bovine rumen has a number of microorganism, suchas bacteria, protozoa, fungi, archaea and viruses. Thesecomplex microbial communities have adapted to thebovine rumen and show symbiotic relationship with thecows. These rumen microbes can easily hydrolyzecellulose, starch, protein and lipid and help host to digesttheir foods. These hydrolase has been extensively used invarious industrie. In this study, we isolated rumen bacteriaproducing various useful hydrolases based on activity-based screening method. Various aerobic and anaerobicmicroorganisms were isolated from bovine rumen by itscellulolytic, lypolytic, and proteolytic activity on NB agarplate containing AZO-CM-cellulose, tributyrin and skimmilk, respectively. Isolated microbes identified by thecomparison of full-sequences of 16S rRNA. Genesencoding cellulase, lipase and protease in isolated strainswere cloned with shotgun method and characterized. Thiswork was supported by a high throughput screening(HTS)-based Integrated Technology Development grantfrom the MEST, and a basic research grant from KRIBB.(International Meeting of the Federation of KoreanMicrobiological Societies : Seoul, 2009)

Keywords : screening, hydrolase, cellulase, lipase, protease,cloning

G-4Elimination of by-products formation duringproduction of 1,3-propanediol from glycerol in

Molecular Bioprocess Research Center

Korea Research Institute of Bioscience and Biotechnology 86

Klebsiella pneumoniae by inactivation of theoxidative pathway for glycerol metabolism

Mi-Young Seo, Jeong-Woo Seo, Sun-Yeon Heo, Jin-OhBaek, Dina Rairakhwada, Baek-Rock Oh, Pil-Soo Seo,Min Ho Choi, and Chul Ho Kim

Molecular Bioprocess Research Center, KRIBB

Klebsiella pneumoniae is a typical microbial strain capableof producing 1,3-propanediol that is a valuable chemicalthat is used mainly for the synthesis of polymethyleneterephalates by polymerization with terephthalates (1). Themicrobial production of 1,3-PD by K. pneumoniaeinvolves the formation of various by-products which aresynthesized through the oxidative pathway. 2,3-Butanediol, one of the main by-products, may serve as anobstacle for obtaining a high purity of 1,3-PD indownstream processes because of its similar boiling point(2). Hence, to eliminate the by-products synthesis, theoxidative branch of glycerol metabolism was inactivatedby deleting from the chromosomal DNA the genesinvolved in the synthesis of by-products. The by-productformation except for acetate was eliminated in theresultant strains, giving the production yield of 0.57mol/mol which was higher as compared to that of the wildtype strain which gave the production yield of 0.47mol/mol. (KSBB Spring Meeting and InternationalSymposium : Pohang, 2009)

Keywords : Klebsiella pneumoniae, 1,3-propanediol,glycerol metabolism, by-product, geneticengineering

G-5Statistical optimization of two-step fermentationfor 1,3-propanediol production by recombinantKlebsiella pneumoniae

Baek-rock Oh1,2, Jeong-Woo Seo1, Mi-Young Seo1, Sun-Yeon Heo1, Jin-Oh Baek1, Pil-Soo Seo1, Min Ho Choi1,Don-Hee Park2,3, and Chul Ho Kim1

1Molecular Bioprocess Research Center, KRIBB2School of Biological Sciences and Technology, ChonnamNational University

3Interdisciplinary Program of Graduate School forBioenergy & Biomaterials, Chonnam National University

Klebsiella pneumoniae is an excellent 1,3-PD producer,but the simultaneous production of many by-products(2,3-butandiol, Ethanol, Succinic acid, etc.) have greatlyreduced the fermentation efficiency of 1,3-PD. Hence, toeliminate the by-products synthesis, the oxidative branchof glycerol metabolism was inactivated by deleting fromthe chromosomal DNA the genes involved in the

synthesis of by-products. The microbial production of1,3-PD by recombinant K. pneumoniae was investigatedin a two-step fermentation process. The first step, includethe growth of the cells in LB broth, till absorbance at 600nm reached 1.5~2.0. The second step, involved theglycerol feeding at this stage, using this strategy glycerolwas rapidly converted to 1,3-PD. In the present study, weattempted to statistically optimize the two-stepfermentation conditions for 1,3-PD by the engineeredstrain of K. pneumoniae using response surfacemethodology (RSM) based on a 25factorial centralcomposite design (CCD) (1). The optimum conditions fortwo-step fermentation were: 6.5 pH; 0.7 vvm aerationvolume; 25.0 g/L glycerol concentration; 15.0 hfermentation time and 35.74C temperature. Theexperimental production of 1,3-PD was 11.29 g/L, whichwas in close agreement with the model prediction. In thisresults, the productivity of 1,3-PD and the yield of 1,3-PDrelative to glycerol reached 0.63 g/Lh and 0.66 mol/mol,respectively, which were about 2 fold higher than those ofone-step fermentation. (KSBB Spring Meeting andInternational Symposium : Pohang, 2009)

Keywords : Klebsiella pneumoniae, 1,3-propanediol,glycerol metabolism, response surfacemethodology, central composite design

G-6Identification and utilization of a 1,3-propanediol oxidoreductase isoenzyme forproduction of 1,3-propanediol from glycerol inKlebsiella pneumoniae

Sun-Yeon Heo1, Jeong-Woo Seo1, Mi-Young Seo1, Baek-Rock Oh1,2, Jin-Oh Baek1,3, Dina Rairakhwada, Lian HuaLuo1,4, Won-Kyung Hong1, and Chul Ho Kim1

1Molecular Bioprocess Research Center, KRIBB2School of Biological Sciences and Technology, ChonnamNational University

3School of Life sciences and Biotechnology, Korea University4Institute for Molecular Biology and Genetics, ResearchCenter of Bioactive Materials, Chonbuk National University

As a potential candidate for another putative 1,3-propanediol (1,3-PD) oxidoreductase of Klebsiellapneumoniae involved in production of 1,3-PD fromglycerol, we identified and characterized a homolog ofEscherichia coli yqhD (88% homology in amino acidsequence), which encodes an alcohol dehydrogenase1).Introduction of multiple copies of the yqhD homologrestored 1,3-PD production in the mutant K. pneumoniaestrain defective in 1,3-PD oxidoreductase DhaT2). Anincrease in NADP-dependent 1,3-PD oxidoreductaseactivity was observedin the recombinant strain harboring

multiple copies of the yqhD homolog, whereas NAD-dependent DhaT activity was not affected. The level of1,3-PD production during batch fermentation in therecombinant strain was comparable to that of the parentstrain, suggesting that further engineering can generate annovel 1,3-PD producing industrial strain connected todifferent cellular metabolic network. (InternationalSymposium & Annual Meeting of the Korean Societyfor Microbiology and Biotechnology : Daejeon, 2009)

Keywords : Klebsiella pneumoniae, 1,3-propanediol,glycerol metabolism, 1,3-PD oxidoreductase

G-7Cloning and expression of Bacillusamyloliquefaciens type 1 levansucrase gene inBacillus megaterium and optimization offermentation conditions for the production oflevansucrase by recombinant strain

