Evaluating the importance of metal ions on inhibitor screening for the SARS-CoV-2 RNA replication complex

The dramatic impact of the COVID-19 pandemic on public health highlights the importance of developing a better understanding of the basic mechanisms of viral replication and the discovery of more effective antiviral therapeutics for coronaviruses and other emerging viral pathogens. Because the virally encoded RNA-dependent RNA polymerase plays a critical function in replicating the genome of all RNA viruses, it is one of the most attractive targets for developing direct-acting antiviral therapeutics. Divalent cations like magnesium ions play key roles as essential cofactors for the nucleotidyl-transfer reaction in DNA and RNA polymerases. Using a previously developed system to produce enzymatically active core replication complexes for SARS-CoV-2 and to measure RNA synthesis activities using in vitro primer extension assays, we have evaluated the effects of magnesium ions on RNA synthesis in the presence and absence of inhibitors. Our preliminary results reveal interesting patterns of sensitivity to magnesium ion concentration that vary depending on the presence and absence of different types of inhibitors. These preliminary findings suggest future directions for research into understanding the potential roles of magnesium and other metal ions in the structure and function of viral polymerases, as well as for understanding the effects of metal ions on the mechanisms of small-molecule inhibitors.