Products and Drug Discovery Services July 2013

Home , UBE1L, UBE1L2, UBE2B, UBE2E3, UBE2G1, UBE2H

Ubiquigent Ltd provides high quality Services / HTS Assay Development reagents, kits, and drug discovery services to the research community Ubiquigent offers expert custom services to researchers interested in ubiquitin/ to help investigators unravel the ubiquitin-like protein (Ubl) cascade drug discovery. We have access to a suite of complexity of, and pursue new thera- automation robotics to help us deliver the highest quality services to you spe- peutic target opportunities across the cifically designed to help accelerate your drug discovery efforts in this field. 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, profiler™ NEW HTS Compatible Assay Kits DUB UK. This provides Ubiquigent with access to Compound Screening Service state-of-the-art services and capabilities HTS compatible assay kits have been devel- within both the PPU and other departments oped to therapeutically relevant ubiquitin sys- Ubiquigent can help you with your DUB (deu- of the College of Life Sciences. The PPU is tem targets including: Inducer of LDL (Low biquitylase) drug discovery programmes by Density Lipoprotein) Receptor (IDOL)-cataly- one of Ubiquigent’s founding partners. determining the selectivity and potency of your sed LDLR (LDL Receptor) substrate ubiquity- inhibitors. We offer the DUBprofiler service in ei- lation and CHIP autoubiquitylation assays. ther single concentration profiling or IC50 analysis mode. HTS Compatible WHAT YOU DO: INSIDE Custom Assay Development • Send us your compound(s). Services As pathways and functions are unraveled, • Request single concentration profiling and/ new and attractive ubiquitin/Ubl cascade drug or IC50 analysis (electronic submission). Kits discovery targets are coming to light with rel- evance across many key therapeutic areas. WHAT WE DO: With access to a wide range of reagents and NEW >> HTS Assay Kits benefiting from our experience working with • Determine the selectivity and specificity of these pathways, Ubiquigent’s Custom Assay your deubiquitylase (DUB) inhibitor lead E1 Activating Enzymes Development team supports pharmaceutical compound(s). and biotechnology companies in translating • Electronically report fully quality con- E2 Conjugating Enzymes these discoveries into lead programmes. trolled data to you (single concentration %

activities or IC50s). E3 Ligases We will work closely and collaboratively with you throughout our partnership. Our approach SERVICE HIGHLIGHTS: is highly flexible; from providing high quality Deconjugating Enzymes activity validated reagents to full-scale proj- • 35 DUBs on the panel and expanding. ect outsourcing developing required screen- • Any scale and combination of screening Ubiquitin/Ubl Binding Proteins ing and deconvolution assays and supplying available; compound(s) vs DUB(s). the reagents to enable you to conduct your • 2 week service cycle. Ubiquitin/Ubl Modifiers, subsequent HTS campaign in-house. Derivatives and Substrates Ubiquigent have successfully developed and DUBprofiler™-HT Cell Lysates delivered HTS-compatible assays and kits enabling leading pharmaceutical compa- Ubiquigent offers DUBprofiler-HT for the screen- nies to conduct screening campaigns against ing of larger numbers of compound (>50) ver- Kinases ubiquitin system targets (DUBs, E3s and oth- sus the DUBprofiler panel. In this service we can ers) of high therapeutic value. Furthermore offer high precision and accuracy ultra-low Antibodies Ubiquigent has developed and supplied many volume compound dispensing (down to 2.5 deconvolution and specificity profiling as- nl) to enable addition of the assay reagents says to help characterize the hits from these directly to the compound in 100% DMSO. This screens. A number of our assays have been Please visit us at is particularly suited to screens where com- developed from first principles starting with pound solubility is a concern. www.ubiquigent.com little or no data from the literature to guide the for the latest information on construction of the assay. Ubiquigent can re- new products and services. search, design, manage and deliver the entire programme to help you to achieve success in your ubiquitin system HTS campaigns.

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

Product Categories are web-interactive Kinases

E1 E2 E3 DCE

Ub/Ubl Anti-Ub-conjugate

Anti-Ub-conjugate DCE

UBP

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

Protein Ubiquitylation:

Ubiquitylation, like phosphorylation, de- uitin chains and deubiquitylases may act on Although still a post-translational modi- scribes a reversible post-translational pro- either mono-ubiquitylated or poly-ubiqui- fication, albeit involving a functional protein tein modification. Ubiquitylation or ‘ubiquit- tylated substrates to remove the ubiquitin rather than a functional group, ubiquityla- ination’ may control the protein substrate’s monomers or chains respectively. tion is a much more complex process than destiny – in respect of its turnover – or its The enzymes of the ubiquitylation pathway phosphorylation mainly due to the ability of signalling functionality. It is a process that play a pivotal role in a number of cellular ubiquitin to form polyubiquitin chains of a refers to the covalent attachment - through processes including, but not exclusively, the variety of different linkage types and com- it’s C-terminus - of a small, 76 amino acid targeted proteasomal degradation of sub- plexity, but also because there are further protein called ubiquitin to the epsilon-amino strate proteins. Three classes of enzymes are related ubiquitin-like (Ubl; including SUMO, group of a lysine residue residing within a involved in the process of substrate ubiqui- NEDD8, ISG15, and FAT10) proteins that substrate protein – which may also be anoth- tylation; activating enzymes (E1s), conjugat- may each follow a similar specific enzymatic er ubiquitin molecule. This results in either ing enzymes (E2s) and protein ligases (E3s). cascade but resulting in different outcomes mono- or poly-ubiquitylation of the substrate; Ubiquitylation of substrate proteins depends for the Ubl modified target substrate. At a the latter being where chains of ubiquitin are on the sequential action of these three en- further level of system complexity modifica- attached to the substrate protein. Ubiquitin zymes. In an ATP-dependent first step, an E1 tions of either the E3 ligase or the substrate may also form chains through attachment enzyme forms a thioester linkage with ubiq- may alter their ability to modify substrates to the amino terminus of a further ubiquitin uitin which is then transferred to the sulphy- or themselves be modified; for example by to generate linear poly-ubiquitin chains. The dryl group of the active-site cysteine on an NEDDylation (NEDD8 being a further Ubl) in structure of the chain determines whether E2 enzyme forming a ubiquitin-thioester in- the case of E3 ligases or phosphorylation in a ubiquitylated protein modulates a specific termediate. An E3 then acts as an adaptor to the case of both E3 ligases and substrates. signalling cascade or may become degraded bind both substrate protein and E2 ‘loaded’ in a proteasomal or lysosomal dependent with ubiquitin. The E3 facilitates isopeptide manner. Mono-ubiquitylated proteins may bond formation between ubiquitin and the be further ubiquitylated to form polyubiq- substrate protein.

2 Ubiquigent Ltd - Linking Ubiquitin Research to Drug Discovery www.ubiquigent.com E2scan™ Kit version 2 NEW - now containing 34 E2s The E2scan Kit is designed to facilitate the identification of E2 conjugating Why use the E2scan kit? enzymes which support the ubiquitylation of substrate proteins catalysed by an E3 ligase and/or the auto-ubiquitylation of the E3 ligase itself. • E2 conjugating enzymes can be critical in determining the ubiquitin chain linkage type(s) generated in a ubiquitylation 1 2 3 4 5 6 7 8 9 10 11 12 reaction. A 2A 2B 2C 2D1 2D2 2D3 2D4 2E1 2E2 2E3 2F 2G1 Kit Enzymes • The linkage type can determine the fate of the ubiquitylated B 2G2 2H 2I 2J1 2J2 2K 2L3 2L6 2M 2N 2N/2V1 2N/2V2 substrate; for example to be degraded or to function in a sig- Kit Controls C 2Q 2Q2 2R1 2R2 2S 2T 2V1 2V2 2W 2Z nalling capacity (Ye and Rape (2009)). BFR Buffer scan D • Discovery of novel aspects to E2 functionality: The E2 kit 2D4 Positive Control E 2A 2B 2C 2D1 2D2 2D3 2D4 2E1 2E2 2E3 2F 2G1 was used to identify E2s that partner RNF4 in polySUMO ubiquitylation and it was discovered that Ube2W (Ubc16) is F 2G2 2H 2I 2J1 2J2 2K 2L3 2L6 2M 2N 2N/2V1 2N/2V2 an E2 with - a previously unknown - specific protein amino G 2Q 2Q2 2R1 2R2 2S 2T 2V1 2V2 2W 2Z terminal monoubiquitylation activity (Tatham et al. 2013).

H BFR BFR 2D4 2D4 E2scan experiments facilitate:

• the rapid identification of the E2s that couple with your E3 The kit contains an E2scan plate - a panel of 34 E2 conjugating enzymes arrayed (auto and/or substrate ubiquitylation modes) 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 • new insights regarding the E3 ligase and/or the substrate substrate ubiquitylation or auto-ubiquitylation assay using your E3 ligase and/ • the development of assays for the E3 ligase 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 References E2scan plate, screen the E2 conjugating enzyme panel in just one hour. Reaction Ye and Rape (2009) Nat Rev Mol Cell Biol 10, 755-764. products may be analysed as required, for example by SDS-PAGE or Western Tatham MH, Plechanovová A, Jaffray EG, Salmen H, Hay RT (2013) Biochem J 453, 137-145. blotting. E2scan Kit version 2 Cat. # 67-0005-001 1 Kit £345

