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SARS-CoV-2 is the causative agent of the COVID-19 pandemic that has proven to be a menacing nemesis to public health infrastructure across the globe. Since its advent in December 2019, the RNA virus has exhibited a high rate of mutation (~8.066 × 10−4 nucleotides per site per year) and the accumulation of mutations in key elements of its genome has led to the emergence of distinct SARS-CoV-2 variants such as α, β, γ, δ, ο, λ, and µ, which showed varying levels of infectivity and spreadability. The structural changes induced by non-synonymous mutations in major drug targets such as Spike protein and Main-protease (Mpro) of the virus are known to alter the binding of potential inhibitors designed against it. In this study, we have analyzed the effects of non-synonymous mutations in major targets of SARS-CoV-2 in response to potential peptide inhibitors, wherein we have screened 12 peptide inhibitors reported to have antiviral properties from three distinct classes (hACE2 receptor-based de-novo designed inhibitors, Antimicrobial) against Receptor Binding Domain (RBD) region and 5 antimicrobial peptides against Mpro of SARS-CoV-2 variants using molecular docking and simulation approaches. The mutational landscape of RBD among SARS-CoV-2 variants had a total of 21 non-synonymous mutations across 18 distinct sites. Among these, 14 mutations were present in the RBM region directly interacting with the hACE-2 receptor. However, Only 3 non-synonymous mutations were observed in Mpro among the 7 variants analyzed with one each in Beta, Omicron, and Lambda variants. We found that LCB1 - a de novo-synthesized peptide has the highest binding affinity to RBD despite non-synonymous mutations in variants and engages key residues of RBD-hACE2 interaction such as K417, E484, N487, and N501. Similarly, an antimicrobial peptide; 2JOS, was identified against Mpro with high binding affinity as it interacts with key residues in dimerization sites such as E166 and F140 crucial for viral replication. Molecular Dynamics simulations affirm the stability of RBD-LCB1 and Mpro-2JOS complexes with an average RMSD of 1.902 and 2.476 respectively. We ascertain that LCB1 and 2JOS peptides are promising inhibitors to combat emerging variants of SARS-CoV-2 and thus warrant further investigations using in-vitro and in-vivo analysis.

Protein-protein interactions between virus and host are crucial for infection. SARS-CoV-2, the causative agent of COVID-19 pandemic is an RNA virus prone to mutations. Formation of a stable binding interface between the Spike (S) protein Receptor Binding Domain (RBD) of SARS-CoV-2 and Angiotensin-Converting Enzyme 2 (ACE2) of host actuates viral entry. Yet, how this binding interface evolves as virus acquires mutations during pandemic remains elusive. Here, using a high fidelity bioinformatics pipeline, we analysed 31,403 SARS-CoV-2 genomes across the globe, and identified 444 non-synonymous mutations that cause 49 distinct amino acid substitutions in the RBD. Molecular phylogenetic analysis suggested independent emergence of these RBD mutants during pandemic. In silico structure modelling of interfaces induced by mutations on residues which directly engage ACE2 or lie in the near vicinity revealed molecular rearrangements and binding energies unique to each RBD mutant. Comparative structure analysis using binding interface from mouse that prevents SARS-CoV-2 entry uncovered minimal molecular determinants in RBD necessary for the formation of stable interface. We identified that interfacial interaction involving amino acid residues N487 and G496 on either ends of the binding scaffold are indispensable to anchor RBD and are well conserved in all SARS-like corona viruses. All other interactions appear to be required to locally remodel binding interface with varying affinities and thus may decide extent of viral transmission and disease outcome. Together, our findings propose the modalities and variations in RBD-ACE2 interface formation exploited by SARS-CoV-2 for endurance.ImportanceCOVID-19, so far the worst hit pandemic to mankind, started in January 2020 and is still prevailing globally. Our study identified key molecular arrangements in RBD-ACE2 interface that help virus to tolerate mutations and prevail. In addition, RBD mutations identified in this study can serve as a molecular directory for experimental biologists to perform functional validation experiments. The minimal molecular requirements for the formation of RBD-ACE2 interface predicted using in silico structure models may help precisely design neutralizing antibodies, vaccines and therapeutics. Our study also proposes the significance of understanding evolution of protein interfaces during pandemic.