USP28 [GST-tagged]


Catalogue Number
64-0020-050
Product Size
50 µg
Price £
£325
Accession Number
NP_065937
Residues Expressed
1-1077
Certificate of Analysis Size
50 µg
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
~149 kDa
Stability
12 months at -70°C; aliquot as required
Protein Sequence
Accession number: NP_065937. 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 GST-USP28 was validated by determining the increase in fluorescence measured as a result of the enzyme catalysed cleavage of the fluorogenic substrate Ubiquitin-Rhodamine110-Glycine generating Ubiquitin and Rhodamine110-Glycine. Incubation of the substrate in the presence or absence of GST-USP28 was compared confirming the deubiquitylating activity of GST-USP28.


Background

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 signaling 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. Ubiquitin carboxyl-terminal hydrolase 28 (Ubiquitin Specific Protease 28; USP28) is a member of the cysteine protease enzyme family and cloning of the human gene was first described by Nagase et al. (2000). USP28 shares 51.4% identity with USP25 and is highly expressed in the heart and skeletal muscle (Valero et al., 2001). Both USP28 and Checkpoint kinase Chk2 are required for DNA-damage-induced apoptosis, and they accomplish this partly through regulation of the p53 induction of proapoptotic genes. USP28 is required to stabilize Chk2, p53 binding protein 53BP1 and claspin in response to DNA double strand breaks (Zhang et al., 2006). USP28 stabilizes the MYC proto oncogene by antagonising the activity of the SCF-FBW7 ubiquitin ligase complex. MYC is a regulator of cell growth, proliferation and apoptosis, and enhanced levels are thought to contribute to the genesis of human tumours such as breast and colon carcinomas. Inhibition of USP28 may be a pharmacologically feasible approach to interfere with MYC function in human tumours (Popov et al., 2007).


References

Komander D, Clague MJ, Urbe S (2009) Breaking the chains: structure and function of the deubiquitinases. Nat Rev Mol Cell Biol 10, 550-563.

Nagase T, Kikuno R, Ishikawa K, Hirosawa M, Ohara O (2000) Prediction of the coding sequences of unidentified human genes. XVII. The complete sequences of 100 new cDNA clones from brain which code for large proteins in vitro. DNA Res 7, 143-150.

Popov N, Wanzel M, Madiredjo M, Zhang D, Beijersbergen R, Bernards R, Moll R, Elledge SJ, Eilers M (2007) The ubiquitin-specific protease USP28 is required for MYC stability. Nat Cell Biol 9, 765-774.

Reyes-Turcu FE, Ventii KH, Wilkinson KD (2009) Regulation and cellular roles of ubiquitin-specific deubiquitinating enzymes. Ann Rev Biol 78, 363-397.

Valero R, Bayes M, Francisca Sanchez-Font M, Gonzalez-Angulo O, Gonzalez-Duarte R, Marfany G (2001) Characterization of alternatively spliced products and tissue-specific isoforms of USP28 and USP25. Genome Biol 2, RESEARCH0043. Zhang D, Zaugg K, Mak TW, Elledge SJ (2006) A role for the deubiquitinating enzyme USP28 in control of the DNA-damage response. Cell 126, 529-542.