Proteolytic processing, deubiquitinase and interferon antagonist activities of Middle East respiratory syndrome coronavirus papain-like protease

Abstract

The emerging Middle East respiratory syndrome coronavirus (MERS-CoV) causes severe pulmonary disease in humans and represents the second example of a highly pathogenic coronavirus (CoV) following severe acute respiratory syndrome coronavirus (SARS-CoV). Genomic studies revealed that two viral proteases, papain-like protease (PLpro) and 3C-like protease (3CLpro), process the polyproteins encoded by the MERS-CoV genomic RNA. We previously reported that SARS-CoV PLpro acts as both deubiquitinase (DUB) and IFN antagonist, but the function of the MERS-CoV PLpro was poorly understood. In this study, we characterized MERS-CoV PLpro, which is a protease and can recognize and process the cleavage sites (CS) of nsp1-2, nsp2-3 and nsp3-4. The LXGG consensus cleavage sites in the N terminus of pp1a/1ab, which is generally essential for CoV PLpro-mediated processing, were also characterized in MERS-CoV. MERS-CoV PLpro, like human SARS-CoV PLpro and NL63-CoV PLP2, is a viral deubiquitinating enzyme. It acts on both K48-and K63-linked ubiquitination and ISG15-linked ISGylation. We confirmed that MERS-CoV PLpro acts as an IFN antagonist through blocking the phosphorylation and nuclear translocation of IFN regulatory factor 3 (IRF3). These findings indicate that MERS-CoV PLpro acts as a viral DUB and suppresses production of IFN-b by an interfering IRF3-mediated signalling pathway, in addition to recognizing and processing the CS at the N terminus of replicase polyprotein to release the non-structural proteins. The characterization of proteolytic processing, DUB and IFN antagonist activities of MERS-CoV PLpro would reveal the interactions between MERS-CoV and its host, and be applicable to develop strategies targeting PLpro for the effective control of MERS-CoV infection.

Thiopurine analogs and mycophenolic acid synergistically inhibit the papain-like protease of Middle East respiratory syndrome coronavirus

“…In addition to proteolytic activity, which is similar to that of other coronaviruses, MERS-CoV PL pro has both deubiquitination (DUB) and ISGylation (ISG = interferon-stimulated gene) activity (Yang et al, 2014). The enzyme is able to deubiquitinate interferon regulatory factor 3, which prevents its nuclear translocation; this results in a suppression of interferon b production; the end result is immune suppression of host cells (Chen et al, 2007;Clementz et al, 2010;Yang et al, 2014;Zheng et al, 2008).…”

Inhibitor Recognition Specificity of MERS-CoV Papain-like Protease May Differ from That of SARS-CoV

“…16, 21 Researchers have discovered that the MERS-PLpro also exhibits deubiquitination and de-ISGylation functions, blocking the interferon regulatory factor 3 (IRF3) pathway. 22, 23 Both 3CLpro and PLpro are known to be essential for viral replication, making them attractive targets in antiviral drug discovery. 20, 24 In this study we characterized MERS-PLpro as a drug target by solving its complete structure followed by thorough analysis with four known SARS-PLpro lead inhibitors.…”

Crystal structure of the papain-like protease of MERS coronavirus reveals unusual, potentially druggable active-site features

“…In spite of the accumulating knowledge on the essential roles of the coronavirus PL pro in virus replication and evasion of the hostcell innate immune response (Devaraj et al, 2007;Frieman et al, 2009;Yang et al, 2013;Mielech et al, 2014;Lindner et al, 2007;Clementz et al, 2010), the three-dimensional structures of only two of these enzymes have been reported so far, i.e., that of the PL pro from SARS-CoV (Ratia et al, 2006) and that of the PL1 pro from Transmissible Gastroenteritis Virus (TGEV) (Wojdyla et al, 2010). Here we present the crystal structure of the MERS-CoV PL pro at 2.50 Å resolution, in order to unravel the structural basis of the activities of the enzyme and facilitate structure-based drug design efforts.…”