Fig 1.
Structure of the SARS-CoV-2 spike protein trimer.
(A) Left: side view of the trimeric spike ectodomain with 3 RBDs in the down-conformation; right: top view of the trimeric spike protein showing RBDs in gray, forest green, and orchid (PDB: 6VXX). (B) Left: side view of the trimeric spike ectodomain with 1 RBD in the up-conformation; right: top view of the trimeric spike protein showing 1 up RBD in gray (PDB: 7BNN). (C) A schematic layout of the spike protein is shown at the top. Right: structure of a monomer displaying the RBD in the open conformation. Spike protein structure shows the receptor-binding subunit S1 and the membrane-fusion subunit S2 separated by the furin-like protease site (S1/S2). Different subdomains of the spike protein are the NTD in green, RBD in gray containing RBM in cyan at their top, the fusion peptide in pink, second cleavage site S2’ in red, and HR1 and HR2 in olive (PBB: 7BNN). The scissors represent the S1/S2 boundary at amino acid position 685. Left: The open conformation RBD highlights the 3 different regions: receptor-binding ridge, flat surface, and 443–450 loop of the RBM that form the ACE2-binding region. ACE2, angiotensin-converting enzyme 2; FP, fusion peptide; NTD, N-terminal domain; RBD, receptor-binding domain; RBM, receptor-binding motif; SARS-CoV-2, Severe Acute Respiratory Syndrome Coronavirus 2.
Fig 2.
Schematic overview of the SARS-CoV-2 variants.
The variant being monitored B.1.1.7 (Alpha), B.1.351 (Beta), P1 (Gamma), and B.1.427 (Epsilon) and the variant of concern B.1.617.2 (Delta) and B.1.1.529 (Omicron) showing amino acid modifications in comparison to the ancestral Wuhan-Hu-1 sequence (NC_045512.2). FP, fusion peptide; NTD, N-terminal domain; RBD, receptor-binding domain; RBM, receptor-binding motif; SARS-CoV-2, Severe Acute Respiratory Syndrome Coronavirus 2.
Table 1.
Efficacy of clinical mAbs, lineages, and canonical mutations in SARS-CoV2 VOCs.
REGN-CoV (REGN10987/Imdevimab and REGN10933/Casirivimab) by Regeneron, VIR-7832 and VIR-7831 (Sotrovimab) by GSK and Vir biotechnology, and LY-CoV555 (Bamlanivimab) and LY-CoV016 (Etesevimab) by Eli Lilly and Company are some of the approved antibody cocktails for emergency use in mild-to-moderate COVID-19 patients. These antibodies were obtained from the people who got infected with SARS-CoV-2 during the first wave of the pandemic. These therapeutic antibody cocktails targeting noncompeting epitopes have varied degrees of neutralizing efficacies against VOCs. The color of highlighted text represents the levels of efficacy mAb retains against each VOC: green represents the mAb retains efficacy; light-green represents reduced efficacy; and orange represents that the mAb is ineffective. ADG-20 developed by Adagio binds to an epitope located in between the Class 1 and Class 4 sites.
Fig 3.
Antibody-mediated neutralization of envelope virus.
SARS-CoV-2 virus entry into human cells is initiated by virus binding to the ACE2-cell surface receptors (point 1). The virus neutralization largely depends on the epitope targeted by antibodies. Some of the antibodies target the RBM (point 2) or NTD (point 3) or other regions of the spike protein, which can inhibit the virus spike protein and host ACE2–receptor interactions, and they are considered among the most potent nAbs. A few rare antibodies can inactive the fusion machinery by activating the premature fusion pathway, thus inhibiting the virus entry into the host cell (point 4). Some of the non-nAbs bind to the viral antigens and activate the Fc-mediated antibody effector functions for their killing or phagocytosis (point 5). The figure was prepared using BioRender. ACE2, angiotensin-converting enzyme 2; Fc, fragment crystallizable; nAb, neutralizing antibody; NTD, N-terminal domain; RBM, receptor-binding motif; SARS-CoV-2, Severe Acute Respiratory Syndrome Coronavirus 2.
Fig 4.
Structural analysis and classification of nAbs of the SARS-CoV-2 spike protein.