Dina Rairakhwada, Jeong-Woo Seo, Baek-Rock Oh, andChul-Ho Kim

Molecular Bioprocess Research Center, KRIBB

An expression system of levansucrase gene from Bacillusamyloliquefaciens type 1 was conducted in Bacillusmegaterium MS9411). Prominent expression of levansucrasewas obtained through xylose induction (0.5%) at 37ºC inB. megaterium, wherein recombinant His6taggedlevansucrase was secreted up to 3 hours after inductionshowing maximum enzyme activity of 12,906 U/l andfurther incubation up to 6 and 9 hours led to degradedlevansucrase showing 10 fold and 31 folds lower enzymeactivity (1210 U/l and 410 U/l respectively). Responsesurface methodology (RSM) was further employed tooptimize the fermentation condition and to improve thelevel of levansucrase production. Maximum levansucraseactivity of 20,250 U/l was obtained in 12 hours of thefermentation carried out at 28ºC, starting the inductionwith 0.735% xylose when the O.D.600 nm was 1.2, whichwas 1.7 and 40 fold higher level as compared to thoseobtained by the non-optimized condition and the nativestrain, respectively. (International Symposium & AnnualMeeting of the Korean Society for Microbiology andBiotechnology : Daejeon, 2009)

Keywords : Bacillus amyloliquefaciens, levansucrase,Bacillus megaterium, response surfacemethodology

G-8Expression of human papillomavirus Type 16,18, 33, 58 L1 proteins and production of virus-like particles for vaccination in Sf-9 cells

Jin-Oh Baek1,2, Jeong-Woo Seo1, Shin-Je Ghim3, Ik-HwanKim2, and Chul Ho Kim1*

1Molecular Bioprocess Research Center, KRIBB2School of Life sciences and Biotechnology, KoreaUniversity

3Department of Pathology, Georgetown UniversityMedical Center, USA

Major capsid protein L1 genes of Human papilloma virus(HPV) type 16, 18, 33, 58 were expressed in Sf-9 insectcells using a baculovirus system. Each L1 genespreviously isolated from Korean patients were ligated intotransfer plasmid pBlueBac to construct the expressionvector and transfected into Sf-9 cells via recombining withlinearized Autographa californica multicapsid nuclearpolyhedrosis virus (AcMNPV) DNA as a helper DNA.Introduction of insert DNA was analyzed by PCR methodusing DNA isolated from recombinant baculovirus as atemplate, and the titer of virus was calculated by plaqueassay. Expression and immunological activity of HPVtype 16, 18, 33, 58 L1 proteins were confirmed by SDS-PAGE, Western blotting and ELISA using mouse anti-HPV16 L1 monoclonal antibody as a primary antibody.Hemagglutination inhibition (HAI) assay was performedto detect the neutralization ability of VLPs between mousered blood cells (MRBCs) and HPV antibody. To assessformation of virus-like particles (VLPs) by self-assemblyof HPV type 16, 18, 33, 58 L1 proteins in the insect cells,immunoactive fractions against the monoclonal antibodywere prepared through ultracentrifugation (10~50% ofsucrose gradient, 36,000rpm, 3 hr, 4ºC) and analyzed byTransmission electron microscopy (TEM). As a result,VLPs of approximately 50~60nm in diameter wereidentified, which could be useful for development ofcervical cancer vaccine. (International Meeting of theFederation of Korean Microbiological Societies : Seoul,2009)

Keywords : human papilloma virus, major capsid proteinL1, virus-like particles, Sf-9 insect cell,baculovirus

G-9Influence of cellular fatty acids composition onethanol tolerance in Escherichia coli

Lian Hua Luo1, Pil-Soo Seo1, Jeong-Woo Seo1, Sun-YeonHeo1, Dae-Hyuk Kim2, and Chul Ho Kim1

87 3rd KRIBB Poster Festival

Korea Research Institute of Bioscience and Biotechnology 88

1Molecular Bioprocess Research Center, KRIBB2Institute for Molecular Biology and Genetics, Research

Center of Bioactive Materials, Chonbuk National UniversityTo investigate effect of cellular fatty acids composition onethanol tolerance in Escherichia coli, Bacillus subtilis desencoding fatty acid desaturase, E. coli fabA encoding ß-hydroxydecanoyl thio ester dehydrase or both genes wereoverexpressed. E. coli recombinant cells harboring fabAshowed elevated tolerance against ethanol compared towild type strain. In contrast, desencoding fatty aciddesaturase resulted in a decreased resistance againstethanol and co-expression of des with fabA complementedthe ethanol tolerance of E. coli. The result could give aclue to engineer bacterial strains resistant to higherconcentration of ethanol. (Fall Meeting & InternationalSymposium of the Korean Society for Biotechnologyand Bioengineering : Daejeon, 2009)

Keywords : Escherichia coli, fatty acids composition,ethanol tolerance, desaturase, fabA

Members

DirectorJae-Jun Song (jjsong@kribb.re.kr)

Principal Researcher Chul-Ho Kim (kim3641@kribb.re.kr)Joong-Su Kim (joongsu@kribb.re.kr)Hyo-Kon Chun (hkchun@kribb.re.kr)

Senior Researcher Jong-Hyun Choi (jhchoi@kribb.re.kr)Jeong-Woo Seo (jwseo@kribb.re.kr)

Post-DocMin-Ho Choi (choimh@kribb.re.kr)Soon-Ok Rim (rimso@kribb.re.kr)Kyong-Cheol Ko (kcko@kribb.re.kr)Jang-Yeol Choi (yeol11@kribb.re.kr)

ResearcherWon-Kyung Hong (hongwk@kribb.re.kr)Sun-Yeon Heo (heosy@kribb.re.kr)Baek-Rock Oh (5backwon@kribb.re.kr)Jin-Oh Baek (seraph@kribb.re.kr)Luo-Lian hua (lotus@kribb.re.kr)Seung-Yong Park (dilko@kribb.re.kr)Soo-Hyun Kim (shyoun99@kribb.re.kr)Bong-Seok Shin (highelf7@kribb.re.kr)Dong-Ho Son (hoyanigg@kribb.re.kr)Bum-Jin Kwon (cooluk20@kribb.re.kr)Dong-Min Jeong (dmchung@kribb.re.kr)Yu-Mi Song (songyumi@kribb.re.kr)

G-10Antiviral effect of isolated flavonols fromRhodiola rosea on influenza A virus

Hyung-Jun Kwon, Young Bae Ryu, Hyung Jae Jeong,Jang Hoon Kim, Su-Jin Park, Mun-Chual Rho, and WooSong Lee

Bioindustry Research Center, KRIBB

Influenza virus is highly contagious and can cause severerespiratory illness as like pandemics. In this study,kaempferol (1), gossypetin (2), rhodionin (3), rhodiosin(4), and rhodiolinin (5) isolated from Rhodiola rosea wereevaluated for their antiviral effect. Five isolated flavonolcompounds (1-5) were tested in neuraminidase (rvH1N1)inhibition assay and influenza A virus (H1N1) in MDCKcells. In evaluation of neuraminidase inhibitory effects, theIC50 values of all compounds (1-5) showed a ranged of2.5-64.2 mM, respectively. Also, compounds 1-5 showedhighly biological activities against influenza A virus(H1N1) with IC50 values of 62.4, 7.9, 16.6, 13.9, and 19.4mM, respectively. Interstingly, gossypetin 2 exhibited themost potent inhibitor in both assay systems. As a result,biological activities of flavonols 1-5 have proven to bedepending upon the number of the hydroxy group onflavonol backbone. (

: Gwangju, 2009)

Keywords : antiviral, flavonol, neuraminidase, Rhodiolarosea

G-11Characterization of bacteriocin produced byLactococcus lactis ET-45 isolated from Kimchi

