products and drug discovery services - ubiquigent

DUBprofiler™ Compound Screening Service

Ubiquigent can help you with your DUB (deu-biquitylase) drug discovery programmes by determining the selectivity and potency of your inhibitors. We offer the DUBprofiler service in ei-ther single concentration profiling or IC50 analy-sis mode.

DUBprofiler™-HT

Ubiquigent offers DUBprofiler-HT for the screen-ing of larger numbers of compound (>25) ver-sus the DUBprofiler panel. In this service we can offer high precision and accuracy ultra-low volume compound dispensing (down to 2.5 nl) to enable addition of the assay reagents directly to the compound in 100% DMSO. This is particularly suited to screens where com-pound solubility is a concern.

Ubiquigent Ltd provides high quality reagents, kits, and drug discovery services to the research community to help investigators unravel the complexity of, and pursue new thera-peutic target opportunities across the ubiquitin cascade.

The Company is located adjacent to the Medical Research Council Protein Phos-phorylation and Ubiquitylation Unit (PPU), based at the University of Dundee, Scotland, UK. This provides Ubiquigent with access to state-of-the-art services and capabilities within both the PPU and other departments of the College of Life Sciences. The PPU is one of Ubiquigent’s founding partners.

Products and Drug Discovery Services October 2014

Web-interactive PDF

Ubiquigent Ltd - Linking Ubiquitin Research to Drug Discovery www.ubiquigent.com

INSIDEServices ..............................................................1

Cell Lines and Kits .............................................3

Assay Kits - HTS, USP14, E2scan, DUBscan...........4

E1 Activating Enzymes ......................................6

E2 Conjugating Enzymes ...................................6

E3 Ligases ........................................................10

Deconjugating Enzymes ..................................11

Proteasome ......................................................12

Ubiquitin/Ubl Modifiers, Derivatives and Substrates .............................12

Ubiquitin/Ubl Binding Proteins .......................13

Substrates ........................................................13

Kinases .............................................................14

Antibodies ........................................................15

Cell Lysates ......................................................16

Please visit us at www.ubiquigent.com

for the latest information on new products and services.

Ubiquigent offers expert custom services to researchers interested in ubiquitin/ubiquitin-like protein (Ubl) cascade drug discovery. We have access to a suite of automation robotics to help us deliver the highest quality services to you spe-cifically designed to help accelerate your drug discovery efforts in this field.

WHAT YOU DO:

• Send us your compound(s).

• Request single concentration profiling and/ or IC50 analysis (electronic submission).

WHAT WE DO:

• Determine the selectivity and specificity of your deubiquitylase (DUB) inhibitor lead compound(s).

• Electronically report fully quality con-trolled data to you (single concentration % activities or IC50s).

SERVICE HIGHLIGHTS:

• 38 different DUBs (42 assays) on the panel and expanding.

• Any scale and combination of screening available; compound(s) vs DUB(s).

• 2 week service cycle.

• StoragePod® system for DUBprofilerTM compound management.

NEW HTS Compatible Assay Kits

HTS compatible assay kits have been devel-oped to therapeutically relevant ubiquitin sys-tem targets including: Inducer of LDL (Low Density Lipoprotein) Receptor (IDOL)-cataly-sed LDLR (LDL Receptor) substrate ubiquity-lation and MuRF1 assays.

HTS Compatible Custom Assay Development

As pathways and functions are unraveled, new and attractive ubiquitin/Ubl cascade drug discovery targets are coming to light with rel-evance across many key therapeutic areas. With access to a wide range of reagents and benefiting from our experience working with these pathways, Ubiquigent’s Custom Assay Development team supports pharmaceutical and biotechnology companies in translating these discoveries into lead programmes.

We will work closely and collaboratively with you throughout our partnership. Our approach is highly flexible; from providing high quality activity validated reagents to full-scale proj-ect outsourcing developing required screen-ing and deconvolution assays and supplying the reagents to enable you to conduct your subsequent HTS campaign in-house.

Ubiquigent have successfully developed and delivered HTS-compatible assays and kits enabling leading pharmaceutical compa-nies to conduct screening campaigns against ubiquitin system targets (DUBs, E3s and oth-ers) of high therapeutic value. Furthermore Ubiquigent has developed and supplied many deconvolution and specificity profiling as-says to help characterize the hits from these screens. A number of our assays have been developed from first principles starting with little or no data from the literature to guide the construction of the assay. Ubiquigent can re-search, design, manage and deliver the entire programme to help you to achieve success in your ubiquitin system HTS campaigns.

Services / HTS Assay Development

JUMP TO WEBSITE JUMP TO PAGEUbiquitin (synthetic)* 60-0200-050 50 µg £67 €80

Ubiquitin (pSer57) 60-0208-050 50 µg £150 €180


Biotin-Ahx-Ubiquitin (synthetic)* 60-0201-050 50 µg £83 €100

Biotin-Ahx-Ubiquitin (pThr7) 60-0203-050 50 µg £183 €220

Biotin-Ahx-Ubiquitin (pThr12) 60-0204-050 50 µg £183 €220

Biotin-Ahx-Ubiquitin (pSer57) 60-0205-050 50 µg £183 €220

Biotin-Ahx-Ubiquitin (pTyr59) 60-0206-050 50 µg £183 €220


Description Cat. # Size Price Description Cat. # Size Price

Pre-Press PRODUCT LIST Addendum April 2015

NEW Parkin-Related Phospho-Ubiquitins



Ubiquitin (synthetic)* 60-0200-050 50 µg £67 €80

Biotin-Ahx-Ubiquitin (synthetic)* 60-0201-050 50 µg £83 €100

Antibodies

Parkin (human; full length), pAb* 68-0018-100 100 µg £215 €259

Parkin pSer65 (human; residues 60 - 68-0056-100 100 µg £215 €259

PINK1 (human; residues 175 – 250), pAb* 68-0019-100 100 µg £215 €259

PINK1 pThr257 (human; residues 250 68-0057-100 100 µg £215 €259

Recombinant Proteins

Parkin 63-0048-025 25 µg £295 €356

PINK1 66-0043-050 50 µg £295 €356

PINK1 (D359A) 66-0044-050 50 µg £295 €356

USP30 CD(57-517) 64-0057-050 50 µg £285 €345

NEWPublications

click to download

pUbiquitins PINK1/Parkin

*All phospho-ubiquitins have been synthetically generated so it is recommended to use these control reagents in experiments as appropriate.

Phosphorlyated ubiquitin and derivatives are offered by Ubiquigent under an exclusive license from

UbiQ Bio BV, Amsterdam, the Netherlands.

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http://ubiquigent.com/services/overview

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http://ubiquigent.com/products/kits

http://ubiquigent.com/products/e1_activating_enzymes

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http://ubiquigent.com/products/e3_ligases

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http://ubiquigent.com/products/ubiquitin_ubl_modifiers_derivatives_and_substrates


http://ubiquigent.com/products/ubiquitin_binding_proteins

http://ubiquigent.com/products/substrates

http://ubiquigent.com/products/kinases

http://ubiquigent.com/products/antibodies

http://ubiquigent.com/products/lysates



http://ubiquigent.com/images/uploads/Files/60-0200-050_30359_v1-0-0.pdf









http://ubiquigent.com/images/uploads/Files/UBQ_pUbiquitins_Flyer_FINAL.pdf

http://ubiquigent.com/images/uploads/Files/UBQ_PINK1_Parkin_Flyer_FINAL.pdf





http://ubiquigent.com/products/antibody/parkin-human-full-length-pab-100

http://ubiquigent.com/products/antibody/parkin-pser65-human-residues-60-72-pab

http://ubiquigent.com/products/antibody/pink1-human-residues-175-250-pab-100

http://ubiquigent.com/products/antibody/pink1-pthr257-human-residues-250-262-pab

http://ubiquigent.com/products/e3_ligases/parkin-untagged

http://ubiquigent.com/products/kinases/pink1-mbp-tagged

http://ubiquigent.com/products/kinases/pink1-d359a-mbp-tagged

http://ubiquigent.com/products/deconjugating_enzymes/usp30_cd57_517_untagged

http://ubiquigent.com/images/uploads/Files/UBQ_pUbiquitins_Flyer_FINAL.pdf

http://ubiquigent.com/images/uploads/Files/UBQ_PINK1_Parkin_Flyer_FINAL.pdf


DUBprofiler™ Compound Screening Service

Ubiquigent can help you with your DUB (deu-biquitylase) drug discovery programmes by determining the selectivity and potency of your inhibitors. We offer the DUBprofiler service in ei-ther single concentration profiling or IC50 analy-sis mode.

