proforma – i proforma for submission of project...

PROFORMA – I

PROFORMA FOR SUBMISSION OF PROJECT PROPOSALS ON RESEARCH

AND

DEVELOPMENT, PROGRAMME SUPPORT

(To be filled by the applicant)

PART I: GENERAL INFORMATION

1. Name of the Institute/University/Organization submitting the Project Proposal:

Tumkur University, Tumkur

2. State: Karnataka

3. Status of the Institute: State govt funded/UGC recognized

(Please see Annexure-I)

4. Name and designation of the Executive Authority of the Institute/University

forwarding the

application: Registrar, Tumkur University Tumkur

5. Project Title: Molecular cloning and expression of anticoagulant and antiplatelet

protease (s) from cucumber sap extract: Understanding the probable molecular

mechanism and therapeutic application in thrombotic disorders.

6. Category of the Project (Please tick): R&D

7. Specific Area (Please see Annexure - II): Life science

8. Duration: 03 Years

9. Total Cost (Rs.): 97.128 lakhs

10. Is the project Single Institutional or Multiple-Institutional (S/M)? : Single

institutional

11. If the project is multi-institutional, please furnish the following:

Name of Project Coordinator: Affiliation:

Address:

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12. Scope of application indicating anticipated product and processes

The most common cause of death world wide is thrombotic disorders such as, myocardial

infarctions, acute arterial thrombosis, atrial fibrillation (clot in heart), unstable angina,

deep vein thrombosis, pulmonary embolism. Several anti-coagulants, anti-platelets agents

are being used to treat thrombotic disorders besides their health threatening side effects.

Consequently, identifying the novel protein(s) exhibiting strong anti-coagulant anti-

platelet properties along with fibrin clot hydrolyzing activity (fibrinolytic activity) from

the plants or fruits helps in the better management of thrombotic disorders. Therefore,

The development of new antithrombotic agents has been inspired by clinical needs and

by progress in biotechnology that have made it potential to fabricate drugs that target

specific steps in thrombogenesis. It is well known that, Heparin has pharmacokinetic and

biophysical limitations that are overcome by new anticoagulants. Of these, low-

molecularweight heparin and direct inhibitors of thrombin have been evaluated clinically.

Coumarins require careful laboratory monitoring because of concerns about safety.

Orally active direct inhibitors of thrombin and factor Xa may replace coumarins. Aspirin

is of limited efficacy because it inhibits only one pathway of platelet activation.

Therefore, identifying the effective and safer anticoagulant and antiplatelet agent helps in

the better management of thrombotic disorders. Moreover, target specific (Factor Xa and

thrombin inhibitors) proteins/enzymes with strong anticoagulant and antiplatelet

properties along with fibrin clot hydrolyzing activities would be the better therapeutic

molecules to treat thrombotic disorders. Thus, purification pharmacological

characterization of protease that exhibit anticoagulant, antiplatelet and fibrinolytic

protease from cucumber sap extract becomes gateway to the researchers/scientist to

further explore the importance of unexplored/ignored medicinal plants. Our preliminary

studies indicated that proteolytic activity of cucumber sap extract found to cause anti-

coagulation, inhibition of agonist ADP and collagen induced platelet aggregation of

human platelet rich plasma and washed human platelets. Hence, this study ultimately

contributes for both academicians and researchers who are working on thrombotic

disorders. Anti-coagulants and anti-platelets agents and fibrin clot hydrolyzing enzymes

have enormous importance and they contribute for better management of thrombotic

disorders. Above all, they may serve as prototypes for designing anti-thrombotic drugs

and they can be used as therapeutic agents to treat hypercoagubility of blood leads to fatal

heart attack and strokes that leads to high rate of death.

13. Project Summary (Not to exceed one page. Please use separate sheet).

The main aim of the project is purification, cloning, expression and pharmacological

characterization of proteases (s) that exhibit anticoagulant, antiplatlelet and fibrinolytic

activities. In addition, establish the possible molecular mechanism of anticoagulant and

antiplatelet activities of purified protease (s). According to our preliminary studies,

cucumber sap extract showed two isoforms of proteases in casein zymogram, it

selectively hydrolyzed human fibrinogen and fibrin clot. Further more, cucumber sap

extract showed strong anticoagulant activity in plasma recalcification time and inhibited

agonist collagen and epinephrine induced platelet aggregation of human platelet rich

plasma in a dose dependent manner. Anticoagulants and antiplatelet agents have an

immense therapeutic value because the most common cause of death world wide is

thrombotic disorders that include myocardial infarctions, acute arterial thrombosis, atrial

fibrillation (clot in heart), unstable angina, deep vein thrombosis and pulmonary

embolism. Anti-coagulant and anti-platelet therapy is the effective therapy for the

prevention and treatment of thrombotic disorders. The currently established

anticoagulants and anti-platelet agents are having numerous limitations with life

threatening side effects. Thus currently available anti-coagulant and anti-platelet therapy

fails to recommend protection against thrombotic disorders. Hence, identifying the novel

anti-coagulant and anti-platelet agents from the natural sources with least side effects is

the challenging task to the researchers. In addition, proteases exhibit anticoagulant,

antiplatelet and fibrin clot hydrolyzing activities helps in the better management of

thrombotic disorders.

PART II: PARTICULARS OF INVESTIGATORS

(One or more co-investigators are preferred in every project. Inclusion of co-

investigator(s) is

mandatory for investigators retiring before completion of the project)

Principal Investigator:

14. Name : Dr. Devaraja. S

Date of Birth : 24-04-1976 Sex (M/F): M

Designation : Assistant Professor

Department : Biochemistry

Institute/University : Tumkur University

Address: Dr. Devaraja. S, Assistant Professor, Department of Studies and

Research in Biochemistry, Tumkur University, University Constituent College

Campus, B.H, Road, Tumkur, PIN: 572103, Telephone: 919945765028, 0816-

2254546 Fax: 0816-2270719 E-mail: [email protected]

Number of research projects being handled at present: 02

Co-Investigator

15. Name : Dr.Kemparaju.K

Date of Birth : 20-07- 1964 Sex (M/F): M

Designation : Associate Professor

Department : DOS in Biochemistry,

Institute/University : University of Mysore

Address : DOS in Biochemistry, University of Mysore, Manasagangothry,

Mysore,

Karnataka, India. PIN: 570 006

Telephone: 91-9945996543

E-mail: [email protected]

Number of Research projects being handled at present: 02

Co-Investigator

16. Name : Dr. K.S. Girish

Date of Birth : 22-06-1976 Sex (M/F): M


Department : DOS in Biochemistry

Institute/University : University of Mysore

Address: DOS in Biochemistry, University of Mysore, Manasagangothry,

Mysore,

Karnataka,

PIN :570006.

Telephone : 9972268633

E-mail : [email protected]


Co-Investigator

16. Name : Dr. Sudhrshan. B,G

Date of Birth : 18-04-1975 Sex(M/F) : M

Designation : Senior Medical Officer and Assistant Professor

Department : Biomedical and instrumentation

Institute/University : R.V. Engineering College

Address : R.V. Engineering College, Mysore road, Banglore, Karnataka,

PIN : 5600059

Telephone: 08067178100



Co-Investigator

16. Name : T.G.Thippeswamy

Date of Birth : 28.06.1971 Sex (M/F): M



Institute/University : Tumkur University,

Address : DOS in Biochemistry, Tumkur University, Constituent college

campus,

B.H.Road, Tumkur, Karnataka, India.

PIN: 572103

Telephone: 0816-2254546 Fax: 0816-2270719



Co-Investigator

16. Name : Bhagyalakshmi.M

Date of Birth :16-08-1981 Sex (M/F): F



Institute/University : Tumkur University,

Address : DOS in Biochemistry, Tumkur University, Constituent college

campus,

B.H.Road, Tumkur, Karnataka, India.

PIN: 572103

Telephone: 0816-2254546 Fax: 0816-2270719



PART III : TECHNICAL DETAILS OF PROJECT

(Under the following heads on separate sheets)

16. Introduction (not to exceed 2 pages or 1000 words)

In healthy people, homeostatic balance exists between procoagulant (clotting) forces and

anticoagulant and fibrinolytic forces. Numerous genetic, acquired and environmental

factors can incline the balance in favor of coagulation, leading to the pathologic

formation of thrombi in veins (deep venous thrombosis), arteries (myocardial infarction,

ischemic stroke), or cardiac chambers. Thrombi can obstruct blood flow at the site of

formation or detach and embolize to block a distant blood vessel (eg, pulmonary

embolism, embolic stroke). As such,the diagnosis and management of thrombotic

disorders are increasingly falling down within the realm of the practice of hematology.

There are about 10 million cases annually reported world wide exceeding the number of

deaths from cancer and other diseases. It is estimated that annual mortality rate of 60,000

in United States and more than 50,000 in India. Thus thrombotic disorders represents a

major health problem world wide including India. Anti-platelet and anti-coagulant agents

have been increasingly becomes an important tool for preventing cardiovascular events.

While aspirin is the most widely used and best studied anti-platelet agent, the use of

clopidogrel (Plavix) and combination dipyridamole-aspirin (Aggrenox) have increased

substantially in recent years, and a 2007 study introduced a new agent, prasugrel (Effient).

Despite the lack of efficacy and safety heavy marketing to physicians and direct-to-

consumer advertising have made Plavix the second best-selling drug in the world. The

main side effect of Plavix is at low dose it increases the risk of unwanted clot formation

and at high dose the coagulation mechanisms in the blood are shut down too severely and

there is increased risk of internal and external bleeding episodes. Thus currently available

anti-coagulant and anti-platelet therapy fails to recommend protection against thrombotic

disorders. Hence, the researchers are extremely enthused about the usage of proteins

those exhibit anti-coagulant, anti-platelet and fibrin clot hydrolyzing proteins/proteases

from naturally occurring medicinal plants and they may complement the existing anti-

platelet and anti-coagulant therapy. Hence, these proteases may be act as powerful

weapons in the better management of thrombotic disorders.

There are several natural unexplored medicinal plants and fruits whose pharmacological

properties are not well established. Cucumber is one such fruit from the creeper Cucumis

sativa L and belongs to Cucurbitaceae family, which is native to Indian subcontinent.

Cucumber has been found to possess water content of about 96% while it is Scarce in

calorific value. However, it has been reported to contain wide varieties of nutritional

components in trace amount such as lipids, proteins, carbohydrates as macronutrients and

potassium, calcium, magnesium, iron, vitamins (Vit-C, Vit-B complex) as micronutrients.

It has got broad spectrum of health beneficial applications & found to protect from heat,

inflammation, of skin, acidity, gastritis, ulcer, arthritis, gout, high and low BP and

elimination of toxic substances from the blood. It has been associated with healing

properties in relation to diseases of the kidney, urinary bladder, liver & pancreas. Sap is a

juicy circulating fluid of cucumber & its biological importance has not been studied so

far. Our preliminary studies established the proteolytic activity of cucumber sap found to

exhibit fibrin (ogen) lytic acitivty, anti-coagulant and anti-platelet property in platelet rich

plasma. Therefore, this project has been under taken to dissect the therapeutic role and

molecular mechanism of protease (s) those exhibit antiplatelet, anticoagulant and fibrin

clot hydrolyzing properties followed by their cloning and expression.

16.1 Origin of the proposal

Blood coagulation cascade is the co-ordinated activation of plasma serine proteases and

their co-factors. The end product is the protease thrombin, which cleaves fibrinogen to

generate a fibrin clot. In addition, platelets which circulating in the blood plays an

important role in the hemostasis or formation of fibrin clot (Furie et al., 2005, 2008). If

the number of platelets are too low in circulating blood that leads to excessive bleeding

and too high leads to the formation of blood clots or thrombosis respectively and it may

obstruct blood vessels and results in stroke, myocardial infarction, pulmonary embolism

or the blockage of blood vessels (Savage et al., 2001: William et al., 2010). In addition,

the abnormality in haemostatic system leads to excessive bleeding or thrombosis. The

human body has a complex mechanism that causes blood to clot if a wound occurs.

Under normal circumstances this is a desirable response that enables the body to heal

itself, but under certain clinical conditions, called “blood clot disorders or thrombotic

disorders", this same mechanism can cause an unwanted clot or "thrombus" that can be

life threatening. Thrombosis the formation of a blood clot within a blood vessel or cavity

of the heart is a maladaptive process of vascular occlusion and remains a primary cause

of cardiovascular morbidity and mortality. The primary etiology of myocardial

infarctions, and approximately 80% of strokes, is acute arterial thrombosis. In addition,

atrial fibrillation (clot in heart), myocardial infarction (heart attack), unstable angina,

deep vein thrombosis, pulmonary embolism and replacements of heart valves also leads

to the formation of clot with in the heart and hence represents the most common cause of

death world wide (Webster et L., 2005:Canhão et al., 2005:Hatzinikolaou-Kotsakou

et al., 2005: Lars Maegdefessel et al., 2010).

