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 (>50) 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 July 2013

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

INSIDE

Services

Kits

NEW >> HTS Assay Kits

E1 Activating Enzymes

E2 Conjugating Enzymes

E3 Ligases

Deconjugating Enzymes

Ubiquitin/Ubl Binding Proteins

Ubiquitin/Ubl Modifiers, Derivatives and Substrates

Cell Lysates

Kinases

Antibodies

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:

• 35 DUBs on the panel and expanding.

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

• 2 week service cycle.

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 CHIP autoubiquitylation 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

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

Ubiquitylation, like phosphorylation, de-scribes a reversible post-translational pro-tein modification. Ubiquitylation or ‘ubiquit-ination’ may control the protein substrate’s destiny – in respect of its turnover – or its signalling functionality. It is a process that refers to the covalent 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-ubiquitylation of the substrate; the latter being where chains of ubiquitin are attached to the substrate protein. Ubiquitin may also form chains through attachment to the amino terminus of a further ubiquitin to generate linear poly-ubiquitin chains. The structure of the chain determines whether a ubiquitylated protein modulates a specific signalling cascade or may become degraded in a proteasomal or lysosomal dependent manner. Mono-ubiquitylated proteins may be further ubiquitylated to form polyubiq-

uitin chains and deubiquitylases may act on either mono-ubiquitylated or poly-ubiqui-tylated substrates to remove the ubiquitin monomers or chains respectively. The enzymes of the ubiquitylation pathway play a pivotal role in a number of cellular processes including, but not exclusively, the targeted proteasomal degradation of sub-strate proteins. Three classes of enzymes are involved in the process of substrate ubiqui-tylation; activating enzymes (E1s), conjugat-ing enzymes (E2s) and protein ligases (E3s). Ubiquitylation of substrate proteins depends on the sequential action of these three en-zymes. In an ATP-dependent first step, an E1 enzyme forms a thioester linkage with ubiq-uitin which is then transferred to the sulphy-dryl group of the active-site cysteine on an E2 enzyme forming a ubiquitin-thioester in-termediate. An E3 then acts as an adaptor to bind both substrate protein and E2 ‘loaded’ with ubiquitin. The E3 facilitates isopeptide bond formation between ubiquitin and the substrate protein.

Although still a post-translational modi-fication, albeit involving a functional protein rather than a functional group, ubiquityla-tion is a much more complex process than phosphorylation mainly due to the ability of ubiquitin to form polyubiquitin chains of a variety of different linkage types and com-plexity, but also because there are further related ubiquitin-like (Ubl; including SUMO, NEDD8, ISG15, and FAT10) proteins that may each follow a similar specific enzymatic cascade but resulting in different outcomes for the Ubl modified target substrate. At a further level of system complexity modifica-tions of either the E3 ligase or the substrate may alter their ability to modify substrates or themselves be modified; for example by NEDDylation (NEDD8 being a further Ubl) in the case of E3 ligases or phosphorylation in the case of both E3 ligases and substrates.

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

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

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 £245Buy 1 DUBscan Kit, get a second kit at 1/2 price!!

E2scan™ Kit version 2 NEW - 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, con-trol 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 sub-strate 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 catalyse 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 •ubiquitin 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•

References

Ye 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 ubiquitin chain binding proteins 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

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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

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B

C

D

E

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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

NEW HTS Assay KitsUbiquitin system target HTS compatible assay kits are available from Ubiquigent. HTS assays developed to therapeutically relevant targets include the Inducer of LDL (Low Density Lipoprotein) Receptor (IDOL)-ca-talysed LDLR (LDL Receptor) substrate ubiquitylation and CHIP (Carboxy Terminus of HSC70-Interacting Protein)-catalysed autoubiquitylation as-says.

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

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

E2 Conjugating EnzymesE2 conjugating enzymes form a ubiquitin/Ubl-thioester intermediate in the ubiquitylation/Ubl pathway. There are predicted to be 38 E2 conjugat-ing 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 transthio-lation 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.