DUBprofiler™-HT

Ubiquigent offers DUBprofiler-HT for the screen-ing of larger numbers of compound (>25) ver-sus the DUBprofiler panel. In this service we can offer high precision and accuracy ultra-low volume compound dispensing (down to 2.5 nl) to enable addition of the assay reagents directly to the compound in 100% DMSO. This is particularly suited to screens where com-pound solubility is a concern.

Ubiquigent Ltd provides high quality reagents, kits, and drug discovery services to the research community to help investigators unravel the complexity of, and pursue new thera-peutic target opportunities across the ubiquitin cascade.

The Company is located adjacent to the Medical Research Council Protein Phos-phorylation and Ubiquitylation Unit (PPU), based at the University of Dundee, Scotland, UK. This provides Ubiquigent with access to state-of-the-art services and capabilities within both the PPU and other departments of the College of Life Sciences. The PPU is one of Ubiquigent’s founding partners.

Products and Drug Discovery Services October 2014

Web-interactive PDF

Ubiquigent Ltd - Linking Ubiquitin Research to Drug Discovery www.ubiquigent.com

INSIDEServices ..............................................................1

Cell Lines and Kits .............................................3

Assay Kits - HTS, USP14, E2scan, DUBscan...........4

E1 Activating Enzymes ......................................6

E2 Conjugating Enzymes ...................................6

E3 Ligases ........................................................10

Deconjugating Enzymes ..................................11

Proteasome ......................................................12

Ubiquitin/Ubl Modifiers, Derivatives and Substrates .............................12

Ubiquitin/Ubl Binding Proteins .......................13

Substrates ........................................................13

Kinases .............................................................14

Antibodies ........................................................15

Cell Lysates ......................................................16

Please visit us at www.ubiquigent.com

for the latest information on new products and services.

Ubiquigent offers expert custom services to researchers interested in ubiquitin/ubiquitin-like protein (Ubl) cascade drug discovery. We have access to a suite of automation robotics to help us deliver the highest quality services to you spe-cifically designed to help accelerate your drug discovery efforts in this field.

WHAT YOU DO:

• Send us your compound(s).

• Request single concentration profiling and/ or IC50 analysis (electronic submission).

WHAT WE DO:

• Determine the selectivity and specificity of your deubiquitylase (DUB) inhibitor lead compound(s).

• Electronically report fully quality con-trolled data to you (single concentration % activities or IC50s).

SERVICE HIGHLIGHTS:

• 38 different DUBs (42 assays) on the panel and expanding.

• Any scale and combination of screening available; compound(s) vs DUB(s).

• 2 week service cycle.

• StoragePod® system for DUBprofilerTM compound management.

NEW HTS Compatible Assay Kits

HTS compatible assay kits have been devel-oped to therapeutically relevant ubiquitin sys-tem targets including: Inducer of LDL (Low Density Lipoprotein) Receptor (IDOL)-cataly-sed LDLR (LDL Receptor) substrate ubiquity-lation and MuRF1 assays.

HTS Compatible Custom Assay Development

As pathways and functions are unraveled, new and attractive ubiquitin/Ubl cascade drug discovery targets are coming to light with rel-evance across many key therapeutic areas. With access to a wide range of reagents and benefiting from our experience working with these pathways, Ubiquigent’s Custom Assay Development team supports pharmaceutical and biotechnology companies in translating these discoveries into lead programmes.

We will work closely and collaboratively with you throughout our partnership. Our approach is highly flexible; from providing high quality activity validated reagents to full-scale proj-ect outsourcing developing required screen-ing and deconvolution assays and supplying the reagents to enable you to conduct your subsequent HTS campaign in-house.

Ubiquigent have successfully developed and delivered HTS-compatible assays and kits enabling leading pharmaceutical compa-nies to conduct screening campaigns against ubiquitin system targets (DUBs, E3s and oth-ers) of high therapeutic value. Furthermore Ubiquigent has developed and supplied many deconvolution and specificity profiling as-says to help characterize the hits from these screens. A number of our assays have been developed from first principles starting with little or no data from the literature to guide the construction of the assay. Ubiquigent can re-search, design, manage and deliver the entire programme to help you to achieve success in your ubiquitin system HTS campaigns.

Services / HTS Assay Development

JUMP TO WEBSITE JUMP TO PAGE



















2 Ubiquigent Ltd - Linking Ubiquitin Research to Drug Discovery www.ubiquigent.com

Protein Ubiquitylation:

The Ubiquitin Cascade

DCE: deconjugating enzyme Ub: ubiquitin Ubl: ubiquitin-like protein

Product Categories are web-interactive Kinases

DCE E1 E2 E3

DCE

UBP

Ub/Ubl Anti-Ub-conjugate

Anti-Ub-conjugate

A growing number of excellent reviews are being published, which offer well-referenced summaries of work in particular disease areas, pathways or amongst the members of certain ubiquitin system protein families. For researchers considering the ubiquitin system as a potential new route to investigating

disease mechanisms, and possibly a new source of drug dis-covery targets to consider, we present a guide and a resource to enable you to begin identifying and grouping such associations.

Read more: ubiquigent.com/resources/ubiquitin-and-therapeutic-areas

Ubiquitylation, like phosphoryla-tion, describes a reversible post-translational protein modification. Ubiquitylation or ‘ubiquitination’ may control the protein substrate’s destiny – in respect of its turnover – or its signalling functionality. It is a process that refers to the co-valent attachment - through it’s C-terminus - of a small, 76 amino acid protein called ubiquitin to the epsilon-amino group of a lysine residue residing within a substrate protein – which may also be anoth-er ubiquitin molecule. This results in either mono- or poly-ubiquity-lation of the substrate; the latter being where chains of ubiquitin are attached to the substrate protein. Ubiquitin may also form...

Read more: www.ubiquigent.com/products

Therapeutic Areas and the Ubiquitin System

http://ubiquigent.com/resources/ubiquitin-and-therapeutic-areas/











http://ubiquigent.com/resources/ubiquitin-and-therapeutic-areas/

www.ubiquigent.com/products


Ubiselect™-MEF_Bio-Ub (P0) Cell Line NEW mouse embryonic fibroblasts 66-5011-001 2.5x106 cells £745

Ubiselect™-MEF_BirA (P0) Cell Line NEW mouse embryonic fibroblasts 66-5010-001 2.5x106 cells £385

Proteaselect™-HEK293_Bio-Rpn11 Cell Line NEW embryonic kidney 66-5012-001 2-4 x106 cells £745

Description Source Cat. # Size Price

Cell Lines and KitsThrough close collaborations with leading academic laboratories Ubiquigent offers specialist cell lines to enable and support ubiquitin system re-search. Each cell line has been fully application validated.

Applications: The presence of N-terminally biotinylated ubiquitin in the Ubiselect -MEF_Bio-Ub cell line enables rapid and high affinity capture of ubiquitylated proteins from mouse embryonic fibroblasts. The system facilitates the capture, separation and analysis of ubiquitylated proteins from ubiquitylated protein binding proteins and ubiquitin chain binding proteins - not usually feasible using conventional Ubiquitin Binding Domain-based systems such as Tandem Ubiquitin Binding Entities (TUBEs). See below for more information.

The Proteaselect cell line enables rapid purification of complete proteasome complexes from HEK293 cells for analysis, direct assay or for USP14 (deubiquitylase; DUB) activation. See USP14 kits and USP14 DUB products.

Refer to the Cell Lines and Application Notes sections of the website for detailed information.

The Ubiselect Kit is designed to facilitate the isolation and identification of your ubiquitylated proteins of interest from a transgenic mouse-derived embryonic fibroblast (MEF) cell line expressing ubiquitin tagged with an N-terminal biotinylation signal (which is biotinylated by co-expression of the BirA enzyme). By employing a powerful capture system which utilises the transgenic MEF cell line expressing ubiquitin - which is biotinylated in the cell - and the high affinity biotin-avidin interaction alongside a high stringency washing protocol you can capture your ubiquitylated target(s) of interest without any associated ubiquitin binding proteins and/or ubiq-uitylated protein binding proteins. The Ubiselect Kit contains an aliquot of each of the control Ubiselect-MEF_BirA (P0) and the Ubiselect-MEF_Bio-Ub (P0) cell lines and an anti-ubiquitin conjugate antibody for the detection of ubiquitylated proteins. Eluted captured products may be analysed by Western blotting using the ubiquitin conjugate specific antibody, an anti-biotin antibody or an antibody specific to your protein of interest.

Ubiselect™ Kit Cat. # 67-0018-001 1 Kit £995

Ubiselect™ Kit NEW

What does the Ubiselect Kit contain?