16.2 (a) Rationale of the study supported by cited literature (b) Hypothesis (c) Key

questions.

Thrombotic disorders increase the risk of formation of fibrin clot with in a blood vessel

or cavity of the heart and it is a leading cause of death and disability world wide. About

20 to 30% of patients hospitalized for an acute medical illness is due to arterial

thrombosis, atrial fibrillation, myocardial infarction, unstable angina, deep vein

thrombosis, pulmonary embolism. Anti-coagulants those inhibits the clot formation by

blocking the action of clotting factors/coagulation factors and anti-platelets agents those

blocks the formation of blood clot by preventing the clumping of platelets are extensively

used in treatment of thrombotic disorders (Leng et al.,1996: Anand et al.,

2010). Thus Anti-coagulant and anti-platelet therapy is the effective therapy for the

prevention and treatment of thrombotic disorders. While, the currently established

anticoagulants such as, warfarin, dabigatran, unfractionated heparin (UFH), enoxaparin,

fondaparinux, bivalirudin (thrombin inhibitors), low molecular weight heparin and anti-

platelet agents such as, aspirin, thienopyridines, dipyridamole, dlopidogrel, dpoprostenol,

abciximab, eptifibatide and tirofiban (glycoprotein IIb/IIIa inhibitors) are having

numerous limitations with several side effects, including lack of reversibility, a sheer

dose response, food and multiple drug-drug interactions, narrow therapeutic index and

cause bleeding (Rosendaal et al.,1993: CAPRIE Steering Committee 1996: van et al.,

2002: Hurlen et al., 2002: Antman et al., 2004). Therefore, identifying the novel anti-

coagulant and anti-platelet agents from the natural sources with least side effects is the

challenging task to the researchers. In addition, the other hypothesis is that targeting

proteases upstream from the common pathway provides a reduction in thrombin

generation slow down the formation of fibrin clot or thrombosis. Investigating the target

specific (factor Xa and thrombin) anti-coagulant, anti-platelet and fibrin clot hydrolyzing

proteins from the naturally occurring plants and fruits with least side effects help in the

better management of thrombotic disorders.

16.5 Current status of research and development in the subject (both international

and

national status)

Plants sustain our daily life, as they are the important source of food, agricultural

crops, and medicines. The value of medicinal plants to human livelihood is essentially

infinite. Medicinal plants are the most exclusive sources of life saving drugs for the

majority of the world’s population. The world health organization has estimated that

more than 80% of the world’s population in developing countries depends primarily

on herbal medicines for basic health care needs. Plant products also play an important

role in the health care systems of the remaining 20% of the population, mainly

residing in developed countries. A large proportion of drugs used in modern medicine

are either directly isolated from plants or synthetically modified from a lead

compound of natural origin. Medicinal plants and their extracts are often used as an

alternative to specifically targeted drugs in the treatment and prevention of many

diseases.

Finding healing powers in plants is an ancient idea. People on all continents have long

applied poultices and imbibed infusions of hundreds of indigenous plants dating back

to prehistory. Plants have the richest source of several products which are immense

use to man. Noteworthy among them and next to those of food value are substances of

medicinal importance. An estimate suggests that about 25 million plant species known

for their medicinal uses. Phytochemical tests have been performed in about 5000 and

nearly 1100 species are extensively exploited in 80% of ayurvedic, 46% of unani and

33% of allopathic medicines.

The world market of pharmaceutical products operates a business turnover of

US$ 320,000,000/year from which US$ 20,000,000 originates from vegetal active

substances. The current world market of phytomedicines operates a business turnover

of US$ 40 billion/year from which US$ 2 billion originates from India.

In more recent history, the use of plants as medicines has involved the isolation of

active compounds, beginning with the isolation of morphine from opium in the early

19th century (Kinghorn, 2001; Samuelsson, 2004). Drug discovery from medicinal

plants led to the isolation of early drugs such as cocaine, codeine, digitoxin, and

quinine, in addition to morphine, of which some are still in use (Newman et al., 2000;

Butler, 2004; Samuelsson, 2004). Isolation and characterization of pharmacologically

active compounds from medicinal plants continue today. More recently, drug

discovery techniques have been applied to the standardization of herbal medicines, to

elucidate analytical marker compounds. The search for new molecules, nowadays, has

taken a slightly different route where the science of ethnobotany and

ethnopharmacology are being used as guide to lead the chemist towards different

sources and classes of compounds. Current research in molecular medicine includes

the drug discovery from medicinal plants involves a multifaceted approach combining

botanical, phytochemical, biological, and molecular techniques. Medicinal plant drug

discovery continues to provide new and important leads against various

pharmacological targets including cancer, inflammation, wound healing, HIV/AIDS,

Alzheimer’s, malaria, pain, cardiovascular disorders including thrombolytic disorders.

Several natural product drug(s) of plant origin have either recently been introduced to

the world market.

16.6 The relevance and expected outcome of the proposed study

1. This project may provide preliminary data about the efficacy and safety use of

proteolytic enzymes that exhibit anticoagulant, antiplatelet properties along with

fibrinolytic activities from cucumber sap extract.

2. Cloning and expression of this protease will be evaluated for antithrombotic

activities along with its mechanism of action

3. Expressed protease will be tested for its non- toxicity

4. The success in cloning and expression of proteases with anticoagulant, antiplatelet

and fibrinolytic activities could definitely helps in the development of safer and

effective antithrombotic drugs with least side effects.

5. The results obtained in this project will be published in the peer reviewed

international journals.

16.7 Preliminary work done so far

According to our preliminary studies the cucumber sap extract found to show varied

proteins bands from the molecular weight range from 200 to 16 kDa (Fig 1). In addition,

the cucumber sap extract found to cause anti-coagulation as it increased normal clotting

time 250 sec to 430 sec (Fig 2). Cucumber sap extracts selectively hydrolyzed Aα and Bβ

chains of fibrinogen with out affecting γ chain over the incubation of 8h, suggesting its

specificity on Aα and Bβ chains of fibrinogen (Fig 3). Interestingly, cucumber extracts

specifically degraded α-polymer and α-chain with out affecting γ- γ dimmer and β-chain

of fibrin clot (Fig 4). Further more, the extract inhibited the collagen and epinephrine

induced aggregation of platelet rich human plasma dose dependently (Fig.5 and 6).

RESULTS

Fig.1.SDS-PAGE pattern of cucumber sap extract.

Cucumber sap extract (100 µg) under non-reduced condition (1), Cucumber sap extract

(100 µg ) under reduced condition (2).Lane M: Molecular mass markers (M) in kDa from

top to bottom: myosin-H-chain (200), phosphorylase b (97.8), BSA (68), ovalbumin (43)

carbonic anhydrase (29), β-lactalbumin (18.4) and lysozyme (14.3).Fig.1b.Activity

staining on casein zymogram

Fig. 2. Effect of cucumber sap extarct on re-calcification time of human plasma.

Cucumber sap extract ranging from 10 to 80 µg was pre-incubated with 300 µl of human

plasma in the presence of 30µl tris-Hcl buffer(10 mM; pH 7.4) for 1 min at 37˚C, 30 µl of

CaCl2 (0.25 M) was added to the pre-incubated mixture. The time for clot formation was

recorded

Fig. 3. Fibrinogenolytic activity of cucumber sap extract: Fibrinogen incubated for 8

h at 37o and run in 10% SDS-PAGE under reduced condition. Fibrinogen (50 µg) alone

(1), fibrinogen (50 µg), + 4µg cucumber sap extract (2), fibrinogen (50 µg), + 8µg

cucumber sap extract (3), fibrinogen (50 µg) + 12µg cucumber sap extract (4), fibrinogen

(50 µg) + 16µg cucumber sap extract(5),fibrinogen (50 µg) + 24µg cucumber sap extract

(6), fibrinogen (50 µg) + 24µg cucumber sap extract (7). M represents the molecular

weight marker in kDa from top to bottom: myosin-H-chain (200), phosphorylase b (97.8),

BSA (68), ovalbumin (43) carbonic anhydrase (29), β-lactalbumin (18.4) and lysozyme

(14.3).

Fig.4. Fibrinolytic activity cucumber sap extract: Washed plasma clot was incubated

with cucumber sap extract for 8 h and then separated on 7.5 % SDS-PAGE under reduced

condition. Plasma clot alone (1), plasma clot + 8 µg cucumber extract (2), 12 µg (3), 16

µg (4) 20 µg (5), 24µg (60), 30µg (7) M represents the molecular weight markers in kDa

from top to bottom: myosin H chain (200), phosphorylase b (97.8), BSA (68), ovalbumin

(45), carbonic anhydrase (31), and trypsin inhibitor (21.5).

Fig. 5. Effect of cucumber sap extract on collagen induced aggregation of platelet rich human

plasma.

Fig. 5. Effect of cucumber sap extract on epinephrine induced aggregation of platelet rich human plasma.

17. Specific objectives (should be written in bulleted form, a short paragraph

indicating the

methods to be followed for achieving the objective and verifiable indicators of

progress shouldfollow each specific objective)

• Isolation, biochemical and pharmacological characterization of anti-

coagulant, antiplatelet and fibrin clot hydrolyzing protease (s) from

cucumber sap extract

• Establish the site of action of purified protease(s) on blood coagulation

cascade using specific clotting factor /deficient clinical

plasma/reconstituted plasma samples

• Cloning and expression of protease (s) that possess anticoagulant,

antiplatelet and fibrinolytic activities

• Investigate the kinetic effects of protease(s) in the various agonist induced

platelet function of human platelet rich plasma and washed human

platelets

• Establish the probable molecular mechanism involved in the anti-coagulant

and anti-platelet properties of protease (s).

18. Work Plan: should not exceed 3-4 pages (the section can be divided according to

the specific aims and under each specific aim, the following should be stated clearly

as sub headings)

18.1 Work plan (methodology/experimental design to accomplish the stated aim

Preparation of sap extract from cucumber.

Cucumbers will be purchased from the Market; both the ends will cut to obtain the sap

material. The ends will be squeezed and the juicy material will be collected. The sap

material will be dissolved further in water and centrifuge at 2000 rpm and the supernatant

will be stored at -20OC until use.

Protein estimation:

The protein estimation will be done according to the method of Lowry et al., (1951) using

the bovine serum albumin (BSA) as standard.

Coagulant activity:

The plasma re-calcification time will be determined according to the method of Quick et

al. (1935). Briefly, the various concentration of cucumber sap extract will be pre-

incubated with 0.2 ml of citrated human plasma in the presence of 10 mM Tris-HCl (20

ml) buffer pH 7.4 for 1 min at 37o C. Twenty microlitres of 0.25 M CaCl2 was added to

the pre-incubated mixture and clotting time will be recorded.

Preparation of platelet rich plasma (PRP) and platelet poor plasma (PPP).

The method of Ardlie and Han (1974) will be employed. Nine volumes of human blood

from healthy donors (who are non-smokers and non-medicated at least for the previous

15 days) in to one volume of acid citrate dextrose (93 mM sodium citrate, 7 mM citric

acid and 140 mM glucose pH 6.5) followed by centrifugation at 90 g for 10 min at room

temperature. The supernatant is called platelet rich plasma (PRP). The remaining blood

will be centrifuged at 500 g for 15 min and the supernatant obtained is the platelet poor

plasma (PPP). The platelet concentration of PRP will be adjusted to 3.1 X 108

platelets/ml with PPP. The PRP maintained at 37o C will be used with in 2 hr. All the

above preparations will be carried out using plastic wares or siliconized glassware’s.

Preparation of washed platelets

The washed platelets will be prepared according the method described by Cazenave et al.

Briefly, platelet rich plasma (PRP) will be obtained by mixing fresh blood sample with

acid citrate dextrose solution –ACD (85 mM sodium citrate; 71 mM citric acid; 111 mM

dextrose) in a 6:1volume ratio followed by centrifugation at 120 × g for 15 min. The

supernatant portion obtained is the PRP and is transferre to plastic tube. Then the PRP

will be centrifuged at 1700 × g for 15 min at 37°C, the supernatant obtaine is to be

discarded and the platelet pellet will suspended in Tyrode's albumin buffer (145 mM

NaCl, 5 mM KCl, 10 mM HEPES, 0.5 mM Na2HPO4, 1 mM MgCl2, 6 mM glucose,

0.3 % human serum albumin) pH 6.5 containing 10 U/mL heparin and 0.5 µM

prostacyclin. After 10 min of incubation at 37°C, the suspension is centrifuge at 1700 ×

g for 15 min, the platelet pellet obtained is again suspended in Tyrode’s albumin buffer

containing 0.5 µM prostacyclin and is again centrifuge at 1700 × g for 15 min. Finally the

platelet pellets obtaine are suspended in Tyrode’s albumin buffer containing 0.02 U/mL

apyrase pH 7.4 and platelets will be counted using a Neubauer chamber and adjuste to

the concentration required using Tyrode's albumin buffer pH 7.4.

