Pre-Press PRODUCT LIST Addendum April 2015

Products and Drug Discovery Services October 2014

Web-interactive PDF

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 ResearchPre-Press Council Protein PRODUCT Phos- LIST Addendum April 2015 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 MuRF1 assays. ther single concentration profiling or IC50 analysis mode. Description Cat. # Size PriceHTS Compatible Description Cat. # Size Price JUMP TO WEBSITE JUMP TO PAGE WHAT YOU DO: Ubiquitin (synthetic)* INSIDE60-0200-050 50 µg £67 Custom €80 AssayNEW Development Parkin-Related Phospho-Ubiquitins • Send us your compound(s). Ubiquitin (pSer57) 60-0208-050 50 µg £150 €180 Ubiquitin (pSer65) 60-0202-050 50 µg £150 €180 Services...... 1 As pathways and functions are unraveled, Ubiquitin (pSer65) 60-0202-050 50 µg £150 €180 • Request single concentration profiling and/ new and attractiveBiotin-Ahx-Ubiquitin ubiquitin/Ubl cascade (pSer65) drug 60-0207-050 50 µg £183 €220 or IC50 analysis (electronic submission). Biotin-Ahx-UbiquitinCell Lines and Kit (synthetic)*s...... 60-0201-050 50 µg3 £83 discovery€100 targets Ubiquitinare coming (synthetic)* to light with rel- 60-0200-050 50 µg £67 €80 Biotin-Ahx-Ubiquitin (pThr7) 60-0203-050 50 µg £183 evance€220 across manyBiotin-Ahx-Ubiquitin key therapeutic (synthetic)* areas. 60-0201-05WHAT WE0 DO:50 µg £83 €100 Assay Kits - HTS, USP14, E2scan, DUBscan...... 4 With access to a wide range of reagents and Biotin-Ahx-Ubiquitin (pThr12) 60-0204-050 50 µg £183 €220 benefiting from ourAntibodies experience working with • Determine the selectivity and specificity of Biotin-Ahx-Ubiquitin (pSer57) 60-0205-050 50 µg £183 €220 your deubiquitylase (DUB) inhibitor lead E1 Activating Enzymes...... 6 these pathways, Parkin Ubiquigent’s (human; full Custom length), pAb* Assay 68-0018-100 100 µg £215 €259 compound(s). Biotin-Ahx-Ubiquitin (pTyr59) 60-0206-050 50 µg £183 Development€220 team supports pharmaceutical E2 Conjugating Enzymes...... 6 and biotechnologyParkin companies pSer65 (human; in translating residues 60 - 68-0056-100 100 µg £215 €259 Biotin-Ahx-Ubiquitin (pSer65) 60-0207-050 50 µg £183 €220 • Electronically report fully quality con- these discoveriesPINK1 into lead (human; programmes. residues 175 – 250), pAb* 68-0019-10trolled0 data100 to µg you£215 (single €259 concentration % E3 Ligases...... 10 PINK1 pThr257 (human; residues 250 68-0057-10activities0 or100 IC µg50s).£215 €259 We will work closely and collaboratively with Deconjugating Enzymes...... 11 you throughout ourRecombinant partnership. Proteins Our approach SERVICE HIGHLIGHTS: is highly flexible; from providing high quality Parkin 63-0048-02• 38 different5 25 DUBs µg £295(42 assays) €356 on the panel Proteasome...... NEW 12 activity validated reagents to full-scale proj- Publications ect outsourcing developingPINK1 required screen- 66-0043-05and expanding.0 50 µg £295 €356 Ubiquitin/Ubl Modifiers, ing and deconvolutionPINK1 (D359A) assays and supplying 66-0044-05• Any scale0 and50 µg combination£295 €356 of screening the reagents to enable you to conduct your available; compound(s) vs DUB(s). Derivatives and Substratesclick...... to 12 USP30 CD(57-517) 64-0057-050 50 µg £285 €345 download subsequent HTS campaign in-house. *All phospho-ubiquitins have been synthetically• generated2 week so service it is recommended cycle. to use

