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Acrylamide fragment inhibitors that induce unprecedented conformational distortions in enterovirus 71 3C and SARS-CoV-2 main protease.

ABSTRACT: RNA viruses are critically dependent upon virally encoded proteases to cleave the viral polyproteins into functional proteins. Many of these proteases exhibit a similar fold and contain an essential catalytic cysteine, offering the opportunity to inhibit these enzymes with electrophilic small molecules. Here we describe the successful application of quantitative irreversible tethering (qIT) to identify acrylamide fragments that target the active site cysteine of the 3C protease (3C^pro) of Enterovirus 71, the causative agent of hand, foot and mouth disease in humans, altering the substrate binding region. Further, we re-purpose these hits towards the main protease (M^pro) of SARS-CoV-2 which shares the 3C-like fold and a similar active site. The hit fragments covalently link to the catalytic cysteine of M^pro to inhibit its activity. We demonstrate that targeting the active site cysteine of M^pro can have profound allosteric effects, distorting secondary structures to disrupt the active dimeric unit.

SUBMITTER: Qin B

PROVIDER: S-EPMC9181371 | biostudies-literature | 2022 Oct

REPOSITORIES: biostudies-literature

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Acrylamide fragment inhibitors that induce unprecedented conformational distortions in enterovirus 71 3C and SARS-CoV-2 main protease.

Qin Bo B Craven Gregory B GB Hou Pengjiao P Chesti Julian J Lu Xinran X Child Emma S ES Morgan Rhodri M L RML Niu Wenchao W Zhao Lina L Armstrong Alan A Mann David J DJ Cui Sheng S

Acta pharmaceutica Sinica. B 20220609 10

RNA viruses are critically dependent upon virally encoded proteases to cleave the viral polyproteins into functional proteins. Many of these proteases exhibit a similar fold and contain an essential catalytic cysteine, offering the opportunity to inhibit these enzymes with electrophilic small molecules. Here we describe the successful application of quantitative irreversible tethering (qIT) to identify acrylamide fragments that target the active site cysteine of the 3C protease (3C<sup>pro</sup> ...[more]

PMID: 35702321

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Project description:BackgroundEnterovirus 71 (EV71) is one of the major causative agents of hand, foot, and mouth disease (HFMD), which is sometimes associated with severe central nervous system disease in children. There is currently no specific medication for EV71 infection. Quercetin, one of the most widely distributed flavonoids in plants, has been demonstrated to inhibit various viral infections. However, investigation of the anti-EV71 mechanism has not been reported to date.MethodsThe anti-EV71 activity of quercetin was evaluated by phenotype screening, determining the cytopathic effect (CPE) and EV71-induced cells apoptosis. The effects on EV71 replication were evaluated further by determining virus yield, viral RNA synthesis and protein expression, respectively. The mechanism of action against EV71 was determined from the effective stage and time-of-addition assays. The possible inhibitory functions of quercetin via viral 2Apro, 3Cpro or 3Dpol were tested. The interaction between EV71 3Cpro and quercetin was predicted and calculated by molecular docking.ResultsQuercetin inhibited EV71-mediated cytopathogenic effects, reduced EV71 progeny yields, and prevented EV71-induced apoptosis with low cytotoxicity. Investigation of the underlying mechanism of action revealed that quercetin exhibited a preventive effect against EV71 infection and inhibited viral adsorption. Moreover, quercetin mediated its powerful therapeutic effects primarily by blocking the early post-attachment stage of viral infection. Further experiments demonstrated that quercetin potently inhibited the activity of the EV71 protease, 3Cpro, blocking viral replication, but not the activity of the protease, 2Apro, or the RNA polymerase, 3Dpol. Modeling of the molecular binding of the 3Cpro-quercetin complex revealed that quercetin was predicted to insert into the substrate-binding pocket of EV71 3Cpro, blocking substrate recognition and thereby inhibiting EV71 3Cpro activity.ConclusionsQuercetin can effectively prevent EV71-induced cell injury with low toxicity to host cells. Quercetin may act in more than one way to deter viral infection, exhibiting some preventive and a powerful therapeutic effect against EV71. Further, quercetin potently inhibits EV71 3Cpro activity, thereby blocking EV71 replication.

Dataset Information

Acrylamide fragment inhibitors that induce unprecedented conformational distortions in enterovirus 71 3C and SARS-CoV-2 main protease.

Publications

Acrylamide fragment inhibitors that induce unprecedented conformational distortions in enterovirus 71 3C and SARS-CoV-2 main protease.

Similar Datasets

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