Platelet aggregation

The turbidometric method of Born (1962) will be followed for using a Chronolog dual

channel aggregometer connected to an omniscrible dual pen recorder to record the light

transmission as a function of time. Aliquotes of PRP (0.45 ml)/human platelet rich

plasma will be pre-incubated with different concentrations of various concentration of

sap extract for 3 min in a cylindrical glass cuvette under constant stirring. The

aggregation will be initiated by the addition of agonist such as collagen, ADP, thrombin

and epinephrine followed for 8 min. As platelets aggregate in response to an added

agonist, light transmission increases progressively producing an aggregation trace on the

recorder paper. The aggregation trace is the plot of light transmission between platelet

rich plasma (PRP) and platelet poor plasma (PPP) base line, which represent 0 % and

100 % aggregation respectively.

cDNA library construction

The cucumber sap extract cDNA library will be constructed accoding to the method of

Chaim et al. Briefly, cucumber sap extract mRNAs will be purified using the FastTrack

2.0 Mrna isolation kit (Invitrogen). The cDNAs will be then synthesized using the Super

Script Plasmid System with Gateway Technology for cDNA Synthesis and Cloning

(Invitrogen), cloned into NotI and SalI pre-cut pSPORT1 vector and transformed into

Escherichia coli DH5α cells. Transformants will be selected on LB (Luria– Bertani) agar

plates containing 100 µg/ml ampicillin.

cDNA library screening

Randomly chosen colonies (approx. 100 clones) will be inoculated into LB broth

containing 100 µg/ml ampicillin and grow overnight at 37◦C (with aeration). From this,

recombinant plasmids were purified using QIAprep Spin Miniprep Kit (Qiagen). The

cloned cDNAs will be sequenced on both strands using ABI PRISM® BigDye

Terminator Cycle Sequencing Ready Reaction Kit (Applied Biosystems). Reactions will

be analysed using an ABI 377 automatic sequencer (Applied Biosystems). The promoter

regions will be used for priming the sequencing reactions. The cDNA sequences will be

analysed, and the putative protein products from these sequences.

Expression and purification of recombinant protease

Selected transfermants will be pregrown at 30 ℃ in 0.5 L baffled shake flasks

containing 0.15 L rich medium (20 g/L tryptone, 13.4 g/L yeast nitrogen base, 4 × 10-4

g/L biotin, 1 % glycerol, 0.1 mol/L potassium phosphate buffer, pH 6.0). The cells will

be grown for 24 h with an additional 1 % glycerol after 12 h. The culture will be

centrifuged and the pellet containing the cells will be resuspended in rich medium with

methanol (20 g/L tryptone, 13.4 g/L yeast nitrogen base, 4 × 10-4 g/L biotin, 1 %

methanol, 0.1 mol/L potassium phosphate buffer, pH 6.0). After induction for 72 h, the

culture will be collected and adjusted pH to 5.0 and then centrifuged to remove cells and

precipitates. Exchanging buffer to 20 mmol/L methylpiperazin-HCl, pH 5.0, by using

Sephedex G 25, the supernatant will be applied to Q-Sepharose Fast Flow (Pharmacia

Biotech AB, Sweden) and the target protein will be eluted at the gradient of 0-0.5 mol/L

NaCl in 20 mmol/L methylpiperazin-HCl, pH 5.0 in 20 column volume. The activity

peak will be pooled and then applied to Phenyl HP (Amersham Pharmacia Biotech AB,

Sweden) in a start buffer of 1 mol/L ammonium sulfate in 20 mmol/L Tris-HCl, pH 7.2.

The active fractions collected were concentrated by ultralfiltration (Amicon, USA) up to

5 times and 0.5 ml of them was applied onto Superdex 200 column. The active material

obtained will be loaded to ConA Hitrap (Pharmacia Biotech AB, Sweden), eluted by 0.5

mol/L NaCl and 0.5 mol/L methyl- a -D-glycopyanoside in 20 mmol/L Tris-HCl, pH 7.4

Proteolytic activity:

Proteolytic activity will be determined according to the method of Satake et al., (1963)

using 2 % casein in 0.2 M Tris-HCl buffer, pH 8.5. Various concentration of purified

protein/s from ion exchange chromatography will be incubated separately with 0.4 mL of

substrate in a total volume of 1 mL for 2 hr and 30 min at 37OC. Undigested casein will

be precipitated by adding 1.5 ml of 0.44 M trichloroacetic acid. The digested casein in

supernatant (1 ml)will be determined using Folin-Ciocalteu’s reagent. The method is

initially standardized using increasing amount of venom samples in the assay mixture.

One unit of activity was defined as the amount of enzyme required to cause an increase in

O.D. by 0.01 at 660 nm/min.

Substrate gel assay (zymogram):

SDS-PAGE (10%) will be prepared according to the method of Laemmli (1970). and

polymerized at a final concentration of 0.2 % with casein/gelatin.purified protein (10 µg

each) will be prepared under non-reduced condition. Electrophoresis will be carried out

using Tris (25 mM), glycine (192 mM) and SDS (0.1%) for 3h at 90V at room

temperature. After electrophoresis, gels will be washed with 10mM sodium phosphate

buffer containing 2.5%of Triton X-100 with shaking for 1h to remove SDS and incubated

overnight at 37oC in Tris-HCl buffer (50 mM) pH 7.6 containing 10 mM CaCl2 and 150

mM NaCl. Gels are then stained with Coomassie brilliant blue R-250 as described by

Nagaraju et al., (2006).

Coagulant activity:

Recalcification time will be determined according to the method of Quick (1935). by

incubating different concentration of purified protease in a total volume of 0.2 ml of

citrated human plasma with of 10 Mm Tris Hcl buffer pH 7.4 for 1 min, and then clotting

time will be determined by adding 20µl of 0.25 M CaCl2.

Fibrinogenolytic activity:

Fibrinogenolytic activity will be determined according to the method of Ouyang and

Teng (1976). Different concentrations of purified protein/s (0-20 µg) from ion exchange

column chromatography incubated independently with 50 µg human fibrinogen in 40 µL

of 10 mM Tris-HCl buffer pH 7.4. After 8 h, reaction was terminated by adding 20 µL

denaturing buffer containing 1 M urea, 4% SDS and 4% β-mercaptoethanol. It was

analyzed by 10% SDS-PAGE.

Fibrinolytic activity

Method of Rajesh et al (2006) will be employed for fibrinolytic activity. EDTA (2

mg/ml) treated blood is centrifuge for 15 min at 500g to separate plasma. Plasma (100 µl)

will be mixed with equal volume of 10 mM CaCl2 for 15 min at 37O C to get soft fibrin

clot. The fibrin clot is washe thoroughly 5-6 times with phosphate buffered saline (PBS)

and suspended in a final volume of 40 µl 10 mM Tris-HCl buffer pH 7.4 containing

different concentrations of purified protein of cucumber. The reaction will be stopped by

adding 20 µl of sample buffer containing 4% SDS, 1 M urea and 4% β-mercaptoethanol.

It is then boile for 3 min and centrifuge to settle the debris of plasma clot. An aliquot of

20 µl supernatant will be analyzed in 7.5% SDS-PAGE for fibrin degradation studied.

Platelet aggregation

The turbidometric method of Born (1962) will be followed for using a Chronolog dual

channel aggregometer connected to an omniscrible dual pen recorder to record the light

transmission as a function of time. Aliquotes of PRP (0.45 ml)/human platelet rich

plasma will be pre-incubated with different concentrations of various concentration of

sap extract for 3 min in a cylindrical glass cuvette under constant stirring. The

aggregation will be initiated by the addition of agonist such as collagen, ADP, thrombin

and epinephrine followed for 8 min. As platelets aggregate in response to an added

agonist, light transmission increases progressively producing an aggregation trace on the

recorder paper. The aggregation trace is the plot of light transmission between platelet

rich plasma (PRP) and platelet poor plasma (PPP) base line, which represent 0 % and

100 % aggregation respectively.

Platelet adhesion assay

Platelet adhesion assay will be performed according the method described by Bellavite et

al. Briefly, 20 µg each of collagen type I, fibrinogen and fibronectin in 200 µl PBS will

be added independently to 96-well polystyrene microplates and kept for 16 h at 4oC. The

wells will be blocked by adding 200 µl of 1% BSA in PBS for 1 h at 37oC, and will be

washed three times with 200 µl PBS. This will be followed by the addition of 100 µl

aliquot of washed platelets stock (1 x 108 washed platelets suspension was pretreated with

10 µg of purified protein from cucumber sap or anti-integrin α2β1 mAb 6F1 or PBS

alone in a final volume of 1 ml Tyrode’s buffer for 10 min at 37oC). In case of purified

cucumber protease, the activity will be inhibited by EDTA at a final concentration of 5

mM before aliquoting. The reaction mixture was incubated at 37oC for 90 min and this

will be followed by washing three times with 200 µl PBS. The adherent platelets are then

treated with 150 µl lysis buffer (0.1 M citrate buffer pH 5.4 containing 5 mM p-

nitrophenyl phosphate and 0.1% Triton X-100) and incubated at 37oC for 90 min.

Addition of 2 N NaOH (100 µl) terminated the reaction by inactivating the platelet

membrane acid phosphatase activity and the optical density is measure at 405 nm.

Platelets adhesion is expressed as % adhesion taking the platelets suspension treated with

PBS as 100%.

In vitro platelet viability and in vivo platelet count

The method of Da Silva will be used to test the platelet viability. Briefly, the human

platelet rich plasma is incubate with increasing dose (0 – 8 µg) of purified protein of

cucumber sap in PBS for 30 min at 37°C in a volume of 100 µl containing 1×105 platelets.

Aliquots of 20 µl are mixe independently with 20 µl trypan blue (0.4%) and the platelet

morphology is observe with a microscope using a Neubauer chamber. The data expressed

as % viability, considering 100% viability in the absence of purified protein (control).

For in vivo platelet count, the method of Loria et al will be followed. Briefly, the purified

protein (µg/g body weight; defibrinogenating dose) will be injected intravenously in to

groups of mice (n=5) in 50 ml PBS, and then the animals will be allowed to bleed by

cardiac puncture at different time intervals of 1, 3 and 5 h after anaesthetized by diethyl

ether in to sodium citrate solution. The platelet counts/µl is determined by using

Neubauer chamber and a group of mice received the PBS alone served as control.

18.2 Connectivity of the participating institutions and investigators (in case of

multiinstitutional projects only)

Not applicable

18.3 Alternate strategies (if the proposed experimental design or method does not

work

what is the alternate strategy)

The principal investigator has the basic knowledge and background for carrying out this

project as he acquainted with the methodologies used for plasma coagulation and platelet

aggregation including protein purification techniques and has publications addressing

thrombosis and hemostasis. The co-principal investigators has enormous experience on

thrombosis and hemostasis related research work and got several publications on that area

and their assistance is essential to complete the project successfully. The team has

investigators with vast research knowledge on the objectives of the project. Therefore, we

are very confident in successful completion of the proposed work with a meaningful

outcome. Certainly, the proposed work will definitely bring out the meaning full

results/product.

Timelines: (Please provide quantifiable outputs) Period of study Achievable targets

First six months Second six months

Year 1

Chemicals and proposed

instruments will be purchased. All

the proposed parameters will be

standardized.

Aim 1: Isolation, biochemical and

pharmacological characterization of

anti-coagulant, anti-platelet and fibrin

clot hydrolyzing protease(s) from

cucumber sap

Year 2

Aim 2: Establish the site of action

of purified protease(s) on blood

coagulation cascade using specific

clotting factor deficient clinical

plasma samples and reconstituted

plasma

Aim 4: Investigate the kinetic

effects of protease(s) in the various

agonist induced platelet function of

human platelet rich plasma and

washed human platelets.

Aim 4. Cloning and expression of

protease(s) that possess anticoagulant,

antiplatelet and fibrinolytic activities

&

Manuscript will be prepared for the

publication.

Year 3

Aim 5: Establish the probable

molecular mechanism involved in

the anti-coagulant and anti-platelet

properties of purified protease (s).

Any unfinished work from previous

period will be done. The second/third

manuscript will be prepared for the

publication.

20. Name and address of 5 experts in the field

Sl no Name Designation Address

1 Dr. V.R. Devaraj

Associate Professor, Banglore University, Central college Campus,

Banglore, Karnataka, India.

2 Dr. Narender, Scientist E, Central Drug Research Institute,

Luknow, UP, India.

3 Dr. Raghu, P, Senior Scientist, National Institute of Hyderbad, AP,

India.

4 Prof, Sreeramulu, Department of Biochemistry, Gulbarga

University, Gulabarga, Karnataka, India.

5 Shylaja Dharmesh, Senior Scientist, Central Food Technological

Research Institute, Mysore, Karnataka, India.

