title: The DUSP-Ubl domain of USP4 enhances its catalytic efficiency by promoting ubiquitin exchange.
abstract: Ubiquitin-specific protease USP4 is emerging as an important regulator of cellular pathways, including the TGF-β response, NF-κB signalling and splicing, with possible roles in cancer. Here we show that USP4 has its catalytic triad arranged in a productive conformation. Nevertheless, it requires its N-terminal DUSP-Ubl domain to achieve full catalytic turnover. Pre-steady-state kinetics measurements reveal that USP4 catalytic domain activity is strongly inhibited by slow dissociation of ubiquitin after substrate hydrolysis. The DUSP-Ubl domain is able to enhance ubiquitin dissociation, hence promoting efficient turnover. In a mechanism that requires all USP4 domains, binding of the DUSP-Ubl domain promotes a change of a switching loop near the active site. This 'allosteric regulation of product discharge' provides a novel way of regulating deubiquitinating enzymes that may have relevance for other enzyme classes.
authors.affiliation: Department of Biochemistry, The Netherlands Cancer Institute, Plesmanlaan 121, 1066CX Amsterdam, The Netherlands.
authors.forename: Marcello
authors.initials: M
authors.lastname: Clerici
authors.affiliation: Department of Biochemistry, The Netherlands Cancer Institute, Plesmanlaan 121, 1066CX Amsterdam, The Netherlands.
authors.forename: Mark P A
authors.initials: MP
authors.lastname: Luna-Vargas
authors.affiliation: Department of Biochemistry, The Netherlands Cancer Institute, Plesmanlaan 121, 1066CX Amsterdam, The Netherlands.
authors.forename: Alex C
authors.initials: AC
authors.lastname: Faesen
authors.affiliation: Department of Biochemistry, The Netherlands Cancer Institute, Plesmanlaan 121, 1066CX Amsterdam, The Netherlands.
authors.forename: Titia K
authors.initials: TK
authors.lastname: Sixma
mesh_terms: D000494:Allosteric Regulation; D002384:Catalysis; D018360:Crystallography, X-Ray; D006801:Humans; D007700:Kinetics; D008958:Models, Molecular; D011485:Protein Binding; D025801:Ubiquitin / Q000737:chemistry / Q000378:metabolism*; D043222:Ubiquitin Thiolesterase / Q000737:chemistry* / Q000378:metabolism*; D064570:Ubiquitin-Specific Proteases
chemical_list: C486203:USP4 protein, human; D025801:Ubiquitin; D043222:Ubiquitin Thiolesterase; D064570:Ubiquitin-Specific Proteases