Ubiquitin/Ubl Binding Proteins...... 13 these control reagents in experiments as appropriate. TM Ubiquigent have successfully developed and • StoragePod® system for DUBprofiler delivered HTS-compatible assays and kits compound management. Substrates...... 13 enabling leading pharmaceutical compa-

nies to conduct screening campaigns against profiler™ Kinases...... 14 DUB Phosphorlyated-HT ubiquitin and derivatives are offered pUbiquitins PINK1/Parkinubiquitin system targets (DUBs, E3s and oth- by Ubiquigent under an exclusive license from UbiQ Bio BV, Amsterdam, the Netherlands. ers) of high therapeutic value. Furthermore profiler Antibodies...... 15 Ubiquigent offers DUB -HT for the screen- Ubiquigent has developed and supplied many ing of larger numbers of compound (>25) ver- deconvolution and specificity profiling as- profiler Cell Lysates...... 16 sus the DUB panel. In this service we can says to help characterize the hits from these offer high precision and accuracy ultra-low screens. A number of our assays have been volume compound dispensing (down to 2.5 developed from first principles starting with nl) to enable addition of the assay reagents Please visit us at little or no data from the literature to guide the directly to the compound in 100% DMSO. This www.ubiquigent.com construction of the assay. Ubiquigent can re- is particularly suited to screens where com- for the latest information on search, design, manage and deliver the entire pound solubility is a concern. new products and services. programme to help you to achieve success in your ubiquitin system HTS campaigns.

Ubiquigent Ltd - Linking Ubiquitin Research to Drug Discovery www.ubiquigent.com Products and Drug Discovery Services October 2014

Web-interactive PDF

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 MuRF1 assays. ther single concentration profiling or IC50 analysis mode. HTS Compatible JUMP TO WEBSITE JUMP TO PAGE WHAT YOU DO: INSIDE Custom Assay Development • Send us your compound(s). Services...... 1 As pathways and functions are unraveled, • Request single concentration profiling and/ new and attractive ubiquitin/Ubl cascade drug or IC50 analysis (electronic submission). Cell Lines and Kits...... 3 discovery targets are coming to light with rel- evance across many key therapeutic areas. WHAT WE DO: Assay Kits - HTS, USP14, E2scan, DUBscan...... 4 With access to a wide range of reagents and benefiting from our experience working with • Determine the selectivity and specificity of E1 Activating Enzymes...... 6 these pathways, Ubiquigent’s Custom Assay your deubiquitylase (DUB) inhibitor lead Development team supports pharmaceutical compound(s). E2 Conjugating Enzymes...... 6 and biotechnology companies in translating • Electronically report fully quality con- these discoveries into lead programmes. trolled data to you (single concentration % E3 Ligases...... 10 activities or IC50s). We will work closely and collaboratively with Deconjugating Enzymes...... 11 you throughout our partnership. Our approach SERVICE HIGHLIGHTS: is highly flexible; from providing high quality • 38 different DUBs (42 assays) on the panel Proteasome...... 12 activity validated reagents to full-scale proj- ect outsourcing developing required screen- and expanding. Ubiquitin/Ubl Modifiers, ing and deconvolution assays and supplying • Any scale and combination of screening Derivatives and Substrates...... 12 the reagents to enable you to conduct your available; compound(s) vs DUB(s). subsequent HTS campaign in-house. • 2 week service cycle.

