N-terminal domain antigenic mapping reveals a site of vulnerability for SARS-CoV-2.
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ABSTRACT:
SUBMITTER: McCallum M
PROVIDER: S-EPMC7962585 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Antibodies that potently neutralize SARS-CoV-2 target mainly the receptor-binding domain or the N-terminal domain (NTD). Over a dozen potently neutralizing NTD-directed antibodies have been studied structurally, and all target a single antigenic supersite in NTD (site 1). Here, we report the cryo-EM structure of a potent NTD-directed neutralizing antibody 5-7, which recognizes a site distinct from other potently neutralizing antibodies, inserting a binding loop into an exposed hydrophobic pocket between the two sheets of the NTD β sandwich. Interestingly, this pocket was previously identified as the binding site for hydrophobic molecules, including heme metabolites, but we observe that their presence does not substantially impede 5-7 recognition. Mirroring its distinctive binding, antibody 5-7 retains neutralization potency with many variants of concern (VOCs). Overall, we reveal that a hydrophobic pocket in NTD proposed for immune evasion can be used by the immune system for recognition. Graphical abstract Cerutti et al. report the cryo-EM structure of potent neutralizing antibody 5-7 bound to SARS-CoV-2 spike. While most NTD-directed neutralizing antibodies target the NTD supersite, 5-7 binds to a conserved hydrophobic pocket on NTD and neutralizes many variants of concern.
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Project description:An essential step for SARS-CoV-2 infection is the attachment to the host cell receptor by its Spike receptor-binding domain (RBD). Most of the existing RBD-targeting neutralizing antibodies block the receptor-binding motif (RBM), a mutable region with the potential to generate neutralization escape mutants. Here, we isolated and structurally characterized a non-RBM-targeting monoclonal antibody (FD20) from convalescent patients. FD20 engages the RBD at an epitope distal to the RBM with a KD of 5.6 nM, neutralizes SARS-CoV-2 including the current Variants of Concern such as B.1.1.7, B.1.351, P.1, and B.1.617.2 (Delta), displays modest cross-reactivity against SARS-CoV, and reduces viral replication in hamsters. The epitope coincides with a predicted 'ideal' vulnerability site with high functional and structural constraints. Mutation of the residues of the conserved epitope variably affects FD20-binding but confers little or no resistance to neutralization. Finally, in vitro mode-of-action characterization and negative-stain electron microscopy suggests a neutralization mechanism by which FD20 destructs the Spike. Our results reveal a conserved vulnerability site in the SARS-CoV-2 Spike for the development of potential antiviral drugs.
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Project description:During the SARS-CoV-2 pandemic, multiple variants with differing amounts of escape from pre-existing immunity have emerged, causing concerns about continued protection. Here, we use antigenic cartography to quantify and visualize the antigenic relationships among 16 SARS-CoV-2 variants titrated against serum samples taken post-vaccination and post-infection with seven different variants. We find major antigenic differences caused by substitutions at spike positions 417, 452, 484, and possibly 501. B.1.1.529 (Omicron BA.1) showed the highest escape from all sera tested. Visualization of serological responses as antibody landscapes shows how reactivity clusters in different regions of antigenic space. We find changes in immunodominance of different spike regions depending on the variant an individual was exposed to, with implications for variant risk assessment and vaccine strain selection.One sentence summaryAntigenic Cartography of SARS-CoV-2 variants reveals amino acid substitutions governing immune escape and immunodominance patterns.
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Project description:Antibodies are crucial to immune protection against SARS-CoV-2, with some in emergency use as therapeutics. Here, we identify 377 human monoclonal antibodies (mAbs) recognizing the virus spike and focus mainly on 80 that bind the receptor binding domain (RBD). We devise a competition data-driven method to map RBD binding sites. We find that although antibody binding sites are widely dispersed, neutralizing antibody binding is focused, with nearly all highly inhibitory mAbs (IC50 < 0.1 μg/mL) blocking receptor interaction, except for one that binds a unique epitope in the N-terminal domain. Many of these neutralizing mAbs use public V-genes and are close to germline. We dissect the structural basis of recognition for this large panel of antibodies through X-ray crystallography and cryoelectron microscopy of 19 Fab-antigen structures. We find novel binding modes for some potently inhibitory antibodies and demonstrate that strongly neutralizing mAbs protect, prophylactically or therapeutically, in animal models.
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Project description:In 2002, severe acute respiratory syndrome-associated coronavirus (SARS-CoV) emerged in humans, causing a global epidemic. By phylogenetic analysis, SARS-CoV is distinct from known CoVs and most closely related to group 2 CoVs. However, no antigenic cross-reactivity between SARS-CoV and known CoVs was conclusively and consistently demonstrated except for group 1 animal CoVs. We analyzed this cross-reactivity by an enzyme-linked immunosorbent assay (ELISA) and Western blot analysis using specific antisera to animal CoVs and SARS-CoV and SARS patient convalescent-phase or negative sera. Moderate two-way cross-reactivity between SARS-CoV and porcine CoVs (transmissible gastroenteritis CoV [TGEV] and porcine respiratory CoV [PRCV]) was mediated through the N but not the spike protein, whereas weaker cross-reactivity occurred with feline (feline infectious peritonitis virus) and canine CoVs. Using Escherichia coli-expressed recombinant SARS-CoV N protein and fragments, the cross-reactive region was localized between amino acids (aa) 120 to 208. The N-protein fragments comprising aa 360 to 412 and aa 1 to 213 reacted specifically with SARS convalescent-phase sera but not with negative human sera in ELISA; the fragment comprising aa 1 to 213 cross-reacted with antisera to animal CoVs, whereas the fragment comprising aa 360 to 412 did not cross-react and could be a potential candidate for SARS diagnosis. Particularly noteworthy, a single substitution at aa 120 of PRCV N protein diminished the cross-reactivity. We also demonstrated that the cross-reactivity is not universal for all group 1 CoVs, because HCoV-NL63 did not cross-react with SARS-CoV. One-way cross-reactivity of HCoV-NL63 with group 1 CoVs was localized to aa 1 to 39 and at least one other antigenic site in the N-protein C terminus, differing from the cross-reactive region identified in SARS-CoV N protein. The observed cross-reactivity is not a consequence of a higher level of amino acid identity between SARS-CoV and porcine CoV nucleoproteins, because sequence comparisons indicated that SARS-CoV N protein has amino acid identity similar to that of infectious bronchitis virus N protein and shares a higher level of identity with bovine CoV N protein within the cross-reactive region. The TGEV and SARS-CoV N proteins are RNA chaperons with long disordered regions. We speculate that during natural infection, antibodies target similar short antigenic sites within the N proteins of SARS-CoV and porcine group 1 CoVs that are exposed to an immune response. Identification of the cross-reactive and non-cross-reactive N-protein regions allows development of SARS-CoV-specific antibody assays for screening animal and human sera.
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