Seong-Yeop Jeong, Nack-Shick Choi, Chan-Sun Park,Keug-Hyun Ahn, Hee-Jong Yang, Byung-Dae Yoon, andMin-Soo Kim

Bioindustry Research Center, KRIBB

Bacteriocin producing lactic acid bacteria were isolatedfrom kimchi using a MRS selective plate with Bacilluscereus as an indicator strain. 16S rDNA sequence andsugar utilization test identified that ET-45 was aLactococcus lactis strain. The bacteriocin exhibitedinhibitory activity against Bacillus cereus, Leuconostocmesenterides, Leuconostoc carnosum, Leuconostoc lactis,Leuconostoc mesenteroides subsp. cremoris, Lactobacillusplantarum, Lactobacillus acidophilus, Lactobacillusviridesceus, Pediococcus dextrinicus, and Enterococcusfacecium. Optimal production of the bacteriocin wasobtained by growing the cells on MRS medium at pH 7.5and 30ºC for 18 ~ 24 hrs. Sucrose and protease peptone

Bioindustry Research Center

are essential for bacteriocin production as carbon sourceand nitrogen source, respectively. Antibacterial activity ofthe bacteriocin was completely disappeared by proteinaseK, which indicates it’s proteinous nature. The bacteriocinwas inactivated by protease such as trypsin, -chymotrysin,Subtilisin A. The bacteriocin was fully stable at 121ºC for60 min. Solvents such as chloroform, ethanol, acetone,acetonitrile, hexane, isopropanal did not effect on theactivity. (International Symposium & Annual Meetingof the Korean Society for Microbiology and Biotechnology: Daejeon, 2009)

Keywords : bateriocin, Lactococus lactis, kimchi

G-12The pure ß-glucan production in Aureobasidiumpullulans IMS-822 KCTC11179BP by Pullulansynthetase gene disruption

Hee-Jong Yang1,2, Chan-Sun Park1, Nack-Shick Choi1,Keug-Hyun Ahn1, Kyu-Woong Hahn2, and Min-Soo Kim1

1Bioindustry Research Center, KRIBB2Department of Biological Sciences, Hannam University

By disruption of pullulan synthetase gene in Aureobasidiumpullulans IMS822 KCTC11179BP, we constructed amutant strain, A.pullulansNP1221, which can produce apure ß-glucan exopolysaccharide. The mutant NP 1221was white-colored cell morphology, whereas the wild typestrain produced a black dye within 36 hrs on Czapek agarplate. When compared the fermentation kinetics betweenwide type and mutant strains, the mutant NP 1221 did notproduce pullulan, which confirmed by production of ß-1,3glycosidic linkage with laminarinase treatment. Thesubstrate uptake rate and ß-glucan production were shownsimilar patterns in both strains. On the other hand, thebiomass yield of mutant NP 1221 was 2.3 fold (9.2 g·l-1)higher than that of wild type. To optimize the cultivationof the mutant NP 1221 various culture conditions (carbon,nitrogen and ascrobic acid) were applied. Based on theresponse surface method(RSM), the optimal concentrationsof sucrose, KNO3 and oxalic acid were 4.74%, 0.19% and0.036%, respectively. Plus, temperature and pH were alsooptimized. Finally, the yield of ß-glucan was 10 foldsincreased. (International Symposium & Annual Meetingof the Korean Society for Microbiology and Biotechnology: Daejeon, 2009)

Keywords : bateriocin, Lactococus lactis, kimchi

Members

Director Woo-Song Lee (wslee@kribb.re.kr)

Principal Researcher Byung-Dae Yoon (bdyoon@kribb.re.kr)Mun-Chual Rho (rho-m@kribb.re.kr)

Senior Researcher Jong-Sun Chang (changjs@kribb.re.kr)Su-Jin Park (sjpark@kribb.re.kr)Min-Soo Kim (ms5732@kribb.re.kr)Cha-Young Kim (kimcy@kribb.re.kr)Young-Bae Ryu (ybryu@kribb.re.kr)

Post-DocNack-Shick Choi (nschoi@kribb.re.kr)Seung-Woong Lee (swn2000@kribb.re.kr)Hyung-Jae Jeong (hjeong21@kribb.re.kr)Young-Min Kim (u9897854@kribb.re.kr)

ResearcherChan-Sun Park (chansun@kribb.re.kr(Hyung-Jun Kwon (hjkwon@kribb.re.kr)Byung-Dae Yoon (bdyoon@kribb.re.kr)Keug-Hyun Ahn (keughyun@kribb.re.kr)Seong-Yeop Jeong (jsyoung@kribb.re.kr)Hee-Jong Yang (godfilts@kribb.re.kr)

89 3rd KRIBB Poster Festival

Open Innovation

Cooperation Between Medicine and Biotechnology

BINT Convergence Technology

3 r d K R I B B P o s t e r F e s t i v a l

91 3rd KRIBB Poster Festival

H-1 Sentinel node detection with PET/MR/fluorescentmulti-modal imaging using 68Ga-SCN-NOTAsilica nanoparticle

Young-Hwa Kim1,2,3,4, Hyewon Youn1,4, KeonWook Kang1,3,4,JunSung Kim5, Yun-Sang Lee1,4, JaeMin Jeong1,3,4, DongSooLee1,4, and June-Key Chung1,2,3,4

1Department of Nuclear Medicine, Seoul National UniversityCollege of Medicine

2Department of Biomedical Sciences, Seoul National UniversityCollege of Medicine

3Laboratory of Molecular Imaging and Therapy, CancerResearch Institute Seoul National University

4Institute of Radiation Medicine, Medical Research Center,Seoul National University

5Biterials Co., Ltd

Purpose: Recently, nanoparticles have shown greatpromise and powerful tools for bio-imaging. We examinedthe feasibility of PET/MR/fluorescent multi-modal imagingsystem for sentinel node detection by using functionalizedsilica-nanoparticle. Materials and Methods: We developed50 nm sized amino PEGylated silica nanoparticlescontaining near infrared fluorescent dye, NIR-797 and Coferrite core (MNP-SiO2). MNP-SiO2 were conjugated witha chelating agent to amine, labeled with the 68Ga,andinjected subcutaneously into foot-pad of mice. Whole-body images were obtained using an animal PET/CTscanner, microMR imaging system, and Maestro opticalimaging system. After acquiring PET and MR images,fusion of PET and MR images was performed using fourfiducial markers in the Functional Image Registration(FIRE) program. Results: PET, MR and fluorescentsignals were observed from the lymph nodes drainingfoot-pad in living mice. PET/fluorescent signals were alsoobserved ex vivo after dissecting the lymph node. And wealso observed respectively images of PET/MR are correlatedwith fusion images. Conclusion: We demonstrated thatour novel probes, 68Ga-SCN-NOTA-MNP-SiO2nanoparticlescan define sentinel nodes in mice following footpadinjection. Multi-modal silica nanoparticles may have apotential for visualizing sentinel nodes using the PET,MR, and fluorescent scanner during the operation andperi-operation. (The 2009 World Molecular ImagingCongress : Montreal (Canada), 2009)

Keywords : imaging agents, nanoparticles, near-infraredfluorescence, magnetic resonance imaging,positron emission tomography

