The Differential Modulation of USP Activity by Internal Regulatory Domains, Interactors and Eight Ubiquitin Chain Types

Division of Biochemistry and Center for Biomedical Genetics, The Netherlands Cancer Institute, Plesmanlaan 121, 1066 CX Amsterdam, The Netherlands.
Chemistry & biology (Impact Factor: 6.65). 12/2011; 18(12):1550-61. DOI: 10.1016/j.chembiol.2011.10.017
Source: PubMed


Ubiquitin-specific proteases (USPs) are papain-like isopeptidases with variable inter- and intramolecular regulatory domains. To understand the effect of these domains on USP activity, we have analyzed the enzyme kinetics of 12 USPs in the presence and absence of modulators using synthetic reagents. This revealed variations of several orders of magnitude in both the catalytic turnover (k(cat)) and ubiquitin (Ub) binding (K(M)) between USPs. Further activity modulation by intramolecular domains affects both the k(cat) and K(M), whereas the intermolecular activators UAF1 and GMPS mainly increase the k(cat). Also, we provide the first comprehensive analysis comparing Ub chain preference. USPs can hydrolyze all linkages and show modest Ub-chain preferences, although some show a lack of activity toward linear di-Ub. This comprehensive kinetic analysis highlights the variability within the USP family.

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Available from: Dharjath Ahamed Shahul Hameed, Aug 04, 2014
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    • "In contrast, loss of USP8 had no impact on, for example, mitochondrial dynamics, depolarization or PINK1 accumulation, further supporting that USP8 controls parkin activity via deubiquitination of the E3 ligase. A key finding of the paper is that USP8 selectively removed K6-linked ubiquitin chains from parkin, although it shows no preference for this linkage type when processing unattached ubiquitin dimers in vitro (Faesen et al, 2011). Expression of an ubiquitin mutant that cannot assemble K6-linked ubiquitin conjugates (Ub K6R ) in USP8- depleted cells was able to rescue delayed parkin recruitment to damaged mitochondria and impaired mitophagy as compared to wildtype ubiquitin. "
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    • "In contrast to the characterized USPs, which have promiscuous linkage preferences (Faesen et al., 2011), the H. sapiens OTU DUBs have more strict linkage specificities (Mevissen et al., 2013). The distinct linkage specificities associated with OTU DUBs could be exploited in restriction analyses to determine the linkage types of the ubiquitin chains conjugated on endogenous substrates (Fiil et al., 2013; Hospenthal et al., 2013; Mevissen et al., 2013), to purify ubiquitylated substrates with specific linkage types, and to assemble UB chains with specific linkage(s) (Bremm et al., 2010). "
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