K-E10

Material Data Sheet

Deconjugating Enzyme Set

Price: $295.00

Catalog #: K-E10

Deconjugating Enzyme Set

Price:

$295

Quantity:

1 kit

Data Sheet:

KE10_Deconjugating Enzyme set.pdf

Stock:

In 50mM HEPES pH 8.0, 150mM NaCl, 10% glycerol, 0.5mM EDTA, 1mM DTT

Purity:

>95% by SDS-PAGE

Background:

This kit contains the following deconjugating enzymes: Isopeptidase T (USP5, IsoT), human recombinant - Isopeptidase T is a member of the high molecular weight ubiquitin C-terminal hydrolases (UCHs). This enzyme primarily disassembles free poly-ubiquitin chains linked through isopeptide bonds. Accession # NP_003472.Ubiquitin C-terminal Hydrolase L3 (UCH-L3), human recombinant- UCH-L3 is a member of the low molecular weight UCHs that processes ubiquitin precursors and ubiquitinated proteins to generate monomeric ubiquitin. Accession # NP_005993. Ubiquitin C-terminal Hydrolase L1 (UCH-L1), human recombinant- UCH-L1 is a member of the low molecular weight UCHs that processes ubiquitin precursors and ubiquitinated proteins to generate monomeric ubiquitin. Accession # NP_004172. His6-Ataxin-3 (MJD protein 1), human recombinant- Ataxin-3 protein belongs to a novel group of cysteine proteases similar to USP-type ubiquitin proteases and has deubiquitinating activity in vitro. The full-length protein contains an N-terminal Josephin domain, two ubiquitin interacting motifs (UIMs), and a variable C-terminus consisting of a polyglutamine stretch and tail. (Accession # NP_004984). His6-A20CD (TNFAIP3 catalytic domain), human recombinant - A20 (TNFa-induced protein 3) is a cytoplasmic zinc finger protein that inhibits NFkB activity and tumor necrosis factor-mediated programmed cell death. A20 contains an N-terminal domain which has deubiquitinating enzyme activity and removes ubiquitin chains from receptor-interacting protein (RIP), thus mediating distinct regulatory effects in the down-regulation of NFkB signaling. Accession # NP_006281. His6-BAP1, human recombinant- BAP1 (BRCA1 Associated Protein 1) interacts with the RING finger domain of the E3 ligase BRCA1 (Breast Cancer1, early onset protein) which functions as a tumor suppressor in the BRCA1 growth control pathway. It has been demonstrated that BAP1 and BRCA1 associate in vivo and have overlapping sub-nuclear localization patterns. BAP1 appears to be a key regulator of the BRCA1 growth control pathway and has been proposed to be a novel candidate tumor suppressor. Accession # NP_004647. His6-USP2CD (UBP41 catalytic domain), human recombinant - USP2 (Ubiquitin Specific Protease 2) is a cysteine protease over-expressed in prostate cancer, is androgen-regulated and interacts with and prolongs the half-life of fatty acid synthase (FAS). FAS stabilization is associated with the malignancy of a subset of aggressive prostate cancers. Mdm2 is also a substrate for USP2 indicating that the enzyme may regulate p53-dependent pathways. Accession # NP_004196. His6-USP7FL (HAUSP), human recombinant- USP7 (Herpes-Associated Ubiquitin Specific Protease) is a nuclear protein that was initially identified as a novel p53-interacting protein. The enzyme deubiquitinates p53 thus stabilizing the levels of this key tumor suppressor, and inducing p53-dependent cell growth repression and apoptosis. Accession # NP_003461. His6-USP8FL (UBPY), human recombinant- USP8 (Ubiquitin Specific Protease8) is a growth-regulated deubiquitinating enzyme (DUB) with a role in endosomal sorting of receptor tyrosine kinases (RTKs) such as EGFR in vivo. USP8 has DUB activity on Ub chains in vitro and does not discriminate between K48- and K63-linked isopeptide bonds. His6-Otubain1, human recombinant - Otubains belong to the ovarian tumor (OTU) protein super-family present in eukaryotes, viruses and pathogenic bacterium. Accession # NP_060140. USP14, human recombinant- USP14 (Ubiquitin Specific Protease 14) is the human homolog of the yeast Ubp6 protein and is member a of the family of thiol proteases involved in the hydrolysis of ubiquitin C-terminal protein derivatives. It has been reported to have a low-affinity association with the proteasome (PA700, 19S cap particle), which increases its basal enzyme activity. Accession # NP_005142. <table border="0"><tbody><tr><td>Protein</td><td>MW</td><td>Quantity</td><td>Stock Concentration</td><td>Final Concentration </td></tr><tr><td>Isopeptidase T</td><td>97 kDa</td><td>5μg</td><td>X mg/ml (X μM)</td><td>0.05-5 nM</td></tr><tr><td>UCH-L3</td><td>26 kDa</td><td>5μg</td><td>X mg/ml (X μM)</td><td>0.05-5 nM</td></tr><tr><td>UCH-L1</td><td>25 kDa</td><td>5μg</td><td>X mg/ml (X μM)</td><td>0.05-5 nM</td></tr><tr><td>His6-Ataxin-3</td><td>42 kDa</td><td>5μg</td><td>X mg/ml (X μM)</td><td>1-5 μM</td></tr><tr><td>His6-A20CD</td><td>46 kDa</td><td>5μg</td><td>X mg/ml (X μM)</td><td>1-5 μM</td></tr><tr><td>His6-BAP1</td><td>81 kDa</td><td>5μg</td><td>X mg/ml (X μM)</td><td>1-5 μM</td></tr><tr><td>His6-USP2CD</td><td>42 kDa</td><td>5μg</td><td>X mg/ml (X μM)</td><td>1-5 μM</td></tr><tr><td>His6-USP7FL</td><td>128 kDa</td><td>5μg</td><td>X mg/ml (X μM)</td><td>1-5 μM</td></tr><tr><td>His6-USP8FL </td><td>131 kDa</td><td>5μg</td><td>X mg/ml (X μM)</td><td>1-5 μM</td></tr><tr><td>His6-Otubain1</td><td>33.2 kDa</td><td>5μg</td><td>X mg/ml (X μM)</td><td>1-5 μM</td></tr><tr><td>USP14</td><td>56 kDa</td><td>5μg</td><td>X mg/ml (X μM)</td><td>1-5 μM</td></tr></tbody></table>