H-2HSP90 regulates angiotensin II-induced

hypertrophy via STAT pathway and IL-6secretion in vascular smooth muscle cell

Seok-Min Kang, Ji Hyung Chung, Kyung-Hye Lee, Ji-WonChoi, and Yangsoo Jang

Cardiovascular Research Institute Yonsei University Collegeof Medicine

Heat shock protein 90 (HSP 90), one of the most abundantproteins in the eukaryotic cells, is essential for cellsurvival. Vascular smooth muscle cells (VSMCs)hypertrophy which is characterized increased proteinsynthesis is critical in vascular remodeling associated withhypertension, atherosclerosis, and restenosis. It has beenknown that angiotensin II plays a critical role in thecellular hypertrophy. Moreover, HSP 90 can directlyinteract with STAT proteins. So, we assessed thehypothesis that HSP 90 regulates Ang II-induced VSMCshypertrophy through nuclear translocation of STATs. Weobserved protein/DNA content was increased by Ang II ina concentration-dependent manner in the VSMCs. Asimilar result was obtained using the [3H] leucineincorporation assay. During that period, nucleartranslocation of STAT 1, 3 and IL-6 secretion wereobserved. Interestingly, treatment with HSP 90 inhibitorgeldanamycin (GA) significantly inhibited the effect ofAng II on hypertrophy of VSMCs. A same result wasobtained using HSP 90 siRNA. When VSMCs weretreated with GA or HSP 90 siRNA, Ang II-inducednuclear translocation of STAT 1 and STAT 3 wasinhibited. Transcription and extracellular level of IL-6were also decreased. These results indicate that Ang II-induced cellular hypertrophy via STAT nucleartranslocation following IL-6 secretion and this pathwaycan be regulated by modulating HSP 90 activity. So HSP90 is another target protein for cardiovascular disease,especially vascular remodeling. (The 73rd Annual Meetingof the Japanese Circulation Society : Osaka (Japan), 2009)

Keywords : HSP90, hypertrophy, vascular smooth musclecell

H-3Clinical significance of L1 expression in breastand thyroid cancer

Koon Soon Kim1, Song Mei Huang2, Yi Sun Jang1, Hye SooKim1, Jong Min Lee1, Hyo Jeong Hong3, Min Hee Lee4, MinhoShong4, Young Suk Jo4, and Jin Man Kim2

1Department of Internal Medicine, College of Medicine,The Catholic University of Korea,

2Department of Pathology, Chungnam National UniversitySchool of Medicine

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3Therapeutic Antibody Research Center, KRIBB4Department of Internal Medicine, Chungnam NationalUniversity School of Medicine

L1 cell adhesion molecule (L1 CAM) has been regarded asa poor prognostic factor of human cancers such as ovarian,uterine and colon cancer. However, the expression leveland clinical impact of L1 CAM in breast and thyroidcancer have been remained to be elucidated. In this study,we have investigated L1 expression by immunohistochemistryand analyzed prognostic impact of L1 expression in breastand thyroid cancer to verify the biological role of L1. Inbreast cancer, 19.6% of tumors showed positive L1 CAMexpression which was exclusively observed in invasivefront of tumors, whereas normal breast tissue adjacent totumor cells did not show L1 expression. Furthermore,positive L1 expression showed significant relationshipwith poor prognostic clinico-pathological features such aspositive axillary lymph node metastasis, higher histologicgrade, negative expression of estrogen receptor andprogesterone receptor. In thyroid cancer, aberrant L1expression was also observed in invasive front of poorlydifferentiated papillary thyroid cancer. Moreover, thetumor cells showing L1 expression showed higher nuclearß-catenin expression and elevated ERK phosphorylation.Taken together, these data suggested that L1 CAM mighthave a role in tumor aggressiveness of breast and thyroidcancers. Diagnostic and therapeutic application of L1CAM to these cancers should be investigated.

Keywords : L1 cell adhesion molecule (L1 CAM), breastcancer, thyroid cancer.

H-4The function of mitochondrial in granular cornealdystrophy type 2

Tae-im Kim, Hyun seok Ahn, Woosuk Chung, KyoungYul Seo, and Eung kweon Kim

Department of Ophthalmology, Yonsei University Collegeof Medicine,

Introduction: The corneal manifestation of granular cornealdystrophy type 2 (GCD 2) aggravates age dependentmanner. To assess mitochondrial function in the tissue andcell of the corneal dystrophy type II is helpful to understandpathology of disease. Methods: The corneal tissues fromheterozygote of GCD 2 were examines using electronmicroscope. Keratocytes from normal, heterozygote, andhomozygote were cultured and evaluated with mitotrackerand cytochrome C stain, and mitochondrial function test,western blot about complex group. After treatment ofH2O2,MitoProbeTMJC-1 Assay was done. Results:Damaged or degenerative mitochondria in keratocyte were

surrounded deposit in corneal stroma of heterozygote.Increased mitochondrial activity and increased expressionof complex group was detected in homozygote cell. However,decreased expression of complex shows in heterozygoteand homozygote in late passage. Membrane potential ofmitochondria decreased after application of H2O2 inkeratocyte of homozygote. Conclusion: The change ofmitochondrial function may influence on the pathogenesisof GCD 2.

Keywords : granular corneal dystrophy type 2, GCD 2

H-515-hydroxy prostagladin dehydrogenase (15-PGDH) is a possible early detection marker incoloncancer : analysis of prostagladin pathwayin human tissue

Dong-Hoon Yang1, Seung-Jae Myung1, In-Wha Kim2, Ho JuneSong1, Yeon Mi Ryu1, Sun Mi Lee1, Jeung Hye Han1, Jong-Soo Lee1, Soon Man Yoon1, Kyung Jo Kim1, Byung Duk Ye1,Jeong-Sik Byeon1, Suk-Kyun Yang1, and Jin-Ho Kim1

1Department of Internal Medicine, Asan Medical Center,University of Ulsan College of Medicine, Asan DigestiveDisease Research Institute

2Asan Institute for Life Sciences

Introduction : Prostaglandin E2 (PGE2) produced bycyclooxygenase-2 (COX-2) promotes colon carcinogenesis.Recently, novel enzymes to degrade (15-hydroxyprostaglandindehydrogenase,15-PGDH) or generate (microsomal PGE2

synthase 1, mPGEs-1) PGE2 have been introduced. Wehave previously demonstrated that 15-PGDH is an in vivotumor suppressor in colon tumorigenesis1. However, therehad been few reports about this novel prostaglandinpathway in human colon carcinogenesis. Aims & Methods :We aimed to investigate the level of PGE2 and theexpression of its regulating genes in the sporadic humancolon tumors. Twenty colonic adenomas and 22 coloniccarcinomas were evaluated. COX-2 and 15-PGDH expressionwas quantified with real-time PCR, and the level of PGE2

and mPGEs-1 was measured with enzyme-linkedimmunosorbent assay and western blot. The relativeexpression of each gene in the neoplastic tissues wasobtained by comparing with the matched normal colonictissues. Results: The decrease of 15-PGDH expressionwas noted in 80% of adenomas and 92.6% of carcinomas.COX-2 was overexpressed in 68.4% of adenomas and48% of carcinomas. 15-PGDH expression was reduced(P=0.000) and the levels of PGE2 and mPGEs-1 wereincreased in colonic neoplasms compared with normaltissues (P=0.013and P=0.000, respectively). However,COX-2 expression did not differ between neoplastic andnormal tissues (P=0.062).In the subgroup analysis, the

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decreased expression of 15-PGDH in neoplastic tissue wasobservedin the adenoma group, but other molecules did notshow any significant difference between neoplastic andnormal tissue. On the other hands, the expression of bothPGE2 and mPGEs-1 was significantly increased (P=0.020and P=0.000, respectively), and 15-PGDH expression wassignificantly reduced (P=0.000) in the carcinoma group.Conclusion: The expression of enzymes involved inPGE2production and degradation are significantly alteredalong with the human colon carcinogenesis. Early inactivationof 15-PGDH followed by COX-2 and mPGEs-1 activationcontributes to PGE2 production leading to human coloncarcinogenesis. 15-PGDH might be a novel candidate forearly detection markers and chemoprevention targets incolon carcinogenesis.

