Linking Ubiquitin Research to Drug Discovery

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  • Name
    Catalogue Number
    Size
    Price
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  • Name:
    TRABID CD(245-697) [6His-tagged]
    Catalogue Number:
    64-0049-050
    Size:
    50 µg
    Price:
    £195
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  • Species
    human
  • Source
    E. coli
  • Quantity
    50 µg
  • Storage
    -70°C
  • Concentration
    0.5 mg/ml
  • Formulation
    50 mM HEPES pH 7.5, 150 mM sodium chloride, 2 mM dithiothreitol, 10% glycerol
  • Molecular Weight
    ~54.8kDa
  • Stability
    12 months at -70°C; aliquot as required
  • Protein Sequence
    Accession number: NP_060050. For full protein sequence information download the Certificate of Analysis pdf.
  • QA; Protein Identification
    Confirmed by mass spectrometry.
  • QA; Activity
    Deubiquitylase Enzyme Assay: The activity of His-TRABID CD(245-697) was validated by the monitoring of mono-ubiquitin generation as a result of the enzyme catalysed cleavage of K63-linked di-ubiquitin. Incubation of the substrate in the presence or absence of His-TRABID CD(245-697) was compared confirming the deubiquitylating activity of His-TRABID CD(245-697).

Deconjugating enzymes (DCEs) are proteases that process ubiquitin or ubiquitin-like gene products, reverse the modification of proteins by a single ubiquitin or ubiquitin-like protein (UBL) and remodel polyubiquitin (or poly-UBL) chains on target proteins (Reyes-Turcu et al., 2009). The deubiquitylating – or deubiquitinating – enzymes (DUBs) represent the largest family of DCEs and regulate ubiquitin dependent signalling pathways. The activities of the DUBs include the generation of free ubiquitin from precursor molecules, the recycling of ubiquitin following substrate degradation to maintain cellular ubiquitin homeostasis and the removal of ubiquitin or ubiquitin-like proteins (UBL) modifications through chain editing to rescue proteins from proteasomal degradation or to influence cell signalling events (Komander et al., 2009). There are two main classes of DUB, cysteine proteases and metalloproteases. TRABID is a cysteine protease and is a member of the OTU (ovarian tumour) superfamily of proteins (Balakirev et al., 2003). Cloning of the human gene was first described by Evans et al. (1992). TRABID was recently reported to specifically and positively regulate the Wnt signalling pathway. TRABID is composed of a tumour necrosis factor receptor associated factor (TRAF) binding domain in the C-terminus and three Zinc-finger (ZnF) motifs at the N-terminus. TRABID preferentially binds to K63-linked polyubiquitin chains but not K48-linked polyubiquitin chains and specifically cleaves K63 chains. K63-linked ubiquitylation is known to fulfil diverse proteasome-independent roles, including DNA repair, endocytosis and NFκB signalling (Shi et al., 2012). Proteins that mediate specific assembly and disassembly of atypical K6, K27, K29 and K33 linkages are mainly unknown. Recent work on these atypical di ubiquitins has shown that TRABID specifically hydrolyses both K29 and K33-linked di-ubiquitin (Licchesi et al., 2012) and in fact cleaves the K29 linkage with 40-fold more efficiency than the K63 linkage (Virdee et al., 2010).

References:

Komander D, Clague MJ and Urbe S (2009) Breaking the chains: structure and function of the deubiquitinases. Nat Rev Mol Cell Biol 10, 550-563.
Licchesi JD, Mieszczanek J, Mevissen TE, Rutherford TJ, Akutsu M, Virdee S, et al. (2012) An ankyrin-repeat ubiquitin-binding domain determines TRABID’s specificity for atypical ubiquitin chains. Nature Structural & Molecular Biology 19, 62-71.
Nakamura T, Hillova J, Mariage-Samson R, Onno M, Huebner K, Cannizzaro LA, et al. (1992) A novel transcriptional unit of the tre oncogene widely expressed in human cancer cells. Oncogene 7, 733-741
Reyes-Turcu FE, Ventii KH and Wilkinson KD (2009) Regulation and cellular roles of ubiquitin-specific deubiquitinating enzymes. Ann Rev Biochem 78, 363-397.
Shi T, Bao J, Wang NX, Zheng J and Wu D (2012) Identification Of Small Molecule TRABID Deubiquitinase Inhibitors By Computation- Based Virtual Screen. BMC Chem Biol 12, 4.
Virdee S, Ye Y, Nguyen DP, Komander D and Chin JW (2010) Engineered diubiquitin synthesis reveals Lys29-isopeptide specificity of an OTU deubiquitinase. Nature Chemical Biology 6, 750-757.