(A) Structural footprints of ACE2 in blue on the RBM (cyan) (PDB: 6M0J). (B, C, D) The C102, C105, and REGN-10933 antibodies in complex with either isolated RBD or the spike trimer illustrate a conserved mode of binding to RBD in the up-conformation. Binding of C102 to the RBM on the RBD in red overlaps with the ACE2-binding site (PDB: 7K8M). (E) COVA2-39 (PDB: (7JMP) with a relatively longer CDRH3 loop binds the RBD in an alternative binding mode, as compared to the antibodies with a shorter CDRH3 loop, such as COVA2-04. (F) Fab222 binds RBD (PDB: 7NX6) with a relatively shorter CDRH3 loop. Residues K417, E484, and N501 are mutated in Alpha, Beta, and Gamma variants and are highlighted in red on the cyan ACE2-binding interface. (G) Structural footprints of a Class 2 antibody C002 on the RBD in red illustrates that the binding occurs toward the outer edge of the RBM (PDB: 7K8S). (H) The C144-spike protein complex structure revealed 3 C144 binding to a closed spike with all down-conformation RBDs (PDB: 7K90). The C144 epitope (red) spans between 2 adjacent RBDs on the surface of the trimeric spike. Close-up view of a quaternary epitope of C144 (red) bridging 2 adjacent monomers by the CDRH3 loop (blue). Class 1 and class 2 antibodies have significant overlap, but class 2 antibodies have fewer structural footprints on the ACE2-binding site at the RBM than class 1 antibodies (see B and G) (I) Structural footprints of a Class 3 antibody C110 on the RBD in red illustrates that C110 recognize a conserved epitope away from the receptor binding site and toward the outer face of the RBD. (J) The C309-spike protein structure illustrates that they bind outside of the ACE2-binding site. (PDB: 6WPS). (K) REGN-CoV2 is an antibody cocktail comprised of REGN10933 and REGN10987. REGN10933 blocks the ACE2-binding site like Class 1 antibodies, and REGN10987 sterically interferes with the ACE2 interactions like Class 3 antibodies. (L, M) Structural footprints of Class 4 antibodies CR3022 and COVA1-16 on the RBD in red, which illustrate that a conserved cryptic epitope accessible only in the up-conformation is recognized. The binding site of Class 4 antibodies is located away from the receptor binding site and toward the inner face of the RBD. (N, O) The CR3022 and COVA1-16 target a similar region on the RBD but with different angle, which explain their potency differences. (P) The structure of the 4A8-spike protein complex revealed that the NTD of each protomer is bound with 4A8 Fab. The structural footprints of 4A8 on the NTDs are shown in red. (Q) Summary of RBD-directed antibodies based on the binding pattern and competition profiles. The footprint residues on the RBD and NTD have been defined as those residues, which were within 4 Å of a Fab atom. ACE2, angiotensin-converting enzyme 2; nAb, neutralizing antibody; NTD, N-terminal domain; RBD, receptor-binding domain; RBM, receptor-binding motif; SARS-CoV-2, Severe Acute Respiratory Syndrome Coronavirus 2.
Fig 5.
SARS-CoV-2 RBD mutations that escape antibody binding.
Total escape at each site was measured using a yeast library screening followed by deep sequencing for distinct RBD antibodies (https://jbloomlab.github.io/SARS2_RBD_Ab_escape_maps/). The escape map identifies mutations that escape antibody binding and were mapped on the RBD-Fab structures. The red indicates the site with maximum and white with minimum resistance for antibody binding. (A, C, E, G, I) The structural footprint of C105, C144, C002, C135, and C110 antibodies on the RBM is in red. (B, D, F, H, J) Mapping of the SARS-CoV-2 virus escapes for different antibodies. Mutations that escape antibody binding were broadly located at their binding site in the RBD (see A-B, C-D, E-F, G-H, and I-J pairs). The footprint residues on the RBD have been defined as those residues, which were within 4 Å of a Fab atom. RBD, receptor-binding domain; SARS-CoV-2, Severe Acute Respiratory Syndrome Coronavirus 2.
Fig 6.
Structural analysis of bnAbs binding to the RBD.
The epitope surface of the A23-58.1 antibody is shown in red. A23-58.1 targets the supersite with minimal contacts to major mutational hotspots (K417, L452R, E484, S494P, and N501) of current VOCs, which are shown in blue. The binding mode of A23-58.1 is very similar to those of class 1 antibodies. The CDR H3 of A23-58.1 contains 14 residues and can only bind an RBD in the up-conformation (PDB: 7LRT). The RBD is shown in gray containing the RBM in cyan at their top. (B, C) S309 is a class 3 antibody while 7D6 binds proximal to the S309 epitope. S309 recognizes an epitope containing a glycan in white (N343 in SARS-CoV-2). 7D6 binds a novel cryptic site located behind the receptor-binding ridge of the RBD that faces toward the NTD. The S309 and 7D6 binding site residues are conserved within the Sarbecovirus subgenus. Both antibodies are resistant to mutations that emerged in the SARS-CoV-2 variants. The footprint residues on the RBD have been defined as those residues, which were within 4 Å of a Fab atom. bnAb, broadly neutralizing antibody; NTD, N-terminal domain; RBD, receptor-binding domain; RBM, receptor-binding motif; SARS-CoV-2, Severe Acute Respiratory Syndrome Coronavirus 2; VOC, variant of concern.