Protein modification by ubiquitin (Ub) is a reversible process. These deconjugation reactions are performed by specific cysteine proteases which generate monomeric Ub from a variety of C-terminal adducts. These enzymes process precursor Ub forms to their mature or active protein forms, they can process linear Ub fusion proteins, they can disassemble Ub isopeptide bonds present in poly chain species, and also remove Ub modifications from protein substrates. Deubiquitinating enzymes (DUBs) are one of the largest enzyme families in the Ub system with diverse functions, making them key regulators of ubiquitin-mediated pathways and they often function by direct or indirect association with the proteasome. DUBs regulate and maintain optimal Ub cellular levels. The activity of DUBs has been implicated in several important pathways including cell growth and differentiation, development, oncogenesis, neuronal disease and transcriptional regulation. In humans, there are ~100 putative members of the DUB super-family and they are divided into 5 broad categories based on Ub-protease domain structure and mechanism of action. These include the ubiquitin-specific protease (USP), ubiquitin C-terminal hydrolase (UCH), ovarian tumor-related protease or otubain (OTU), Machado-Joseph disease (MJD) and JAB1/MPN/Mov34 metalloprotease (JAMM) enzymes. The catalytic domains contain two short and well-conserved Cys and His box motifs forming a catalytic triad with these Cys and His residues together with either an Asp or Asn. DUBs contain ubiquitin-interaction motif (UIM) domains which bind to and recognize Ub/Ub chains helping to recruit poly-ubiquitinated substrates. UCHs have a preference for small substrates and their primary role is Ub recycling from C-terminal fusions such as lysine, glutathione, or peptide remnants. High MW DUBs (such as USPs) primarily remove Ub from larger proteins and disassemble poly-Ub chains, and regulate rates of proteasomal degradation. Several DUBs function freely or in association with the 26S proteasome. DUBs also act on mono-ubiquitinated substrates regulating protein-protein interactions and signaling pathways.

Use & Storage

Use:

Final concentration provided is for use in vitro and depends on conditions and substrate.

Storage:

Store at -80°C. Avoid multiple freeze/thaw cycles.