Keywords : 15-hydroxy prostagladin dehydrogenase (15-PGDH), coloncancer

H-6Identification of characteristic molecularsignature of acute myocardial infarction fromhuman peripheral blood

Hun-Jun Park1, Ji heon Noh2, Jung Woo Eun2, Jeong KyuKim2, Kwang Hwa2, Hyun JIn Bae2, Hongjian Xie2, Pum-Jun Kim1, and Suk Woo Nam2,*

1Cardiovascular Center, Seoul St. Mary's Hospital, Collegeof Medicine, Catholic University of Korea

2Department f Pathology, College of Medicine, TheCatholic University of Korea

Recent studies suggested that gene expression profiling ofblood cells can reflect the status of diseases, and can beapplied for classification or prediction in various fields,particularly in clinical purposes. Thus we assessed thehypothesis that gene expression changes in patients’peripheral blood can distinguish disease or disease-freegroups in the category of coronary artery diseases. Formicroarray analysis, we used the Illumina SentrixHumanRef-6 Expression BeadChipv2 containingapproximately 46,000 probe set to profile peripheral bloodRNA from 7 patients diagnosed with ST segmentelevation myocardial infarction (AMI) and treated withpercutaneous coronary intervention (n = 7) (follow-upgroup), and compared their mRNA profiles with those of10 healthy control subjects. Differential gene expressionanalysis resulted in two major clusters on dendrogram, aAMI and follow-up, control groups, suggesting thereexsist characteristic molecular signature for AMI.Supervised analysis identified 546 outlier genes asmolecular signature for AMI. Among these, we validatedspecific gene expressions of AMI by realtime-PCR. Ourresults suggest that the AMI caused large-scale gene

expression changes and this characteristic molecularsignatures of AMI may be applicable to discerning myocardialinfarction from other categorized heart diseases as well asnormal.

Keywords : acute myocardial infarction, gene expression,prediction, molecular signature

H-7Development of pH-sensitive MRI imagingcarriers and evaluation the molecular imagingtechniques using ovarian cancer animal models

Young Taik Oh1,* and Kyung Taek Oh2

1Department of Radiology, Yonsei University College ofMedicine

2College of Pharmacy, CAU

Ovarian cancer is the leading cause of death ingynecologic malignancy with advanced stage at diagnosis.To improve the survival rate of ovarian cancer, an earlydetecting method and effective chemotherapy are needed.Recently, molecular imaging makes biological processimage possible at a cellular level. In other hand,nanotechnology has been focused on a promising systemfor the prevention and overcoming of cancer with earlydetection and chemotherapy. Among them, pH-sensitivepolymeric carriers have been used in targeted antitumordrug delivery and based on the intrinsic differences betweenvarious solid tumors and the surrounding normal tissues.Typical tumor pH ranges from 7.0 to 6.5 in a xenograftanimal model and appears to be more diffuse in clinicaltumors. We established ovarian cancer model with a humanovarian cell line (The NIH:OVCAR-3) and in vivo MRimaging and are developing the pH sensitive carrier system.

Keywords : magnetic resonance imaging, ovary cancer,pH sensitive, drug delivery system

H-8Gene polymorphism study for early detectionof gastric precancerous lesions and search ofgastric cancer high-risk group

Chang Soo Eun1, Hyun Ja Kim2, Dong Soo Han1, KyuSang Song3, and Bo Youl Choi2

1Department of Internal Medicine, Hanyang UniversityCollege of Medicine

2Department of Preventive Medicine, Hanyang UniversityCollege of Medicine

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3Department of Pathology, Chungnam National UniversityCollege of Medicine

Atrophic gastritis and intestinal metaplasia of gastric mucosahave been considered to be precancerous lesions of gastriccarcinoma. However, the key susceptible genes to gastricprecancerous lesions are still poorly understood. The aimsof this study were to determine genetic polymorphismsassociated with gastric precancerous lesions and to investigatepossible interaction between susceptible genes andenvironmental factors in gastric carcinogenesis. Todetermine the genetic polymorphisms associated withgastric precancerous lesions, we performed a pilot genomewide association study (GWAS) using blood DNA fromatrophic gastritis patients (n=20), intestinal metaplasiapatients (n=20), and normal controls (n=20). To validatethe association between candidate genes from GWAS andgastric precancerous lesions, we performed SNP genotypingof candidate genes using blood DNA from 700 subjects ina community-based cohort. And we aimed to further explorepossible gene-gene interaction and gene-environmentalinteraction in gastric precancerous lesions. At present, thestudy results are yet to be announced.

Keywords : gastric precancerous lesion, genetic polymorphism,genome-wide association study, gene-environmentalinteraction

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H-10Proteome analysis for searching key markerassociated with psoriasis vulgaris

Sung-ae Kim1, Sun-young Cho1, Joohyun Ryu2, ByoungChul Park2, and Jae-We Cho1

1Department of Dermatology, School of Medicine,Keimyung University

2Medical Proteomics Research Center, KRIBB

Psoriasis is characterized by epidermal proliferationcombined with incomplete terminal differentiation, as wellas an inflammatory response responsible for the chronicnature of the lesions. The way to understand psoriasis istherefore to reach a better appreciation of the messagesthat enable the skin cells to initiate an inflammatoryresponse, and by better understanding the way in whichthe inflammatory cells responsible for innate and acquiredimmune responses are capable of bringing about proliferationand abnormal epidermal differentiation. Psoriasis researchis entering a new era. Progress in delineaspog immunogeneticsand pathomechanisms of disease brings with it a need tounderstand fully the clinical spectrum of disease andintegrate phenotype with genomics and proteomics. In thisstudy we analyzed the proteome from non-lesional andlesion skin of psoriasis usiog proteomics. Our data showedthat several proteins involviog DNA synthesis or redoxsystem were increased in lesional skin compared to non-lesional skin; TF (skrotransferrin), Tubulin beta-2C chain,ATP5B, thymidinie phosphorylase, dihydropyrimidinase-related protein 2, protein disulfide-isomerase A3, aldehydedehydrogenase, isoform 1 of 14-3-3 protein sigma, serumamyloid P-component, heat shock protein beta-1, glutathione

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S-transferase P, peroxiredoxin-2. In conclusion, modulatoryproteins for DNA synthesis and redox system play roles inpathogenesis in psoriasis, and we have tried to study thefunctional roles of these proteins in psoriasis. (

: Seoul, 2009)

Keywords : psoriasis, proteomics, redox system, DNAsynthesis

H-11Synthesis and characterization of fluorescentmagneto polymeric nanoparticles for bimodalimaging probes

Joseph Park1, Jaemoon Yang2, Jin-Suck Suh2, SeungjooHaam1, and Yong-Min Huh2

1Department of Chemical and Biomolecular Engineering,College of Engineering, Yonsei University

2Department of Radiology, College of Medicine, YonseiUniversity

Novel bifunctional fluorescent magneto polymericnanoprobes (FMPNs) were synthesized to providesimultaneous diagnostic information via magneticresonance imaging (MRI) and optical imaging. FMPNsconsist of ultra-sensitive magnetic nanocrystals thatfunction as MR probes combined with Nile Red, whichfunctions as a fluorescent probe. FMPNs were encapsulatedby a nano-emulsion method in polyvinyl alcohol (PVA,87-89% hydrolyzed) through a matrix of polymethylmethacrylate (PMMA). FMPNs exhibited excellent colloidalstability and monodispersity. The production of MR andoptical images demonstrated that FMPNs have potential asdual-mode imaging agents.