The kit contains control Ubiselect-MEF_BirA (P0) cell line (Cat# 66-5010-001), Ubiselect-MEF_Bio-Ub (P0) cell line (Cat# 66-5011-001), and an anti-Mono and Polyubiquitylated conjugates antibody (Cat# 68-0122-025) for detection of your ubiquitylated protein. Also included in the kit is a protocol insert which pro-vides you with culture conditions and a detailed capture protocol for the isolation of ubiquitylated proteins from the cell lines.

Ubiselect Kit experiments facilitate:

Selective isolation of ubiquitylated proteins from Ubi• select-MEF_Bio-Ub cells through the removal of ubiquitin binding proteins and ubiquitylated protein binding proteins that inter-act with your captured ubiquitylated protein of interest.

The manipulation of Ubi• select-MEF_Bio-Ub cells as per ex-isting MEF cell protocols. Identification of mono or poly-ubiquitin attached to your protein of interest.

Exploration of the formation of E1, E2, E3 and substrate •ubiquitin-conjugates.

Developing cell or cell-lysate based target specific ubiquity-•lation assays (eg FRET) utilising the biotin on the ubiquitin attached to your protein of interest.

Isolation of candidate cysteine-ubiquitylated proteins and •determination of their identity using mass spectrometry.

Transgenes in transgenic mouse from which Ubiselect-MEF_BirA (P0) and Ubiselect-MEF_Bio-Ub (P0) have been derived

Figure 1a: Ubiselect™-MEF_Bio-Ub (P0): This is a hemizygous cell line which expresses three moieties of biotin-accepting ubiquitin plus the E. coli enzyme BirA. (Lectez et al., 2014)

Figure 1b: Ubiselect™-MEF_BirA (P0): This is a hemizygous cell line which expresses the E. coli enzyme BirA. (Lectez et al., 2014)

Key Utility and Applications:

High affinity and selective capture of ubiquitylated proteins from •Mouse Embryonic Fibroblast (MEF) cells

High sensitivity detection of ubiquitylated proteins only and NOT other •interacting proteins

Cell-lysate based target specific ubiquitylation assays•

Reference:Lectez B, Migotti R, Lee SY, Ramirez J, Beraza N et al. (2014) Journal of Proteome Research 13, 3016-3026.



Kits

Many enzymes from across the ubiquitin system have been identified as potential drug discovery targets; see our Therapeutic Area pages for more information. With reference to these therapeutic area opportunities, Ubiquigent high-throughput screening (HTS) compatible assay kits are developed and optimised for your convenience and are validated specifically for HTS applications. Everything that you require is provided in the kit: enzymes, substrates, buffers, protocols and access to ongoing support. Kits are available in 5,000 and 30,000 data point sizes, larger kits (for 1 million+ data points have been and can be supplied too). We also offer customisation to fit your precise needs: number of data points required, budget, assay design, configuration and logistics or specialist therapeutic area requirements.

Mammalian proteasomes are associated with three DUBs: USP14, UCHL5 (UCH37) and RPN11 (POH1). UCHL5 and USP14 reside on the regulatory particle and remove ubiquitin from the substrate before substrate degradation whereas RPN11’s activity is delayed until the proteasome is committed to degrading the substrate. The DUB activity of USP14 is known to be activated by proteasomes. Ubiquigent offers two USP14 kits: The USP14 Activa-tion kit allows you to deter mine the optimal ratio of USP14:26S Proteasome [Ub-VS treated] appropri ate for your assay. The Activated USP14 DUB assay Kit comprises Ubiquigent’s pre-formu lated product USP14 & 26S Protea some [Ubiquitin-Vinyl Sulfone (Ub-VS) treated] (Cat# 64-1010-096) at an op timised molar ratio ready for use in a USP14 deubiquitylase assay.

E2 conjugating enzyme assay kits 70-2000 Custom p. o. a.

IDOL (E3 ligase)-catalysed LDLR substrate ubiquitylation assay kit 70-1010 5,000 and 30,000 data points p. o. a.

IDOL (E3 ligase)-catalysed MRLC substrate ubiquitylation assay kit 70-1020 5,000 and 30,000 data points p. o. a.

IDOL (E3 ligase)-catalysed auto-ubiquitylation assay kit 70-1030 5,000 and 30,000 data points p. o. a.

MuRF1 (E3 ligase)-catalysed MyLC1 substrate ubiquitylation assay kit 70-1050 5,000 and 30,000 data points p. o. a.

MuRF1 (E3 ligase)-catalysed MyLC2 substrate ubiquitylation assay kit 70-1060 5,000 and 30,000 data points p. o. a.

MuRF1 (E3 ligase)-catalysed auto-ubiquitylation assay kit 70-1070 5,000 and 30,000 data points p. o. a.

CHIP (E3 ligase)-catalysed auto-ubiquitylation assay kit 70-1040 5,000 and 30,000 data points p. o. a.

E3 enzyme assay kits (excluding those listed above) 70-1500 Custom p. o. a.

DUB enzyme assay kits 70-4000 Custom p. o. a.

Activated USP14 DUB assay Kit * 67-0015-096 1 kit £225

USP14 Activation Kit * 67-0014-001 1 kit £160

5x DUB Assay Buffer 64-2001-500 500 µl £25

5x DUB Assay Stop Buffer 64-2002-500 500 µl £25

4x Ubiquitin-Rhodamine 110 60-0122-500 500 µl £50

Description Cat. # Format Price

Description Cat. # Format Price

HTS Assay Kits

USP14 Assay Kits

* Limited Use License The Biotin AviTag™ technology is covered by U.S. Pat. Nos: 5,723,584; 5,874,239; 5,932,433; and 6,265,552 and includes any and all materi-als, methods, kits and related derivatives claimed by these patents. The purchase of the UBIQUIGENT’s AviTagged products confers to the pur-chaser the limited right to use the AviTag™ technology for research use. Commercial use of the AviTag™ technology to manufacture a commer-cial product, or use of the AviTag™ technology to facilitate or advance research which will be applied to the development of a commercial prod-uct requires a license from Avidity, LLC. Examples of Commercial use include, but are not limited to biosensors, diagnostics, therapeutic ap-plications, proximity assays, and drug screening assays. Users desiring a commercial license should contact Avidity, LLC in the US @ 720-859-6111 or by email to [email protected].


The E2scan Kit is designed to facilitate the identification of E2 conjugating enzymes which support the ubiquitylation of substrate proteins catalysed by an E3 ligase and/or the auto-ubiquitylation of the E3 ligase itself.

The DUBscan kit has been flexibly designed for many potential applications. One application that might be of interest is the identification of DUB enzymes which catalyse the deubiquitylation of a substrate protein.

E2scan Kit version 2 Cat. # 67-0005-001 1 Kit £345

DUBscan Kit Cat. # 67-0006-001 1 Kit £245

E2scan™ Kit version 2 - now containing 34 E2s

DUBscan™ Kit

The kit contains an E2scan plate - a panel of 34 E2 conjugating enzymes arrayed in duplicate across a 96-well plate - plus all the components (E1 enzyme, control E3 ligase, ubiquitin, ATP, assay buffer) required to perform a complete substrate ubiquitylation or auto-ubiquitylation assay using your E3 ligase and/or E3 ligase/substrate combination of choice. Through the addition of a substrate mix (E1, E3, substrate, and ubiquitin) and then ATP to the wells of an E2scan plate, screen the E2 conjugating enzyme panel in just one hour. Reaction products may be analysed as required, for example by SDS-PAGE or Western blotting.

The kit contains a DUBscan plate (a panel of DUBs arrayed across 47 wells of a 96 well plate) plus a control ubiquitylation reaction mix for performing both positive and negative control deubiquitylation reactions.

In respect of one kit application - the identification of DUB enzymes which catal-yse the deubiquitylation of a substrate protein; through the addition of your own DUB substrate of interest to the wells of a DUBscan plate you can screen the DUB enzyme panel in just one hour. Reaction products may be analysed as required, for example by SDS-PAGE and Western blotting using specific antibodies. One kit contains all the reagents and buffers necessary to perform a ‘DUBscan’ of the panel of 47 DUB assay wells.

Why use the E2scan kit?

E2 conjugating enzymes can be critical in determining the ubiq-•uitin chain linkage type(s) generated in a ubiquitylation reaction.

The linkage type can determine the fate of the ubiquitylated •substrate; for example to be degraded or to function in a sig-nalling capacity (Ye and Rape (2009)).

