High-Throughput Activity Assay for Screening Inhibitors of the SARS-CoV-2 Mac1 Macrodomain

ACS Chem Biol. 2022 Jan 21;17(1):17-23. doi: 10.1021/acschembio.1c00721. Epub 2021 Dec 14.


Macrodomains are a class of conserved ADP-ribosylhydrolases expressed by viruses of pandemic concern, including coronaviruses and alphaviruses. Viral macrodomains are critical for replication and virus-induced pathogenesis; therefore, these enzymes are a promising target for antiviral therapy. However, no potent or selective viral macrodomain inhibitors currently exist, in part due to the lack of a high-throughput assay for this class of enzymes. Here we developed a high-throughput ADP-ribosylhydrolase assay using the SARS-CoV-2 macrodomain Mac1. We performed a pilot screen that identified dasatinib and dihydralazine as ADP-ribosylhydrolase inhibitors. Importantly, dasatinib inhibits SARS-CoV-2 and MERS-CoV Mac1 but not the closest human homologue, MacroD2. Our study demonstrates the feasibility of identifying selective inhibitors based on ADP-ribosylhydrolase activity, paving the way for the screening of large compound libraries to identify improved macrodomain inhibitors and to explore their potential as antiviral therapies for SARS-CoV-2 and future viral threats.

Publication types

  • Letter

MeSH terms

  • Antiviral Agents / pharmacology*
  • Dasatinib / pharmacology
  • High-Throughput Screening Assays / methods*
  • N-Glycosyl Hydrolases / antagonists & inhibitors*
  • Protein Domains
  • SARS-CoV-2 / drug effects*
  • SARS-CoV-2 / enzymology


  • Antiviral Agents
  • N-Glycosyl Hydrolases
  • ADP-ribosylarginine hydrolase
  • Dasatinib