PART IV: BUDGET PARTICULARS

Budget (In Rupees)

A. Non-Recurring (e.g. equipments, accessories, etc.)

Sl

No

Equipment required

1st year 2

nd year 3

rd year Total

1

2

3

4

5

6

7

8

9

10

11

12

FPLC (SCL10 Shimadzu)

Flurimetry (F5301PCShimadzu)

Lyophilizer (CRIST ALPHA)

Microscope with camera

Western blot unit

Electrophoresis unit

Ph meter

Incubator

weighing balance

Hot air oven

Computer, printer and Xerox

machine

Yes

Yes

Yes

----------------

Yes

Yes

--------------

-------------

--------------

-------------

Yes

-----------

--------------

--------------

---------------

Yes

--------------

------------

Yes

-----------

-----------

Yes

-------------

Yes

-------------

-------------

-------------

--------------

--------------

--------------

Yes

Yes

Yes

-----------

------------

------------

Rs. 18,00000

Rs, 10,00000

Rs, 4,00000

Rs, 700000

Rs, 50,000

Rs, 50,000

Rs, 20,000

Rs, 50,000

Rs, 100000

Rs, 50,000

Rs, 200000

Rs,300000

13

14

15

16

17

18

Laminar air flow

Centrifuge with variable rotors

PCR

Gel doc

Gel dryer

Transiluminator

Electrophorator

Yes

--------------

-------------

-------------

-------------

-------------

------------

Yes

Yes

Yes

Yes

yes

-------------

-------------

------------

-------------

-------------

------------

Rs,200000

Rs, 400000

Rs,300000

Rs, 200000

Rs, 100000

Rs, 200000

Total Rs. 61,20,000

Budget A Rs. 61,20,000

Overhead Costs (@20% of

project cost)

---------------- --------------- -------------- Rs, 12,24,000

Sub-Total (A) = Rs, 61, 20,000

B. Recurring

B.1 Manpower (See guidelines at Annexure-III)

Sl no Position

Consolidated

Emolument

Year 1

(Lakhs)

Year 2

(Lakhs)

Year 3

(Lakhs)

Total

(Lakhs)

1 JRF 2 14,000+HRA 3.696 3.696 3.696 11.088

Total -------------

---

------------- 3.696 3.696 3.696 11.088

Sub-Total (B.1) = 11.088 lakhs

B.2 Consumables

Sl

no

Items Year 1

(Lakhs)

Year 2

(Lakhs)

Year 3

(Lakhs)

Total

(Lakhs)

01 Chemicals &

Glassware/plastic

wares

3 3 3 9

Sub-Total (B.2) =9 lakhs

B.3 Travel

Travel Type Year 1

(Lakhs)

Year 2

(Lakhs)

Year 3

(Lakhs)

Total

(Lakhs)

Total 0.2 0.2 0.2 0.6

Sub –Total (B.3) = 0.6 lakhs

B.4 Contingency

Travel

Type

Year 1

(Lakhs)

Year 2

(Lakhs)

Year 3

(Lakhs)

Total

(Lakhs)

Total 1.0 1.0 1.0 3.00

Sub-total (B.4)= 3 lakhs

B.5 Overhead (If applicable)

Item Year 1

(Lakhs)

Year 2

(Lakhs)

Year 3

(Lakhs)

Total

(Lakhs)

4.08 4.08 4.08 12.24

Sub-total of B.5= 12.24lakhs

(B.1+B.2+B.3+B.4+B.5)

(11.088 lakh+9.0 lakh+0.6lakh+ 3.0 lakh+ 12.24lakh)= 35.928 lakhs

Sub-total of B= 35.928 lakhs

Grand Total (A + B)

(61.20 lakh+ 35.928) = 97.128 lakhs

Grand total=97.128 lakhs

Note: Please give justification for each head and sub-head separately mentioned in

the above table.

Financial Year: April - March

In case of multi-institutional project, the budget estimate to be given separately for each

institution.

JUSTIFICATION:

The principle investigator himself works at bench, in addition proposed staff is

essential to share the responsibilities to achieve the proposed project in time.

Consumable proposed are essential, chemicals such as reagent kits, factor deficient

plasma, human fibrinogen, thrombin, collagen, ADP, epinephrine, pepstatin,

leupeptin, column materials etc and other glassware’s.

Traveling grant is essential for attending symposia/ seminar/works shops/ training

programmes/consulting rare references in nearest universities/ research institutions.

Contingency is essential to meet expenses such as postal charges, typing of reports

and research articles, acquisition of books and documents of relevance, photo and

copying of reports and reprints etc.

Although, the preliminary work was standardized with the help of co-principle

investigators lab in Mysore University. Mysore, Karnataka, India. Currently working

Tumkur University is newly established and Department of PG studies and research

in Biochemistry lack all the basic instruments. Hence, all equipments are very much

essential for carrying out this type of work.

PART V: EXISTING FACILITIES

Resources and additional information

1. Laboratory: Available

a. Manpower: 2 JRFs will be recruited

b. Equipments

2. Other resources such as clinical material, animal house facility, glass house.

Experimental garden, pilot plant facility etc.

Basic facilities are available in the host institute (Tumkur University) Animal studies

will be carried out with the help of co-investigators from University of Mysore.

Sl No Equipments available

1 pH meter

2 Spectrophotometer

3 Centrifuge

4 Electronic balance

5 Colorimeter

6 Water bath

7 Incubator

8 Hot air oven

9 Electrophoresis unit

10 ELISA reader

11 Western blot

PART VI: DECLARATION/CERTIFICATION

It is certified that a) The research work proposed in the scheme/project does not in any way duplicate

the work already done or being carried out elsewhere on the subject. b) The same project proposal has not been submitted to any other agency for financial

support. c) The emoluments for the manpower proposed are those admissible to persons of

corresponding status employed in the institute/university or as per the Ministry of Science & Technologyguidelines (Annexure-III)

d) Necessary provision for the scheme/project will be made in the Institute/University/State budget in anticipation of the sanction of the scheme/project.

e) if the project involves the utilisation of genetically engineered organisms, we agree to submitan application through our Institutional Biosafety Committee. We also declare that while conducting experiments, the Biosafety Guidelines of the Department of Biotechnology would be followed in toto.

f) If the project involves field trials/experiments/exchange of specimens, etc. we will ensure that ethical clearances would be taken from concerned ethical Committees/Competent authorities and the same would be conveyed to the Department of Biotechnology before implementing the project.

g) it is agreed that any research outcome or intellectual property right(s) on the invention(s) arising out of the project shall be taken in accordance with the instructions issued with the approval of the Ministry of Finance, Department of Expenditure, as contained in Annexure-V.

h) We agree to accept the terms and conditions as enclosed in Annexure-IV. The same is signed and enclosed.

i) The institute/university agrees that the equipment, other basic facilities and such other administrative facilities as per terms and conditions of the grant will be extended to Investigator throughout the duration of the project.

j) The Institute assumes to undertake the financial and other management responsibilities of the project.

Signature of Principal Investigator: Signature of Executive

Authority

Date: Institute/University with

seal

Date:

Signature of Co-Investigator Signature of Co-

Investigator

Date: Date:

Signature of Co-Investigator Signature of Co-

Investigator

Date: Date:

Signature of Co-Investigator

Date :

PART VII: PROFORMA FOR BIOGRAPHICAL SKETCH OF

INVESTIGATORS

Provide the following information for the key personnel in the order listed on PART II.

Follow this format for each person. DO NOT EXCEED THREE PAGES

Name : Dr. Devaraja. S


Department/Institute/University: Tumkur University

Date of Birth : 24-04-1976 Sex (M/F): M, SC/ST: SC

Education (Post-Graduation onwards & Professional Career)

A. Position and Honors

Position and Employment (Starting with the most recent employment)

Sl

No

Institution

Place

Position From (Date) To

(date)

Degree Year University/Institute Field of Specialization

M. Sc., 2003 University of Mysore Biochemistry

Ph. D., 2010 University of Mysore Biochemistry

Post doc 2010 Cleveland Clinic,

Ohio State University, USA

Molecular cardiology

1 University of

Mysore

Department of

Biochemistry

Junior Research

Fellow

July 2005-august

2006

2 University of

Mysore

Department of

Biochemistry

Senior Research

Fellow

August 2006-January

2009

3 Learners Research

Institute ,

Cleveland clinic,

Ohio, USA

Department of

Molecular

Cardiology

Post doctoral

Fellow

14th May2010 to 2nd

July 2010

4 Tumkur University Department of

Biochemistry

Assistant

Professor

5th July2010 to till

date

Honors/Awards

Title of Award Year

1. Best teacher award from Dayananda Sagar College of 2003- 2004 Life Sciences, Bangalore 2. UPG Fellowship (Mysore University) 2005-2006 3. University Grand Commission Fellowship (UGC, Govt of India) 2006-2009 4. Post doctoral Fellowship, Learners Research Institute, 2010 Cleveland Clinic, Cleveland, Ohio, USA.

Professional Experience and Training relevant to the Project

The principal investigator has h vast research knowledge on the objectives of the

project which is accounted by their peer reviewed publications. Therefore, we are very

confident in successful completion of the proposed work with a meaningful outcome.

More over, Dr. K. Kemparaju and Dr. K.S. Girish, University of Mysore has lot of

expertise in the field of hemostatsis and platelet biology research. They have

standardized several protocols related to hemostasis. In addition, they have isolated

enzymatic toxins and elucidated their involvement in the hemostasis (coagulation

cascade). The data generated by the investigators are well appreciated by scientific

community as suggested by the citations for their peer reviewed publications in

international reputed journals. In addition, other co-investigators namely, Dr,

Sudharshan, Senior medical Officer and Thippeswamy and Bagyalakshmi also

committed to involve in the successful completion of this piece of work.

B. Publications (Numbers only): 08

Books : Nil, . Research Papers, Reports:16 , General articles :Nil

Patents : Nil, .Others (Please specify) : Nil

Selected peer-reviewed publications (Ten best publications in chronological order)

1. Devaraja S, Girish K.S, Gowtham Y.M.J, Kemparaju K. The Hag-protease-II is a

fibrin(ogen)ase from Hippasa agelenoides spider venom gland extract:

Purification, characterization and its role in hemostasis.. Toxicon. 2011.

2. Devaraja. S., Nagaraju. S., Mahadeswaraswamy. Y.H., Girish. K.S., Kemparaju.

K., A low molecular weight serine protease: purification and characterization

from Hippasa agelenoides spider venom gland extract. Toxicon 2008 52(1): 130-

138.

3. Nagaraju. S., Devaraja. S., Kemparaju. K., Purification and properties of

hyaluronidase from Hippasa partita (funnel web spider) venom. Toxicon 2007

50(3):383-393.

4. S.Devaraja, K. S. Girish, V. R. Devaraj and K. Kemparaju. Factor Xa-like and

fibrinogenolytic activities of a serine protease from Hippasa agelenoides spider

venom gland extract. J of Thromb Thrombolysis, 2010 Jan; 29(1): 119-126.

5. R.Sharma, Y.H. Mahadeswaraswamy, K. Harish Kumar, S. Devaraja, K.

Kemparaju, B.S. Vishwanath and K.S. Girish. Effect of Anticoagulants on the

Plasma Hyaluronidase activities. Journal of Clinical Laboratory Analysis 2008,

22, 1–5.

6. Mahadeswaraswamy YH, Devaraja S, Kumar MS, Goutham YNJ and Kemparaju

K. Inhibition of local effects of Indian Daboia/Vipera russellii venom by the

methanolic extract of grape seeds (Vitis vinifera L.). Indian Journal of

Biochemistry and Biophysics, 2009, 46, 154-160.

7. Hemshekhar M, Sunitha, Sebastin Santhosh M, Devaraja S, Kemparaju K, Girish

K.S. An overview on genus garcinia: phytochemical and therapeutical aspects.

Phytochemistry review 2011.

8. Y. H. Mahadeswaraswamy, B. Manjula, S. Devaraja, K.S. Girish and K.

Kemparaju: Daboia Russelli Venom Hyaluronidase: Purification, Charcterization

and inhibiton by β-3-(3-hydroxy – 4 – oxopyridyl) α-amino- propionic acid.

Current Topics in medicinal chemistry 2011.

List maximum of five recent publications relevant to the proposed area of work.

1. Devaraja S, Girish K.S, Gowtham Y.M.J, Kemparaju K. The Hag-protease-II is a

fibrin(ogen)ase from Hippasa agelenoides spider venom gland extract:

Purification, characterization and its role in hemostasis.. Toxicon. 2011.

2. Devaraja. S., Nagaraju. S., Mahadeswaraswamy. Y.H., Girish. K.S., Kemparaju.

K., A low molecular weight serine protease: purification and characterization

from Hippasa agelenoides spider venom gland extract. Toxicon 2008 52(1): 130-

138.

