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 stable structural stability (RMSD: ∼2Å), Cα fluctuations (RMSF: ∼3Å) and strong protein-ligand interactions and the same were confirmed 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 (MMGBSA = -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 structural insights having 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.
The coronavirus disease 2019 (COVID-19) pandemic is caused by newly discovered severe acute respiratory syndrome-coronavirus 2 (SARS-CoV-2). One of the striking targets amongst all the proteins in coronavirus is the main protease (M pro ), as it plays vital biological roles in replication and maturation of the virus, and hence the potential target. The aim of this study is to repurpose the Food and Drug Administration (FDA) approved molecules via computer-aided drug designing against M pro (PDB ID: 6Y2F) of SARS CoV-2 due to its high x-ray resolution of 1.95 Å as compared to other published M pro structures. High Through Virtual Screening (HTVS) of 2456 FDA approved drugs using structurebased docking were analyzed. Molecular Dynamics simulations were performed to check the overall structural stability (RMSD), Ca fluctuations (RMSF) and protein-ligand interactions. Further, trajectory analysis was performed to assess the binding quality by exploiting the protein-residue motion cross correlation (DCCM) and binding free energy (MM/GBSA). Tenofovir, an antiretroviral for HIV-proteases and Terlipressin, a vasoconstrictor show stable RMSD, RMSF, better MM/GBSA with good cross correlation as compared to the Apo and O6K. Moreover, the results show concurrence with Nelfinavir, Lopinavir and Ritonavir which have shown significant inhibition in in vitro studies. Therefore, we conclude that Tenofovir and Terlipresssin might also show protease inhibition but are still open to clinical validation in case of SARS-CoV 2 treatment.
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