Characterization of the ubiquitin-specific protease activity of the mouse/human Unp/Unph oncoprotein.
The ubiquitin-specific proteases (Ubps) are a family of largely dissimilar enzymes with two major conserved sequence regions, containing either a conserved cysteine residue or two conserved histidine residues, respectively. The murine Unp oncoprotein and its human homologue, Unph, both contain regions similar to the conserved Cys and His boxes common to all the Ubps. In this study we show that Unp and Unph are active deubiquitinating enzymes, being able to cleave ubiquitin from both natural and engineered linear ubiquitin-protein fusions, including the polyubiquitin precursor. Mutation of the conserved Unp Cys and His residues abolishes this activity, and identifies the likely His residue in the catalytic triad. Unp is tumorigenic when overexpressed in mice, leading to the suggestion that Unp may play a role in the regulation of ubiquitin-dependent protein degradation. We have demonstrated here that the high-level expression of Unp in yeast does not disrupt the degradation of the N-end rule substrate Tyr-beta-galactosidase (betagal), the non-N-end rule substrate ubiquitin-Pro-betagal, or the degradation of abnormal, canavanine-containing proteins. These data suggest that Unp is not a general modulator of ubiquitin-dependent proteolysis. However, Unp may have a role in the regulation of the degradation of a specific, as yet undescribed, substrate(s).
Gilchrist
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Molecular Genetics Group, John Curtin School of Medical Research, Australian National University, GPO Box 334, ACT 2601, Canberra, Australia.
Baker
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D000818:Animals; D002172:Canavanine / Q000378:metabolism / Q000494:pharmacology; D018076:DNA, Complementary / Q000096:biosynthesis; D010450:Endopeptidases / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics; D005786:Gene Expression Regulation; D006801:Humans; D051379:Mice; D016297:Mutagenesis, Site-Directed; D015513:Oncogene Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D010957:Plasmids; D011518:Proto-Oncogene Proteins; D012441:Saccharomyces cerevisiae / Q000235:genetics; D013379:Substrate Specificity; D043222:Ubiquitin Thiolesterase; D064570:Ubiquitin-Specific Proteases; D001616:beta-Galactosidase / Q000378:metabolism
D018076:DNA, Complementary; D015513:Oncogene Proteins; D011518:Proto-Oncogene Proteins; C486203:USP4 protein, human; C082325:Usp4 protein, mouse; D002172:Canavanine; D001616:beta-Galactosidase; D010450:Endopeptidases; D043222:Ubiquitin Thiolesterase; D064570:Ubiquitin-Specific Proteases