Discovery of novel aspects to E2 functionality: The E2• scan kit was used to identify E2s that partner RNF4 in polySUMO ubiquitylation and it was discovered that Ube2W (Ubc16) is an E2 with - a previously unknown - specific protein amino terminal monoubiquitylation activity (Tatham et al. 2013).

E2scan experiments facilitate:

The rapid identification of the E2s that couple with your E3 •(auto and/or substrate ubiquitylation modes)

New insights regarding the E3 ligase and/or the substrate•

The development of assays for the E3 ligase•

ReferencesYe and Rape (2009) Nat Rev Mol Cell Biol 10, 755-764.Tatham MH, Plechanovová A, Jaffray EG, Salmen H, Hay RT (2013) Biochem J 453, 137-145.

Why use the DUBscan kit?

Examine the cleavage of • in vitro ubiquitylated substrates generated using different E2/E3 combinations (refer to the Ubiquigent E2scan kit for identifying E2s that ‘couple’ with your E3 of interest)

Examine the cleavage of • in vivo ubiquitylated substrates af-ter capture from a cell lysate (refer to Ubiquigent Ubiselect for capturing ubiquitylated proteins)

Investigate DUB ubiquitin-ubiquitin linkage cleavage speci-•ficity (refer to Ubiquigent di-ubiquitin and ubiquitin chains for use as substrates)

Examine the relative position of cleavage within ubiquitin •chains (distal versus proximal)

Explore mono and poly-deubiquitylation DUB specificity (re-•fer to specific Ubiquigent E2s – such as Ube2W – for mono-ubiquitylating E2s)

Explore potential DUB inhibitors and/or activators•

Identify novel DUB binding proteins•

Explore how DUBs may interact with and modify the activity of •other ubiquitin system proteins such as E2s and vice versa

1 2 3 4 5 6 7 8 9 10 11 12

A

B

C

D

E

F

G

H

2A 2B 2C 2D1 2D2 2D3 2D4 2E1 2E2 2E3 2F 2G1

2G2 2H 2I 2J1 2J2 2K 2L3 2L6 2M 2N 2N/2V1 2N/2V2

2Q 2Q2 2R1 2R2 2S

BFR

BFR

BFR 2D4

2D4

2D4

2A 2B 2C 2D1 2D2 2D3 2D4 2E1 2E2 2E3 2F 2G1

2G2 2H 2I 2J1 2J2 2K 2L3 2L6 2M 2N 2N/2V1 2N/2V2

2Q 2Q2 2R1 2R2 2S

2Z

2Z

2T

2T

2V1

2V1

2V2

2V2

2W

2W

1 2 3 4 5 6 7 8 9 10 11 12

A

B

C

D

E

F

G

H

USP1/UAF1 USP2 USP4 USP5 USP6 USP7 USP9X BFR BFR AMSH OTUB1

USP11 USP14USP14/UbVS-Ptsm

UbVS-Ptsm Ptsm USP15 USP19 USP20 USP21 USP25 BFR

USP27x USP28 USP30 USP35 USP36 USP45 CYLD UCHL1 UCHL3 UCHL5 BFR

BAP1 A20 Cezanne OTU1 OTUB1 OTUB2 OTUD3 OTUD5 OTUD6A OTUD6B BFR

OTULIN TRABID ATXN-3 ATXN-3L JOSD1 JOSD2 AMSH AMSH-LP AMSH+ Zn

AMSH-LP + Zn BFR

Kit Enzymes

Kit Controls

Buffer

Buffer

Positive Control

Negative Control

User Controls

User DUBs

Suggested empty wells to enable user to run DUBscan samples on 12-well gels

BFR

Kit Enzymes

Kit Controls

Buffer

Positive Control

BFR

AMSH

OTUB1


SCAN Kits

Ubiquigent offers two Scan kits: The DUBscan™ kit has been flexibly designed for many potential applications. One application that might be of greatest interest is the identification of DUB enzymes which support the deubiquitylation of substrate proteins. In addition, the E2scan™ Kit version 2 is designed to facilitate the identification of E2 conjugating enzymes which support the ubiquitylation of substrate proteins catalysed by an E3 ligase and/or the auto-ubiquitylation of the E3 ligase itself.

http://www.ncbi.nlm.nih.gov/pubmed/19851334

http://www.ncbi.nlm.nih.gov/pubmed/23560854

APP-BP1/UBA3 NAE1 / HPP1 NP_003896 / NP_003959.3 None 61-0006-010 10 µg £95

61-0006-050 50 µg £270

ATG7 APG7, GSA7 NP_006386 6His 61-0008-010 10 µg £95

61-0008-050 50 µg £270

SAE1/SAE2 AOS1 / UBA2 NP_005491.1 / NP_005490.1 GST/6His 61-0005-010 10 µg £95

61-0005-050 50 µg £270

UBA6 E1-L2, UBE1L2 NP_060697 6His 61-0002-010 10 µg £50

61-0002-050 50 µg £113

UBA7 UBE1L NP_003326.2 6His 61-0007-010 10 µg £95

61-0007-050 50 µg £270

UBE1 A1S9, A1S9T NP_003325 6His 61-0001-010 10 µg £50

61-0001-050 50 µg £113

Description Alternate Name Accession Number Tag Cat. # Size Price


E1 Activating EnzymesE1 activating enzymes initiate the activation and conjugation of ubiquitin and ubiquitin-like modifiers (Ubls) in a cascade of events that modifies sub-strate proteins resulting in proteasomal or lysosomal degradation, changes in intracellular localisation, activity or other signaling events. To date eight E1 enzymes have been identified. E1 enzymes utilise ATP to activate the terminal glycine residue of ubiquitin/Ubl generating a covalent thioester link-age between ‘activated’ ubiquitin/Ubl and the E1 enzyme itself. The ubiquitin/Ubl is then transferred to the sulphydryl group of the active-site cysteine on an E2 conjugating enzyme in a transthiolation reaction.

E2 Conjugating EnzymesE2 conjugating enzymes form a ubiquitin/Ubl-thioester intermediate in the ubiquitylation/Ubl pathway. There are predicted to be 38 E2 conjugating enzymes and four classes of the enzyme exist, all of which contain a catalytic (Ubc) domain with an active site cysteine. After transfer of ubiquitin/Ubl from the E1 activating enzyme to the E2 via a transthiolation reaction, the loaded E2 interacts with E3 ligases to transfer the activated ubiquitin/Ubl to the substrate lysine acceptor residue; either directly from the E2 or via a lysine reside on the E3 ligase itself.


UBE2A HR6A NP_003327 None 62-0002-020 20 µg £65

62-0002-100 100 µg £135

6His 62-0071-020 20 µg £65

62-0071-100 100 µg £135

GST 62-0001-020 20 µg £65

62-0001-100 100 µg £135

UBE2B HR6B NP_003328 None 62-0004-020 20 µg £65

62-0004-100 100 µg £135

6His 62-0072-020 20 µg £65

62-0072-100 100 µg £135

GST 62-0003-020 20 µg £65

62-0003-100 100 µg £135

UBE2C UbcH10 NP_008950 None 62-0007-020 20 µg £65

62-0007-100 100 µg £135

6His 62-0005-020 20 µg £65

62-0005-100 100 µg £135

GST 62-0006-020 20 µg £65

62-0006-100 100 µg £135

UBE2D1 UbcH5a NP_003329 None 62-0010-020 20 µg £65

62-0010-100 100 µg £135

6His 62-0008-020 20 µg £65

62-0008-100 100 µg £135

GST 62-0009-020 20 µg £65

62-0009-100 100 µg £135

E2 Conjugating Enzymes continued...



