USP2 (isoform 4) [GST-tagged]


Catalogue Number
64-0014-050
Product Size
50 µg
Price £
£330
Accession Number
NP_741994
Residues Expressed
1-396
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
~73 kDa
Stability
12 months at -70°C; aliquot as required
Protein Sequence
Accession number: NP_741994. 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-USP2 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-USP2 was compared confirming the deubiquitylating activity of GST-USP2.


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 specific protease 2 (USP2) is a member of the cysteine protease enzyme family and cloning of the human gene was first described by Baek et al. (1997). USP2 influences the nuclear translocation of NF-κB transcription factors in response to inflammation, injury and other environmental changes. In resting cells, NF-κB is bound to the inhibitory protein IκBα, and therefore maintained in an inactive state in the cytoplasm. The proteasomal degradation of IκBα is prevented by USP2 deubiquitylase activity. This regulates the expression of genes that are involved in immune responses, cell proliferation, differentiation and apoptosis (Metzig et al., 2011). Deregulation of NF-κB activity is involved in the pathology of many diseases including chronic inflammation and cancer. USP2 expression itself is frequently downregulated in breast carcinomas (Metzig et al., 2011). USP2 deubiquitylase activity induces cell death via the apoptosis inducing factor (AIF). AIF is a mitochondrial oxidoreductase that becomes truncated when the cell is under stress and the truncated AIF (tAIF) becomes translocated into the nucleus, and induces caspase-independent cell death. USP2 deubiquitylates and stabilizes tAIF, thus promoting AIF-mediated cell death. In contrast, the E3 ligase CHIP ubiquitylates and destabilizes tAIF, thus preventing cell death (Oh et al., 2011).


References

Baek SH, Choi KS, Yoo YJ, Cho JM, Baker RT, Tanaka K, Chung CH (1997) Molecular cloning of a novel ubiquitin-specific protease, UBP41, with isopeptidase activity in chick skeletal muscle. J Biol Chem 272, 25560-25565.

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

Metzig M, Nickles D, Falschlehner C, Lehmann-Koch J, Straub BK, Roth W, Boutros M (2011) An RNAi screen identifies USP2 as a factor required for TNF-alpha-induced NF-kappaB signaling. Int J Cancer 129, 607-618.

Oh KH, Yang SW, Park JM, Seol JH, Iemura S, Natsume T, Murata S, Tanaka K, Jeon YJ, Chung CH (2011) Control of AIF-mediated cell death by antagonistic functions of CHIP ubiquitin E3 ligase and USP2 deubiquitinating enzyme. Cell Death Differ 18, 1326-1336.

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