DUBscan™ Kit The DUBscan kit has been flexibly designed for many potential applications. One Why use the DUBscan kit? application that might be of interest is the identification of DUB enzymes which catalyse the deubiquitylation of a substrate protein. • Examine the cleavage of in vitro ubiquitylated substrates generated using different E2/E3 combinations (refer to the 1 2 3 4 5 6 7 8 9 10 11 12 Kit Enzymes Ubiquigent E2scan kit for identifying E2s that ‘couple’ with your

USP1/ USP2 USP4 USP5 USP6 USP7 USP9X BFR BFR AMSH OTUB1 A UAF1 Kit Controls E3 of interest)

USP14/ UbVS- USP11 USP14 Ptsm USP15 USP19 USP20 USP21 USP25 BFR BFR Buffer B UbVS- Ptsm Ptsm • Examine the cleavage of in vivo ubiquitylated substrates af- Positive Control C USP27x USP28 USP30 USP35 USP36 USP45 CYLD UCHL1 UCHL3 UCHL5 BFR AMSH ter capture from a cell lysate (refer to Ubiquigent ubiquitin chain binding proteins for capturing ubiquitylated proteins) D BAP1 A20 Cezanne OTU1 OTUB1 OTUB2 OTUD3 OTUD5 OTUD6A OTUD6B BFR OTUB1 Negative Control

OTULIN TRABID ATXN-3 ATXN-3L JOSD1 JOSD2 AMSH AMSH-LP AMSH AMSH-LP BFR • Investigate DUB ubiquitin-ubiquitin linkage cleavage speci- E + Zn + Zn User Controls ficity (refer to Ubiquigent di-ubiquitin and ubiquitin chains F BFR Buffer for use as substrates) G User DUBs

H • Examine the relative position of cleavage within ubiquitin Suggested empty wells to enable user to run DUBscan chains (distal versus proximal) samples on 12-well gels • Explore mono and poly-deubiquitylation DUB specificity (re- scan The kit contains a DUB plate (a panel of DUBs arrayed across 47 wells of fer to specific Ubiquigent E2s – such as Ube2W – for mono- a 96 well plate) plus a control ubiquitylation reaction mix for performing both ubiquitylating E2s) positive and negative control deubiquitylation reactions. • Explore potential DUB inhibitors and/or activators In respect of one kit application - the identification of DUB enzymes which catalyse the deubiquitylation of a substrate protein; through the addition of • Identify novel DUB binding proteins 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 • Explore how DUBs may interact with and modify the activity of analysed as required, for example by SDS-PAGE and Western blotting using other ubiquitin system proteins such as E2s and vice versa specific antibodies. One kit contains all the reagents and buffers necessary to perform a ‘DUBscan’ of the panel of 47 DUB assay wells.

scan Buy 1 DUB Kit, get a second kit at 1/2 price!! DUBscan Kit Cat. # 67-0006-001 1 Kit £245

3 Ubiquigent Ltd - Linking Ubiquitin Research to Drug Discovery www.ubiquigent.com NEW HTS Assay Kits E2 Conjugating Enzymes E2 conjugating enzymes form a ubiquitin/Ubl-thioester intermediate in Ubiquitin system target HTS compatible assay kits are available from the ubiquitylation/Ubl pathway. There are predicted to be 38 E2 conjugat- Ubiquigent. HTS assays developed to therapeutically relevant targets ing enzymes and four classes of the enzyme exist, all of which contain include the Inducer of LDL (Low Density Lipoprotein) Receptor (IDOL)-ca- a catalytic (Ubc) domain with an active site cysteine. After transfer of talysed LDLR (LDL Receptor) substrate ubiquitylation and CHIP (Carboxy ubiquitin/Ubl from the E1 activating enzyme to the E2 via a transthio- Terminus of HSC70-Interacting Protein)-catalysed autoubiquitylation as- lation reaction, the loaded E2 interacts with E3 ligases to transfer the says. activated ubiquitin/Ubl to the substrate lysine acceptor residue; either Price directly from the E2 or via a lysine reside on the E3 ligase itself. HTS Assay Catalogue # Datapoints GBP Alternate Accession Vial Price E2 Name Number Tag Catalogue # Size GBP IDOL-catalysed LDLR Substrate Ubiquitylation Assay Kit 70-1010 5,000 and 30,000 p.o.a UBE2A HR6A NP_003327 None 62-0002-020 20 µg £65 IDOL-catalysed MRLC Substrate 62-0002-100 100 µg £135 Ubiquitylation Assay Kit 70-1020 5,000 and 30,000 p.o.a 6His 62-0071-020 20 µg £65 62-0071-100 100 µg £135 IDOL-catalysed GST 62-0001-020 20 µg £65 Autoubiquitylation Assay Kit 70-1030 5,000 and 30,000 p.o.a 62-0001-100 100 µg £135

