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

Abstract

The pandemic caused by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) continues to expand. Papain-like protease (PLpro) is one of two SARS-CoV-2 proteases potentially targetable with antivirals. PLpro is an attractive target because it plays an essential role in cleavage and maturation of viral polyproteins, assembly of the replicase-transcriptase complex, and disruption of host responses. We report a substantive body of structural, biochemical, and virus replication studies that identify several inhibitors of the SARS-CoV-2 enzyme. We determined the high resolution structure of wild-type PLpro, the active site C111S mutant, and their complexes with inhibitors. This collection of structures details inhibitors recognition and interactions providing fundamental molecular and mechanistic insight into PLpro. All compounds inhibit the peptidase activity of PLpro in vitro, some block SARS-CoV-2 replication in cell culture assays. These findings will accelerate structure-based drug design efforts targeting PLpro to identify high-affinity inhibitors of clinical value.

SARS-CoV-2 Papain-Like Protease Potential Inhibitors—In Silico Quantitative Assessment

“…It may be further divided into three subdomains: thumb, palm, and fingers (Figure 1A). The active site is located between palm and thumb, utilizing three main residues, called the catalytic triad: Cys111, His272, and Asp286 (Figure 1B) [24]. Although SARS-CoV-2 PLpro is a relatively short known protein, its great significance for the virus, as well as being an immensely valid molecular target for novel potential drugs, led to great attention on it in the scientific world.…”

“…As such, it has already been put in a spotlight by the scientific community. This includes research on its structure [24][25][26], functions [11,27], and similarity to PLpro from other related coronaviruses, most notably SARS-CoV [28]. The search for PLpro inhibitors has already begun.…”

“…The search for PLpro inhibitors has already begun. So far, most studies focus on trying to utilize previously developed noncovalent SARS-CoV inhibitors or their derivatives [7,24,29] or to design specific, covalent inhibitors [30,31]. However, covalent protease inhibitors come with risks of toxicity due to high reactivity and, in some cases, low selectivity and nonspecific binding off the target [32][33][34].…”

“…There is a considerable amount of data on such compounds with regards to SARS-CoV PLpro, which will facilitate the design of analogical inhibitors for the novel coronavirus, as these two strains share considerable similarity in terms of PLpro structure [24]. Alas, the recently developed and studied SARS-CoV-2 PLpro inhibitors exhibit only moderate binding affinity toward this enzyme [7,24,35,36]. Furthermore, they lack consideration of potential toxicity, most importantly in terms of potential off binding to similar deubiquitinating enzymes, most notably to UCH-L1.…”

Potent, Novel SARS-CoV-2 PLpro Inhibitors Block Viral Replication in Monkey and Human Cell Cultures

“…CPI-169 represents a novel chemical scaffold and a novel addition to the very limited PLpro inhibitor chemotypes identified in the literature. 33,50 Given the availability of co-crystal structures of GRL0617:PLpro (SARS-CoV-2), structure-guided drug design was pursued exploring this chemical scaffold. Five binding sites on PLpro were explored by modification of the benzamide scaffold to identify additional interactions to increase inhibitor potency.…”