Structural basis for enhanced infectivity and immune evasion of SARS-CoV-2 variants

Several fast-spreading variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have become the dominant circulating strains that continue to fuel the COVID-19 pandemic despite intensive vaccination efforts throughout the world. We report here cryo-EM structures of the full-length spike (S) trimers of the B.1.1.7 and B.1.351 variants, as well as their biochemical and antigenic properties. Mutations in the B.1.1.7 protein increase the accessibility of its receptor binding domain and also the binding affinity for receptor angiotensin-converting enzyme 2 (ACE2). The enhanced receptor engagement can account for the increased transmissibility and risk of mortality as the variant may begin to infect efficiently infect additional cell types expressing low levels of ACE2. The B.1.351 variant has evolved to reshape antigenic surfaces of the major neutralizing sites on the S protein, rendering complete resistance to some potent neutralizing antibodies. These findings provide structural details on how the wide spread of SARS-CoV-2 enables rapid evolution to enhance viral fitness and immune evasion. They may guide intervention strategies to control the pandemic.

Download Full-text

Structural basis for enhanced infectivity and immune evasion of SARS-CoV-2 variants

Science ◽

10.1126/science.abi9745 ◽

2021 ◽

pp. eabi9745

Author(s):

Yongfei Cai ◽

Jun Zhang ◽

Tianshu Xiao ◽

Christy L. Lavine ◽

Shaun Rawson ◽

...

Keyword(s):

Immune Evasion ◽

Neutralizing Antibodies ◽

Viral Fitness ◽

Structural Basis ◽

Amino Acid Substitutions ◽

Receptor Binding Domain ◽

Angiotensin Converting Enzyme 2 ◽

Antigenic Properties ◽

Structural Details ◽

Fast Spreading

Several fast-spreading variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have become the dominant circulating strains in the COVID-19 pandemic. We report here cryo-EM structures of the full-length spike (S) trimers of the B.1.1.7 and B.1.351 variants, as well as their biochemical and antigenic properties. Amino acid substitutions in the B.1.1.7 protein increase the accessibility of its receptor binding domain and also the binding affinity for receptor angiotensin-converting enzyme 2 (ACE2). The enhanced receptor engagement may account for the increased transmissibility. The B.1.351 variant has evolved to reshape antigenic surfaces of the major neutralizing sites on the S protein, making it resistant to some potent neutralizing antibodies. These findings provide structural details on how SARS-CoV-2 has evolved to enhance viral fitness and immune evasion.

Download Full-text

Amino acids 484 and 494 of SARS-CoV-2 spike are hotspots of immune evasion affecting antibody but not ACE2 binding

10.1101/2021.04.22.441007 ◽

2021 ◽

Author(s):

Marta Alenquer ◽

Filipe Ferreira ◽

Diana Lousa ◽

Mariana Valério ◽

Mónica Medina-Lopes ◽

...

Keyword(s):

Amino Acids ◽

Amino Acid ◽

Angiotensin Converting Enzyme ◽

Immune Evasion ◽

Neutralizing Antibodies ◽

Receptor Binding Domain ◽

Converting Enzyme ◽

Antibody Neutralization ◽

Angiotensin Converting Enzyme 2 ◽

The Core

AbstractUnderstanding SARS-CoV-2 evolution and host immunity is critical to control COVID-19 pandemics. At the core is an arms-race between SARS-CoV-2 antibody and angiotensin-converting enzyme 2 (ACE2) recognition, a function of the viral protein spike and, predominantly, of its receptor-binding-domain (RBD). Mutations in spike impacting antibody or ACE2 binding are known, but the effect of mutation synergy is less explored. We engineered 22 spike-pseudotyped lentiviruses containing individual and combined mutations, and confirmed that E484K evades antibody neutralization elicited by infection or vaccination, a capacity augmented when complemented by K417N and N501Y mutations. In silico analysis provided an explanation for E484K immune evasion. E484 frequently engages in interactions with antibodies but not with ACE2. Importantly, we identified a novel amino acid of concern, S494, which shares a similar pattern. Using the already circulating mutation S494P, we found that it reduces antibody neutralization of convalescent sera. This amino acid emerges as an additional hotspot for immune evasion and a target for therapies, vaccines and diagnostics.One-Sentence SummaryAmino acids in SARS-CoV-2 spike protein implicated in immune evasion are biased for binding to neutralizing antibodies but dispensable for binding the host receptor angiotensin-converting enzyme 2.