Ubiquitin/Ubl Binding Proteins...... 13 TM Ubiquigent have successfully developed and • StoragePod® system for DUBprofiler delivered HTS-compatible assays and kits compound management. Substrates...... 13 enabling leading pharmaceutical companies to conduct screening campaigns against DUBprofiler™-HT Kinases...... 14 ubiquitin system targets (DUBs, E3s and oth- ers) of high therapeutic value. Furthermore profiler Antibodies...... 15 Ubiquigent offers DUB -HT for the screen- Ubiquigent has developed and supplied many ing of larger numbers of compound (>25) ver- deconvolution and specificity profiling as- profiler Cell Lysates...... 16 sus the DUB panel. In this service we can says to help characterize the hits from these offer high precision and accuracy ultra-low screens. A number of our assays have been volume compound dispensing (down to 2.5 developed from first principles starting with nl) to enable addition of the assay reagents Please visit us at little or no data from the literature to guide the directly to the compound in 100% DMSO. This www.ubiquigent.com construction of the assay. Ubiquigent can re- is particularly suited to screens where com- for the latest information on search, design, manage and deliver the entire pound solubility is a concern. new products and services. programme to help you to achieve success in your ubiquitin system HTS campaigns.

Ubiquigent Ltd - Linking Ubiquitin Research to Drug Discovery www.ubiquigent.com Protein Ubiquitylation: Ubiquitylation, like phosphoryla- The Ubiquitin Cascade tion, describes a reversible post- Product Categories are web-interactive translational protein modification. Kinases Ubiquitylation or ‘ubiquitination’ may control the protein substrate’s destiny – in respect of its turnover – or its signalling functionality. It E1 E2 E3 DCE is a process that refers to the covalent attachment - through it’s C-terminus - of a small, 76 amino Ub/Ubl Anti-Ub-conjugate acid protein called ubiquitin to the epsilon-amino group of a lysine Anti-Ub-conjugate residue residing within a substrate DCE protein – which may also be anoth- er ubiquitin molecule. This results in either mono- or poly-ubiquity- UBP lation of the substrate; the latter being where chains of ubiquitin are attached to the substrate protein. Ubiquitin may also form... DCE: deconjugating enzyme Ub: ubiquitin Ubl: ubiquitin-like protein Read more: www.ubiquigent.com/products

Therapeutic Areas and the Ubiquitin System

A growing number of excellent reviews are being published, disease mechanisms, and possibly a new source of drug dis- which offer well-referenced summaries of work in particular covery targets to consider, we present a guide and a resource to disease areas, pathways or amongst the members of certain enable you to begin identifying and grouping such associations. ubiquitin system protein families. For researchers considering the ubiquitin system as a potential new route to investigating Read more: ubiquigent.com/resources/ubiquitin-and-therapeutic-areas

2 Ubiquigent Ltd - Linking Ubiquitin Research to Drug Discovery www.ubiquigent.com Cell Lines and Kits

Through close collaborations with leading academic laboratories Ubiquigent offers specialist cell lines to enable and support ubiquitin system research. 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.

Description Source Cat. # Size Price

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

Ubiselect™ Kit NEW

Key Utility and Applications: What does the Ubiselect Kit contain? • High affinity and selective capture of ubiquitylated proteins from The kit contains control Ubiselect-MEF_BirA (P0) cell line (Cat# Mouse Embryonic Fibroblast (MEF) cells 66-5010-001), Ubiselect-MEF_Bio-Ub (P0) cell line (Cat# 66- 5011-001), and an anti-Mono and Polyubiquitylated conjugates • High sensitivity detection of ubiquitylated proteins only and NOT other antibody (Cat# 68-0122-025) for detection of your ubiquitylated interacting proteins protein. Also included in the kit is a protocol insert which pro- • Cell-lysate based target specific ubiquitylation assays vides you with culture conditions and a detailed capture protocol for the isolation of ubiquitylated proteins from the cell lines. 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 Ubiselect Kit experiments facilitate: N-terminal biotinylation signal (which is biotinylated by co-expression of the BirA enzyme). By employing a powerful capture system which utilises • Selective isolation of ubiquitylated proteins from Ubiselect- the transgenic MEF cell line expressing ubiquitin - which is biotinylated MEF_Bio-Ub cells through the removal of ubiquitin binding in the cell - and the high affinity biotin-avidin interaction alongside a high proteins and ubiquitylated protein binding proteins that inter- stringency washing protocol you can capture your ubiquitylated target(s) act with your captured ubiquitylated protein of interest. of interest without any associated ubiquitin binding proteins and/or ubiquitylated protein binding proteins. The Ubiselect Kit contains an aliquot of • The manipulation of Ubiselect-MEF_Bio-Ub cells as per ex- each of the control Ubiselect-MEF_BirA (P0) and the Ubiselect-MEF_Bio-Ub isting MEF cell protocols. Identification of mono or poly- (P0) cell lines and an anti-ubiquitin conjugate antibody for the detection ubiquitin attached to your protein of interest. of ubiquitylated proteins. Eluted captured products may be analysed by Western blotting using the ubiquitin conjugate specific antibody, an anti- • Exploration of the formation of E1, E2, E3 and substrate biotin antibody or an antibody specific to your protein of interest. ubiquitin-conjugates. • Developing cell or cell-lysate based target specific ubiquity- Transgenes in transgenic mouse from which lation assays (eg FRET) utilising the biotin on the ubiquitin Ubiselect-MEF_BirA (P0) and Ubiselect-MEF_Bio-Ub (P0) have been derived attached to your protein of interest.

