Title : Molecular docking and dynamics simulation of effects of key mutations in SARS-CoV-2 spike protein RBD in complex with human ACE2 with withanolide A inhibitor
Abstract:
Background and objectives: Different countries, including India witnessed sharp increase in SARS-CoV-2 cases and deaths associated with a rising proportion of B.1.617 variant. The strain was characterized by L452R and E484Q primary mutations in the receptor binding domain (RBD) described as the mutation of concern. However, the biophysical basis for comprehending the molecular mechanism causing the increase in the infectivity rate and immune evasion ability of the mutant in relation to wild-type (wt) SARS-CoV-2, are very scanty. The aim of the current study was to evaluate the effect of mutation on the molecular dynamics, energetics, antigenicity, interactions and binding affinity of RBD of the spike (S) glycoprotein in complex with human ACE2 receptor with withanolide A ligand.
Methods: The C-terminal domain of SARS-CoV-2 S protein RBD in complex with human ACE2 (PDB ID: 6LZG) and SARS-COV-2 P2B-2F6 murine monoclonal antibodies (mAbs) with RBD (PDB ID: 7BWJ) were retrieved from RCSB PDB (http://www.rcsb.org//pdb). The 3D structure of withanolide A (PubChem CID: 11294368), used as ligand, was retrieved from PubChem database (https://pubchem.ncbi.nlm.nih.gov/). The L452R and E484Q point mutations were introduced into the RBD chain using VMD (http://www.ks.uiuc.edu/Research/vmd/). The binding affinities towards mutant and wildtype RBD were compared for withanolide A, ACE2, P2B-2F6 mAbs using AutoDock Vina (https://www.cgl.ucsf.edu/chimera/) and Haddock (www.bonvinlab.org). The ADMET and drug‐likeness properties of withanolide A was determined using SwissADME (http://www.swissadme.ch/) and pkCSM (http://biosig.unimelb.edu.au/pkcsm/prediction). Antigenic propensity of the wt and mutant proteins were predicted using antigenic peptides tools (http://imed.med.ucm.es/Tools/antigenic.pl). Molecular dynamic simulation and post trajectory analysis was done for: the mutant and wildtype RBD bound with or without ACE2 in complex with withanolide A, and the free withanolide A (in water) using Gromacs 2021 (www.gromacs.org). Free energy of binding was computed using MM-PBSA approach. The above computations were performed in Ubuntu 20.04.2 LTS 64-bit OS, 3.36.8 Gnome version.
Results: Molecular docking demonstrated binding energy -10.3 and -11.2 kcal/mol for withanolide A with wt and mutant RBD bound to ACE2, respectively. The wt RBD not bound to ACE2 displayed binding energy -7.7 kcal/mol with withanolide A. The ACE2 and P2B-2F6 mAb showed binding affinity of -12.4 and -9.6 kcal/mol, respectively with wt RBD. No violation of Lipinski’s RO5, favourable ADMET properties and bioavailability scores (0.55) signify the suitability of drug-likeness for withanolide A. Molecular dynamic simulation revealed the root mean square deviation (RMSD) and root mean square fluctuations (RMSF) of ~0.05 nm for withanolide A. The RMSD and RMSF of wt and mutant RBD not bound to ACE2 in complex with withanolide A were ~1.00 and ~0.5 nm, respectively. The RMSD and RMSF of wt and mutant RBD bound to ACE2 in complex with withanolide A were ~2.00 and ~1.4 nm, respectively. The net free energy of binding for the wt and mutant RBD bound to ACE2 were -104.18 and -108.21 kJ/mol, respectively, with withanolide A. The net free energy of binding for the wt and mutant RBD not bound to ACE2 were -16.66 and -17.68 kJ/mol, respectively, with withanolide A. The key amino acid player in protein-ligand interactions was Ile291 through H-bond in both wt and mutant RBD bound to ACE2 in complex with withanolide A consisting fewer hydrophobic interactions in the latter. Following structure analysis, P2B-2F6 mAb identified a linear epitope located in residues E466 - D492, which overlapped with ACE2 binding sites in mutant and wt RBD.
Interpretation and conclusions: The mutant RBD bound to ACE2 in complex with withanolide A was energetically more stable than the wild type strain, and the mutant/wild type RBD not bound to ACE2 in complex with withanolide A. Insignificant changes in the antigenic propensity of RBD protein with or without ACE2 receptor and their mutants were found. The results suggested that the L452R and E484Q mutations in RBD of the SARS-CoV-2 B.1.617 variant improved the stability of S protein, with future implications for vaccine development and application.