Download Full-text

Structural Evaluation of the Spike Glycoprotein Variants on SARS-CoV-2 Transmission and Immune Evasion

International Journal of Molecular Sciences ◽

10.3390/ijms22147425 ◽

2021 ◽

Vol 22 (14) ◽

pp. 7425

Author(s):

Mohd Zulkifli Salleh ◽

Jeremy P. Derrick ◽

Zakuan Zainy Deris

Keyword(s):

Immune Responses ◽

Immune Evasion ◽

Social Economic ◽

Viral Fitness ◽

Neutralising Antibodies ◽

B Cell Epitopes ◽

Potential Reduction ◽

Angiotensin Converting Enzyme 2 ◽

Structural Evaluation ◽

Alpha Variant

Abstract: The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) presents significant social, economic and political challenges worldwide. SARS-CoV-2 has caused over 3.5 million deaths since late 2019. Mutations in the spike (S) glycoprotein are of particular concern because it harbours the domain which recognises the angiotensin-converting enzyme 2 (ACE2) receptor and is the target for neutralising antibodies. Mutations in the S protein may induce alterations in the surface spike structures, changing the conformational B-cell epitopes and leading to a potential reduction in vaccine efficacy. Here, we summarise how the more important variants of SARS-CoV-2, which include cluster 5, lineages B.1.1.7 (Alpha variant), B.1.351 (Beta), P.1 (B.1.1.28/Gamma), B.1.427/B.1.429 (Epsilon), B.1.526 (Iota) and B.1.617.2 (Delta) confer mutations in their respective spike proteins which enhance viral fitness by improving binding affinity to the ACE2 receptor and lead to an increase in infectivity and transmission. We further discuss how these spike protein mutations provide resistance against immune responses, either acquired naturally or induced by vaccination. This information will be valuable in guiding the development of vaccines and other therapeutics for protection against the ongoing coronavirus disease 2019 (COVID-19) pandemic.

Download Full-text

Signatures in SARS-CoV-2 spike protein conferring escape to neutralizing antibodies

PLoS Pathogens ◽

10.1371/journal.ppat.1009772 ◽

2021 ◽

Vol 17 (8) ◽

pp. e1009772

Author(s):

Marta Alenquer ◽

Filipe Ferreira ◽

Diana Lousa ◽

Mariana Valério ◽

Mónica Medina-Lopes ◽

...

Keyword(s):

Amino Acid ◽

Immune Evasion ◽

Neutralizing Antibodies ◽

Immune Escape ◽

Host Immunity ◽

Arms Race ◽

Spike Protein ◽

Antibody Neutralization ◽

Angiotensin Converting Enzyme 2 ◽

The Core

Understanding SARS-CoV-2 evolution and host immunity is critical to control COVID-19 pandemics. At the core is an arms-race between SARS-CoV-2 antibody and angiotensin-converting enzyme 2 (ACE2) recognition, a function of the viral protein spike. Mutations in spike impacting antibody and/or ACE2 binding are appearing worldwide, imposing the need to monitor SARS-CoV2 evolution and dynamics in the population. Determining signatures in SARS-CoV-2 that render the virus resistant to neutralizing antibodies is critical. We engineered 25 spike-pseudotyped lentiviruses containing individual and combined mutations in the spike protein, including all defining mutations in the variants of concern, to identify the effect of single and synergic amino acid substitutions in promoting immune escape. We confirmed that E484K evades antibody neutralization elicited by infection or vaccination, a capacity augmented when complemented by K417N and N501Y mutations. In silico analysis provided an explanation for E484K immune evasion. E484 frequently engages in interactions with antibodies but not with ACE2. Importantly, we identified a novel amino acid of concern, S494, which shares a similar pattern. Using the already circulating mutation S494P, we found that it reduces antibody neutralization of convalescent and post-immunization sera, particularly when combined with E484K and with mutations able to increase binding to ACE2, such as N501Y. Our analysis of synergic mutations provides a signature for hotspots for immune evasion and for targets of therapies, vaccines and diagnostics.

