Structural, Dynamical, and Entropic Differences Between SARS-CoV and SARS-CoV-2 s2m Elements Using Molecular Dynamics Simulations

Abstract

The functional role of the highly conserved stem-loop II motif (s2m) in SARS-CoV and SARS-CoV-2 in the viral lifecycle remains enigmatic and an intense area of research. Structure and dynamics of the s2m are key to establishing a structure-function connection, yet a full set of atomistic resolution coordinates is not available for SARS-CoV-2 s2m. Our work constructs three-dimensional coordinates consistent with NMR solution phase data for SARS-CoV-2 s2m and provides a comparative analysis with its counterpart SARS-CoV s2m. We employed initial coordinates based on PDB ID: 1XJR for SARS-CoV s2m and two models for SARS-CoV-2 s2m: one based on 1XJR in which we introduced the mutations present in SARS-CoV-2 s2m and the second based on the available SARS-CoV-2 NMR NOE data supplemented with knowledge-based methods. For each of the three systems, 3500 ns molecular dynamics simulations were used to sample the structure and dynamics, and principal component analysis (PCA) reduced the ensembles to hierarchal conformational substates for detailed analysis. Dilute solution simulations of SARS-CoV s2m, demonstrate that the GNRA-like terminal pentaloop is rigidly defined by base stacking uniquely positioned for possible kissing dimer formation. However, the SARS-CoV-2 s2m simulation did not retain the reported crystallographic SARS-CoV motifs and the terminal loop expands to a highly dynamic “nonaloop.” Increased flexibility and structural disorganization are observed for the larger terminal loops, where an entropic penalty is computed to explain the experimentally observed reduction in kissing complex formation. Overall, both SARS-CoV and SARS-CoV-2 s2m elements have a similarly pronounced L-shape due to different motif interactions. Our study establishes the atomistic three-dimensional structure and uncovers dynamic differences that arise from s2m sequence changes, which sets the stage for the interrogation of different mechanistic pathways of suspected biological function.

Host microRNA interactions with the SARS-CoV-2 viral genome 3’-untranslated region

“…We and others have analyzed the SARS-CoV-2 3'-UTR for potential binding sites for host miRs (6,7,11,33,54,55) and identified that miR-760-3p has a predicted binding site located at nt 29,833 -29,856 in the reference SARS-CoV-2 genome (NCBI: NC_045512.2) (Fig. 1, nt 272-313 in the 3'-UTR, turquoise dashed line) and that miR-34a-5p and miR-34b-5p both have the same predicted binding site, downstream of the s2m, extending into the lower stem of the s2m (Fig.…”

“…Furthermore, this motif was also shown to bind host cellular microRNA (miR)-1307-3p in vitro. (6,7) With this in mind, we investigated whether the SARS-CoV-2 viral genome 3'-UTR could undergo additional interactions with host miRs, based on secondary structural elements.…”