Single-molecule assay reveals the impact of composition, RNA duplex, and inhibitors on the binding dynamics of SARS-CoV-2 polymerase complex.

ABSTRACT: The genome replication of SARS-CoV-2, the causative agent of COVID-19, involves a multi-subunit replication complex consisting of non-structural proteins (nsps) 12, 7 and 8. While the structure of this complex is known, the dynamic behavior of the subunits interacting with RNA is missing. Here we report a single-molecule protein-induced fluorescence enhancement (SM-PIFE) assay to monitor binding dynamics between the reconstituted or co-expressed replication complex and RNA. Increasing binding times were observed, in this order, with nsp7 (none) nsp8 and nsp12, in nsp8-nsp12 mixtures and in reconstituted mixtures bearing all three proteins. Unstable, transient, and stable binding modes were recorded in the latter case, indicating that complexation is dynamic, and the correct conformation mu

SUBMITTER: Lovell TC 

PROVIDER: S-EPMC11741280 | biostudies-literature | 2025 Jan

REPOSITORIES: biostudies-literature