UBE2D2 UbcH5b NP_003330 None 62-0012-020 20 µg £65

62-0012-100 100 µg £135

6His 62-0073-020 20 µg £65

62-0073-100 100 µg £135

GST 62-0011-020 20 µg £65

62-0011-100 100 µg £135

UBE2D3 UbcH5c NP_003331 None 62-0014-020 20 µg £65

62-0014-100 100 µg £135

6His 62-0074-020 20 µg £65

62-0074-100 100 µg £135

GST 62-0013-020 20 µg £65

62-0013-100 100 µg £135

UBE2D4 UbcH5d NP_057067 None 62-0017-020 20 µg £65

62-0017-100 100 µg £135

6His 62-0015-020 20 µg £65

62-0015-100 100 µg £135

GST 62-0016-020 20 µg £65

62-0016-100 100 µg £135

UBE2E1 UbcH6 AAH09139 None 62-0019-020 20 µg £65

62-0019-100 100 µg £135

6His 62-0075-020 20 µg £65

62-0075-100 100 µg £135

GST 62-0018-020 20 µg £65

62-0018-100 100 µg £135

UBE2E2 UbcH8 NP_689866.1 None 62-0066-020 20 µg £65

62-0066-100 100 µg £135

6His 62-0020-020 20 µg £65

62-0020-100 100 µg £135

GST 62-0087-020 20 µg £65

62-0087-100 100 µg £135

UBE2E3 UbcH9 NP_006348 None 62-0022-020 20 µg £65

62-0022-100 100 µg £135

6His 62-0076-020 20 µg £65

62-0076-100 100 µg £135

GST 62-0021-020 20 µg £65

62-0021-100 100 µg £135

UBE2F NCE2 NP_542409 None 62-0025-020 20 µg £65

62-0025-100 100 µg £135

6His 62-0023-020 20 µg £65

62-0023-100 100 µg £135

GST 62-0024-020 20 µg £65

62-0024-100 100 µg £135

UBE2G1 Ubc7 NP_003333 None 62-0028-020 20 µg £65

62-0028-100 100 µg £135

6His 62-0026-020 20 µg £65

62-0026-100 100 µg £135

GST 62-0027-020 20 µg £65

62-0027-100 100 µg £135

UBE2G2 Ubc7 NP_003334 None 62-0030-020 20 µg £65

62-0030-100 100 µg £135

6His 62-0077-020 20 µg £65

62-0077-100 100 µg £135

GST 62-0029-020 20 µg £65

62-0029-100 100 µg £135





UBE2H UbcH2 NP_003335 None 62-0032-020 20 µg £65

62-0032-100 100 µg £135

6His 62-0078-020 20 µg £65

62-0078-100 100 µg £135

GST 62-0031-020 20 µg £65

62-0031-100 100 µg £135

UBE2I Ubc9 NP_003336 None 62-0034-020 20 µg £65

62-0034-100 100 µg £135

6His 62-0033-020 20 µg £65

62-0033-100 100 µg £135

GST 62-0065-020 20 µg £65

62-0065-100 100 µg £135

UBE2J1 NCUBE1 NP_057105.2 6His 62-0096-020 20 µg £65

62-0096-100 100 µg £135

UBE2J2 NCUBE2 NP_477515.2 None 62-0067-020 20 µg £65

62-0067-100 100 µg £135

6His 62-0035-020 20 µg £65

62-0035-100 100 µg £135

GST 62-0036-020 20 µg £65

62-0036-100 100 µg £135

UBE2K UbcH1 NP_005330 None 62-0039-020 20 µg £65

62-0039-100 100 µg £135

6His 62-0079-020 20 µg £65

62-0079-100 100 µg £135

GST 62-0038-020 20 µg £65

62-0038-100 100 µg £135

UBE2L3 UbcH7 AAH53368 None 62-0042-020 20 µg £65

62-0042-100 100 µg £135

6His 62-0040-020 20 µg £65

62-0040-100 100 µg £135

GST 62-0041-020 20 µg £65

62-0041-100 100 µg £135

UBE2L6 UbcH8 NP_004214 None 62-0044-020 20 µg £65

62-0044-100 100 µg £135

6His 62-0080-020 20 µg £65

62-0080-100 100 µg £135

GST 62-0043-020 20 µg £65

62-0043-100 100 µg £135

UBE2M [pAb available - p. 16] Ubc12 AAH58924 None 62-0068-020 20 µg £65

62-0068-100 100 µg £135

6His 62-0048-020 20 µg £65

62-0048-100 100 µg £135

GST 62-0088-020 20 µg £65

62-0088-100 100 µg £135

UBE2N Ubc13 NP_003339 None 62-0047-020 20 µg £65

62-0047-100 100 µg £135

6His 62-0045-020 20 µg £65

62-0045-100 100 µg £135

GST 62-0046-020 20 µg £65

62-0046-100 100 µg £135

UBE2Q1 NICE-5 NP_060052 6His 62-0081-020 20 µg £65

62-0081-100 100 µg £135

GST 62-0049-020 20 µg £65

62-0049-100 100 µg £135





UBE2Q2 NP_775740 None 62-0069-020 20 µg £65

62-0069-100 100 µg £135

6His 62-0051-020 20 µg £65

62-0051-100 100 µg £135

GST 62-0089-020 20 µg £65

62-0089-100 100 µg £135

UBE2R1 UbcH3 NP_004350 None 62-0054-020 20 µg £65

62-0054-100 100 µg £135

6His 62-0052-020 20 µg £65

62-0052-100 100 µg £135

GST 62-0053-020 20 µg £65

62-0053-100 100 µg £135

UBE2R2 CDC34B AAH04862 None 62-0098-020 20 µg £65

62-0098-100 100 µg £135

6His 62-0094-020 20 µg £65

62-0094-100 100 µg £135

UBE2S E2-EPF AAH65364 None 62-0056-020 20 µg £65

62-0056-100 100 µg £135

6His 62-0082-020 20 µg £65

62-0082-100 100 µg £135

GST 62-0055-020 20 µg £65

62-0055-100 100 µg £135

UBE2T [pAb available - p. 16] HSPC150 NP_054895 None 62-0070-020 20 µg £65

62-0070-100 100 µg £135

6His 62-0057-020 20 µg £65

62-0057-100 100 µg £135

GST 62-0090-020 20 µg £65

62-0090-100 100 µg £135

UBE2V1 Uev1 AAG24229 None 62-0059-020 20 µg £65

62-0059-100 100 µg £135

6His 62-0084-020 20 µg £65

62-0084-100 100 µg £135

GST 62-0102-020 20 µg £65

62-0102-100 100 µg £135

UBE2V2 Mms2 NP_003341.1 None 62-0100-020 20 µg £65

62-0100-100 100 µg £135

6His 62-0099-020 20 µg £65

62-0099-100 100 µg £135

GST 62-0101-020 20 µg £65

62-0101-100 100 µg £135

UBE2W Ubc16 NP_001001481.1 6His 62-0085-020 20 µg £65

62-0085-100 100 µg £135

GST 62-0091-020 20 µg £65

62-0091-100 100 µg £135

UBE2Z USE1 NP_075567.2 6His 62-0086-020 20 µg £65

62-0086-100 100 µg £135

GST 62-0060-020 20 µg £65

62-0060-100 100 µg £135





Cul1/Rbx1 Cullin1/HRT1 NP_003583.2/NP_055063.1 None 63-1000-025 25 µg £245

Cul1/Rbx1/Skp1 Cullin1/HRT1/p19A NP_003583.2/NP_055063.1/NP_008861.2 None 63-1001-025 25 µg £295

Cul2/Rbx1 NEW Cullin2 /HRT1 AAH09591.1/NP_055063.1 None 63-1004-025 25 µg £245

Cul3/Rbx1 [Cullin3 pAb available - p. 15] KIAA0617/HRT1 NP_003581.1/NP_055063.1 None 63-1003-025 25 µg £245

Cul5/Rnf7 [Cullin5 & RNF7 pAb’s - p.15-16] VACM/ROC2 NP_003469.2 /NP_055060.1 None 63-1002-025 25 µg £245

DCNL1 FLJ10704 NP_065691.2 6His 63-2000-025 25 µg £95

GST 63-2001-025 25 µg £95

DCNL2 C13orf17 NP_001014305.1 6His 63-2004-025 25 µg £95

GST 63-2002-025 25 µg £95

HUWE1 NEW LASU1, UREB1 NP_113584.3 GST 63-0042-025 25 µg £215

ITCH AIF4 NP_1136713 GST 63-0006-025 25 µg £150

NLEL NEW Non-LEE-encoded Ligase NP_309587.1 GST 63-0038-025 25 µg £215

SMURF1 KIAA1625 NP_065162.1 GST 63-0027-025 25 µg £225

SMURF2 NEW MGC138150 AAG45422.1 GST 63-0046-025 25 µg £215

WWP1 AIP5 NP_008944 None 63-0034-025 25 µg £222

GST 63-0033-025 25 µg £225

BIRC2 NEW API1, cIAP1, HIAP2 NP_001157 GST 63-0015-025 25 µg £215

IPAH9.8 AAP79029.1 AAP79029.1 GST 63-0018-025 25 µg £92

CBL RNF55 NP_005179.2 GST 63-0004-010 10 µg £110

63-0004-050 50 µg £285

IDOL NEW MIR, MyLIP NP_037394.2 His 63-0044-025 25 µg £245

MuRF1 NEW TRIM63, RNF28, SMRZ NP_115977.2 None 63-0047-025 25 µg £245

Parkin [pAb available - p. 16] NEW PARK2, PRKN NP_004553.2 None 63-0048-025 25 µg £295

RNF8 KIAA0646 NP_003949 GST 63-0021-025 25 µg £150

RNF11 CGI-123 NP_055187 6His 63-0023-025 25 µg £92

GST 63-0024-025 25 µg £92

TRIAD1 [pAb available - p. 16] ARIH2 NP_006312 6His 63-0029-025 25 µg £150

ZNRF1 NEW Zinc and ring finger protein 1

Q8ND25 GST 63-0040-025 25 µg £295

ZNRF2 NEW Zinc and ring finger protein 2

Q8NHG8 GST 63-0041-025 25 µg £295

CHIP STUB1 NP_005852 None 63-0003-020 20 µg £125

63-0003-100 100 µg £250

6His 63-0001-025 25 µg £92

GST 63-0002-025 25 µg £92

E3 LigasesE3 ligases confer specificity to the ubiquitylation pathway by recognising target substrates and mediating transfer of ubiquitin or ubiquitin like proteins (Ubls) from an E2 conjugating enzyme to a substrate. There are estimated to be >700 E3 ligases forming four classes of enzyme; N-end rule ubiquitin li-gases, Homology to E6AP C-Terminus (HECT) domain E3 ligases, Cullin Ring Ligases (CRLs) and Really Interesting New Gene (RING) domain E3 ligases.


