Aided by extensive spike protein mutation, the SARS-CoV-2 Omicron variant overtook the previously dominant Delta variant. Spike conformation plays an essential role in SARS-CoV-2 evolution via ...changes in receptor-binding domain (RBD) and neutralizing antibody epitope presentation, affecting virus transmissibility and immune evasion. Here, we determine cryo-EM structures of the Omicron and Delta spikes to understand the conformational impacts of mutations in each. The Omicron spike structure revealed an unusually tightly packed RBD organization with long range impacts that were not observed in the Delta spike. Binding and crystallography revealed increased flexibility at the functionally critical fusion peptide site in the Omicron spike. These results reveal a highly evolved Omicron spike architecture with possible impacts on its high levels of immune evasion and transmissibility.
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•Omicron S architecture differs from Delta and other variants•Tight packing of Omicron S RBDs results in unique up- and down-state arrangements•3-RBD-down Omicron S stabilizes the rearrangement of the NTD-to-RBD (N2R) linker•S2 subunit conformational changes lead to altered fusion peptide dynamics
Gobeil, Henderson, Stalls et al. identify diverse Omicron S ectodomain conformations demonstrating altered architecture that exhibits tight packing of the 3-RBD-down state, NTD-to-RBD (N2R) linker rearrangements, and changes in fusion peptide conformational dynamics. These distinct conformational features of its S protein may underlie Omicron’s higher transmissibility and immune evasion.
The SARS-CoV-2 spike (S) protein, a primary target for COVID-19 vaccine development, presents its receptor binding domain in two conformations, the receptor-accessible 'up' or receptor-inaccessible ...'down' states. Here we report that the commonly used stabilized S ectodomain construct '2P' is sensitive to cold temperatures, and this cold sensitivity is abrogated in a 'down' state-stabilized ectodomain. Our findings will impact structural, functional and vaccine studies that use the SARS-CoV-2 S ectodomain.
SARS-CoV-2-neutralizing antibodies (NAbs) protect against COVID-19. A concern regarding SARS-CoV-2 antibodies is whether they mediate disease enhancement. Here, we isolated NAbs against the ...receptor-binding domain (RBD) or the N-terminal domain (NTD) of SARS-CoV-2 spike from individuals with acute or convalescent SARS-CoV-2 or a history of SARS-CoV infection. Cryo-electron microscopy of RBD and NTD antibodies demonstrated function-specific modes of binding. Select RBD NAbs also demonstrated Fc receptor-γ (FcγR)-mediated enhancement of virus infection in vitro, while five non-neutralizing NTD antibodies mediated FcγR-independent in vitro infection enhancement. However, both types of infection-enhancing antibodies protected from SARS-CoV-2 replication in monkeys and mice. Three of 46 monkeys infused with enhancing antibodies had higher lung inflammation scores compared to controls. One monkey had alveolar edema and elevated bronchoalveolar lavage inflammatory cytokines. Thus, while in vitro antibody-enhanced infection does not necessarily herald enhanced infection in vivo, increased lung inflammation can rarely occur in SARS-CoV-2 antibody-infused macaques.
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•RBD or NTD antibodies exhibited infection enhancement in vitro but not in vivo•Neutralizing or infection-enhancing NTD antibodies bound distinct epitopes•In vitro infection-enhancing antibodies protected from SARS-CoV-2 in vivo•Cross-reactive RBD-neutralizing antibodies were protective—most potent DH1047
Convalescent human-derived SARS-CoV-2 RBD and NTD antibodies mediated neutralization as well as infection enhancement in vitro, yet infusion of these antibodies in mice or cynomolgus macaques resulted in suppression of virus replication.
The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron BA.2 sub-lineage has gained in proportion relative to BA.1. Because spike (S) protein variations may underlie differences in ...their pathobiology, here we determine cryoelectron microscopy (cryo-EM) structures of the BA.2 S ectodomain and compare these with previously determined BA.1 S structures. BA.2 receptor-binding domain (RBD) mutations induce remodeling of the RBD structure, resulting in tighter packing and improved thermostability. Interprotomer RBD interactions are enhanced in the closed (or 3-RBD-down) BA.2 S, while the fusion peptide is less accessible to antibodies than in BA.1. Binding and pseudovirus neutralization assays reveal extensive immune evasion while defining epitopes of two outer RBD face-binding antibodies, DH1044 and DH1193, that neutralize both BA.1 and BA.2. Taken together, our results indicate that stabilization of the closed state through interprotomer RBD-RBD packing is a hallmark of the Omicron variant and show differences in key functional regions in the BA.1 and BA.2 S proteins.
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•Omicron BA.2 spike (S) protein architecture differs from BA.1 S•Mutation-induced remodeling of interfacial loops results in more stable BA.2 RBD•Remodeled BA.2 RBD shows better interprotomer packing in 3-RBD-down (or closed) S•Fusion peptide in BA.2 S protein is less accessible to antibodies than in BA.1
Stalls et al. determine Omicron BA.2 S structures indicating remodeled RBD loops leading to a more thermostable RBD that is better packed within the 3-RBD-down spike and loss of class 4 RBD directed antibody binding. Enhanced spike stability and immune evasion may contribute to BA.2 efficiently outcompeting BA.1.
Natural antibodies (Abs) can target host glycans on the surface of pathogens. We studied the evolution of glycan-reactive B cells of rhesus macaques and humans using glycosylated HIV-1 envelope (Env) ...as a model antigen. 2G12 is a broadly neutralizing Ab (bnAb) that targets a conserved glycan patch on Env of geographically diverse HIV-1 strains using a unique heavy-chain (VH) domain-swapped architecture that results in fragment antigen-binding (Fab) dimerization. Here, we describe HIV-1 Env Fab-dimerized glycan (FDG)-reactive bnAbs without VH-swapped domains from simian-human immunodeficiency virus (SHIV)-infected macaques. FDG Abs also recognized cell-surface glycans on diverse pathogens, including yeast and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike. FDG precursors were expanded by glycan-bearing immunogens in macaques and were abundant in HIV-1-naive humans. Moreover, FDG precursors were predominately mutated IgM+IgD+CD27+, thus suggesting that they originated from a pool of antigen-experienced IgM+ or marginal zone B cells.
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•Fab-dimerized glycan-reactive (FDG) natural antibodies (Abs) are prevalent in primates•Fab-dimerization can occur independent of VH domain-swapping•Precursor FDG natural Abs acquire neutralization breadth in retroviral infection•FDG Abs recognize a quaternary epitope in the S2 subunit of SARS-CoV-2 spike
Structural and functional analyses identify a category of glycan-reactive antibodies in macaques and humans that are marked by the dimerization of the antigen-binding fragment. These antibodies are involved in HIV neutralization and also recognize the S2 protein of SARS-CoV-2.
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