CHIP-catalysed UBE2B HR6B NP_003328 None 62-0004-020 20 µg £65 Autoubiquitylation Assay Kit 70-1040 5,000 and 30,000 p.o.a 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

E1 Activating Enzymes UBE2C UbcH10 NP_008950 None 62-0007-020 20 µg £65 62-0007-100 100 µg £135 E1 activating enzymes initiate the activation and conjugation of ubiquitin 6His 62-0005-020 20 µg £65 and ubiquitin-like modifiers (Ubls) in a cascade of events that modifies 62-0005-100 100 µg £135 substrate proteins resulting in proteasomal or lysosomal degradation, GST 62-0006-020 20 µg £65 changes in intracellular localisation, activity or other signaling events. 62-0006-100 100 µg £135 To date eight E1 enzymes have been identified. E1 enzymes utilise ATP to activate the terminal glycine residue of ubiquitin/Ubl generating a co- UBE2D1 UbcH5a NP_003329 None 62-0010-020 20 µg £65 valent thioester linkage between ‘activated’ ubiquitin/Ubl and the E1 en- 62-0010-100 100 µg £135 zyme itself. The ubiquitin/Ubl is then transferred to the sulphydryl group 6His 62-0008-020 20 µg £65 of the active-site cysteine on an E2 conjugating enzyme in a transthiola- 62-0008-100 100 µg £135 tion reaction. GST 62-0009-020 20 µg £65 62-0009-100 100 µg £135 Alternate Accession Vial Price E1 Name(s) Number(s) Tag Catalogue # Size GBP UBE2D2 UbcH5b NP_003330 None 62-0012-020 20 µg £65 62-0012-100 100 µg £135 6His 62-0073-020 20 µg £65 APP-BP1/UBA3 NAE1 / HPP1 NP_003896 / None 61-0006-010 10 µg £95 62-0073-100 100 µg £135 NP_003959.3 61-0006-050 50 µg £270 GST 62-0011-020 20 µg £65 62-0011-100 100 µg £135 ATG7 APG7, GSA7 NP_006386 6His 61-0008-010 10 µg £95 61-0008-050 50 µg £270 UBE2D3 UbcH5c NP_003331 None 62-0014-020 20 µg £65 62-0014-100 100 µg £135 SAE1/SAE2 AOS1 / UBA2 NP_005491.1 / GST/6His 61-0005-010 10 µg £95 6His 62-0074-020 20 µg £65 NP_005490.1 61-0005-050 50 µg £270 62-0074-100 100 µg £135 GST 62-0013-020 20 µg £65 UBA6 E1-L2, UBE1L2 NP_060697 6His 61-0002-010 10 µg £50 62-0013-100 100 µg £135 61-0002-050 50 µg £113 UBE2D4 UbcH5d NP_057067 None 62-0017-020 20 µg £65 UBA7 UBE1L NP_003326.2 6His 61-0007-010 10 µg £95 62-0017-100 100 µg £135 61-0007-050 50 µg £270 6His 62-0015-020 20 µg £65 62-0015-100 100 µg £135 UBE1 A1S9, A1S9T NP_003325 6His 61-0001-010 10 µg £50 GST 62-0016-020 20 µg £65 61-0001-050 50 µg £113 62-0016-100 100 µg £135

4 Ubiquigent Ltd - Linking Ubiquitin Research to Drug Discovery www.ubiquigent.com Alternate Accession Vial Price Alternate Accession Vial Price E2 Name Number Tag Catalogue # Size GBP E2 Name Number Tag Catalogue # Size GBP

UBE2E1 UbcH6 AAH09139 None 62-0019-020 20 µg £65 UBE2K UbcH1 NP_005330 None 62-0039-020 20 µg £65 62-0019-100 100 µg £135 62-0039-100 100 µg £135 6His 62-0075-020 20 µg £65 6His 62-0079-020 20 µg £65 62-0075-100 100 µg £135 62-0079-100 100 µg £135 GST 62-0018-020 20 µg £65 GST 62-0038-020 20 µg £65 62-0018-100 100 µg £135 62-0038-100 100 µg £135

UBE2E2 UbcH8 NP_689866.1 None 62-0066-020 20 µg £65 UBE2L3 UbcH7 AAH53368 None 62-0042-020 20 µg £65 62-0066-100 100 µg £135 62-0042-100 100 µg £135 6His 62-0020-020 20 µg £65 6His 62-0040-020 20 µg £65 62-0020-100 100 µg £135 62-0040-100 100 µg £135 GST 62-0087-020 20 µg £65 GST 62-0041-020 20 µg £65 62-0087-100 100 µg £135 62-0041-100 100 µg £135