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

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

Figure 1a: Ubiselect™-MEF_Bio-Ub (P0): This Figure 1b: Ubiselect™-MEF_BirA (P0): This is a hemizygous cell line which expresses is a hemizygous cell line which expresses Reference: three moieties of biotin-accepting ubiquitin the E. coli enzyme BirA. (Lectez et al., 2014) Lectez B, Migotti R, Lee SY, Ramirez J, Beraza N et al. (2014) plus the E. coli enzyme BirA. (Lectez et al., 2014) Journal of Proteome Research 13, 3016-3026.

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

HTS Assay 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.

Description Cat. # Format Price

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.

USP14 Assay Kits

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 determine the optimal ratio of USP14:26S Proteasome [Ub-VS treated] appropriate for your assay. The Activated USP14 DUB assay Kit comprises Ubiquigent’s pre-formulated product USP14 & 26S Proteasome [Ubiquitin-Vinyl Sulfone (Ub-VS) treated] (Cat# 64-1010-096) at an optimised molar ratio ready for use in a USP14 deubiquitylase assay.

Description Cat. # Format Price

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

* 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 commercial product, or use of the AviTag™ technology to facilitate or advance research which will be applied to the development of a commercial product requires a license from Avidity, LLC. Examples of Commercial use include, but are not limited to biosensors, diagnostics, therapeutic applications, 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].

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

E2scan™ Kit version 2 - 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 reaction. 1 2 3 4 5 6 7 8 9 10 11 12 • The linkage type can determine the fate of the ubiquitylated substrate; for example to be degraded or to function in a sig- A 2A 2B 2C 2D1 2D2 2D3 2D4 2E1 2E2 2E3 2F 2G1 Kit Enzymes nalling capacity (Ye and Rape (2009)). B 2G2 2H 2I 2J1 2J2 2K 2L3 2L6 2M 2N 2N/2V1 2N/2V2 scan Kit Controls • Discovery of novel aspects to E2 functionality: The E2 kit C 2Q 2Q2 2R1 2R2 2S 2T 2V1 2V2 2W 2Z was used to identify E2s that partner RNF4 in polySUMO BFR Buffer D ubiquitylation and it was discovered that Ube2W (Ubc16) is 2D4 Positive Control an E2 with - a previously unknown - specific protein amino E 2A 2B 2C 2D1 2D2 2D3 2D4 2E1 2E2 2E3 2F 2G1 terminal monoubiquitylation activity (Tatham et al. 2013). F 2G2 2H 2I 2J1 2J2 2K 2L3 2L6 2M 2N 2N/2V1 2N/2V2 scan G 2Q 2Q2 2R1 2R2 2S 2T 2V1 2V2 2W 2Z E2 experiments facilitate:

BFR BFR 2D4 2D4 H • 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 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 • The development of assays for the E3 ligase

E3 ligase, ubiquitin, ATP, assay buffer) required to perform a complete substrate References ubiquitylation or auto-ubiquitylation assay using your E3 ligase and/or E3 ligase/ Ye and Rape (2009) Nat Rev Mol Cell Biol 10, 755-764. substrate combination of choice. Through the addition of a substrate mix (E1, E3, Tatham MH, Plechanovová A, Jaffray EG, Salmen H, Hay RT (2013) Biochem J 453, 137-145. 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. 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 select C USP27x USP28 USP30 USP35 USP36 USP45 CYLD UCHL1 UCHL3 UCHL5 BFR AMSH ter capture from a cell lysate (refer to Ubiquigent Ubi for capturing ubiquitylated proteins) D BAP1 A20 Cezanne OTU1 OTUB1 OTUB2 OTUD3 OTUD5 OTUD6A OTUD6B BFR OTUB1 Negative Control

AMSH AMSH-LP • Investigate DUB ubiquitin-ubiquitin linkage cleavage speci- E OTULIN TRABID ATXN-3 ATXN-3L JOSD1 JOSD2 AMSH AMSH-LP + Zn + Zn BFR User Controls ficity (refer to Ubiquigent di-ubiquitin and ubiquitin chains F BFR Buffer for use as substrates) G User DUBs • Examine the relative position of cleavage within ubiquitin H Suggested empty wells to chains (distal versus proximal) enable user to run DUBscan samples on 12-well gels • Explore mono and poly-deubiquitylation DUB specificity (refer to specific Ubiquigent E2s – such as Ube2W – for mono- The kit contains a DUBscan plate (a panel of DUBs arrayed across 47 wells of a 96 ubiquitylating E2s) well plate) plus a control ubiquitylation reaction mix for performing both positive • Explore potential DUB inhibitors and/or activators and negative control deubiquitylation reactions. • Identify novel DUB binding proteins In respect of one kit application - the identification of DUB enzymes which catal- • Explore how DUBs may interact with and modify the activity of yse the deubiquitylation of a substrate protein; through the addition of your own other ubiquitin system proteins such as E2s and vice versa 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. DUBscan Kit Cat. # 67-0006-001 1 Kit £245

5 Ubiquigent Ltd - Linking Ubiquitin Research to Drug Discovery www.ubiquigent.com E1 Activating Enzymes

E1 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 covalent thioester linkage 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.

Description Alternate Name Accession Number Tag Cat. # Size Price 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 E2 Conjugating Enzymes

E2 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.

Description Alternate Name Accession Number Tag Cat. # Size Price 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...

6 Ubiquigent Ltd - Linking Ubiquitin Research to Drug Discovery www.ubiquigent.com E2 Conjugating Enzymes continued...

Description Alternate Name Accession Number Tag Cat. # Size Price 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 E2 Conjugating Enzymes continued... GST 62-0029-020 20 µg £65 62-0029-100 100 µg £135

7 Ubiquigent Ltd - Linking Ubiquitin Research to Drug Discovery www.ubiquigent.com E2 Conjugating Enzymes continued...

Description Alternate Name Accession Number Tag Cat. # Size Price 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

E2 Conjugating Enzymes continued... 62-0049-100 100 µg £135

8 Ubiquigent Ltd - Linking Ubiquitin Research to Drug Discovery www.ubiquigent.com E2 Conjugating Enzymes continued...

Description Alternate Name Accession Number Tag Cat. # Size Price 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

9 Ubiquigent Ltd - Linking Ubiquitin Research to Drug Discovery www.ubiquigent.com E3 Ligases

E3 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 ligases, Homology to E6AP C-Terminus (HECT) domain E3 ligases, Cullin Ring Ligases (CRLs) and Really Interesting New Gene (RING) domain E3 ligases.