Keywords : magnetic resonance imaging Optical imagingNile red Magnetic nanocrystal ATRP

H-12VEGF gene therapy combined with mesenchymalstem cell improve neuronal survival

Sung Su An1,2, Hong Lian Jin1, Meng-Lu Liu1,2, Jin Soo Oh1,2,Keung Nyun Kim1, Do Heum Yoon1,2, and Yoon Ha1,*

1Department of Neurosurgery, Spine & Spinal CordInstitute, College of Medicine, Yonsei University,

2Brain Korea 21 Project for Medical Science, College ofMedicine, Yonsei University

Bone marrow derived-mesenchymal stem cells (BMSC)and vascular endothelial growth factor (VEGF) have been

known as therapeutic sources (for what?). In this study, weinvestigated whether the combination of treatment withpEpo-SV-VEGF and rat BMSC has synergetic effects onthe survival of a neuronal N2a cell line. The synergeticeffect of the gene/cell combined therapy on hypoxia-injuryN2a cell model was evaluated, compared with gene therapyalone and cell therapy alone, using phosphate-bufferedsaline (PBS) as control group. In the gene/cell combinedgroup, the survival of N2A cells was significantly increasedmuch more than in the gene or cell therapy alone group,compared with the PBS control. Furthermore, reversetranscription PCR (RT-PCR) analysis and TUNEL assayalso showed that the gene/cell combined treatmentsignificantly inhibited the apoptosis. Consistent with theseresults, the therapeutic effect of VEGF could be reducedby dose-dependent neutralization with a mouse anti-VEGFantibody. Taken together, our results suggested that thecombined treatment with pEpo-SV-VEGF and BMSC cansynergetically improve the survival of the neuronal N2acells. (2nd Termis World Congress in conjunction 2009Seoul Stem Cell Symposium : Seoul, 2009)

Keywords: bone marrow derived-mesenchymal stem cells(BMSC), vascular endothelial growth factor(VEGF), synergetic effect, pEpo-SV-VEGF.

H-13Therapeutic effects of hypoxia-inducible geneexpression system in rat spinal cord injurymodel

Hong Lian Jin1, Meng-Lu Liu1,2, Sung Su An1,2, Jin Soo Oh1,2,Keung Nyun Kim1, Do Heum Yoon1,2, and Yoon Ha1,*

1Department of Neurosurgery, Spine & Spinal CordInstitute, College of Medicine, Yonsei University

2Brain Korea 21 Project for Medical Science, College ofMedicine, Yonsei University

Gene therapy is one of the most important emergingtherapeutic modalities in the field of neuronal regenerativemedicine. However, the safety and lack of efficacy lead tothe major obstacles that restrict this promising therapy toenter the clinical application. In this study, a novel hypoxia-inducible gene expression system has been established forthe development of an effective and safe gene therapy forspinal cord injury. In order to achieve high gene expressionspecific to the injury tissue and avoid unwanted geneoverexpression in the normal neural tissue, both theerythropoietin 3’-untranslated region and the oxygendependent degradation domain were used in this system tocontrol the expression of the reporter luciferase gene or thetherapeutic VEGF gene under hypoxic condition. Theeffect of this hypoxia inducible gene expression system onthe targeted gene expression was evaluated both in vitro in

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the N2a cell line and in vivo in a rat spinal cord injurymodel. This system resulted in an increase of expressionspecifically in the hypoxia conditioned N2a cells and in theinjured spinal cord of model rats. Our immunofluorescencestaining images further confirmed that the increase of thetherapeutic VEGF gene expression by this hypoxia-induciblesystem was specific to both the hypoxic condition and thespinal cord injury, but not to the normoxic condition or theuninjury spinal cord. These results suggested that thehypoxia-inducible gene expression system with VEGFgene might be useful for gene therapy in spinal cordinjury. (2nd Termis World Congress in conjunction 2009Seoul Stem Cell Symposium : Seoul, 2009)

Keywords: gene therapy, hypoxia-inducible, spinal cordinjury

H-14Design and evaluation of microfluidics basedthermoelectric biosensor with split-flowmicrochannel

Yong-Hwan Choi, Seung-Il Yoon, and Yong-Jun Kim

School of Mechanical Engineering, Yonsei University

This study presents design, fabrication, and experimentalresults of a microfluidics based thermoelectric biosensorfor a label-free and real-time detection. The proposedthermoelectric biosensor consists of a thermopile withbismuth and chrome pairs, one reaction chamber, tworeference chambers, and a PDMS microchannel. In thisdevice, the reference chamber on the cold junctions andthe reaction chamber on the hot junctions are integrated ina single chip. Fluidic streams to each chamber are controlledby the split-flow microchannel. The thermoelectric biosensorwith the split-flow microchannel can self-compensate withoutusing an additional heating element, which in turn wouldeliminate the need of thermal calibration. To characterizethe fabricated biosensor, the reaction heat of biotin andstreptavidin pairs was measured. The output signals from5.68 10-12 to 3.85 10-9W sec were measured accordingto the concentrations of streptavidin. And the sensitivity ofthe sensor was 0.8W sec/cal. After all, the biochemicalreaction of BCG and -BCG was detected by using theproposed thermoelectric biosensor. 3 µl BCG with 105 to109CFU was reacted with 3µl -BCG with 300ng/µl in thereaction chamber, only. The measured output signals werevaried from 5.01 10-12 to 5.14 10-10W sec for variousamounts of BCG.

Keywords : thermoelectric, split-flow, BCG, biotin-streptavidin, reaction heat

H-15Novel method for coating the iron oxidenanoparticles with poly(amino acid) derivativesusing poly(succinimide)

Hee-man Yang, Chan Woo Park, Yeong min Jo, and Jong-Duk Kim

Department of Chemical and Biomolecular Engineering,KAIST

Poly(amino acid)s have found increasing interest as carriersof biologically active agent because they have structurethat can be potentially degraded by proteolytic enzymes inthe body. Poly(amino acid)s would seem to have significantadvantages over other polymers owing to their protein-likestructure, water-solubility and absence of toxicity, antigenicityand immunogenicity. The biocompatible and water

BINT Convergence Technology

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dispersible poly(amino acid) derivative, with a hydrophilicbackbone and side chains facilitating linkage to the surfaceof nanoparticles was synthesized to coat hydrophobic ironoxide nanoparticles. Such nanoparticles were coated bycoordinate bonding and hydrophobic interaction withhydrophilic poly(amino acid) derivatives. This is a newmethod for transfer of hydrophobic iron oxide nanoparticlesfrom organic solvents into water. Other poly(amino acid)such as poly(aspartic acid), poly(asparagines), andPoly(dimethylaminoethyl aspartamide) was also used tocoat 10 nm sized iron oxide nanoparticles by our methods.The biocompatible poly(amino acid)-coated iron oxidenanoparticles were smaller than 30 nm in aqueoussolutions, extremely stable, and maintained their stabilityeven after several lyophilizations. The poly(amino acid)-coated nanoparticles showed no cytotoxicity, good saturationmagnetization. The poly(amino acid)-coated iron oxidenanoparticles demonstrate strong potential in bioapplicationssuch as magnetic resonance imaging.