UBE2E3 UbcH9 NP_006348 None 62-0022-020 20 µg £65 UBE2L6 UbcH8 NP_004214 None 62-0044-020 20 µg £65 62-0022-100 100 µg £135 62-0044-100 100 µg £135 6His 62-0076-020 20 µg £65 6His 62-0080-020 20 µg £65 62-0076-100 100 µg £135 62-0080-100 100 µg £135 GST 62-0021-020 20 µg £65 GST 62-0043-020 20 µg £65 62-0021-100 100 µg £135 62-0043-100 100 µg £135

UBE2F NCE2 NP_542409 None 62-0025-020 20 µg £65 UBE2M Ubc12 AAH58924 None 62-0068-020 20 µg £65 62-0025-100 100 µg £135 62-0068-100 100 µg £135 6His 62-0023-020 20 µg £65 6His 62-0048-020 20 µg £65 62-0023-100 100 µg £135 62-0048-100 100 µg £135 GST 62-0024-020 20 µg £65 GST 62-0088-020 20 µg £65 62-0024-100 100 µg £135 62-0088-100 100 µg £135

UBE2G1 Ubc7 NP_003333 None 62-0028-020 20 µg £65 UBE2N Ubc13 NP_003339 None 62-0047-020 20 µg £65 62-0028-100 100 µg £135 62-0047-100 100 µg £135 6His 62-0026-020 20 µg £65 6His 62-0045-020 20 µg £65 62-0026-100 100 µg £135 62-0045-100 100 µg £135 GST 62-0027-020 20 µg £65 GST 62-0046-020 20 µg £65 62-0027-100 100 µg £135 62-0046-100 100 µg £135

UBE2G2 Ubc7 NP_003334 None 62-0030-020 20 µg £65 UBE2Q1 NICE-5 NP_060052 6His 62-0081-020 20 µg £65 62-0030-100 100 µg £135 62-0081-100 100 µg £135 6His 62-0077-020 20 µg £65 GST 62-0049-020 20 µg £65 62-0077-100 100 µg £135 62-0049-100 100 µg £135 GST 62-0029-020 20 µg £65 62-0029-100 100 µg £135 UBE2Q2 NP_775740 None 62-0069-020 20 µg £65 62-0069-100 100 µg £135 UBE2H UbcH2 NP_003335 None 62-0032-020 20 µg £65 6His 62-0051-020 20 µg £65 62-0032-100 100 µg £135 62-0051-100 100 µg £135 6His 62-0078-020 20 µg £65 GST 62-0089-020 20 µg £65 62-0078-100 100 µg £135 62-0089-100 100 µg £135 GST 62-0031-020 20 µg £65 62-0031-100 100 µg £135 UBE2R1 UbcH3 NP_004350 None 62-0054-020 20 µg £65 62-0054-100 100 µg £135 UBE2I Ubc9 NP_003336 None 62-0034-020 20 µg £65 6His 62-0052-020 20 µg £65 62-0034-100 100 µg £135 62-0052-100 100 µg £135 6His 62-0033-020 20 µg £65 GST 62-0053-020 20 µg £65 62-0033-100 100 µg £135 62-0053-100 100 µg £135 GST 62-0065-020 20 µg £65 62-0065-100 100 µg £135 UBE2R2 CDC34B AAH04862 None 62-0098-020 20 µg £65 62-0098-100 100 µg £135 UBE2J1 NCUBE1 NP_057105.2 6His 62-0096-020 20 µg £65 6His 62-0094-020 20 µg £65 62-0096-100 100 µg £135 62-0094-100 100 µg £135

UBE2J2 NCUBE2 NP_477515.2 None 62-0067-020 20 µg £65 UBE2S E2-EPF AAH65364 None 62-0056-020 20 µg £65 62-0067-100 100 µg £135 62-0056-100 100 µg £135 6His 62-0035-020 20 µg £65 6His 62-0082-020 20 µg £65 62-0035-100 100 µg £135 62-0082-100 100 µg £135 GST 62-0036-020 20 µg £65 GST 62-0055-020 20 µg £65 62-0036-100 100 µg £135 62-0055-100 100 µg £135

5 Ubiquigent Ltd - Linking Ubiquitin Research to Drug Discovery www.ubiquigent.com Alternate Accession Vial Price Alternate Accession Vial Price E2 Name Number Tag Catalogue # Size GBP E3 Name(s) Number(s) Tag Catalogue # Size GBP