3. S.Devaraja, K. S. Girish, V. R. Devaraj and K. Kemparaju. Factor Xa-like and

fibrinogenolytic activities of a serine protease from Hippasa agelenoides spider

venom gland extract. J of Thromb Thrombolysis, 2010 Jan; 29(1): 119-126.

4. R.Sharma, Y.H. Mahadeswaraswamy, K. Harish Kumar, S. Devaraja, K.

Kemparaju, B.S. Vishwanath and K.S. Girish. Effect of Anticoagulants on the

Plasma Hyaluronidase activities. Journal of Clinical Laboratory Analysis 2008,

22, 1–5.

C. Research Support, Ongoing Research Projects

Sl

No.

Title of Project Funding

Agency

Amount Date of sanction and

duration

1 Fund for infrastructure

development in science

and technology

Vision Group

on Science

and

Technology,

Bangalore

40 lakh March 2011-March 2013

2 Screening of Indian

medicinal plants for

anticoagulant and

UGC Minor

project

2 lakh March 2012- March 2014

antiplatelet properties

Completed Research Projects (State only major projects of last 3 years)

Sl No. Title of Project Funding Agency Amount Date of

Completion

Not applicable

Place : Signature of Investigator

Date :

Dr. K. Kemparaju Biodata

Name : DR. K. Kemparaju

Designation : Associate Professor of Biochemistry

Date of Birth : 20 th July 1964

Organization : Department of Studies in Biochemistry

University of Mysore, Manasagangotri,

Mysore – 570 006, India.

Permanent address : As above

Educational Qualification:

Ph.D. Degree : Biochemistry (1996) University of Mysore

M.Sc. Degree : Biochemistry (1987) University of Mysore

B.Sc. Degree : CBZ (1985) University of Mysore

Teaching and research experience

2009 Due for : Professor of Biochemistry (yet to declare)

2001 to 2009 : Associate Professor of Biochemistry,

2002 to 2003 : Post doctoral fellow,

Albert Einstein College of Medicine,

Bronx, New York, USA.

1996-2001 : Senior Lecturer in Biochemistry, Univ. of Mysore.

1988-1996 : Lecturer in Biochemistry, Univ. of Mysore.

1987–1988 : Ph.D. student, Dept. of Biochemistry, Indian

Institute of Science, Bangalore.

Supervisor of PhD students

Awarded Submitted Working

8 1 5

Publications:

1. Kumar MS, Devaraj VR, Vishwanath BS, Kemparaju K. Anti-coagulant activity of a

metalloprotease: further characterization from the Indian cobra (Naja naja) venom. J

Thromb Thrombolysis. 2010 Apr;29(3):340-8.

2. Devaraja S, Girish KS, Devaraj VR, Kemparaju K. Factor Xa-like and

fibrin(ogen)olytic activities of a serine protease from Hippasa agelenoides spider

venom gland extract. J Thromb Thrombolysis. 2010 Jan; 29(1):119-26.

3. Ushanandini S, Nagaraju S, Nayaka SC, Kumar KH, Kemparaju K, Girish KS. The

anti-ophidian properties of Anacardium occidentale bark extract. Immunopharmacol

Immunotoxicol. 2009; 31(4):607-15.

4. Shashidharamurthy R, Mahadeswaraswamy YH, Ragupathi L, Vishwanath BS,

Kemparaju K. Systemic pathological effects induced by cobra (Naja naja) venom

from geographically distinct origins of Indian peninsula. Exp Toxicol Pathol. 2009

Sep 2.

5. Girish KS, Kemparaju K, Nagaraju S, Vishwanath BS. Hyaluronidase inhibitors: a

biological and therapeutic perspective. Curr Med Chem. 2009; 16(18):2261-88.

Review.

6. Mahadeswaraswamy YH, Devaraja S, Kumar MS, Goutham YN, Kemparaju. K.

Inhibition of local effects of Indian Daboia/Vipera russelli venom by the methanolic

extract of grape (Vitis vinifera L.) seeds. Indian J Biochem Biophys. 2009 Apr;

46(2):154-60.

7. Chandrashekara KT, Nagaraju S, Nandini SU; Basavaiah, Kemparaju K.

Neutralization of local and systemic toxicity of Daboia russelii venom by Morus alba

plant leaf extract. Phytother Res. 2009 Aug; 23(8):1082-7.

8. Sharma R, Mahadeswaraswamy YH, Harish Kumar K, Devaraja S, Kemparaju K,

Vishwanath BS, Girish KS. Effect of anticoagulants on the plasma hyaluronidase

activities. J Clin Lab Anal. 2009; 23(1):29-33.

9. Devaraja S, Nagaraju S, Mahadeswaraswamy YH, Girish KS, Kemparaju K. A low

molecular weight serine protease: Purification and characterization from Hippasa

agelenoides (funnel web) spider venom gland extract. Toxicon. 2008 Jul; 52(1):130-8.

10. Mahadeswaraswamy YH, Nagaraju S, Girish KS, Kemparaju K. Local tissue

destruction and procoagulation properties of Echis carinatus venom: inhibition by

Vitis vinifera seed methanol extract. Phytother Res. 2008 Jul; 22(7):963-9.

11. Nagaraju S, Girish KS, Fox JW and Kemparaju K. ‘Partitagin’ a hemorrhagic

metalloprotease from Hippasa partite spider venom: Role in tissue necrosis Biochimie

89, 1322-1331, 2007.

12. Nagaraju S, Devaraja S, Kemparaju K. Purification and properties of hyaluronidase

from Hippasa partita (funnel web spider) venom gland extract.Toxicon. 2007 Sep 1;

50(3):383-93. Epub 2007 Apr 24.

13. Girish KS, Kemparaju K. The magic glue hyaluronan and its eraser hyaluronidase: a

biological overview. Life Sci. 2007 May 1; 80(21):1921-43. Review.

14. Shashidharamurthy R, Kemparaju K. Region-specific neutralization of Indian cobra

(Naja naja) venom by polyclonal antibody raised against the eastern regional venom:

A comparative study of the venoms from three different geographical distributions. Int

Immunopharmacol. 2007 Jan; 7(1):61-9.

15. Girish KS, Machiah KD, Ushanandini S, Harish Kumar K, Nagaraju S, Govindappa

M, Vedavathi M, Kemparaju K. Antimicrobial properties of a non-toxic glycoprotein

(WSG) from Withania somnifera (Ashwagandha). J Basic Microbiol. 2006; 46(5):365-

74.

16. Nagaraju S, Mahadeswaraswamy YH, Girish KS, Kemparaju K. Venom from spiders

of the genus Hippasa: biochemical and pharmacological studies. Comp. Biochem

Physiol C Toxicol Pharmacol. 2006 Sep; 144(1):1-9.

17.Ushanandini S, Nagaraju S, Harish Kumar K, Vedavathi M, Machiah DK, Kemparaju

K, Vishwanath BS, Gowda TV, Girish KS. The anti-snake venom properties of

Tamarindus indica (leguminosae) seed extract. Phytother Res. 2006 Oct; 20(10):851-

8.

18. Shashidharamurthy R, Kemparaju K. A neurotoxic phospholipase A2 variant:

isolation and characterization from eastern regional Indian cobra (Naja naja) venom.

Toxicon. 2006 Jun 1; 47(7):727-33.

19. Girish KS, Kemparaju K. Inhibition of Naja naja venom hyaluronidase: role in the

management of poisonous bite. Life Sci. 2006 Feb 23; 78(13):1433-40.

20. Kemparaju K, Girish KS. Snake venom hyaluronidase: a therapeutic target. Cell

Biochem Funct. 2006 Jan-Feb; 24(1):7-12. Review.

21. Girish KS, Kemparaju K. Inhibition of Naja naja venom hyaluronidase by

plant-derived bioactive components and polysaccharides. Biochemistry (Mosc). 2005

Aug; 70(8):948-52.

22. Girish KS, Kemparaju K. A low molecular weight isoform of hyaluronidase:

purification from Indian cobra (Naja naja) venom and partial characterization.

Biochemistry (Mosc). 2005 Jun; 70(6):708-12.

23. Rajesh R, Raghavendra Gowda CD, Nataraju A, Dhananjaya BL, Kemparaju K,

Vishwanath BS. Procoagulant activity of Calotropis gigantea latex associated with

fibrin(ogen)olytic activity. Toxicon. 2005 Jul; 46(1):84-92.

Book chapter

1. K. Kemparaju, K.S. Girish and S. Nagaraju; Hyaluronidases, a Neglected Class of

Glycosidases from Snake Venom, Beyond a Spreading Factor. (Reptile Venoms and

Toxins by S. Meckessy, IRL Press, USA.) 235 to 256, 2009.

Invited talks delivered in national and international symposia

1. K. Kemparaju; Geographical variation of Naja naja venom: an important factor in

making therapeutic antivenom. 2005; Dept. of Biochemistry and Biotechnology,

Annamalai University, Annamalai nagar, Tamilnadu, India.

2. K. Kemparaju; Hyaluronidase : a potential target in the management of

snakebite,2007; School of Biotechnology, Chemical and Biomedical Engineering,

VIT University, Vellore, Tamilnadu, India.

3. K. Kemparaju; The metzincin Partitagin is a unique Beta fibrinogenase from

Hippasa

partita spider venom that inhibits platelet aggregation. 2008; Dept. of

Biochemistry

and Biotechnology, Annamalai University, Annamalai nagar, Tamilnadu, India.

4. K. Kemparaju; NN-PF3 is a fibrin(ogen)olytic enzyme from Naja naja venom.

2009, Dept. Biochemistry and Biotechnology, Annamalai University, Annamalai

nagar, Tamilnadu, India.

5. K.Kemparaju; Local effects of snakebite: possible therapeutic targets for the

better management. 2008, Little flower Hospital and Research Centre ,

Angamaly, Kochin , kerala India.

6. K.Kemparaju; Neutralization Vipera russellii venom and its purified

hemorrhagin by gallic acid, 2009, Medical Biotechnology on Clinical research, J,

N. Tata Auditorium, IISc Campus, Bangalore.

7. K.Kemparaju; The non toxic fibrin(ogen)olytic enzyme from Naja naja venom:

inhibition of collagen induced human platelet aggregation by affecting alpha-2,

beta-1 and GP-VI. 2010, State level Refresher Course for pre-University Teachers,

Suttur, Mysore, India.

Papers/Posters presented at national and international symposia:

1. Kumar M.S., Vishwanath B.S., Kemparaju K. NN-PF3, an anticoagulant and

antiplatelet metalloprotease from the Indian cobra (Naja naja) venom. International

anatomical sciences and cell biology conference. May 2009, Department of anatomy,

National university of Singpore, Singpore.

2. Mahadeswaraswamy YH and Kemparaju K. Screening of Indian medicinal plants for

anti-venom property against Indian Daboia/ Vipera russellii and Echis carinatus

venoms. Dr. Norman E. Borlaug commemoration national conference on “Plant

diversity and plant health”. March 2010, Department of Studies in Botany,

University of Mysore, Mysore. India.

3. Mahadeswraswamy Y.H, Girish K.S and Kemparaju, K. Antivenom potential of

Gallic acid against the Indian Daboia/Vipera russellii venom and its purified

hemorrhagic complex. State level conference on Recent advances in Environmental

Science and Engineering. October, 2009. Department of Biotechnology and

Engineering, Shridevi Institute of Engineering and Technology, Tumkur, India.

4. Devaraja S and Kemparaju K. A low molecular weight serine protease the Hag-

protease from Hippasaagelenoides spider venom gland extract: Role in

fibrin(ogen)olysis andplatelet function. International conference on `cardiovascular

diseasessecondary to the metabolic disorders: Mechanism and therapy`.December,

2009. Department of Studies in Biochemistry, University of Mysore, Mysore. India.

1. Mahadeswaraswamy YH and Kemparaju K. Gallic acid: a potent inhibitor of Indian

Daboia russellii venom and its purified hemorrhagic complex induced local toxicity.

National symposium on Bioactive molecules: from discovery to industry. April 2009.

Department of Studies in Biochemistry, University of Mysore, Mysore. India.

2. Mahadeswaraswamy YH and Kemparaju K. Local tissue destruction property of

Indian Daboia /Vipera russellii venom and its purified hemorrhagic complex:

Inhibition by Gallic acid. National conference on Recent trends in animal

physiology. October, 2009, Department of Studies in Zoology, University of Mysore,

Mysore. India.

3. Devaraja S and Kemparaju K. Local and systemic toxicity of Hag-protease a low

molecular weight serine protease from Hippasa agelenoides spider venom gland

extract. National symposium on Recent Trends in Animal Physiology. October 2009.

Department of Studies in Zoology, University of Mysore, Mysore. India

4. Mahadeswraswamy YH, Girish KS and Kemparaju K. Local toxicity of Indian

Daboia/Vipera russellii venom and its purified hemorrhagic complex: Contribution

for the better management. National conference on Medical Biotechnology and

Clinical research. October 2009. Department of Biotechnology, Sri. M.