Keywords : iron oxide, poly(amino acid), poly(succinimide),magnetic resonance imaging, dual interaction.

H-16Aqueous-processible photoresponsive polymersfor addressable micro-patterning of multipleproteins

Miju Kim1, Jongchul Choi2, Min-Gon Kim3, and JunsangDoh1,2

1School of Interdisciplinary Bioscience and Bioengineering,POSTECH

2Department of mechanical engineering, POSTECH3Biomonitoring Research Center, KRIBB

Addressable patterning of multiple proteins is challengingdue to the difficulties in alignment of each pattern withoutdamaging biological activities of proteins. Here, wedeveloped a new photoresiposible random ter-polymer,poly(2,2-DMNBMA-r-MMA-r-PEGMA) (PDMP), that canbe processed under mild aqueous processing conditions.By optimizing the composition of PDMP, we could makeit soluble in PBS (pH 7.4) after UV exposure. Also, byincorporating short sidechain of poly(ethylene glycol), thispolymer exhibited excellent resistance to proteinadsorption and cell adhesion. Using this polymer andmicroscope projection photolithography, we couldsuccessfully fabricate multiple protein micropatternedsurfaces. This technique will have general applications forthe fabrication of multiplex biosensors based on proteinsand cells.

Keywords : SGM, sasang constitution, SNP markers

H-17Novel ultra-sensitive immunoassay technologyutilizing peroxidase-mimics from nano-biotechnology

Moon Il Kim, Min-Ah Woo, and Hyun Gyu Park*

Chemical and Biomolecular Engineering, KAIST

Inorganic Fe3O4 magnetic nanoparticles (MNPs) gatheredrecent attention due to their extraordinary peroxidasemimicking activity. Herein, a simple, sensitive and specificdetection of Her-2 antigen for the diagnosis of breastcancer has been developed based on MNPs' peroxidase-like activity. A direct detection of breast cancer cellsoverexpressing Her-2 antigen was performed with Her-2antibody-conjugated MNPs. The presence of the antigenon the cell surface can be monitored by colorimetric changeof substrate with the naked eye and without complexinstruments. Number of cells overexpressing Her-2 antigencould also be quantified by MNP-antibody conjugates. Byusing tyramide signal amplification (TSA) system reactiveby MNP conjugates, cells were fluorescently imaged. Wealso developed one-pot multi-catalyst system, so called“nanofactory”, entrapping MNPs and oxidase inmesoporous silica with high loading (40 wt% MNPs and20 wt% enzyme). This nanofactory was used to detectclinically important glucose by incorporating glucoseoxidase. During the reaction, H2O2 was produced byglucose oxidase in the presence of glucose in a sample,and MNPs oxidized substrate into colorful product. Theenzymes in the nanofactory were immobilized by crosslinkedenzyme aggregate (CLEA) mode resulting in enhancedstability and reusability by simple magnet enforcement.

Keywords : magnetic nanoparticle, peroxidase, cell detection,glucose biosensor

H-18pH responsive nanocomposites monitoring drugrelease

Byungkyu Cho, Taekhoon Kim, Juhong Jin, andKwangyeol Lee

Department of Chemistry, Korea University

A number of nanocomposites have been developed fortargeted delivery, concomitant with molecular imaging.The common surmise is that the molecular imaging aidedby the imaging contrast agent also represents the drugreleasing event. However, the evidence utterly lack forsuch event due to the difficulty in material fabrications tointegrate both functions of molecular imaging and drug

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releasing. Herein we wish to report a nanocomposite systemwhich can synchronize the T1 MR signal enhancementand drug releasing event. By providing the in vivo drugreleasing profile, the more accurate description might bepossible for relationship between molecular imaging anddrug efficacy.

Keywords : pH, cancer cell, drug release, molecular imaging.nanocomposite

H-19Fe3O4-Loaded pH-responsive polymer as aMRI probe for cancer diagnosis

Guang Hui Gao1, Min Sang Kim1, Jae Won Lee2, Geun HoIm2, Jung Hee Lee2, and Doo Sung Lee1,*

1Department of Polymer Science and Engineering,Sungkyunkwan University

2Department of Radiology, Samsung Medical Center,School of Medicine, Sungkyunkwan University

Nanometer-scaled polymeric micelles are becomingextensive in biomedical applications, as carriers ofhydrophobic imaging agents for cancer diagnosis. Stimuli-responsive polymeric micelles have attracted extensiveattention as smart materials, because the polymerstructure and property can change responding to externalenvironments such as pH. As for pH-sensitive polymers,the important factor is the presence of ionizable weakacidic or basic units into hydrophobic backbones. Afterionization around physiological pH, the coiled polymericchains extend dramatically as a hydrophilic state due togenerating anionic or cationic charges. Herein, we designpH-responsive biodegradable copolymers encapsulatingmagnetite nanoparticles (Fe3O4) for cancer diagnosis,because tumor tissues have more acidic environment as aresult of lactic acid produced by hypoxia and acidicextracellular organelles. As for cancer diagnosis, the pH-responsive polymeric micelles consist of hydrophilic methylether poly(ethylene glycol) (PEG) and pH-responsivepoly(ß-amino ester) segments, which are ionized andsoluble in the pathological acidic environment, but they aredeionized and insoluble in the blood system as amphiphilicmolecules self-assmebling into micelles. After loadingmagnetite nanopartilces into the polymeric micelles, it canbe used for rapid acidic pH-triggered magnetic nanoparticlesrelease and aggregation at tumor site for MRI diagnosticapplications. We evaluate the potential of this MRIcontrast agent in MR imaging application via relaxivityproperty, in vitro pH-dependent release, and especially, invivo imaging in the tumor areas of the rat. The final in vivoimaging results will test that a new can advance in theacidic targeting and detecting cancer diagnosis.

Keywords : pH sensitivity, polymeric micelles, iron oxide,diagnosis

H-20Development of core technology for neuronalsignal detection using Si nanowire-based nano-biochip

Dong-Joo Kim, Duk-Won Seo, Seung-Yong Lee, andSang-Kwon Lee

Department of Semiconductor Science and Technology,SPRC, Chonbuk National University

We will investigate the core technology of neuron signaldetection using Si nanowire-based field-effect transistors(FETs) prepared by top-down and bottom-up techniques.Though the current research, the core technologies such asculturing technologies of neuron cells, neuron electricalcircuit mechanism, drug delivery test, interface betweenthe neuron cells, etc. will be applied in the field of silicon-based neuron-chip applications for detecting of the earlydetection of the diseases and hand-carried real timediagnostic chip etc. The current project will take threeyears. It has following research contents for each year. Inthe first year, the research is focused on the study of singleneuron cell with a silicon nanowire-based FET. It includesgrowth control of silicon nanowires and fabrication ofnano-biochip, cultivation of single neuron cell. In thesecond year, the interaction and interface phenomenabetween the neurons and nano-biochip will mainly bestudied and examined by optimizing the cultivationcondition of two neurons with synapse format. In last thirdyear, the key technologies of the cultivation of patternedneuron cells on the nano-biochip are mainly optimized bythe network formed neurons on the nano-chips. In our poster,we will present recent results in the fabrication of nano-biochip and cultivation neuron cells. (The 3rd InternationalConference on One-dimensional Nanomaterials : Atlanta(USA), 2009)