UBE2T HSPC150 NP_054895 None 62-0070-020 20 µg £65 Cul5/Rnf7 VACM/ NP_003469.2 / None 63-1002-025 25 µg £245 62-0070-100 100 µg £135 NEW ROC2 NP_055060.1 6His 62-0057-020 20 µg £65 62-0057-100 100 µg £135 DCNL1 FLJ10704 NP_065691.2 6His 63-2000-025 25 µg £95 GST 62-0090-020 20 µg £65 NEW GST 63-2001-025 25 µg £95 62-0090-100 100 µg £135 DCNL2 C13orf17 NP_001014305.1 6His 63-2004-025 25 µg £95 UBE2V1 Uev1 AAG24229 None 62-0059-020 20 µg £65 NEW GST 63-2002-025 25 µg £95 62-0059-100 100 µg £135 6His 62-0084-020 20 µg £65 IPAH9.8 AAP79029.1 AAP79029.1 GST 63-0018-025 25 µg £92 62-0084-100 100 µg £135 GST 62-0102-020 20 µg £65 ITCH AIF4 NP_1136713 GST 63-0006-025 25 µg £150 62-0102-100 100 µg £135 RNF8 KIAA0646 NP_003949 GST 63-0021-025 25 µg £150 UBE2V2 Mms2 NP_003341.1 None 62-0100-020 20 µg £65 62-0100-100 100 µg £135 RNF11 CGI-123 NP_055187 6His 63-0023-025 25 µg £92 6His 62-0099-020 20 µg £65 GST 63-0024-025 25 µg £92 62-0099-100 100 µg £135 GST 62-0101-020 20 µg £65 SMURF1 KIAA1625 NP_065162.1 GST 63-0027-025 25 µg £225 62-0101-100 100 µg £135 TRIAD1 ARIH2 NP_006312 6His 63-0029-025 25 µg £150 UBE2W Ubc16 NP_001001481.1 6His 62-0085-020 20 µg £65 62-0085-100 100 µg £135 WWP1 AIP5 NP_008944 None 63-0034-025 25 µg £222 GST 62-0091-020 20 µg £65 GST 63-0033-025 25 µg £225 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 Deconjugating Enzymes Deconjugating enzymes (DCE) comprise a large group of hydrolases and isopeptidases that regulate the ubiquitin/ubiquitin-like protein (Ubl) path- E3 Ligases ways by cleaving ubiquitin/Ubl-ubiquitin/Ubl, or ubiquitin/Ubl-substrate bonds. There are >70 DCEs – termed deubiquitinases or deubiquitylases E3 ligases confer specificity to the ubiquitylation pathway by recognis- (DUBs) – encoded by the human genome that deconjugate ubiquitin-ubiq- ing target substrates and mediating transfer of ubiquitin or ubiquitin like uitin chains or ubiquitin-substrates. These DUBs may be divided into five proteins (Ubls) from an E2 conjugating enzyme to a substrate. There are subfamilies: ubiquitin-specific proteases (USPs), ovarian tumour proteas- estimated to be >700 E3 ligases forming four classes of enzyme; N-end es (OTUs), ubiquitin C-terminal hydrolases (UCHs), Josephins, and JAB1/ rule ubiquitin ligases, Homology to E6AP C-Terminus (HECT) domain E3 li- MPN/MOV34 (JAMMs). gases, Cullin Ring Ligases (CRLs) and Really Interesting New Gene (RING) domain E3 ligases. Alternate Accession Vial Price DCE Name Number Tag Catalogue # Size GBP Alternate Accession Vial Price E3 Name(s) Number(s) Tag Catalogue # Size GBP DUB/USP USP1/UAF1 NP_003359/NP_065890 6His/None 64-0040-050 50 µg £285 CBL RNF55 NP_005179.2 GST 63-0004-010 10 µg £110 NEW

63-0004-050 50 µg £285 USP2 (isoform 4) UBP41 NP_741994 GST 64-0014-050 50 µg £125

CHIP STUB1 NP_005852 None 63-0003-020 20 µg £125 USP4 NP_003354 6His 64-0001-050 50 µg £285 63-0003-100 100 µg £250 6His 63-0001-025 25 µg £92 USP5 Isopeptidase T P45974 6His 64-0002-050 50 µg £125 GST 63-0002-025 25 µg £92

USP6 CD(529-1406) TRE2 NP_004496 GST 64-0045-050 50 µg £285 Cul1/Rbx1 Cullin1/ NP_003583.2/ None 63-1000-025 25 µg £245 NEW NEW HRT1 AAH17370.2 USP7 HAUSP CAA96580 6His 64-0003-050 50 µg £195

