UCHL1 [6His-tagged]


Catalogue Number
64-0007-050
Product Size
50 µg
Price £
£330
Accession Number
AAH00332.1
Residues Expressed
1-223
Certificate of Analysis Size
50 µg
Species
Human
Source
E. coli expression
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
~27 kDa
Stability
12 months at -70°C; aliquot as required
Protein Sequence
Accession number: AAH00332.1. For full protein sequence information download the Certificate of Analysis pdf.
QA; Protein Identification
Confirmed by mass spectrometry.
QA Activity

Deubiquitylating Enzyme Assay: The activity of His-UCHL1 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 His-UCHL1 was compared confirming the deubiquitylating activity of His-UCHL1.


Background

The deubiquitylating enzymes (DUBs) 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 protein (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 enzyme the cysteine proteases and metalloproteases. UCHL1 is a cysteine protease and member of the UCH family of ubiquitin C-terminal hydrolases. Cloning of the human UCHL1 gene was first described by Day and Thompson (1987). UCHL1 contains a catalytic triad consisting of a cysteine (Cys90), a histidine (His161), and an aspartate (Asp176) residue. The overall structure of UCHL1 is very similar to that of its nearest UCH relative UCHL3, with which it shares 51% sequence identity (Das et al., 2006). UCHL1 is expressed predominantly in neurons, testis and ovary (Osaka et al., 2003). In vivo UCHL1 has been shown to be involved in the regulation of the ubiquitin pool, apoptosis, learning and memory, and its absence in mice because of spontaneous intragenic deletions yields phenotypes with neurological defects (Saigoh et al., 1999). Mutations in UCHL1 have been implicated in Parkinson disease (PD); a point mutation near the active site that changes Ile93 to Met (I93M) has been linked to an increased risk of developing an autosomal-dominant form of PD (Leroy et al., 1998).


References

Das C, Hoang QQ, Kreinbring CA, Luchansky SJ, Meray RK, Ray SS, Lansbury PT, Ringe D, Petsko GA (2006) Structural basis for conformational plasticity of the Parkinson's disease-associated ubiquitin hydrolase UCH-L1. Proc Natl Acad Sci USA 103, 4675-80.

Day IN, Thompson RJ (1987) Molecular cloning of cDNA coding for human PGP 9.5 protein. A novel cytoplasmic marker for neurones and neuroendocrine cells. FEBS Lett 210, 157-60.

Komander D, Clague MJ, Urbe S (2009) Breaking the chains: structure and function of the deubiquitinases. Nat Rev Mol Cell Biol 10, 550-63.

Leroy E, Boyer R, et al. (1998) The ubiquitin pathway in Parkinson's disease. Nature 395, 451-2.

Osaka H, Wang YL, et al. (2003) Ubiquitin carboxy-terminal hydrolase L1 binds to and stabilizes monoubiquitin in neuron. Hum Mol Genet 12, 1945-58.

Saigoh K, Wang YL, et al. (1999) Intragenic deletion in the gene encoding ubiquitin carboxy-terminal hydrolase in gad mice. Nat Genet 23, 47-51.