Selectivity in ISG15 and ubiquitin recognition by the SARS coronavirus papain-like protease.
The severe acute respiratory syndrome coronavirus papain-like protease (SARS-CoV PLpro) carries out N-terminal processing of the viral replicase polyprotein, and also exhibits Lys48-linked polyubiquitin chain debranching and ISG15 precursor processing activities in vitro. Here, we used SDS-PAGE and fluorescence-based assays to demonstrate that ISG15 derivatives are the preferred substrates for ... the deubiquitinating activity of the PLpro. With k(cat)/K(M) of 602,000 M(-1)s(-1), PLpro hydrolyzes ISG15-AMC 30- and 60-fold more efficiently than Ub-AMC and Nedd8-AMC, respectively. Data obtained with truncated ISG15 and hybrid Ub/ISG15 substrates indicate that both the N- and C-terminal Ub-like domains of ISG15 contribute to this preference. The enzyme also displays a preference for debranching Lys48- over Lys63-linked polyubiquitin chains. Our results demonstrate that SARS-CoV PLpro can differentiate between ubiquitin-like modifiers sharing a common C-terminal sequence, and that the debranching activity of the PLpro is linkage type selective. The potential structural basis for the demonstrated specificity of SARS-CoV PLpro is discussed.
Mesh Terms:
Binding Sites, Cysteine Endopeptidases, Cytokines, Enzyme Activation, Protein Binding, Substrate Specificity, Ubiquitin, Ubiquitins, Viral Proteins
Binding Sites, Cysteine Endopeptidases, Cytokines, Enzyme Activation, Protein Binding, Substrate Specificity, Ubiquitin, Ubiquitins, Viral Proteins
Arch. Biochem. Biophys.
Date: Oct. 01, 2007
PubMed ID: 17692280
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