Cul1/Rbx1/Skp1 Cullin1/ NP_003583.2/ None 63-1001-025 25 µg £295 USP9x CD(1554-1995) DFFRX NP_001034680 GST 64-0017-050 50 µg £195 NEW HRT1/ AAH17370.2/ p19A NP_008861.2 USP14 AAH03556 6His 64-0018-050 50 µg £125 NEW 64-0018-005 5 µg £50 Cul3/Rbx1 KIAA0617/ NP_003581.1/ None 63-1003-025 25 µg £245 NEW 96 assay HRT1 AAH17370.2 USP14 & 26S Proteasome [Ub-VS treated] 6His/None 64-1010-096 50points µg £135 NEW

6 Ubiquigent Ltd - Linking Ubiquitin Research to Drug Discovery www.ubiquigent.com Alternate Accession Vial Price DCE Name(s) Number Tag Catalogue # Size GBP Modifiers/Ubls Although ubiquitin is the most well studied and understood functional

USP15 (isoform 2) Unph2 NP_006304 None 64-0026-050 50 µg £125 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 USP19-TM (1-1290) ZMYND9 NP_006668 GST 64-0022-050 50 µg £285 Ubl protein family include ubiquitin, SUMO, NEDD8, ISG15, URM1, FAT10,

USP20 LSFR3A AAH39593 GST 64-0039-050 50 µg £285 UFM1, and ATG12. These related proteins are structurally similar, and NEW are activated, conjugated, and released from conjugates in a mechanism akin to that for ubiquitin. There is cross-talk between conjugation path- USP21 CD(196-565) USP23 NP_001014443 GST 64-0037-050 50 µg £285 NEW ways with some substrate proteins becoming targeted by more than one type of modifier. USP25 NP_037528 GST 64-0023-050 50 µg £125

USP27x USP22L NP_001138545 DAC 64-0046-050 50 µg £285 Alternate Accession Vial Price NEW MOD/Ubl Name Number Tag Catalogue # Size GBP 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 MODIFIERS ISG15 G1P2, IFI15 NP_005092 None 60-0008-500 500 µg £160 USP45 MGC14793 NP_001073950 GST 64-0025-050 50 µg £285

LC3b MAP1LC3B NP_073729.1 None 60-0112-500 500 µg £160 CYLD KIAA0849 NP_056062 6His 64-0010-050 50 µg £125 6His 60-0110-500 500 µg £100 GST 60-0111-500 500 µg £100

DUB/OTU NEDD8 Q15843 None 60-0009-500 500 µg £75 A20 CD(1-366) TNFAIP3 NP_006281 GST 64-0047-050 50 µg £125 NEW NEW SUMO1 SMT3C NP_003343 None 60-0006-500 500 µg £150 OTU1 YOD1 NP_061036 GST 64-0036-050 50 µg £285 6His 60-0002-500 500 µg £108

OTUB1 FLJ20113 NP_060140 GST 64-0011-050 50 µg £125 NEW SUMO2 HSMT3 NP_008868 None 60-0007-500 500 µg £150 6His 60-0003-500 500 µg £108 OTUB2 OTU2; OTB2 NP_075601.1 GST 64-0012-050 50 µg £125

Ubiquitin RPS27A P62990.1 None 60-0001-010 10 mg £50 OTUD3 KIAA0459 NP_056022 GST 64-0035-050 50 µg £285

OTUD5 (p117S) DUBA AAH09917 GST 64-0043-050 50 µg £285 NEW FLUOROGENIC SUBSTRATES Ubiquitin RPS27A P62990.1 AMC 60-0116-050 50 µg £127 OTUD6A DUBA-2 NP_997203 6His 64-0038-050 50 µg £285 NEW Lys-TAMRA 60-0118-050 50 µg £190 Rhod110 60-0117-050 50 µg £160 OTUD6B DUBA-5 NP_057107 GST 64-0028-050 50 µg £285

OTULIN FAM105B NP_612357 GST 64-0048-050 50 µg £125 NEW UBIQUITIN CHAINS Di-Ubiquitin, K6-Linked None 60-0101-010 10 µg £105 TRABID CD(245-697) ZRANB1 NP_060050 6His 64-0049-050 50 µg £125 NEW Di-Ubiquitin, K11-Linked None 60-0102-010 10 µg £105 Di-Ubiquitin, K27-Linked None 60-0103-010 10 µg £105 DUB/UCH Di-Ubiquitin, K29-Linked None 60-0104-010 10 µg £105 Di-Ubiquitin, K33-Linked None 60-0105-010 10 µg £105 UCHL1 PARK5 AAH00332.1 6His 64-0007-050 50 µg £125 Di-Ubiquitin, K48-Linked None 60-0106-010 10 µg £50 Di-Ubiquitin, K48-Linked None 60-0106-050 50 µg £125 UCHL3 AAH18125 GST 64-0027-050 50 µg £125 Di-Ubiquitin, K63-Linked None 60-0107-010 10 µg £50 Di-Ubiquitin, K63-Linked None 60-0107-050 50 µg £125 UCHL5 UCH37 AAH15521.1 GST 64-0008-050 50 µg £195 Di-Ubiquitin, Linear GST-cleaved 60-0115-010 10 µg £65 GST-cleaved 60-0115-050 50 µg £160 DUB/Josephin Ataxin-3 AAH33711 GST 64-0033-050 50 µg £125 Poly-Ubiquitin , Ub n2-7; K63-linked None 60-0109-100 100 µg £75