CRL

DCUN/DCNL

HECT

IAP

IPAH

RING/RBR

UBOX


USP1/UAF1 NP_003359/NP_065890 6His/None 64-0040-050 50 µg £285

USP2 (isoform 4) UBP41 NP_741994 GST 64-0014-050 50 µg £125

USP4 NP_003354 6His 64-0001-050 50 µg £285

USP5 Isopeptidase T P45974 6His 64-0002-050 50 µg £125

USP6 CD(529-1406) TRE2 NP_004496 GST 64-0045-050 50 µg £285

USP7 HAUSP CAA96580 6His 64-0003-050 50 µg £195

USP8 NEW NP_005145 None 64-0053-050 50 µg £285

USP9x CD(1554-1995) DFFRX NP_001034680 GST 64-0017-050 50 µg £195

USP14 AAH03556 6His 64-0018-050 50 µg £125

64-0018-005 5 µg £50

USP14 & 26S Proteasome [Ub-VS treated] 6His/None 64-1010-096 96 assay points

£135

USP15 (isoform 2) [pAb available p. 16] Unph2 NP_006304 None 64-0026-050 50 µg £125

USP19-TM (1-1290) ZMYND9 NP_006668 GST 64-0022-050 50 µg £285

USP20 LSFR3A AAH39593 GST 64-0039-050 50 µg £285

USP21 CD(196-565) USP23 NP_001014443 GST 64-0037-050 50 µg £285

USP25 NP_037528 GST 64-0023-050 50 µg £125

USP27x USP22L NP_001138545 DAC 64-0046-050 50 µg £285

USP28 KIAA1515 NP_065937 GST 64-0020-050 50 µg £285

USP30 CD(57-517) NEW NP_116052 None 64-0057-050 50 µg £285

USP36 CD(81-461) KIAA1453 AAH27992.1 GST 64-0006-050 50 µg £285

USP45 MGC14793 NP_001073950 GST 64-0025-050 50 µg £285

CYLD [pAb available - p. 15] KIAA0849 NP_056062 6His 64-0010-050 50 µg £125

Deconjugating EnzymesDeconjugating enzymes (DCE) comprise a large group of hydrolases and isopeptidases that regulate the ubiquitin/ubiquitin-like protein (Ubl) pathways by cleaving ubiquitin/Ubl-ubiquitin/Ubl, or ubiquitin/Ubl-substrate bonds. There are >70 DCEs – termed deubiquitinases or deubiquitylases (DUBs) – encoded by the human genome that deconjugate ubiquitin-ubiquitin chains or ubiquitin-substrates. These DUBs may be divided into five subfamilies: ubiquitin-specific proteases (USPs), ovarian tumour proteases (OTUs), ubiquitin C-terminal hydrolases (UCHs), Josephins, and JAB1/MPN/MOV34 (JAMMs).

USP

A20 CD(1-366) TNFAIP3 NP_006281 GST 64-0047-050 50 µg £125

OTU1 YOD1 NP_061036 GST 64-0036-050 50 µg £285

OTUB1 [pAb available - p. 16] FLJ20113 NP_060140 GST 64-0011-050 50 µg £125

OTUB2 OTU2, OTB2 NP_075601.1 GST 64-0012-050 50 µg £125

OTUD1 CD(270-481) NEW DUBA-7 NP_001138845 6His 64-0054-050 50 µg £285

OTUD3 KIAA0459 NP_056022 GST 64-0035-050 50 µg £285

OTUD5 (p177S) DUBA AAH09917 GST 64-0043-050 50 µg £285

OTUD6A DUBA-2 NP_997203 6His 64-0038-050 50 µg £285

OTUD6B DUBA-5 NP_057107 GST 64-0028-050 50 µg £285

OTULIN [pAb available - p. 16] FAM105B NP_612357 GST 64-0048-050 50 µg £125

TRABID CD(245-697) ZRANB1 NP_060050 6His 64-0049-050 50 µg £125

VCPIP CD(25-561) NEW VCPIP135 NP_079330 GST 64-0055-050 50 µg £285

OTU

UCHL1 PARK5 AAH00332.1 6His 64-0007-050 50 µg £125

UCHL3 AAH18125 GST 64-0027-050 50 µg £125

UCHL5 UCH37 AAH15521.1 GST 64-0008-050 50 µg £195

UCH



Deconjugating Enzymes continued...


Ataxin-3 AAH33711 GST 64-0033-050 50 µg £125

Ataxin-3L NP_001129467 6His 64-0034-050 50 µg £195

JOSD1 KIAA0063 NP_055691 6His 64-0031-050 50 µg £195

JOSD2 SBBI54 NP_612207 6His 64-0032-050 50 µg £195

AMSH CD(252-424) STAMBP AAD05037 GST 64-0042-050 50 µg £125

AMSH-LP CD(264-436) NP_065850 GST 64-0029-050 50 µg £195

Ubiquitin RPS27A P62990.1 None 60-0001-010 10 mg £50

Di-Ubiquitin, K6-Linked None 60-0101-010 10 µg £105






None 60-0106-050 50 µg £125


None 60-0107-050 50 µg £125

Di-Ubiquitin, Linear GST-cleaved 60-0115-010 10 µg £65

GST-cleaved 60-0115-050 50 µg £160

Poly-Ubiquitin, Ub n2-7; K63-linked None 60-0109-100 100 µg £75

Josephin

JAMM

Ubiquitin, Ubiquitin Chains

Ubiquitin/Ubl Modifiers, Derivatives and SubstratesAlthough ubiquitin is the most well studied and understood functional protein post-translational modifier, there is a growing family of ubiquitin-like proteins (Ubls; Modifiers) that may also modify cellular targets to influence diverse biological processes. Members of the ubiquitin and Ubl protein family include ubiquitin, SUMO, NEDD8, ISG15, URM1, FAT10, UFM1, and ATG12. These related proteins are structurally similar, and are activated, conjugated, and released from conjugates in a mechanism akin to that for ubiquitin. There is cross-talk between conjugation pathways with some substrate proteins becoming targeted by more than one type of modifier.




ProteasomeThe ubiquitin–proteasome system (UPS) targets selected proteins for degradation by the 26S proteasome. The initial steps in this pathway generate proteins that are covalently tagged with a polyubiquitin chain that is then recognized by ubiquitin receptors of the 26S proteasome. This is a large complex composed of a 20S catalytic core particle and two 19S regulatory particles that catalyse the final step in the pathway. While the 20S particle is composed of a catalytic chamber for protein degradation, collectively the proteins that comprise the 19S particle perform several proteasomal functions that include recognition of ubiquitylated substrates, cleavage of the polyubiquitin chain for ubiquitin recycling, control of access to the 20S proteolytic chamber, and substrate unfolding and subsequent translocation into the 20S core particle for degradation. Mammalian proteasomes are associated with three DUBs: USP14, UCHL5 (UCH37) and RPN11 (POH1). UCHL5 and USP14 reside on the regulatory particle and remove ubiquitin from the substrate before substrate degradation whereas RPN11’s activity is delayed until the proteasome is committed to degrading the substrate. The DUB activity of USP14 is known to be activated by proteasomes.

26S Proteasome NEW 65-1010-010 10 µg £50

26S Proteasome [Ub-VS treated] NEW 65-1020-010 10 µg £95

Description Cat. # Size Price

Deconjugating Enzymes continued...