Visvesvaraya Institute of Technology and Sri Krishnadevaraya Education Trust, JN

Tata Auditorium, Indian Institute of Science campus, Bangalore, India.

5. Yashonandana J Gowtham, Sadiq Saleh Ahmed, Naveen H M and K. Kemparaju.

The Indian King Cobra (Ophiophagus hannah) Venom: A preliminary study.

National symposium on Bioactive molecules: from discovery to industry. April 2009.

Department of Studies in Biochemistry, University of Mysore, Mysore. India.

6. Yashonandana J Gowtham and K. Kemparaju. The Indian King Cobra

(Ophiophagus hannah) Venom strongly interferes in Hemostasis. National

conference on Recent trends in animal physiology. October, 2009, Department of

Studies in Zoology, University of Mysore, Mysore. India.

7. Devaraja S and Kemparaju K. Hag-protease from Hippasa agelenoides spider

venom extract: Role in tissue necrosis and hemostasis. National symposium on

Bioactive molecules: from discovery to industry. April 2009. Department of Studies

in Biochemistry, University of Mysore, Mysore.

8. Rashmi K, Zahara Ashkavand, Devaraja S and K.Kemparaju. preliminary studies

on proteolytic activity of cucumber sap extract. Nationalsymposium on Bioactive

molecules: from discovery to industry. April2009. Department of Studies in

Biochemistry, University of Mysore, Mysore.

9. Mahadeswaraswamy YH and Kemparaju K. Local effects of snakebite: Possible

therapeutic targets for the better management. International symposium on Snakes,

venoms and snake bites. SNA-CON October 2008, Angamaly, Cochin.

10. Girish. K.S., Nagaraju, S., and Kemparaju, K. Hyaluronidase: a potential target in

the management of snakebite. 3rd National Symposium on Venoms and Toxins. Dept.

of Biochem. Univ. of Mysore (2004).

11. Nagaraju, S., Girish. K.S., and Kemparaju, K. Spider venom: comparative

characterization from three different species (Hippasa). 3rd National Symposium on

Venoms and Toxins. Dept. of Biochem. Univ. of Mysore (2004).

12. Kemparaju K. and Girish.K.S. A pharmacological examination of the Indian saw-

scaled viper venom. 28, Proc. National Symp. On Proteins in Biology, Mysore, India

(1999).

13. Jagadeesha, D.K., Shashidhara murthy, R and Kemparaju, K. Isolation and

characterization of a non-toxic fibrinogenase from Indian cobra (Naja naja) venom.

48, Proc. National Symp. On Proteins in Biology, Mysore, India (1999).

14. Shashidhara murthy, R., Jagadeesha, D.K. and Kemparaju, K. Isolation,

characterization and chemical modification of histidine and lysine residues of a

neurotoxic phospholipase A2 from Indian cobra (Naja naja) venom. 55, Proc.

National Symp. On Proteins in Biology, Mysore, India (1999).

15. Kemparaju, K. and Gowda, T.V. Comparative characterization of phospholipases

A2 from Indian saw-scaled viper (Echis carinatus) venom. 324, 64th Annual Meeting

of SBC, Lucknow, India (1995).

16. Kemparaju, K. and Gowda, T.V. Selective neutralization of enzymatic activity of a

toxic phospholipase A2 from Echis carinatus venom by polyclonal antibodies, P7-82,

XVIth IUBMB meeting, India (1994).

17. Kemparaju, K. and Gowda, T.V. Purification and characterization of a toxic

phospholipase A2 (EC-PLA2-IV) from Indian saw-scaled viper (Echis carinatus)

venom. PEFD 019, 197, 62nd Annual Meeting of SBC, Madurai, Inida (1993).

18. Kemparaju, K. and Gowda, T.V. Preliminary studies on Indian saw-scaled viper

(Echis carinuatus) venom. Abstract No. 365, 61st Annual Meeting of SBC,

Hyderabad, India (1992).

19. Kemparaju, K., Kasturi, S. and Gowda, T.V. Effect of trypsin digestion on the

enzymatic and edema inducing activities of Vipera russelli phospholipase A2 A4-9,

15, Second Asia-Pacific congress on Animal, plant and microbial Toxins, Varanasi,

India (1990).

Resource person in academic programmes

1.Immune system: an overview.

Science and Technology Rejuvenation program for Pre-University Teachers organized by

Government Pre-University College Board, (2010) .

2. Mechanisms of signal transduction.

Refresher Course for Zoology Teachers. Organized by DOS in Zoology at Academic

Staff College, Univ. of Mysore, Mysore (2002).

3. Immunological techniques.

Refresher Course for Zoology Teachers. Organized by DOS in Zoology at Academic

Staff College, Univ. of Mysore, Mysore (2000).

4. SDS-PAGE; Detection and Analysis of proteins.

Work Shop on “Techniques in protein purification” in DOS in Biochem., July 15th to 24th

1999.

5. Proteins: structure-function relationships.

In-service Training Course in Chemistry for Postgraduate Teachers of Kendriya

Vidyalaya Sanghatan, Organized by KVS, New Delhi conducted at Regional Institute of

Education, Mysore (1997).

6. Fundamentals of Biochemistry- special emphasis on enzymes and metabolism.

In-service Training Course in Chemistry for Postgraduate Teachers of Kendriya

Vidyalaya Sanghatan, Organized by KVS, New Delhi conducted at Regional Institute of

Education, Mysore (1996).

Workshop/symposia conducted

1. Joint Secretary for Work Shop on “Techniques in protein purification” in DOS in

Biochem., July 15th to 24th 1999.

2. Treasurer, National Symposium on “Proteins in Biology: Structure/Function,

Expression and Specificity of Action” at Dos in Biochem. Univ. of Mysore, Mysore,

November 15th to 16th 1999.

3. Joint Secretary, 3`rd National symposium on venoms and Toxins” at Dos in Biochem.

Univ. of Mysore, Mysore, Jan 23 rd and 24th 2004.

4. Joint Secretary, International Conference on Cardiovascular Diseases Secondary to the

Metabolic Disorder: Mechanisms and Therapy. Dept. of Biochemistry, University of

Mysore, Mysore, Dec. 17 – 19,

External Reviewer for the Journals

Toxicon, Comparative Biochemistry and Physiology, Gene, Basic and Clinical

Pharmacology and Toxicology, Molecular and Cellular Biochemistry, Indian Journal of

Biochemistry and Biophysics, Chinese Journal of Medicine, Indian Journal of Medical

Research (ICMR)

Evidence Based complementary Medicine, African Journal of Agricultural Sciences

Memberships

Society of Biological Chemists, India, People for Animals.

Dr. K. S. Girish biodata

Name : Dr. Girish. K.S

Birth Date : June 22, 1976

Home Address : Kesturu, Krishnarajanagara

Taluk, Mysore-Dist,

Karnataka-571602, India

Citizenship : India

Postal Address : Dr. Girish. K.S

Assistant Professor,

DOS in Biochemistry,

Manasagangothri,

University of Mysore,

Mysore, India-560 006.

Email : [email protected]

Current address : DOS in Biochemistry,

University of Mysore, MGM

Educational Qualification:

Ph.D. Degree : Biochemistry (2004) University of Mysore

M.Sc. Degree : Biochemistry (1998) University of Mysore

B.Sc. Degree : BBM (1996) University of Mysore

POST-DOC : Virology/Tendon biology/Cancer biology (2004-

2007)

University of Pittsburgh/University of Virginia,

USA

Appointments and positions

Dates

Attended

Name and Location of Organization Rank/Title

1999-2004

Manasagangothri,

Mysore University, Mysore, India

Post-Graduate Teaching

Assistant

2004-2005 Department of Surgery

University of Pittsburgh

Postdoctoral Associate

2005- 2007 Department of Orthopedics

University of Virginia

Research Associate

2008

DOS in Biochemistry

University of Mysore, India

Assistant Professo

Peer-Reviewed Publications:

1. Girish KS, Hogan M, James R, Balian G, Hurwitz S, Chhabra A. Growth

differentiation factor-5 regulation of extracellular matrix gene expression in murine

tendon fibroblasts. J. Tissue Engineering and Regenerative Med 2010 (Accepted).

Impact factor: 1.6

2. Devaraja S, Girish KS, Devaraj VR, Kemparaju K. Factor Xa-like and

fibrin(ogen)olytic activities of a serine protease from Hippasa agelenoides spider venom

gland extract. J Thromb Thrombolysis 29, 119-126, 2010. Impact factor: 1.8

3. Ushanandini S, Nagaraju S, Chandra Nayaka S, Harish Kumar K, Kemparaju K,

Girish KS. The Anti-ophidian Properties of Anacardium occidentale bark extract.

Immunopharmacology and Immunotoxicology 31, 607-615, 2009. Impact factor: 0.9

4. Girish KS, Kemparaju K, Nagaraju S and Vishwanath BS. Hyaluronidase

Inhibitors: A biological and Therapeutic Perspective. Current Medicinal Chemistry

16, 2261-2288, 2009. Impact factor: 4.9

5. Sharma R, Mahadeshwara swamy YH, Harish kumar K, Devaraja S, Vishwanath

BS, Kemparaju K and Girish KS. Effect of anticoagulants on plasma hyaluronidase

activities. Journal of Clinical Laboratory Analysis 23, 29-33, 2009. Impact factor: 1.2

6. Devaraja S, Nagaraju S, Mahadeswaraswamy YH, Girish KS, Kemparaju K. A

low molecular weight serine protease: Purification and characterization from Hippasa

agelenoides (funnel web) spider venom gland extract. Toxicon 52, 130-138, 2008.

Impact factor: 2.45

7. James R, Kesturu GS, Balian G, Chhabra AB. Tendon: Biology, Biomechanics,

Repair, Growth Factors, and Evolving Treatment Options. American Journal of Hand

Surgery 33, 102-112, 2008. Impact factor: 1.6

8. Mahadeshwara swamy YH, Nagaraju S, Girish KS and Kemparaju K.

Neutralization of Saw-scaled viper (Echis Carinatus) venom induced local effects and

some enzymes by Mimosa pudica root extract. Phytotherapy Research 22, 963-969,

2008. Impact factor: 1.3

9. Nagaraju S, Girish KS, Fox JW and Kemparaju K. ‘Partitagin’ a hemorrhagic

metalloprotease from Hippasa partite spider venom: Role in tissue necrosis Biochimie

89, 1322-1331, 2007. Impact factor: 3.1

10. Girish KS and Kemparaju K. The magic glue hyaluronan and its eraser

hyaluronidase: a biological overview. Life Sciences 80 1921-1943, 2007. Impact factor:

2.6

11. Girish KS, Deepa M, Ushanandini S, Harish Kumar K, Nagaraju S, Govindappa

M, Vedavathi M and Kemparaju K: Antimicrobial properties of a non-toxic

glycoprotein (WSG) from Withania somnifera (Ashwagandha). Journal of Basic

Microbiology 46 (5) 365-374, 2006. Impact factor: 0.8

12. Nagaraju S, Mahadeshwaraswamy YH, Girish KS and Kemparaju K:

Biochemical and pharmacological characterization of venom from the spiders H. partita,

H. aglenoides and H. mahabaleshwarensis tikadar and malhotra. Communicated to

Comparitive Biochemistry and Physiology (Part C) 144 (1), 1-9, 2006. Impact

factor: 2.4

13. Ushanandini S, Nagaraju S, Harish Kumar K, Vedavathi M, Deepa M, Kemparaju

K, Vishwanath BS, Gowda TV, Girish KS: The anti snake venom properties of

Tamarindus indica (Leguminosae) seed extract. Phytotherapy Research 20 (10) 851-

858, 2006. Impact factor: 1.3

14. Deepa K. Machiah, Girish KS and Gowda TV: A glycoprotein from a folk

medicinal plant, Withania somnifera, inhibits hyaluronidase activity of snake venoms.

Comparative Biochemistry and Physiology (Part C) 143, 158-161, 2006. Impact

factor: 2.4

15. Girish S Kesturu, Colleton BA, Liu Y, Heath L, Shakil OS, Rinaldo Jr CR,

Shankarappa R: Minimization of genetic distances by the Consensus, Ancestral and

Center-of-Tree (COT) sequences for HIV-1 variants within an infected individual and

the design of reagents to test immune reactivity. Virology 348 (2) 437-448, 2006.

