Conformational dynamics and energetic landscape of the SARS-CoV-2 nsp12 with and without accessory proteins: new insights on this potential therapeutic target against COVID-19

The SARS-CoV-2 replication-transcription complex (RTC) represents a key antiviral target, yet the conformational dynamics underlying its function remain insufficiently characterised. While cryo-EM structures resolve the architecture of the nsp12 polymerase and its nsp7/nsp8 cofactors, they do not capture the motions governing catalysis and RNA engagement. In this study, independent triplicate 300 ns all-atom molecular dynamics simulations (nine trajectories, 2.7 µs aggregate) were conducted for nsp12 in three assembly states: isolated nsp12-RNA, the central RTC and the nsp13-containing mini-RTC. Across replicas, cofactors consistently reduced nsp12 flexibility, particularly in the interface domain and thumb subdomain, while maintaining catalytic-site geometry. MM/GBSA analysis identified electrostatic interactions as the primary determinants of RNA recognition and delineated a reproducible set of RNA-contact hotspots. When the receptor was fixed as nsp12, cofactors produced only a modest increase in nsp12-RNA interaction strength, within the range of replica-to-replica variability. Thus, their principal influence appears to be on the dynamics rather than on static binding energetics. The simulations also consistently predicted R914 as a persistent favourable contact with the product-RNA strand, a feature not highlighted in prior static analyses. This observation is presented as a hypothesis for experimental validation and structure-based antiviral design.