Models and complexes construction
Building the NAb/SARS-CoV-2 RBD reference model. We used the model PDB ID: 7YAD downloaded from RCSB Protein Data Bank (RCSB PDB) (https://www.rcsb.org/structure/7YAD), to generate our reference model representing the interaction of Sotrovimab variable domain (Fv) with Omicron SARS-CoV-2 spike RBD. The selection criteria of 7YAD model [15] are the generation of the 3D structure by electron microscopy, the high resolution of 2.66Å and the relatively good validation report. In addition, it represent the interaction with SARS-COV-2 RBD in the open state. The 7YAD structure shows two RBD-S309 units (S309-RBD-RBD-S309). Upon downloading the structure, only one unit was selected representing one S309 Fv domain binding to one spike RBD (Chains A, B, M). The complex was extracted, cleaned from any heteroatoms and used as reference model to generate the different variant complexes by RBD replacement.
SARS-CoV and SARS-CoV-2 variants sequence retrieval, modifications and modeling. The amino acid sequence of the extracellular domain of SARS-CoV and SARS-CoV-2 spike protein were acquired from the National Center for Biological Information (NCBI) protein ID: YP_009825051.1 and ID: YP_009724390.1 respectively. SARS-CoV-2 variants-specific mutations were introduced to the collected sequence to generate the different variant sequences based on published mutations on databases such as CoVariants (https://covariants.org/) and Stanford University SARS-CoV-2 Variants database (https://covdb.stanford.edu/variants/omicron_ba_1_3/) [39]. The sequences corresponding to the spike protein of SARS-CoV, and total of 25 variants of SARS-CoV-2 Wuhan (ALPHA, BETA, GAMMA, DELTA-21J, and KAPPA) in addition to Omicron’s subvariants (BA.1, BA.2, BA.4/BA.5, BA.2.12.1, BA.2.75, BQ1, XBB, XBB.1) were used to build spikes 3D monomer models. The monomers were modeled in an open state form using the SWISS-MODEL server-User Template Mode (Waterhouse et al., 2018) (https://swissmodel.expasy.org/interactive#structure). The template for each monomer was selected and extracted from Research Collaboratory for Structural Bioinformatics Protein Data Base (https://www.rcsb.org/). Selection’s criteria were based on resolution, chain quality, sequence gaps, furin site and proline modifications and validation report. The templates used for each model are listed in Supplementary Table S1. The monomer chain representing the open state RBD was extracted from each model, cleaned from any heteroatoms and saved using PyMol software [42] into new Pdb file. Each monomer was introduced in the SWISS-MODEL server-User Template Mode to generate an open-state monomer spike protein for SARS-CoV, SARS-CoV-2 variants and Omicron sub variants.
RBD/S309 complexes construction. RBDs of the different SARS-CoV, SARS-CoV-2 variants and Omicron’s sub variants, were extracted from the generated models and the complexes with S309 was constructed by molecular replacement. The reference crystalized RBD chain M of PBD 7YAD was replaced with the modeled RBD. The complex was saved and energy minimized. Energy minimization was done in one-steps using Swiss-pdb Viewer 4.1.0 (http://www.expasy.org/spdbv/) [43]. This was applied to all the generated models.
Interactions and complex binding affinity analysis. The interactions between the RBD of spike protein of SARS-CoV, SARS-CoV-2 variants and Omicron subvariants with neutralizing antibody S309 were analyzed based on polar and hydrophobic interaction using the LigPlot + software [44]. The stability and affinity were assisted based on thermodynamic measure of the formed complex energy, Gibbs free energy, (ΔG), this was performed using an antibody-antigen binding affinity online tool, CSM-AB (https://biosig.lab.uq.edu.au/csm_ab/prediction) [45]. Binding affinity percentage was calculated in reference to Wuhan /S309 complex binding affinity.
Testing the generated method by analyzing newly reported Omicron subvariants and some experimentally tested mutations. Several reports have been discussing the neutralization effect of NAbs and possible antibody escape of some new Omicrons subvariants [32, 36–38, 40, 41]. Here we used our developed method to evaluate the binding affinity of several of these new subvariants including AY.1, XBB.1.5, BF.7, BQ.1.1, BA.1.1, BA.3, BA.2.3.20, BM.1.1.1, BA.5.6.2, BA.2.75.2, and CH.1.1 (Orthrus) ,with NAb S309. Additionally, the effect of several amino acids exchange in the NAb epitope have been tested experimentally by ELISA and/or pseudovirus neutralization assays. Several mutations showed to be resistance to inhibition by S309 leading to an antibody escape. These key residues include R346 and P337, G339, N440 and S371 [23, 25]. Therefore, we apply our method to computationally test the effect of some mutations on these residues. As we already generated parent’s RBD sequences, newly emerged mutations were introduced, new models and complexes were built and the mutation’s effect on binding energy with the NAb was predicted by recalculating complex’s Gibbs free energy (ΔG) in reference to parent’s complex and Wuhan binding affinity.