Impact factor: 3.2

16. Girish KS and Kemparaju K: The hyaluronidase enzyme a prime target for better

management of snakebite. Life sciences 78, 1433-1440, 2006. Impact factor: 2.6

17. Kemparaju K and Girish KS: Snake venom hyaluronidase: s therapeutic target.

Cell Biochemistry and Function 24 (1), 7-12, 2006. Impact factor: 1.5

18. Vedavathi M, Girish KS and Karunakumar M: A novel low molecular weight

alanine aminotransferase from fasted rat liver. Biochemistry (Moscow) 71 (Suple 1),

S105-S112, 2006. Impact factor: 1.2

19. Girish KS and Kemparaju K: Inhibition of Naja naja venom hyaluronidase by

plant derived bioactive components and polysaccharides. Biochemistry (Moscow) 70

(8), 948-952, 2005. Impact factor: 1.2

20. Girish KS and Kemparaju K: A low molecular weight isoform of hyaluronidase:

purification from Indian cobra (Naja naja) venom and partial characterization.

Biochemistry (Moscow) 70 (6), 708-712, 2005. Impact factor: 1.2

21. Vedavathi M, Girish KS and Karunakumar M: Isolation and characterization of

alanine aminotransfarase isoforms from starved male rat liver. Molecular and Cellular

Biochemistry 267, 13-23, 2004. Impact factor: 1.56

22. Girish KS, Mohan Kumari HP, Nagaraju S, Vishwanath BS and Kemparaju K.

Hyaluronidase and Protease activities from Indian snake venoms: Neutralization by

Mimosa pudica root extract. Fitotherapia, 75 (3-4), 378-380, 2004. Impact factor: 1.25

23. Girish KS, Shashidhara murthy R, Nagaraju S, Veerabasappa Gowda TV and

Kemparaju K. Isolation and characterization of hyaluronidase a “spreading factor” from

Indian cobra (Naja naja) venom. Biochimie, 86(3), 193-202, 2004. Impact factor: 3.1

24. Girish KS, Jagadeesh DK, Rajeev KB and Kemparaju K., Snake venom

hyaluronidase: An evidence for isoforms and extracellular matrix degradation.

Molecular and Cellular Biochemistry, 240, 105-110, 2002. Impact factor: 1.56

25. Jagadeesh DK, Shashidharamurthy R, Girish KS and Kemparaju K. A non-toxic

anticoagulant metalloprotease: purification and characterization from Indian cobra

(Naja naja naja) venom. Toxicon 40 (6), 667-675, 2002. Impact factor: 2.45

26. Shashidharamurthy R, Jagadeesh DK, Girish KS and Kemparaju K. Variations

in biochemical and pharamacological properties of Indian cobra (Naja naja naja) venom

due to geographical distribution. Molecular and Cellular Biochemistry, 229, 93-101,

2002. Impact factor: 1.56

Book Chapters:

Kemparaju K, Girish KS and Nagaraju S. Hyaluronidases, a neglected class of

glycosydases from snake venom: beyond a spreading factor in a book “ Hand Book of

Venoms and Toxins of Reptiles” (Cat # 9165), August 2009, CRC Press LLC.

Hemshekhar M, Kemparaju K, and Girish KS. An overview on remedial qualities of

Tamarindus indica seeds. Book Chapter in “Nuts and Seeds in Health and Disease”

Edited by Victor R Preedy, Academic Press-Elsevier, 2010.

National/International Symposia Presentation:

1. Girish KS, James R, Park A, Hogan MV, Balian G, Chhabra AB. Expression

of Matrix and Tendon Specific Cellular Markers by Rat Adipose Derived

Mesenchymal Stem Cells Treated with Growth Differentiation Factor-5. International

Conference on Anatomical and Cell Biology Sciences. National University of

Singapore, Singapore, May 26th to May 30th 2010.

2. Mahadeswraswamy YH, Girish KS and Kemparaju K. Local toxicity of

Indian Daboia/Vipera russellii venom and its purified hemorrhagic complex:

Contribution for the better management. National conference on Medical

Biotechnology and Clinical research. October 2009. Department of Biotechnology, Sri.

M. Visvesvaraya Institute of Technology and Sri Krishnadevaraya Education Trust, JN

Tata Auditorium, Indian Institute of Science campus, Bangalore, India. (Best poster

presentation award, second prize).

3. Hemshekhar, Sunitha K, Avinash S, Mouna N, Deepthi L, Soumya N, Girish

KS. Guggul proteins: A group of neglected bioactive molecules. National symposium

on “Bioactive molecules: from Discovery to Industry”. Organized by DOS in

Biochemistry, University of Mysore, Manasagangothri, April 2009, Mysore

4. Girish KS, James R, Balian G and Chhabra A. GDF-5 modulates the

synthesis and expression of extracellular matrix and cell adhesion related molecules of

rat Achilles tendon fibroblasts. 54th Orthopedic Research Society meeting, San

Francisco, CA, USA. Feb 2008

5. Hogan MV, Starnes T, Kesturu Girish, James R, Balian G, Chhabra A. Cell-

based Augmentation of Tendon Repair Using a Biodegradable Polymer Scaffold. 2008

AAOS Annual Meeting, San Francisco, CA. (Accepted Podium)

6. Girish KS, Roshan J, Balian G, Chhabra A: Treatment with GDF-5 induces

tendinogenic differentiation of adipose derived stromal cells. 53rd Orthopedic Research

Society meeting, San Deigo, CA, USA. March 2007

7. Hogan MV, Starnes T, Kesturu Girish, James R, Huang D, Balian G,

Chhabra A. GDF-5 Induced Tendon Repair and Regeneration. 2007 National Medical

Association Annual Convention and Scientific Assembly, Honolulu, HI (Podium)

8. Kwoon M, Girish KS, Meyers T and Diduch D: Intra-articular

chondroprotection against septic arthritis with the use of adenosine agonist (ATL-313).

53rd Orthopedic Research Society meeting, San Deigo, CA, USA. March 2007

9. Trevor S, Huang D, Girish KS, Balian G and Chhabra A: Immunolocalization

of GDF-5 in native and surgically repaired Rat Achilles Tendon. 53rd Orthopedic

Research Society meeting, San Deigo, CA, USA. March 2007

10. Trevor S, Huang D, Girish KS, James R, Balian G and Chhabra A:

Potentiation of tendon repair and regeneration. Annual meeting of orthopedic surgeons

(AAOS). San Deigo, CA, USA. March 2007

11. Meyers T, Girish KS, Kwoon M and Diduch D: Intravenous versus intra-

articular chonroprotection against septic arthritis with the use of an adenosine agonist

(ATL-313). 59th Orthopedic annual Virginia Orthopedic Society meeting, Norfolk,

Virginia, USA May 19th to May 21st 2006

12. James R, Huang D, Girish KS, McCulloch MD, O’Neal BL, Balian G,

Chhabra AB: GDF-5 improves collagen fiber organization and increases extracellular

matrix synthesis during tendon repair. 52nd Orthopedic Research Society meeting,

Chicago, IL, USA. March 19th to March 25th 2006

13. Girish Kesturu*, LianFu Wang, Charles R. Rinaldo, Jr., and Raj

Shankarappa: Quantitative Considerations in the Assessment of Hepatitis C Virus

Quasispecies. 3rd Annual Richard L. Simmons Lecture in Surgical Science and

Department of Surgery Research Day. University of Pittsburgh, Pittsburgh, USA. April

27th 2005

14. Girish KS and Kemparaju K. The hyaluronidase enzyme a prime target in

management of snakebite. 3rd National symposium on venoms and toxins. University of

Mysore, Mysore, India, Jan 23th to Jan 26th 2004

15. Nagaraju S, Girish KS and Kemparaju K. Comparative characterization of

three Indian funnel-web spider venoms. 3rd National symposium on venoms and toxins.

University of Mysore, Mysore, Mysore, India, Jan 23th to Jan 26th 2004

16. Dhananjaya BL, Girish KS and Cleatus DM Desouza. Partial characterization

of 5’ nucleotidase from Indian cobra (Naja naja) venom. 3rd National symposium on

venoms and toxins. University of Mysore, Mysore, Mysore, India, Jan 23th to Jan 26th

2004

17. Vedavathi M, Girish KS and Karunakumar M. Comparative characterization

of low and high molecular weight isoforms of alanine aminotransfarases from starved

rat liver. 3rd National symposium on venoms and toxins. University of Mysore, Mysore,

Mysore, India, Jan 23th to Jan 26th 2004

18. Kemparaju K and Girish KS. A pharmacological examination of the Indian

saw scaled viper venom. National Symposium on Proteins (Structure and Function).

Mysore, India, 1999

External Reviewer

Toxicology, Biochimie, Connective tissue research, Journal of Medicinal Chemistry,

Biomaterials, and Pesticide, Biochemistry and Physiology

Number of papers reviewed: 12

Dr.B.G.Sudharshan Biodata

PERSONAL PROFORMA

NATIONALITY : Indian D.O.B. : 18-04-1975 GENDER : MALE LANGUAGES KNOWN : English, Kannada, and Hindi ACADEMIC COURSE : MBBS, PhD (Bio-medical engineering)

ACADEMIC PERFORMANCES:

Course University Institution class Percentage

PhD Avinashlingam university , Coimbatore

CMRTU , R.V.C.E (MAY2007TO DEC 2009).

MBBS Bangalore University SRI SIDDARTHA MEDICAL COLLEGE TUMKUR

PASS

PHD THESIS TITLED:

“Electromagnetic and RF Interference on the Implanted Cardiac Pacemakers and Remedial Measures for their Effective Functioning”

PUBLICATIONS:

� Interference with Cardiac Pacemaker due to Cellular Phones, International Journal of

Computer Applications in Engineering, Technology and Science, issue: April 2009,

pp.289-291

� A Review on Electromagnetic Interference on Cardiac pacemakers, presented and

published at International conference on Emerging Technologies and Applications in

Engineering, Technology and Sciences, held on 13-14 January, 2008

� Electromagnetic Interference due to Mobile Phones on Artificial Cardiac Pacemaker

Patients, XXXII National Systems Conference, 03-11-2008, at Roorkee

� Heart rate variability ;A review paper to be presented in an international conference

in 2011 International Conference on Signal Processing, Communication,

Computing and Networking Technologies to be held on 21-22 july2011

� HRV under stress using two different methods of non-linear analysis in a national

conference held at Dr.AIT WIRELESS CONTROL AND Communication

technology on apr28-29.

� Understanding autonomic and dynamics under stress using non-linear analysis of

data page no page no152-155 National conference on emerging trends in bio-

medical signal processing 26feb -2011

WORK EXPERIENCE: 1. ONE YEAR ROTATORY INTERNSHIP IN DISTRICT HOSPITAL TUMKUR;

• Attending opd

• Emergencies

• Community health programmes

• Serving rural people

• Regular concerned department posting 2. RESIDENT DOCTOR DEPARTMENT OF NEUROLOGY MANIPAL

HOSPITAL;

• Attending neuro opd

• Treating patients in wards in absence of specialists

• Performing procedures like lumbar punctures etc .3. RESIDENT DOCTOR DEPARTMENT OF ICU BANGALORE HOSPITAL;

• Treating critically ill patients

• Treatment terminally ill patients

• Performing procedures like intubation etc 4. MEDICAL OFFICER MINISTRY OF LABOUR, GOVT OF INDIA;

• Regular opd

• Handling emergencies

• Had a mobile dispensing unit where in patients were treated at there working place

• Implementation of national programmes such as immunisation, vaccination 5. MEDICAL OFFICER R.V.C.E TILL TO DATE;

• Treating patients on opd and day care basis

• The strength of students ranges from 1200 in hostels and 4500 students on campus and staff members 450 and their families

• Maintaining hygiene and health of all these people

• Regular vaccination for students mainly hepatitis-a .hepatitis-b and chicken-pox vaccines

• Regular medical checkups for students and food handlers

• Attending emergencies during night time

• Shifting of patients to the nearby tertiary care centres and taking care of them till they are discharged

• Counseling for students

• Arranging health camp for students mainly eye check-up camps

• Oral hygiene camp annually POSITION HELD AT RVCE:

1. MEDICAL OFFICER IN RVCE Health centre and hostel Health centre.

2. ASSISTANT PROFESSOR IN Biomedical Instrumentation Signal Processing Department.

3. RESEARCH AND ACADEMICS; Guiding students of M-tech biomedical instrumentation and engineering for

research projects namely; � Heart rate variability on stress induced subjects by using non-linear

methods. � Heart rate variability in hypothyroid patients. � Introduction classes for bio-medical instrumentation students. � Handling theory classes such as Physiology for Engineers.

FUNDED PROJECTS:

1. As co-investigator, in a project titled insilico analysis of bone-tumors funded by DST.

2. Projects submitted for various funding agencies;

• Synthesis and characterization of plasmin and their thrombolytic applications submitted as CO-PI to ICMR.

• Development of Doppler based foetal ultrasound stethoscope submitted to DBT as CO-PI.

• Development of a healthcare monitoring device for early detection of cardio-vascular diseases. As principal investigator to DBT

3. An ergonomic study of working conditions in garment industry and their health complications and remedial measures. Submitted to ICSSR.

