Structural and conformational analysis of SARS CoV 2 N-CTD revealing monomeric and dimeric active sites during the RNA-binding and stabilization: Insights towards potential inhibitors for N-CTD.

The advent of SARS-CoV-2 has become a universal health issue with no appropriate cure available to date. The coronavirus nucleocapsid (N) protein combines viral genomic RNA into a ribonucleoprotein and protects the viral genome from the host's nucleases. Structurally, the N protein comprises two independent domains: the N-terminal domain (NTD) for RNA-binding and C-terminal domain (CTD) involved in RNA-binding, protein dimerization, and nucleocapsid stabilization. The present study explains the structural aspects associated with the involvement of nucleocapsid C-terminal domain in the subunit assembly that helps the RNA binding and further stabilizing the virus assembly by protecting RNA from the hosts exonucleases degradation. The molecular dynamics (MD) simulations of the N-CTD and RNA complex suggests two active sites (site I: a monomer) and (site II: a dimer) with structural stability (RMSD: ~2 Å), Cα fluctuations (RMSF: ~3 Å) and strong protein-ligand interactions were estimated through the SiteMap module of Schrodinger. Virtual screening of 2456 FDA-approved drugs using structure-based docking identified top two leads distinctively against Site-I (monomer): Ceftaroline fosamil (MM-GBSA = -47.12 kcal/mol) and Cefoperazone (-45.84 kcal/mol); and against Site-II (dimer): Boceprevir, (an antiviral protease inhibitor, -106.78 kcal/mol) and Ceftaroline fosamil (-99.55 kcal/mol). The DCCM and PCA of drugs Ceftaroline fosamil (PC1+PC2 = 71.9%) and Boceprevir (PC1 +PC2 = 61.6%) show significant correlated residue motions which suggests highly induced conformational changes in the N-CTD dimer. Therefore, we propose N-CTD as a druggable target with two active binding sites (monomer and dimer) involved in specific RNA binding and stability. The RNA binding site with Ceftaroline fosamil binding can prevent viral assembly and can act as an antiviral for coronavirus.

In Silico Structure-Based Repositioning of Approved Drugs for Spike Glycoprotein S2 Domain Fusion Peptide of SARS-CoV-2: Rationale from Molecular Dynamics and Binding Free Energy Calculations.

The membrane-anchored spike (S) protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has a pivotal role in directing the fusion of the virus particle mediated by the host cell receptor angiotensin-converting enzyme 2 (ACE-2). The fusion peptide region of the S protein S2 domain provides SARS-CoV-2 with the biological machinery needed for direct fusion to the host lipid membrane. In our present study, computer-aided drug design strategies were used for the identification of FDA-approved small molecules using the optimal structure of the S2 domain, which exhibits optimal interaction ratios, structural features, and energy variables, which were evaluated based on their performances in molecular docking, molecular dynamics simulations, molecular mechanics/generalized Born model and solvent accessibility binding free energy calculations of molecular dynamics trajectories, and statistical inferences. Among the 2,625 FDA-approved small molecules, chloramphenicol succinate, imipenem, and imidurea turned out to be the molecules that bound the best at the fusion peptide hydrophobic pocket. The principal interactions of the selected molecules suggest that the potential binding site at the fusion peptide region is centralized amid the Lys790, Thr791, Lys795, Asp808, and Gln872 residues.IMPORTANCE The present study provides the structural identification of the viable binding residues of the SARS-CoV-2 S2 fusion peptide region, which holds prime importance in the virus's host cell fusion and entry mechanism. The classical molecular mechanics simulations were set on values that mimic physiological standards for a good approximation of the dynamic behavior of selected drugs in biological systems. The drug molecules screened and analyzed here have relevant antiviral properties, which are reported here and which might hint toward their utilization in the coronavirus disease 2019 (COVID-19) pandemic owing to their attributes of binding to the fusion protein binding region shown in this study.