Structure of papain-like protease from SARS-CoV-2 and its complexes with non-covalent inhibitors

Abstract

The number of new cases world-wide for the COVID-19 disease is increasing dramatically, while efforts to contain Severe Acute Respiratory Syndrome Coronavirus 2 is producing varied results in different countries. There are three key SARS-CoV-2 enzymes potentially targetable with antivirals: papain-like protease (PLpro), main protease (Mpro), and RNA-dependent RNA polymerase. Of these, PLpro is an especially attractive target because it plays an essential role in several viral replication processes, including cleavage and maturation of viral polyproteins, assembly of the replicase-transcriptase complex (RTC), and disruption of host viral response machinery to facilitate viral proliferation and replication. Moreover, this enzyme is conserved across different coronaviruses and promising inhibitors have already been discovered for its SARS-CoV variant. Here we report a substantive body of structural, biochemical, and virus replication studies that identify several inhibitors of the enzyme from SARS-CoV-2 in both wild-type and mutant forms. These efforts include the first structures of wild-type PLpro, the active site C111S mutant, and their complexes with inhibitors, determined at 1.60–2.70 Angstroms. This collection of structures provides fundamental molecular and mechanistic insight to PLpro, and critically, illustrates details for inhibitors recognition and interactions. All presented compounds inhibit the peptidase activity of PLpro in vitro, and some molecules block SARS-CoV-2 replication in cell culture assays. These collated findings will accelerate further structure-based drug design efforts targeting PLpro, with the ultimate goal of identifying high-affinity inhibitors of clinical value for SARS-CoV-2.

Molecular docking studies of some selected gallic acid derivatives against five non-structural proteins of novel coronavirus

“…The substrate binding site in PLpro is made up of a canonical cysteine protease catalytic triad (Cys111, His272, and Asp286) (Fig. 1 ) found at the interface between the thumb and palm domains; other amino acids in the binding site are Trp106, Gly266, Gly271, His73, Arg140, and Asn109 [ 7 ]. The in silico docking of PLp with gallic acid derivatives revealed that 3-O-(6-galloylglucoside), 4-O-(6-galloylglucoside), and remdesivir had the utmost binding affinity scores of −6.6, −6.8, and −6.5 kcal/mol respectively (Table 4 ).…”

“…The substrate binding domain is made up of a canonical cysteine protease catalytic triad (Cys111, His272, and Asp286) (Fig. 1 ) found at the interface between the thumb and palm domains; other amino acids in these domain are Trp106, Gly266, Gly271, His73, Arg140, and Asn109 [ 7 ]. It is noteworthy that none of the hit compounds had interactions with the catalytic triad (Cys111, His272, and Asp286) and might be the reason for the characteristics BE values exhibited by them (Fig.…”

“…Main protease (Mpro/nsp5), papain-like protease (Plp/nsp3), RNA-dependent RNA polymerase (RdRp/nsp12), helicase (nsp13), and uridylate-specific endoribonuclease (NendoU/nsp15) of SARS CoV 2 were selected for this study. The 3D crystal structures of the receptors were retrieved from the Protein Data Bank (PDB) ( www.pdb.org/pdb ) with PDB IDs of 6 LU7 [ 8 ], 6W9C [ 7 ], 6 M71 [ 25 ], 6XEZ [ 9 ], and 6VWW [ 10 ] respectively. Chain A of the protein receptors was selected and cleaned from heteroatoms, then prepared for docking using the Dock preparation tool of UCSF-Chimera© (version 1.13) software ( http://www.cgl.ucsf.edu/chimera ).…”

Drug repurposing screen identifies masitinib as a 3CLpro inhibitor that blocks replication of SARS-CoV-2in vitro

“…This cleavage is mediated by two virally encoded proteases: the main viral protease, known as Mpro, 3CLpro or non-structural protein 5 (nsp5) and a second protease known as the papain-like protease, PLpro, a domain within nsp3 3 . There is much interest in developing de-novo inhibitors to target these proteases [5][6][7][8][9][10] but the full clinical development of such drugs is a time-consuming process, usually requiring several years.…”

Interaction of small molecules with the SARS-CoV-2 papain-like protease: In silico studies and in vitro validation of protease activity inhibition using an enzymatic inhibition assay

“…Based on the molecular docking results for 6WUU and the blind docking data, the receptor grid was adjusted for hypericin and its isomer. The residues that were used included K157, E167, D164, Y273, T301, Y264, P247, P248, Y268 and Q269 ( 52 , 96 ). In regards to 7JRN, the positive control compound GRL-0617 was docked using the QPLD protocol ( 90 , 91 , 92 , 93 , 94 , 95 ).…”