Severe acute respiratory syndrome coronavirus papain-like protease: Structure of a viral deubiquitinating enzyme

Abstract

Replication of severe acute respiratory syndrome (SARS) coronavirus (SARS-CoV) requires proteolytic processing of the replicase polyprotein by two viral cysteine proteases, a chymotrypsin-like protease (3CLpro) and a papain-like protease (PLpro). These proteases are important targets for development of antiviral drugs that would inhibit viral replication and reduce mortality associated with outbreaks of SARS-CoV. In this work, we describe the 1.85-Å crystal structure of the catalytic core of SARS-CoV PLpro and show that the overall architecture adopts a fold closely resembling that of known deubiquitinating enzymes. Key features, however, distinguish PLpro from characterized deubiquitinating enzymes, including an intact zinc-binding motif, an unobstructed catalytically competent active site, and the presence of an intriguing, ubiquitinlike N-terminal domain. To gain insight into the active-site recognition of the C-terminal tail of ubiquitin and the related LXGG motif, we propose a model of PLpro in complex with ubiquitinaldehyde that reveals well defined sites within the catalytic cleft that help to account for strict substrate-recognition motifs.membrane-associated protease ͉ ubiquitin-like domain

Diarylheptanoids from <i>Alnus japonica</i> Inhibit Papain-Like Protease of Severe Acute Respiratory Syndrome Coronavirus

“…pro Inhibition Assay The PL pro inhibition assay applied report by Ratia et al 22) The inhibition assay was optimized in a 96-well plate format to establish suitable assay conditions and incubation times. The fluorogenic peptide substrate, Arg-Leu-Arg-Gly-Gly-AMC (RLRGG-AMC), was purchased from Bachem Bioscience.…”

“…The substrate concentration was 100 nM, and release of AMC was measured in the same manner as for the IC 50 measurements described above. 22) RESULTS AND DISCUSSION SARS-CoV PL pro inhibitory activity-guided fractionation of the ethanol extract led to the isolation of nine diarylheptanoids (1-9) (Fig. 1).…”

Structures of proteases for ubiqutin and ubiquitin-like modifiers

“…The overall architecture of the catalytic core domain of USP7 appears to be conserved in other members of USP family such as USP2 (38), USP8 (39), USP14 (40) and deubiquitinating enzyme from severe acute respiratory syndrome coronavirus (41). In the case of USP14, the r.m.s.d.…”

The search for a structural basis for therapeutic intervention against the SARS coronavirus

“…Nsp3 is a large multidomain protein of 1922 amino acids that is yielded by proteolytic cleavage of the pp1a polyprotein at two sites by the papain-like protease (PL pro ). Two crystal structures of the functional enzymatic domains of nsp3 have been determined: the 'X' domain with proposed ADP-ribose-1 00 -phosphate dephosphorylation (ADRP) activity (Saikatendu et al, 2005;Egloff et al, 2006) and the papain-like protease (PL pro ) domain (Ratta et al, 2006). The 'X' domain, also known as the ADRP domain, is conserved among all CoV (Putics et al, 2005) and is structurally related to macro-H2A-like fold proteins (Fig.…”