OTULIN [GST-tagged]


Catalogue Number
64-0048-050
Product Size
50 µg
Price £
£330
Accession Number
NP_612357
Residues Expressed
1-352
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
~67kDa
Stability
12 months at -70°C; aliquot as required
Protein Sequence
Accession number: NP_612357. 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-OTULIN was validated by the monitoring of mono-ubiquitin generation as a result of the enzyme catalysed cleavage of linear di-ubiquitin. Incubation of the substrate in the presence or absence of GST-OTULIN was compared confirming the deubiquitylating activity of GST-OTULIN.


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 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. OTULIN is a cysteine protease and a member of the OTU (ovarian tumour) superfamily of proteins (Balakirev et al., 2003). Cloning of the human gene was first described by Ota et al. (2004). OTU enzymes play important roles as negative-feedback regulators in NF-kB signalling, interferon signalling and in p97 (cdc48)-mediated processes although the cellular functions of most OTU enzymes remain to be discovered. Ovarian tumour family DUBs contain a papain-like catalytic core of ~180 amino acids. In addition to their catalytic domain, many OTU members have additional ubiquitin-binding domains (UBDs). At least 20 different UBD families have been described, and knowledge of linkage-specific UBDs have provided the means to understand the roles of different ubiquitin linkages in cells (Licchesi et al., 2012). OTULIN (OTU DUB with linear linkage specificity) is the latest OTU to be discovered. It has been proven to specifically cleave linear ubiquitin and antagonize the E3 ligase LUBAC (linear ubiquitin chain assembly complex), thereby regulating NF-κB signalling (Wiener and Wolberger, 2013). It has been shown that the overexpression of OTULIN prevents tumour necrosis factor a (TNFα)-induced NEMO (NFκB Essential Modifier) association with ubiquitylated RIPK1, suggesting that OTULIN regulates linear poly-ubiquitin signalling (Fiil et al., 2013; Keusekotten et al., 2013).


References

Balakirev MY, Tcherniuk SO, Jaquinod M and Chroboczek J (2003) Otubains: a new family of cysteine proteases in the ubiquitin pathway. EMBO Rep 4, 517-522.

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.

Ota T, Suzuki Y, Nishikawa T, Otsuki T, Sugiyama T, Irie R, et al. (2004) Complete sequencing and characterization of 21,243 full-length human cDNAs. Nature Genetics 36, 40-45.

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