Thippeswamy.T.G. Biodata

1. Name : T.G.THIPPESWAMY 2. Date of Birth : 28.06.1971 3. Designation : Assistant Professor

4. Department : Department of Biochemistry 5. Residential Address : c/o Ravisha, Nalanda convent Road, Tumkur. 6. Phone Number: Office: Mobile: 9663417864 7. Email address : [email protected] 8. Personal Homepage:

9. Academic Qualification

Degree University Year of Passing Specialization (if any)

B.Sc Kuvempu 1995 Chemistry, Botany, Zoology

M.Sc Kuvempu 1997 Biochemistry

NET CSIR- UGC 2002 Biochemistry

10. Teaching / Industrial Experience

Teaching and research activities in Biochemistry

Designation Teaching/ Research

Experience

Duration Name of the organization

Technical Assistant

Fish nutrition and extension work

14.12.1998 - 14.01.2003

Central Marine Research Institute (CMFRI), Chennai (Tamil Nadu), Kochi (Kerala), India.

Senior Technical Assistant

Micronutrient Research

15.01.2003 - 08-04-2008

National Institute of Nutrition (NIN), Hyderabad, Andhrapradesh, India.

Technical Officer

Micronutrient Research

08.04.2008 - 24.06.2010

National Institute of Nutrition (NIN), Hyderabad, Andhrapradesh, India.

Assistant Professor

Biochemistry 25.06.2010 - Till date

University College of Science , Tumkur University,Tumkur, Karnataka.

1. Standardization and estimation of ascorbic acid in diets by High

Performance 2. Liquid Chromatography (HPLC) coupled with Electrochemical Detector

and UVdetector.

3. Proximate composition analysis in food 4. Standardization and estimation of 5-methyltetraydrofolate and ascorbic

acid using Dried Blood Spot technique from finger puncture blood samples by HPLC Method.

5. Purification and characterisation of protein

Publications:

1. Pullakhandam R, Nair, MK, Kasula S, Kilari S, Thippande TG: Ferric reductase

activity of low molecular weight human milk fraction is associated with enhanced

iron solubility and uptake in Caco-2 cells. Biochem Biophys Res Commun.2008

374(2):369-72. Impact Factor : 2.72

Papers National Conferences, Seminars, Symposia:

1. T.G.Thippeswaamy: Poster presentation title on “5- Methyl tetrahydofolate in

Dried Blood Spot as an Indicator of Folate status in Humans” at 36th annual

conference of Association of Clinical Biochemists of India (ACBICON 2009),

Kochi.

2. T.G.Thippeswaamy: Oral presentation title on “Stability of Ascorbic acid in

vegetables and fruits under different conditions of temperature and storage” at

Knowledge Utsav, Jain University, Global Campus on 28th August, 2010 at

Bangalore.

3. T.G.Thippeswamy: Poster Presentation entitled “Measurement of Iodine Value in

Vegetable oils: Stability of Shark liver oil” National Level one day conference on

“Recent Trends in Food Science &Nutrition Research ” organised by the Centre

for Nano Science Research in association with Dr.P.Sadananda Maiya Centre for

Food Science and Research, Bangalore and Jain University, Bangalore on15th

December, 2011.

4. T.G.Thippeswamy: Abstract entitled “Role of dihydrofolate reductase and folate

conjugase on folic acid absorption in gestational diabetes, arthritis and

pregnancy induced hypertension: Implication on neural tube defects” National

Level one day conference on “Recent Trends in Food Science &Nutrition

Research ” organised by the Centre for Nano Science Research in association with

Dr.P.Sadananda Maiya Centre for Food Science and Research, Bangalore and Jain

University, Bangalore on15th December, 2011.

5. T.G.Thippeswamy and D.Manjunatha: Abstract submitted entitled

“Nanobiosensors for Detection of Agricultural and food contaminants “at the

Conference on “Social Relevance of Nano Materials and Applications An

Interdisciplinary Approach” organised by the Centre for Nano Science Research

in association with Higher Education Department , Government of Karnataka,

Bangalore on31st December, 2011

6. D.Manjunatha and T.G.Thippeswamy: Abstract submitted entitled “Nanosensors: A

Technology for Biomedical Applications” at the conference on “Social Relevance of

Nano Materials and Applications an Interdisciplinary Approach” organised by the

Centre for Nano Science Research in association with Higher Education Department,

Government of Karnataka, Bangalore on31st December, 2011.

7. T.G.Thippeswamy, P.Raghu, K. Sreenivasalu and K.Madhavan Nair: Abstract

submitted entitled “Purification of iron binding low molecular weight human milk

factor” at the conference on “ Perceptive on health benefits of therapeutic

molecules” organized by Centre for Bioscience and Innovation, Tumkur University

and Karnataka state higher education council, Bangalore on 06-01-2012

8. Sowmyashree, Bhavana, T.G.Thippeswamy, M.Bhagyalaksmi and Dr.S.Devaraja:

Abstract submitted entitled “Anticoagulant properties of endocarp extract of

swietenia Macrophylla and Thespesia populnea sap” at the conference on

“Perceptive on health benefits of therapeutic molecules” organized by Centre for

Bioscience and Innovation, Tumkur University and Karnataka state higher education

council, Bangalore on 06-01-2012

Conferences, Seminars, Symposia, Workshops participated:

International events: 1. Attended International seminar on “Bangalore India Bio-2011” at Bangalore

International Exhibition Centre on May, 4-6,2011

National events: 1. Attended National Level Seminar on “Reforms in Higher Education and Public

Participation” organised by Karnataka State Higher Education Council,

Government of Karnataka, June 18th, 2011

2. Attended 5th National Conference of Ancient Sciences and Archaeology organised

by Post Graduate Department of Studies and Research in History & Archaeology,

Centre for public History and Archaeology , Tumkur University and Ancient

Sciences and Archaeological Society of India on 22- 23, July,2011.

3. Attended two days conference on “Biodiversity and Sustainable development” at

Siddaganga College of Arts, Science and Commerce on 20-21, August, 2011.

4. Attended National Level one day workshop on “Biotechnology for Health and

Environment in Future” organised by the Department of Biotechnology on

Saturday, 22 October, 2011 at Shridevi Institute of Engineering & Technology, Sira

Road, Tumkur- 572106.

5. Attended 28 days “Orientation Course” Batch No.102 conducted by UGC-

Academic Staff College ,University of Madras, Chennai- 600 005 from 02-09-2011

to 29-09-2011.

6. Participated in three days National conference on “VEDANTA: A HOLISTIC

APPROACH” at Tumkur University in association with Vedanta Bharathi,

K.R.Nagara,Mysore from 26th ,27th and 28th December,.2011.

7. Participated three days workshop on “Training on Research process and Writing a

scientific paper” sponsored by Indian Council of Medical Research(ICMR) , New

Delhi and Rajiv Gandhi University of Health Sciences(RDUHS) , Bangalore held

during 11th to 13th February 2012 at Sree Siddaganga College of Pharmacy,

B.H.Road, Tumkur-572102, Karnataka.

Regional events:

1. Attended State level one day workshop on “Recent scenario on biogas from food

wastes” organized by Department of Biotechnology , ShriShridevi institute of

engineering and technology ,10th march 2011,Tumkur-02

2. Attended 2 days work shop on “Handling of analytical instruments – HPLC,

GC, AAS, and UV-Spec & PCR” at Azyme Biosciences Private Limited,

Bangalore on 28, 29 November, 2010.

3. Attended “Inter university subject based conference on Bioscience and

Bioinformatics” on 23-24 May, 2011 at Tumkur University, Tumkur.

4. Attended State Level Conference on “Frontiers of Nanotechnology” organised

by Karnataka State Higher Education Council, Government of Karnataka, 31st

May, 2011

5. Attended “Inter university subject based conference on Chemistry and

Biochemistry” on 10-11June, 2011 at Central College, Bangalore University,

Bangalore.

6. Attended Seminar on “GM Crops –Future Perspectives” organised by Tumkur

University 16th July, 2011

7. Attended 3 days “Faculty development program” conducted at Jnana Sahydri,

Kuvempu University, Shankaragatta, Shimoga from 03-03-2011 to 05-03-2011 in

association with VGST and Colligate Education, Govt. of Karnataka.

8. Attended workshop on “Multi disciplinary project proposals” on 09 June, 2011 at

Tumkur University, Tumkur.

Events Organized (conferences, seminars, symposia, workshops, etc)

1. Work shop conducted on “Biochemical techniques” at Department

of PG Studies & Research in Biochemistry, Tumkur University, Tumkur.

2. Local organising committee member for one day national seminar

on “Recent Trends in Food Science &Nutrition Research ” organised by the

Centre for Nano Science Research in association with Dr.P.Sadananda Maiya

Centre for Food Science and Research, Bangalore and Jain University, Bangalore

on15th December, 2011.

3. Local organizing committee member for one day national conference on

“Perceptive on health benefits of therapeutic molecules” organized by Centre

for Bioscience and Innovation, Tumkur University and Karnataka state higher

education council, Bangalore on 06-01-2012

Membership of Professional Associations, academic bodies, etc.

Life member ship: Nutrition Society of India (NSI), National Institute of Nutrition, Hyderabad.

Bhagyalakshmi.M biodata

1. Name of the applicant : BHAGYALAKSHMI.M

2. Mailing address : Assistant Professor,

Department of Studies in Biochemistry

Tumkur University,Tumkur,Karnataka,

Mobile No: 9481343477

Email: [email protected]

3. Date of Birth : 16-08-1981

Academic Qualification:

Degree University Year of Passing Specialization

B.Sc Kuvempu University 2004 Biochemistry,

Microbiology, Botany

Teaching: 2006-2010- Worked as a Lecturer in the Department of Biochemistry, Kuvempu University. Projects Guided: More than 15 M.Sc students for their project work in various topics of Biochemistry. National Conferences, Seminars, Symposia:

1. Bhagyalakshmi. M and Chinmayee(2010):“Evaluation of tioxidantproperties of E. officinalis, T. chebula and T. bellirica berries”. Knowledge Utsav” National Conference. 28th august,2010. Conferences, Seminars, Symposia, Workshops participated: International events:

1. Bangalore INDIA BIO-2011, at Bangalore International Exhibition Centre(BIEC)

from 04 to 06 May 2011 organized by the Department of Information Technology,

Biotechnology and Science & Technology, Government of Karnataka, Vision Group

on Biotechnology.

National events:

1. Three days national conference on “VEDANTA; A holistic approach” organized

by Tumkur university and Vedanta Bharathi, on 26 to 28 dec 2011.

2. UGC sponsored national level conference on “Biodiversity and sustainable

development ”organized by Sree Siddaganga college on 20 to 21 Aug 2011

3. One day national seminar on “ Reforms in Higher Eduction and Public

participation” organized by Karnataka state higher education council ,on 18

June 2011

4. International symposium on” challenges in drug discovery program -2011

(ISCDDP)”, February 16, 17 in Mysore.

5. State level one day workshop on “Recent scenario on biogas from food

wastes” organized by Department of Biotechnology ,SriShridevi institute of

engineering and technology ,10th march 2011,Tumkur-02

6. “Knowledge Utsav” National Conference organized by Jain University and

Tumkur University, Global Campus, Kanakapura Taluk, Bangalore Rural,

Karnataka. On 28th august 2010.

7. National seminar on “Recent advances and challenges in Biochemistry and

Biotechnology” organized by the Department of Biochemistry, Jnansahyadri,

Kuvempu university, shankaraghatta, Shimoga. On 01, 02 April, 2009

M.Sc Kuvempu University 2006 Biochemistry

8. National seminar on “ Emerging areas in Biomedical Sciences” organized by

Department of Biochemistry, P.G Centre, Shivagangotri, Davangere, on 29 march

2008.

9. National seminar on “ Advances in Biochemistry, Biotechnology and

Nanotechnology Science for 21st Century” organized by Department of

Biochemistry, P.G Centre, Shivagangotri, Davangere, on Nov. 13, 14, 2006.

10. National seminar on “Trends and Technologies in the Biochemical sciences”

organized by Department of Biochemistry, P.G Centre, Shivagangotri,

Davangere, on 10, 11 Feb. 2006

Regional events:

1. One day workshop on formulation vision documentation of strategic road map of

Tumkur university on 14-7-2011

2. One day workshop on formulation of interdisciplinary projects organized by

Tumkur university on 09-07-2011

3. Second State level Science and Technology conference 2011, organized by

Karnataka state council for Science and Technology, Vision Group for Science

and Technology and Department of Science & technology, Govt. of Karnataka, on

26th to 28th may 2011.

4. One day workshop on current scenario on production of Biogas from food wastes

organized by Shridevi Institute of Engineering and Technology, Tumkur-02,on

10-003-2011.

5. Workshop on ‘‘Recent scenario in SCAR’’ (stem cells, cancer, AIDS and Raman

spectroscopy) organized by Department of Biotechnology, Shridevi institute of

engineering and technology, Tumkur-02 on 29-09-2010.

proforma – i proforma for submission of project...

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