Keywords : neuron cell, Si NW, nano-biochip, synapse,neuron cell network

H-218-nitroguanine derivatives as potential candidatesfor selective bioconjugation

Hee-Seung Lee and Jaehoon Eom

Department of Chemistry, KAIST

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The development and applications of selective bioconjugationmethods have undergone significant advances in recentyears. Bioconjugation, where biomolecules are linkedtogether or to solid supports, is an important aspect of thebiological sciences and critical to many applications includingthe fabrication of functional biomolecule microarrays,biosensors, and nanotechnology. The conventional conjugationmethods rely on physicochemical adsorption using non-covalent interactions or chemoligations (NHS, imine,epoxide, etc.) with functional groups that are naturallypresent in biomolecules. However, many problemsinvolved in the conventional methods, such as lack ofrobustness, instability of the reagents, or incomplete couplingefficiency as well as low selectivity, have not been solvedyet. Herein we shows a new example of the fusion ofsynthetic chemistry and chemical biology, which has apotential to create a powerful, efficient, specific conjugationmethods. In this study, we designed and synthesized aseries of 8-nitroguanine derivatives, which were subjectedto nucleophilic substitution reaction with various nucleophiles.The results suggested that 8-nitroguanine derivatives canbe used as selective linker to connect biomolecules in amild condition.

Keywords : bioconugation, nitroguanine, chemoligation

H-22Dural stimuli-responsive dendritic-linear blockcopolymer

Eunyoung Lee and Woo-Dong Jang

Department of Chemistry, College of Science, YonseiUniversity

Recently, a great attention has been focused on the stimuli-sensitive polymers which changes their propensity respond toexternal physical and chemical stimuli, such as temperature,pH, ionic strength, and electromagnetic radiation. Particularly,thermo-responsive polymers are great potential to use assensors, support for catalysts, separation processes, andcarriers for bioactive materials delivery. Poly(2-isopropyl-2-oxazoline) (PiPrOx) is a typical thermo-responsivepolymer that undergoes a rapid and reversible hydration-dehydration change through the lower critical solutiontemperature (LCST). The fast responsiveness of PiPrOx isachieved by the precise control of the well-defined polymericstructures with appreciably narrow molecular weightdistributions through the living polymerization mechanism.Furthermore, PiPrOx is biocompatible, biodegradable, andpossess stealth characteristics in vitro and in vivo comparableto poly(ethylene glycol). Alternatively, pH-responsivepolymers are also attractive for biological applicationsbecause pH gradients play very important roles in biologicalsystem. Polymers having weak acidic or basic functionalities

exhibit a change in the ionization state upon variation ofpH, which leads to solubility change of the polymers. Whilevarious dual stimuli-responsive polymers have been reportedby the conjugation of pH- and temperature-responsivepolymer blocks, most studies were focused on linear-linearblock copolymer structures. We are going to report a dualstimuli responsive dendritic-linear block copolymer (Den-PiPrOX) having pH-responsive dendritic wedge andthermo-responsive PiPrOx chain. The introduction of dendriticbuilding block is particularly attractive because the multivalentmodification of dendritic wedge can provide various potentialapplications.

Keywords : dendrimer, isopropyloxazoline, LCST, stimuli-responsiveness

H-23Liquid crystal-based monitoring of biomolecularinteractions at aqueous-LC interfaces

QiongZheng Hu and Chang-Hyun Jang*

College of BioNano Technology, Kyungwon University

We investigate the orientational behavior of thermotropicliquid crystals (LC) confined at aqueous-LC interfaces.Copper or gold grids supported on glass surfaces treatedwith octadecyltrichlorosilane (OTS) were impregnatedwith LC, 4-cyano-4'-pentylbiphenyl (5CB). The films of5CB confined within the grids were then immersed intoaqueous solutions of sodium dodecyl sulfate (SDS). Dynamicchanges in the orientation of LC relative to the SDSconcentrations were observed using a cross polarizedmicroscope. We also demonstrate the orientational responseof LC to oligopeptide-modified amphiphiles placed at theaqueous-LC interface. Because the action of enzymes thatcleaves the oligopeptides triggers odering transitions ofLC, this system is expected to allow real-time monitoringof interfacial enzymatic interactions.

Keywords : liquid crystals, orientational behavior, aqueous-LC interfaces, real-time monitoring, enzymaticinteractions

H-24Development of nanoplasmonics based biosensorfor the real time, label-free detection of synapticnetwork activity

Young-Jae Oh, Sang-Gil Park, and Ki-Hun Jeong

Department of Bio and Brain Engineering, KAIST

Korea Research Institute of Bioscience and Biotechnology 100

The goal of this research project is label free, real timeoptical measurements of synaptic network activities for theneuroscience and biomedical researches. Especially, opticalsensing on metal nanostructures based nanoplasmonics cangreatly increase the detection sensitivity and resolution. Thisnovel sensing technique is expected to perform biochemicalsynaptic activity analysis and overcome disadvantages oflabeling processes. The goals of the first research year areplasmonic design and fabrication of the metal nanopatternsfor nanoplasmonic substrates. Specifically, nanoplasmonicsubstrates can be developed by the formation of metalnanostructures through colloidal lithography, othernanofabrication techniques, followed by conventionalphotolithography process based hierarchical patterning ofnanostructures. Neurochemicals can be detected by usingthese nanoplasmonic substrates. We have demonstratedneurotransmitter detection researches using surface enhanceRaman scattering(SERS) thorough three ways: (1) usingcrystallographic nanoparticles array; (2) using nanochannels ;(3) using biocompatible hydrogel plasmonic biopatchs.Furthermore, we have developed a highly sensitive plasmonicsubstrate fabrication process through annealing process foreasy and mass production. In 2009, we have presentedfour posters and had one oral presentation at conferences.And two patents related to research are applied. ( : Daejeon, 2009)

Keywords :

H-25Application of optofluidc sensor for highlysensitive bio/environmental detection

Chaesung Lim2, Hyangah Chon1, Byunghee Han1, JooheeKo2, and Jaebum Choo1,2

1Department of Bionano Engineering, Hanyang University2Department of Applied Chemistry, Hanyanhg University

We report a highly sensitive surface-enghanced Ramanscattering (SERS) platform for a highly sensitivebio/environmental detection. In this presentation, tworesearch results will be presented; one is the selective traceanalysis of mercury (II) ions in drinkable water usingaptamer-conjugated metal nanoparticles, and the other isthe SERS-based immunoassay using hollow gold nanospheresand magnetic beads. First, aptamer-conjugated metalnanoparticles specifically binding to HgII ions has beenfabricated. Aggregation property of metal nanoparticlesupon the addition of HgII has been used for its trace analysis.Under optimized assay conditions, the concentration limitof detection was estimated to be 5 nM, and this satisfies alimit of detection below the EPA defined limit of 10 nMin drinkable water. Second, SERS-based sandwich

immunoassay has been performed in a microfluidicchannel. For validation, a well-known lung cancer marker,carcinoembryonic antigen (CEA), was used as a target.Based on experimental results, the limit of detection (LOD)was determined to be 1-10 pg/mL, this value being about100-1000 times more sensitive than the LOD of conventionalenzyme-linked immunosorbent assay (ELISA). Furthermore,the assay takes less than one hour to complete, includingwashing and optical detection steps.

Keywords : surface-enhanced raman scattering, optofluidics,aptamer, biomarkers

3rd KRIBB Poster Festival | Abstracts |