A yeast-based system to study SARS-CoV-2 Mpro structure and to identify nirmatrelvir resistant mutations.

IF 6.7 1区 医学 Q1 Immunology and Microbiology
PLoS Pathogens Pub Date : 2023-08-31 eCollection Date: 2023-08-01 DOI:10.1371/journal.ppat.1011592
Jin Ou, Eric M Lewandowski, Yanmei Hu, Austin A Lipinski, Ali Aljasser, Mariliz Colon-Ascanio, Ryan T Morgan, Lian M C Jacobs, Xiujun Zhang, Melissa J Bikowitz, Paul R Langlais, Haozhou Tan, Jun Wang, Yu Chen, John S Choy
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引用次数: 5

Abstract

The SARS-CoV-2 main protease (Mpro) is a major therapeutic target. The Mpro inhibitor, nirmatrelvir, is the antiviral component of Paxlovid, an orally available treatment for COVID-19. As Mpro inhibitor use increases, drug resistant mutations will likely emerge. We have established a non-pathogenic system, in which yeast growth serves as an approximation for Mpro activity, enabling rapid identification of mutants with altered enzymatic activity and drug sensitivity. The E166 residue is known to be a potential hot spot for drug resistance and yeast assays identified substitutions which conferred strong nirmatrelvir resistance and others that compromised activity. On the other hand, N142A and the P132H mutation, carried by the Omicron variant, caused little to no change in drug response and activity. Standard enzymatic assays confirmed the yeast results. In turn, we solved the structures of Mpro E166R, and Mpro E166N, providing insights into how arginine may drive drug resistance while asparagine leads to reduced activity. The work presented here will help characterize novel resistant variants of Mpro that may arise as Mpro antivirals become more widely used.

一种基于酵母的系统,用于研究严重急性呼吸系统综合征冠状病毒2型Mpro结构并鉴定耐尼马特韦突变。
严重急性呼吸系统综合征冠状病毒2型主要蛋白酶(Mpro)是一个主要的治疗靶点。Mpro抑制剂nirmatrevir是奈玛特韦片/利托那韦片组合包装的抗病毒成分,该药物是新冠肺炎的口服治疗药物。随着Mpro抑制剂使用的增加,耐药突变可能会出现。我们已经建立了一个非致病系统,其中酵母生长是Mpro活性的近似值,能够快速鉴定酶活性和药物敏感性改变的突变体。已知E166残基是耐药性的潜在热点,酵母检测发现了具有强烈尼马特雷韦耐药性的替代物和其他损害活性的替代物。另一方面,奥密克戎变异株携带的N142A和P132H突变对药物反应和活性几乎没有变化。标准酶分析证实了酵母的结果。反过来,我们解决了Mpro E166R和Mpro E166 N的结构,深入了解了精氨酸如何在天冬酰胺导致活性降低的情况下驱动耐药性。本文介绍的工作将有助于表征随着Mpro抗病毒药物的广泛使用,可能出现的Mpro新的耐药性变体。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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来源期刊
PLoS Pathogens
PLoS Pathogens 生物-病毒学
CiteScore
11.40
自引率
3.00%
发文量
598
审稿时长
2 months
期刊介绍: Bacteria, fungi, parasites, prions and viruses cause a plethora of diseases that have important medical, agricultural, and economic consequences. Moreover, the study of microbes continues to provide novel insights into such fundamental processes as the molecular basis of cellular and organismal function.
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