Description Alternate Name Accession Number Tag Cat. # Size Price CRL 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

DCUN/DCNL 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

HECT 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

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

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

RING/RBR 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 Q8ND25 GST 63-0040-025 25 µg £295 protein 1 ZNRF2 NEW Zinc and ring finger Q8NHG8 GST 63-0041-025 25 µg £295 protein 2 UBOX 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

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

Deconjugating 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).

Description Alternate Name Accession Number Tag Cat. # Size Price USP

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 £135 points

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

OTU 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

UCH 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

Deconjugating Enzymes continued...

11 Ubiquigent Ltd - Linking Ubiquitin Research to Drug Discovery www.ubiquigent.com Deconjugating Enzymes continued...

Description Alternate Name Accession Number Tag Cat. # Size Price Josephin 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

JAMM

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

Proteasome

The 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.

Description Cat. # Size Price

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

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

Ubiquitin/Ubl Modifiers, Derivatives and Substrates

Although 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.

Description Alternate Name Accession Number Tag Cat. # Size Price

Ubiquitin, Ubiquitin Chains

Ubiquitin RPS27A P62990.1 None 60-0001-010 10 mg £50 Di-Ubiquitin, K6-Linked None 60-0101-010 10 µg £105 Di-Ubiquitin, K11-Linked None 60-0102-010 10 µg £105 Di-Ubiquitin, K27-Linked None 60-0103-010 10 µg £105 Di-Ubiquitin, K29-Linked None 60-0104-010 10 µg £105 Di-Ubiquitin, K33-Linked None 60-0105-010 10 µg £105 Di-Ubiquitin, K48-Linked None 60-0106-010 10 µg £50 None 60-0106-050 50 µg £125 Di-Ubiquitin, K63-Linked None 60-0107-010 10 µg £50 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

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

12 Ubiquigent Ltd - Linking Ubiquitin Research to Drug Discovery www.ubiquigent.com Ubiquitin/Ubl Modifiers, Derivatives and Substrates continued...

Description Alternate Name Accession Number Tag Cat. # Size Price Fluorogenic Substrates, Biotin Conjugates

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

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

Ub/Ubl Binding Proteins

Ubiquitin 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.

Description Alternate Name Accession Number Tag Cat. # Size Price

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

Substrates

A substrate molecule is modified by enzymes that catalyse chemical reactions involving the substrate. Ubiquitylation is a post translational modification 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.

Description Alternate Name Accession Number Tag Cat. # Size Price

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

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

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. 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.

Description Alternate Name Accession Number Tag Cat. # Size Price

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/ 6His/none/ 66-0041-050 50 µg £215 AAH00358 none

AMPKA2/AMPKB2/AMPKG1 NEW NP_006243/NP_005390/ 6His/none/ 66-0042-050 50 µg £215 [pAb available - p. 15] AAH00358 none

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

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

Ubiquigent 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 polyubiquitylated 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 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.

* This antibody was developed and validated by the Key: Medical Research Council Protein Phosphorylation and WB = Western Blotting; IP = Immunoprecipitation; Ubiquitylation Unit (University of Dundee, Dundee, UK). Hu = Human; Mo = Mouse; Ch = Chicken; Rt = Rat

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

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

DCNL4 (human; full length), pAb* NEW DCUN1D4 WB, IP / Hu 68-0008-100 100 µg £215

DCNL5 (human; full length), pAb* NEW DCUN1D5 WB, IP / Hu 68-0010-100 100 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 pSer472 (mouse; residues 466 - 478), pAb* NEW PPP1R12A WB / Hu, Mo 68-0044-100 100 µg £215

MYPT1 pSer909 (mouse; residues 903 - 917), pAb* NEW PPP1R12A WB / Hu, Mo 68-0046-100 100 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

Antibodies continued...

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

Description Alternate Name Tested Applications / Reactivity 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

Cell Lysates

HEK293 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).

Description Cat. # Size Price HEK293 cell lysate 66-3002-010 10 mg £222

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

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