Protocol to identify flavonoid antagonists of the SARS-CoV-2 main protease

STAR Protoc. 2024 Jun 21;5(2):102990. doi: 10.1016/j.xpro.2024.102990. Epub 2024 Apr 6.

Abstract

Flavonoids are naturally occurring metabolites of plants that can inhibit the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) main protease (Mpro), which is required for viral replication. Here, we present a protocol to identify flavonoid antagonists of the SARS-CoV-2 Mpro. We describe steps for the expression and purification of Mpro and a kinetic enzymatic assay for Mpro activity using a dequenching fluorescence resonance energy transfer peptide substrate. We then detail procedures for using this enzymatic assay to test flavonoid antagonism and reversible inhibition. For complete details on the use and execution of this protocol, please refer to Lin et al.1.

Keywords: High Throughput Screening; Molecular Biology; Protein Biochemistry; Protein expression and purification.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, N.I.H., Extramural

MeSH terms

  • Antiviral Agents / pharmacology
  • COVID-19 / metabolism
  • COVID-19 / virology
  • COVID-19 Drug Treatment
  • Coronavirus 3C Proteases* / antagonists & inhibitors
  • Coronavirus 3C Proteases* / metabolism
  • Flavonoids* / chemistry
  • Flavonoids* / pharmacology
  • Fluorescence Resonance Energy Transfer / methods
  • Humans
  • Kinetics
  • Protease Inhibitors / chemistry
  • Protease Inhibitors / metabolism
  • Protease Inhibitors / pharmacology
  • SARS-CoV-2* / drug effects

Substances

  • Flavonoids
  • Coronavirus 3C Proteases
  • Protease Inhibitors
  • Antiviral Agents
  • 3C-like proteinase, SARS-CoV-2