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

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

7 Ubiquigent Ltd - Linking Ubiquitin Research to Drug Discovery www.ubiquigent.com Ub/Ubl Binding Proteins Antibodies

Ubiquitin signals are decoded in cells by at least 200 ubiquitin bind- The anti-polyubiquitylated conjugates mAb (FK1) and the anti-mono and ing proteins, which interact with different types of polyubiquitin chains polyubiquitylated conjugates mAb (FK2) can be used to demonstrate the and ubiquitin-like modifiers. These interactions induce conformational specific recognition of polyubiquitylated and mono and polyubiquitylated changes that allow these proteins to transmit the ubiquitin signal to ef- protein conjugates respectively, but show no reactivity with free ubiq- fector proteins. The specificity of ubiquitin signaling is achieved by the uitin. These antibodies have been extensively characterised and used not interaction of monoubiquitylated and polyubiquitylated targets with only to investigate ubiquitin chain formation on poly and mono ubiquity- ubiquitin-binding proteins resulting in various biochemical cascades and lated proteins by Western blotting but also in the detection of intracel- outcomes in the cell. lular polyubiquitin chains in immunoassays.

Alternate Accession Vial Price Vial Price UbBP Name(s) Number Tag Catalogue # Size GBP AB Catalogue # Size GBP

NEMO IKBKG AAD38081 GST 66-1002-050 50 µg £205 Polyubiquitylated conjugates, 68-0111-500 500 µg £242 mAb (FK1) NEMO (D311N) IKBKG AAD38081 GST 66-1013-050 50 µg £205 Mono and Polyubiquitylated conjugates, 68-0121-500 500 µg £242 Optineurin NP_001008214 GST 66-1005-050 50 µg £205 mAb (FK2)

Optineurin (D474N) NP_001008214 GST 66-1014-050 50 µg £205 Mono and Polyubiquitylated conjugates, 68-0123-025 25 µg £57 mAb (FK2) Biotin linked TOLLIP IL-1RAcPIP NP_061882.2 GST 66-1016-050 50 µg £195 68-0123-100 100 µg £190 NEW TOM1 Target of myb1 NP_001129204.1 GST 66-1015-050 50 µg £225 Mono and Polyubiquitylated conjugates, 68-0122-025 25 µg £57 NEW mAb (FK2) HRP linked S5a PSMD4; RPN10 NP_002801 None 66-2002-050 50 µg £65 68-0122-100 100 µg £190 NEW GST 66-2001-050 50 µg £65

Cell Lysates Kinases HEK293 Cell Lysates (IL-1 stimulated and non-stimulated) may be Like ubiquitylation, phosphorylation is a reversible post-translational used to demonstrate the ability of ubiquitin binding domain proteins modification. Abnormal protein phosphorylation is a cause or conse- (Optineurin and NEMO) to capture polyubiquitylated proteins including quence of disease, including cancers, diabetes and inflammatory disease, IL-1 receptor-associated kinase 1 (IRAK1). while defects in genes that encode protein kinases and phosphatases underlie a number of inherited disorders. Approximately 500 protein ki- Vial Price nases are encoded in the human genome and they account for nearly LYSATE Catalogue # Size GBP 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- HEK293 cell lysate 66-3002-010 10 mg £222 tion and ubiquitylation. HEK293 cell lysate (IL-1 stimulated) 66-3001-010 10 mg £315

KINASE Alternate Accession Vial Price Name Number Tag Catalogue # Size GBP

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

ORDERS / SALES SUPPORT SERVICES / TECHNICAL SUPPORT North America: Contact Cosmo Bio USA, Inc. North America: +1-888-431-3233 +1-760-431-4600 www.ubiquigent.com [email protected] Other Locales: (UK/EU): +44-(0)1382-381147 Dundee, Scotland, UK (International): +1-617-245-0020 Other Locales: Contact Ubiquigent Ltd., UK. [email protected] (UK/EU): +44-(0)1382-381147 © Ubiquigent 2013. Unless otherwise noted, Ubiquigent, (International): +1-617-245-0020 Email [email protected] for enquiries regarding compound Ubiquigent logo and all other trademarks are the property [email protected] profiling and/or custom assay development services. of Ubiquigent, Ltd.