E2 AlternateName

Accession Number Tag Catalogue #

VialSize

Price GBP

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

APP-BP1/UBA3 NAE1 / HPP1 NP_003896 / None 61-0006-010 10 µg £95 NP_003959.3 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 / GST/6His 61-0005-010 10 µg £95 NP_005490.1 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

IDOL-catalysed LDLR Substrate Ubiquitylation Assay Kit 70-1010 5,000 and 30,000 p.o.a

IDOL-catalysed MRLC Substrate Ubiquitylation Assay Kit 70-1020 5,000 and 30,000 p.o.a

IDOL-catalysed Autoubiquitylation Assay Kit 70-1030 5,000 and 30,000 p.o.a

CHIP-catalysed Autoubiquitylation Assay Kit 70-1040 5,000 and 30,000 p.o.a

E1 Activating EnzymesE1 activating enzymes initiate the activation and conjugation of ubiquitin and ubiquitin-like modifiers (Ubls) in a cascade of events that modifies substrate 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 co-valent thioester linkage between ‘activated’ ubiquitin/Ubl and the E1 en-zyme itself. The ubiquitin/Ubl is then transferred to the sulphydryl group of the active-site cysteine on an E2 conjugating enzyme in a transthiola-tion reaction.

E1 AlternateName(s)

Accession Number(s) Tag Catalogue #

VialSize

Price GBP

HTS Assay Catalogue # DatapointsPrice GBP

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 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

E2 AlternateName

Accession Number Tag Catalogue #

VialSize

Price GBP E2 Alternate

Name Accession Number Tag Catalogue #

VialSize

Price GBP

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

DUB/USPUSP1/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

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 50 µg £135

Deconjugating EnzymesDeconjugating enzymes (DCE) comprise a large group of hydrolases and isopeptidases that regulate the ubiquitin/ubiquitin-like protein (Ubl) path-ways 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-ubiq-uitin chains or ubiquitin-substrates. These DUBs may be divided into five subfamilies: ubiquitin-specific proteases (USPs), ovarian tumour proteas-es (OTUs), ubiquitin C-terminal hydrolases (UCHs), Josephins, and JAB1/MPN/MOV34 (JAMMs).

DCE AlternateName

Accession Number Tag Catalogue #

VialSize

Price GBP

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

CBL RNF55 NP_005179.2 GST 63-0004-010 10 µg £110 63-0004-050 50 µg £285

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

Cul1/Rbx1 Cullin1/ NP_003583.2/ None 63-1000-025 25 µg £245 HRT1 AAH17370.2

Cul1/Rbx1/Skp1 Cullin1/ NP_003583.2/ None 63-1001-025 25 µg £295 HRT1/ AAH17370.2/ p19A NP_008861.2

Cul3/Rbx1 KIAA0617/ NP_003581.1/ None 63-1003-025 25 µg £245 HRT1 AAH17370.2

Cul5/Rnf7 VACM/ NP_003469.2 / None 63-1002-025 25 µg £245 ROC2 NP_055060.1

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

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

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

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

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

TRIAD1 ARIH2 NP_006312 6His 63-0029-025 25 µg £150

WWP1 AIP5 NP_008944 None 63-0034-025 25 µg £222 GST 63-0033-025 25 µg £225

E3 LigasesE3 ligases confer specificity to the ubiquitylation pathway by recognis-ing 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 ligases, Homology to E6AP C-Terminus (HECT) domain E3 li-gases, Cullin Ring Ligases (CRLs) and Really Interesting New Gene (RING) domain E3 ligases.

E3 AlternateName(s)

Accession Number(s) Tag Catalogue #

VialSize

Price GBP

E3 AlternateName(s)

Accession Number(s) Tag Catalogue #

VialSize

Price GBP

UBE2T 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

E2 AlternateName

Accession Number Tag Catalogue #

VialSize

Price GBP

NEW

NEW

NEW

NEW

NEW

NEW

NEW

NEW

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96 assaypoints

MOD/Ubl AlternateName

Accession Number Tag Catalogue #

Vial Size

Price GBP

MODIFIERSISG15 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 RPS27A P62990.1 None 60-0001-010 10 mg £50

