(8) Top candidates are subjected to molecular dynamics simulation for stability and free energy perturbation. Difluprednate due to the relatively lower glycan-shielding and immunodominance of the RBD compared with the other glycan-coated domains of the spike.3 The emergence of variants of concern (VOCs) by the continuous evolution of the virus is of explicit relevance to the efficacy of COVID-19 therapeutics, as mutations in the prototype spike, RBD in particular, have rendered antibodies incapable of neutralization4,5 and substantially dropped the effectiveness of vaccines.6 Although debatable, all VOCs are in general characterized as rapidly transmissible, immune evading, and more pathogenic in some cases, compared with the wild-type strain.2 The trend of increasing host fitness by the VOCs is well set to this end; however, the mechanism by which positively selected mutations in the spike modulate the viral function and ditch the immune response require crucial assessments. Open in a separate window Physique?1 Mutations in the SARS-CoV-2 spike, receptor binding, and immune escape are shown (A) The emergence of VOCs is depicted chronologically, and shared mutations within NTD and RBD are shown over the protruding spikes around the viron surface. The 3D modeled spike protein is shown in the lower panel depicting 35 mutations in Omicron. (B) Mutations in the RBD with increased ACE2 binding affinity (left) are also involved in immune escape (right). Single or multiple mutations in RBD that abolish the neutralization of clinically approved mAbs are shown in the right panel. 3D structures of the VOC-mAbs complexes were built by replacing/mutating RBD in the WT-mAbs complexes. Mutations in the RBD region of Omicron sub-lineages are shown on the left bottom panel for comparison. All VOCs enhance ACE2 binding, but not all mutations in RBD augment ACE2 binding Collectively, three mutations, K417N, E484K, and N501Y, were mainly responsible for the increased ACE2 binding in the Beta strain that caused the second wave of COVID-19 in 2020. The Beta and Gamma variants differ by K417N/T mutations in the spike and share comparable ACE2-binding affinity (Physique?1B). The Delta variant accounted for the deadliest second wave of COVID-19 in April 2021 and harbors ten mutations in its spike, i.e., T19R, G142D? (associated with frequent back mutations, increased viral loads, and immune evasion), 156, 157, R158G, L452R, T478K, D614G, P681R, and D950N, where L452R and T478K substitutions were responsible for increased ACE2 binding. The Omicron variant appeared with the highest quantity of mutations in its spike (Physique?1A), which not Difluprednate only dramatically enhanced its ACE2 binding, infectivity, and transmission but also escaped most of the FDA-approved nAbs and vaccines. Cataloging mutations in the spike protein (N?= 303,250) demonstrate that all 195 amino acids in the RBD are associated with mutations in the spike; however, not all suggested mutations involve ACE2 binding.7 During early COVID-19 pandemic, Starr et?al. conducted a deep mutational analysis and recognized mutations in the RBD that alter ACE2 binding and/or are suitable for antibody-based therapeutics.8 Difluprednate Surprisingly, among the 16 suggested RBD mutations participating in the immune escape of VOCs, only six, i.e., G339D, L452R, S477N, T478K, E484K, and N501Y, are involved in increased ACE2 binding. Alternatively, the positive selection of the VOCs can be attributed to the rise in resistance to the post-vaccinated and convalescent sera by favoring best-fitting mutations within the antigenic epitopes.4,9 Besides, due to its immunodominant nature, 40% of the anti-SARS-CoV2 antibodies bind RBD, whereas many RBD mutations produce conformational alterations by changing IL-16 antibody the charges and hydrophobicity within the epitopes and break.