Characterization\nof the Conformational Hotspots of\nthe RNA-Dependent RNA Polymerase Complex Identifies a Unique Structural\nMalleability of nsp8

Several antiviral therapeutic approaches have been targeted\ntoward\nthe RNA-dependent RNA polymerase (RdRp) complex that is involved in\nviral genome replication. In SARS-CoV-2, although the RdRp is a multiprotein\ncomplex, the focus has been on the ligand binding catalytic core\n(nonstructural protein nsp12), and not the multiprotein functional\ndynamics. In this study, we focus on the conformational ensembles\nof the RdRp complex and their modulation by the presence of RNA, performing\ncomprehensive microsecond-scale atomistic simulations of the apo-\nand RNA-bound complex. We delineate the differential impact of RNA\non the constituent proteins, such as conformational polymorphisms,\ndominant segment-specific fluctuations, and the switch in dynamical\ncrosstalk within the complex. We distinguish dynamical signatures\nof nsp7, nsp8, and nsp12 in the apo-state that are reduced in the\npresence of the RNA and appear to “prime” the complex\nfor activity. Importantly, we identify a unique structural malleability\nof the nsp8 protein with high conformational heterogeneity in the\napo state, especially at three sites (Y71 for nsp8A, and D52 and A66\nfor nsp8B). Our work highlights the functional implications of the\npolymorphism of nsp8 structures and reveals possibilities for the\ndevelopment of allosteric inhibitors.