FLUOROGENIC SUBSTRATESUbiquitin 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 CHAINSDi-Ubiquitin, K6-Linked None 60-0101-010 10 µg £105Di-Ubiquitin, K11-Linked None 60-0102-010 10 µg £105Di-Ubiquitin, K27-Linked None 60-0103-010 10 µg £105Di-Ubiquitin, K29-Linked None 60-0104-010 10 µg £105Di-Ubiquitin, K33-Linked None 60-0105-010 10 µg £105Di-Ubiquitin, K48-Linked None 60-0106-010 10 µg £50Di-Ubiquitin, K48-Linked None 60-0106-050 50 µg £125Di-Ubiquitin, K63-Linked None 60-0107-010 10 µg £50Di-Ubiquitin, K63-Linked None 60-0107-050 50 µg £125Di-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

Modifiers/UblsAlthough ubiquitin is the most well studied and understood functional protein post-translational modifier, there is a growing family of ubiquit-in-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 path-ways with some substrate proteins becoming targeted by more than one type of modifier.

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

USP15 (isoform 2) 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

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 KIAA0849 NP_056062 6His 64-0010-050 50 µg £125

DUB/OTUA20 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 FLJ20113 NP_060140 GST 64-0011-050 50 µg £125

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

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

OTUD5 (p117S) 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 FAM105B NP_612357 GST 64-0048-050 50 µg £125

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

DUB/UCHUCHL1 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

DUB/JosephinAtaxin-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

DUB/JAMMAMSH 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

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www.ubiquigent.comDundee, Scotland, UK

ORDERS / SALES SUPPORT

North America: Contact Cosmo Bio USA, Inc. +1-760-431-4600 info@cosmobiousa.com

Other Locales: Contact Ubiquigent Ltd., UK. (UK/EU): +44-(0)1382-381147 (International): +1-617-245-0020 sales.support@ubiquigent.com

SERVICES / TECHNICAL SUPPORT

North America: +1-888-431-3233 Other Locales: (UK/EU): +44-(0)1382-381147 (International): +1-617-245-0020 tech.support@ubiquigent.com

Email services@ubiquigent.com for enquiries regarding compound profiling and/or custom assay development services.

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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 £222HEK293 cell lysate (IL-1 stimulated) 66-3001-010 10 mg £315

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AntibodiesThe anti-polyubiquitylated conjugates mAb (FK1) and the anti-mono and polyubiquitylated conjugates mAb (FK2) can be used to demonstrate the specific recognition of polyubiquitylated and mono and polyubiquitylated protein conjugates respectively, but show no reactivity with free ubiq-uitin. These antibodies have been extensively characterised and used not only to investigate ubiquitin chain formation on poly and mono ubiquity-lated proteins by Western blotting but also in the detection of intracel-lular polyubiquitin chains in immunoassays.

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

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

Mono and Polyubiquitylated conjugates, 68-0123-025 25 µg £57 mAb (FK2) Biotin linked 68-0123-100 100 µg £190 Mono and Polyubiquitylated conjugates, 68-0122-025 25 µg £57 mAb (FK2) HRP linked 68-0122-100 100 µg £190

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KinasesLike ubiquitylation, phosphorylation is a reversible post-translational modification. Abnormal protein phosphorylation is a cause or conse-quence of disease, including cancers, diabetes and inflammatory disease, while defects in genes that encode protein kinases and phosphatases underlie a number of inherited disorders. Approximately 500 protein ki-nases 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: phosphoryla-tion and ubiquitylation.

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

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

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

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

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

TBK1 NAK NP_037386 GST 66-0016-050 50 µg £195

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NEMO IKBKG AAD38081 GST 66-1002-050 50 µg £205

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

Optineurin 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

Ub/Ubl Binding ProteinsUbiquitin signals are decoded in cells by at least 200 ubiquitin bind-ing 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 ef-fector proteins. The specificity of ubiquitin signaling is achieved by the interaction of monoubiquitylated and polyubiquitylated targets with ubiquitin-binding proteins resulting in various biochemical cascades and outcomes in the cell.

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