Ubiquitin/Ubl Modifiers, Derivatives and Substrates continued...



ISG15 G1P2, IFI15 NP_005092 None 60-0008-500 500 µg £160

LC3b MAP1LC3B NP_073729.1 None 60-0112-500 500 µg £160

6His 60-0110-500 500 µg £100

GST 60-0111-500 500 µg £100

NEDD8 Q15843 None 60-0009-500 500 µg £75

SUMO1 SMT3C NP_003343 None 60-0006-500 500 µg £150

6His 60-0002-500 500 µg £108

SUMO2 HSMT3 NP_008868 None 60-0007-500 500 µg £150

6His 60-0003-500 500 µg £108

Ubiquitin, fluorogenic substrates RPS27A P62990.1 AMC 60-0116-050 50 µg £127

Lys-TAMRA 60-0118-050 50 µg £190

Rhod110 60-0117-050 50 µg £160

Ubiquitin, biotinylated NEW RPS27A P62990.1 Biotin 60-0121-020 20 µg £45

NEW 60-0121-100 100 µg £90

Modifiers

Fluorogenic Substrates, Biotin Conjugates

Ub/Ubl Binding ProteinsUbiquitin signals are decoded in cells by at least 200 ubiquitin binding proteins, which interact with different types of polyubiquitin chains and ubiquitin-like modifiers. These interactions induce conformational changes that allow these proteins to transmit the ubiquitin signal to effector proteins. The specificity of ubiquitin signalling is achieved by the interaction of monoubiquitylated and polyubiquitylated targets with ubiquitin-binding proteins resulting in various biochemical cascades and outcomes in the cell.

NEMO [pAb available - p. 15] IKBKG AAD38081 GST 66-1002-050 50 µg £205

NEMO (D311N) IKBKG AAD38081 GST 66-1013-050 50 µg £205

Optineurin [pAb available - p. 16] NP_001008214 GST 66-1005-050 50 µg £205

Optineurin (D474N) NP_001008214 GST 66-1014-050 50 µg £205

TOLLIP IL-1RAcPIP NP_061882.2 GST 66-1016-050 50 µg £195

TOM1 Target of myb1 NP_001129204.1 GST 66-1015-050 50 µg £225

S5a PSMD4, RPN10 NP_002801 None 66-2002-050 50 µg £65

GST 66-2001-050 50 µg £65




beta Catenin NEW CTNNB NP_001895.1 GST 66-2005-050 50 µg £295

FOXO3 [pAb available - p. 15] NEW FKHRL1 NP_001446 GST 66-2004-050 50 µg £295

HSP27 NEW HSPB1 NP_001531.1 GST 66-2003-050 50 µg £295

MYPT1 [pAb available - p. 15] NEW PPP1R12A NP_001137357.1 GST 66-2006-050 50 µg £215

MYPT3 [pAb available - p. 15] NEW PPP1R16A NP_116291.1 GST 66-2007-050 50 µg £215

PKB alpha [pAb available - p. 16] NEW AKT1 NP_001014431 GST 66-0018-050 50 µg £295

SGK3 [pAb available - p. 16] NEW CISK, SGKL NP_037389 GST 66-0021-050 50 µg £295


SubstratesA substrate molecule is modified by enzymes that catalyse chemical reactions involving the substrate. Ubiquitylation is a post translational modifica-tion where there is a sequential enzymatic cascade involving 3 enzymes, activation (E1s), conjugation (E2s) and ligation (E3s) which results in ubiquitin being attached to a substrate protein. Like ubiquitylation, phosphorylation is a reversible post-translational modification: Phosphorylation reactions involve the transfer of a phosphate group from a high-energy donor molecule, such as ATP, to a substrate protein or lipid, catalysed by enzymes called kinases and is a process central to intracellular signalling.

Ubiquitin/Ubl Modifiers, Derivatives and Substrates continued...



BRSK2 SAD1 AAP97725.1 6His 66-0001-050 50 µg £195

CHK2 NEW CHEK2, RAD53 NP_009125 GST 66-0019-050 50 µg £295

IKK epsilon [pAb available - p. 15] NEW IKBKE, IKKI NP_054721 GST 66-0038-050 50 µg £295

MAPK3 NEW ERK1, p44ERK1, p44MAPK CAA42744.1 none 66-0026-050 50 µg £295

MAPK7 NEW ERK5 NP_002740.2 6His 66-0027-050 50 µg £295

MAPK8 NEW JNK1, SAPK1 AAA36131.1 6His 66-0028-050 50 µg £295

MAPK9 NEW JNK2 NP_002743.3 6His 66-0029-050 50 µg £295

MAPK10 NEW JNK3 NP_002744.1 GST 66-0030-050 50 µg £295

MAPK11 NEW p38-Beta, SAPK2B CAA74792.1 GST 66-0031-050 50 µg £295

MAPK12 NEW ERK6, p38-Gamma CAA71511.1 GST 66-0032-050 50 µg £295

MAPK14 NEW SAPK2A, p38-Alpha AAA57456.1 GST 66-0034-050 50 µg £295

MAPK14 (T106M) NEW SAPK2A, p38-Alpha AAA57456.1 GST 66-0035-050 50 µg £295

MAPK15 [pAb available - p. 15] NEW ERK8 AAL40897.1 6His 66-0036-050 50 µg £295

MARK3 CTAK1 AAC15093.1 6His 66-0004-050 50 µg £195

MARK4 KIAA1860 NP_001186796.1 6His 66-0005-050 50 µg £195

PINK1 [pAb available - p. 16] NEW XP_968367.1 MBP 66-0043-050 50 µg £295

PINK1 (D359A) NEW XP_968367.1 MBP 66-0044-050 50 µg £295

PKA NP_002721.1 GST 66-0014-050 50 µg £195

PKB alpha (S473D) NEW AKT1 NP_001014431 6His 66-0017-050 50 µg £295

B-Raf NEW BRAF, B-RAF1 NP_004324.2 GST 66-0022-050 50 µg £275

B-Raf (V600E) NEW BRAF, B-RAF1 NP_004324.2 GST 66-0023-050 50 µg £295

C-Raf (Y340D; Y341D) NEW c-RAF, RAF1 NP_002871 GST 66-0024-050 50 µg £275

p70 S6 kinase (T412E) NEW P70S6K1 P67999 6His 66-0039-050 50 µg £295

SGK1 (S422D) [pAb available - p. 16] NEW NP_005618 6His 66-0020-050 50 µg £295

TAK1_TAB1 NP_003179.1 / NP_006107.1 6His 66-0007-050 50 µg £195

TBK1 [pAb available - p. 16] NAK NP_037386 GST 66-0016-050 50 µg £195


AMPK alpha 1 (T172D) NEW Tau-protein kinase PRKAA1 AAC52355 GST 66-0040-050 50 µg £215

AMPKA1/AMPKB2/AMPKG1 NEW NP_006242/NP_005390/AAH00358

6His/none/none

66-0041-050 50 µg £215

AMPKA2/AMPKB2/AMPKG1 [pAb available - p. 15]

NEW NP_006243/NP_005390/AAH00358

6His/none/none

66-0042-050 50 µg £215

KinasesLike ubiquitylation, phosphorylation is a reversible post-translational modification. Abnormal protein phosphorylation is a cause or consequence of disease, including cancers, diabetes and inflammatory disease, while defects in genes that encode protein kinases and phosphatases underlie a num-ber of inherited disorders. Approximately 500 protein kinases are encoded in the human genome and they account for nearly 25% of the molecular targets currently being pursued by drug discovery companies. It is becoming increasingly apparent that there is significant cross-talk between these two key intracellular systems: phosphorylation and ubiquitylation.



ABIN1 (human; full length), pAb* NEW Nip40-1 WB, IP / Hu 68-0002-100 100 µg £215

ABIN1 (mouse; full length), pAb* NEW Nip40-1 WB, IP / Mo 68-0001-100 100 µg £215

AMPK alpha 2 (human; residues 352 – 366), pAb* NEW WB, IP / Hu, Mo 68-0055-100 100 µg £215

CAND1 (human; residues 5 – 245), pAb* NEW TIP120 WB, IP / Hu 68-0003-100 100 µg £215

Cullin3 (human; residues 554 - 768), pAb* NEW KIAA0617 WB / Hu 68-0004-100 100 µg £215

Cullin5 (human; residues 577 – 689), pAb* NEW VACM1 WB, IP / Hu, Mo, Ch 68-0005-100 100 µg £215

CYLD (human; full length), pAb* NEW KIAA0849 WB, IP / Hu 68-0006-100 100 µg £215

DCNL3 (human; full length), pAb* NEW DCUN1D3 WB, IP / Hu 68-0007-100 100 µg £215



DCNL5 (mouse; full length), pAb* NEW DCUN1D5 WB, IP / Mo 68-0009-100 100 µg £215

FAF1 (human; full length), pAb* NEW UBXD12 WB, IP / Hu 68-0011-100 100 µg £215

FOXO3A pSer294, (human; residues 289-298), pAb* NEW FKHRL1 WB, IP / Hu 68-0038-100 100 µg £215

FOXO3A pSer344 (mouse; residues 339-348), pAb* NEW FKHRL1 WB, IP / Mo 68-0039-100 100 µg £215

FOXO3A pSer7 (mouse; residues 3-11), pAb* NEW FKHRL1 WB, IP / Mo 68-0037-100 100 µg £215

HOIL1 (human; full length), pAb* NEW RING finger protein 54 WB, IP / Hu 68-0012-100 100 µg £215

HOIP (human; full length), pAb* NEW RNF31 WB, IP / Hu 68-0013-100 100 µg £215

IKK beta (human; full length), pAb* NEW IKK2 IP / Hu, Mo 68-0049-100 100 µg £215

IKK epsilon (human; residues 701-716), pAb* NEW IKBKE, IKKI WB, IP / Hu, Mo 68-0050-100 100 µg £215

IKK epsilon (mouse; residues 702-717), pAb* NEW IKBKE, IKKI WB, IP / Hu, Mo 68-0051-100 100 µg £215

IKK epsilon pSer172 (human; residues 168 – 177), pAb* NEW IKBKE, IKKI WB, IP / Hu, Mo 68-0052-100 100 µg £215

MAPK15 (human; full length), pAb* NEW ERK8 WB / Hu 68-0040-100 100 µg £215

Mono and Polyubiquitylated conjugates (FK2), mAb 68-0121-500 500 µg £242

Mono and Polyubiquitylated conjugates (FK2), Biotin-linked, mAb 68-0123-025 25 µg £57

68-0123-100 100 µg £190

Mono and Polyubiquitylated conjugates (FK2), HRP-linked, mAb 68-0122-025 25 µg £57

68-0122-100 100 µg £190

MYPT1 (human; residues 728 – 838), pAb* NEW PPP1R12A WB / Hu 68-0041-100 100 µg £215

MYPT1 pSer554 (mouse; residues 547 - 618), pAb* NEW PPP1R12A WB / Hu 68-0045-100 100 µg £215

MYPT1 pSer445 (mouse; residues 437 - 452), pAb* NEW PPP1R12A WB / Hu, Mo 68-0043-100 100 µg £215



MYPT1 pThr34 (mouse; residues 27 - 41), pAb* NEW PPP1R12A WB / Hu, Mo 68-0042-100 100 µg £215

MYPT2 (human; residues 350-450), pAb* NEW PPP1R12B WB / Hu 68-0047-100 100 µg £215

MYPT3 (human; full length), pAb* NEW PPP1R16A WB / Hu 68-0048-100 100 µg £215

NEMO (human; full length), pAb* NEW IKBKG WB, IP / Hu, Mo 68-0014-100 100 µg £215

AntibodiesUbiquigent offers antibodies against full length or partial proteins, from across the ubiquitin cascade system. These include E2 conjugating enzymes, E3 ligases, DUBs, ubiquitin binding proteins substrates and conjugate antibodies. The anti-polyubiquitylated conjugates mAb (FK1) and the anti-mono and polyubiquitylated conjugates mAb (FK2) have been extensively characterised and can be used to demonstrate the specific recognition of polyubiq-uitylated and mono and polyubiquitylated protein conjugates respectively, but show no reactivity with free ubiquitin. Ubiquigent also offer a number of kinase antibodies. Like ubiquitylation, phosphorylation is a reversible post-translational modification. Abnormal protein phosphorylation is a cause or consequence of disease, including cancers, diabetes and inflammatory disease, while defects in genes that encode protein kinases and phosphatases underlie a number of inherited disorders. Around 500 protein kinases are encoded in the human genome and they account for around 25% of the mo-lecular targets currently being pursued by drug discovery companies. It is becoming increasingly apparent that there is significant cross-talk between these two key intracellular systems; phosphorylation and ubiquitylation.

Description Alternate Name Tested Applications / Reactivity Cat. # Size Price


Antibodies continued...

* This antibody was developed and vali dated by the Medical Research Council Protein Phosphorylation and Ubiqui tylation Unit (University of Dundee, Dundee, UK).

Key: WB = Western Blotting; IP = Immunoprecipitation; Hu = Human; Mo = Mouse; Ch = Chicken; Rt = Rat


Cell LysatesHEK293 Cell Lysates (IL-1 stimulated and non-stimulated) may be used to demonstrate the ability of ubiquitin binding domain proteins (Optineurin and NEMO) to capture polyubiquitylated proteins including IL-1 receptor-associated kinase 1 (IRAK1).

HEK293 cell lysate 66-3002-010 10 mg £222

HEK293 cell lysate (IL-1 stimulated) 66-3001-010 10 mg £315

Description Cat. # Size Price

Optineurin (human; full length), pAb* NEW IP / Hu, Mo 68-0015-100 100 µg £215

OTUB1 (human; full length), pAb* NEW FLJ20113 WB, IP / Hu 68-0016-100 100 µg £215

OTULIN (human; full length), pAb* NEW FAM105B WB, IP / Hu 68-0017-100 100 µg £215

Parkin (human; full length), pAb* NEW PARK2 WB, IP / Hu, Mo, Rt 68-0018-100 100 µg £215

Parkin pSer65 (human; residues 60 - 72), pAb* NEW PARK2 IP / Hu 68-0056-100 100 µg £215

PINK1 (human; residues 175 – 250), pAb* NEW Park6 WB, IP / Hu, Mo 68-0019-100 100 µg £215

PINK1 pThr257 (human; residues 250 - 262), pAb* NEW Park6 IP / Hu 68-0057-100 100 µg £215

PKB alpha (human; residues 1-147), pAb* NEW AKT1 IP / Hu 68-0030-100 100 µg £215

PKB beta (mouse; residues 455-469), pAb* NEW AKT2 WB, IP / Mo 68-0031-100 100 µg £215

PKB gamma (human; residues 116-127), pAb* NEW AKT3 IP / Hu 68-0032-100 100 µg £215

Polyubiquitylated conjugates (FK1), mAb 68-0111-500 500 µg £242

RNF7 (human; full length), pAb* NEW Rbx2 WB, IP / Hu 68-0020-100 100 µg £215

SGK1 (human; full length), pAb* NEW IP / Hu 68-0033-100 100 µg £215

SGK1 (human; residues 412 – 431), pAb* NEW WB, IP / Hu 68-0034-100 100 µg £215

SGK2 (human; residues 333 – 346), pAb* NEW IP / Hu 68-0035-100 100 µg £215

SGK3 (human; full length), pAb* NEW CISK, SGKL IP / Hu 68-0036-100 100 µg £215

SHARPIN (human; full length), pAb* NEW WB, IP / Hu 68-0021-100 100 µg £215

TBK1 (human; full length), pAb* NEW NAK IP / Hu, Mo 68-0053-100 100 µg £215

TBK1 pSer172 (human; residues 168 – 177), pAb* NEW NAK WB, IP / Hu 68-0054-100 100 µg £215

TRAF6 (mouse; full length), pAb* NEW RNF85 IP / Mo 68-0022-100 100 µg £215

TRIAD1 (human; full length), pAb* NEW ARIH2 WB, IP / Hu, Ch, Mo 68-0023-100 100 µg £215

TRIM65 (human; full length), pAb* NEW 4732463G12Rik WB, IP / Hu, Mo 68-0024-100 100 µg £215

UBE2M (mouse; full length), pAb* NEW Ubc12 WB / Mo 68-0025-100 100 µg £215

UBE2T (human; full length), pAb* NEW HSPC150 WB, IP / Hu 68-0026-100 100 µg £215

UBXD7 (human; full length), pAb* NEW UBXN7 WB, IP / Hu, Mo 68-0027-100 100 µg £215

USP11 (human; full length), pAb* NEW UHX1 WB, IP / Hu 68-0028-100 100 µg £215

USP15 (human; full length), pAb* NEW Unph2 WB, IP / Hu 68-0029-100 100 µg £215

Description Alternate Name Tested Applications / Reactivity Cat. # Size Price

Antibodies continued...

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North America: Contact Cosmo Bio USA, Inc. +1-760-431-4600 [email protected]

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