Download Full-text

Structural basis for bivalent binding and inhibition of SARS-CoV-2 infection by human potent neutralizing antibodies

Cell Research ◽

10.1038/s41422-021-00487-9 ◽

2021 ◽

Author(s):

Renhong Yan ◽

Ruoke Wang ◽

Bin Ju ◽

Jinfang Yu ◽

Yuanyuan Zhang ◽

...

Keyword(s):

Neutralizing Antibodies ◽

Clinical Intervention ◽

Structural Features ◽

Full Length ◽

Structural Basis ◽

Receptor Binding Domain ◽

S Protein ◽

Angiotensin Converting Enzyme 2 ◽

Neutralizing Activity ◽

Potent Neutralization

AbstractNeutralizing monoclonal antibodies (nAbs) to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) represent promising candidates for clinical intervention against coronavirus disease 2019 (COVID-19). We isolated a large number of nAbs from SARS-CoV-2-infected individuals capable of disrupting proper interaction between the receptor binding domain (RBD) of the viral spike (S) protein and the receptor angiotensin converting enzyme 2 (ACE2). However, the structural basis for their potent neutralizing activity remains unclear. Here, we report cryo-EM structures of the ten most potent nAbs in their native full-length IgG-form or in both IgG-form and Fab-form bound to the trimeric S protein of SARS-CoV-2. The bivalent binding of the full-length IgG is found to associate with more RBDs in the “up” conformation than the monovalent binding of Fab, perhaps contributing to the enhanced neutralizing activity of IgG and triggering more shedding of the S1 subunit from the S protein. Comparison of a large number of nAbs identified common and unique structural features associated with their potent neutralizing activities. This work provides a structural basis for further understanding the mechanism of nAbs, especially through revealing the bivalent binding and its correlation with more potent neutralization and the shedding of S1 subunit.

Download Full-text

A noncompeting pair of human neutralizing antibodies block COVID-19 virus binding to its receptor ACE2

Science ◽

10.1126/science.abc2241 ◽

2020 ◽

Vol 368 (6496) ◽

pp. 1274-1278 ◽

Cited By ~ 238

Author(s):

Yan Wu ◽

Feiran Wang ◽

Chenguang Shen ◽

Weiyu Peng ◽

Delin Li ◽

...

Keyword(s):

Neutralizing Antibodies ◽

Immune Escape ◽

Complex Structure ◽

Structural Basis ◽

Virus Binding ◽

Angiotensin Converting Enzyme 2 ◽

Binding Interface ◽

Neutralizing Capacity ◽

Rational Vaccine Design ◽

Therapeutic Study

Neutralizing antibodies could potentially be used as antivirals against the coronavirus disease 2019 (COVID-19) pandemic. Here, we report isolation of four human-origin monoclonal antibodies from a convalescent patient, all of which display neutralization abilities. The antibodies B38 and H4 block binding between the spike glycoprotein receptor binding domain (RBD) of the virus and the cellular receptor angiotensin-converting enzyme 2 (ACE2). A competition assay indicated different epitopes on the RBD for these two antibodies, making them a potentially promising virus-targeting monoclonal antibody pair for avoiding immune escape in future clinical applications. Moreover, a therapeutic study in a mouse model validated that these antibodies can reduce virus titers in infected lungs. The RBD-B38 complex structure revealed that most residues on the epitope overlap with the RBD-ACE2 binding interface, explaining the blocking effect and neutralizing capacity. Our results highlight the promise of antibody-based therapeutics and provide a structural basis for rational vaccine design.

Download Full-text

A bifluorescent-based assay for the identification of neutralizing antibodies against SARS-CoV-2 variants of concern in vitro and in vivo

10.1101/2021.06.28.450214 ◽

2021 ◽

Author(s):

Kevin Chiem ◽

Desarey Morales Vasquez ◽

Jesus Silvas ◽

Jun-Gyu Park ◽

Michael Piepenbrink ◽

...

Keyword(s):

South African ◽

Neutralizing Antibodies ◽

Viral Infections ◽

Rapid Assessment ◽

The United States ◽

Viral Fitness ◽

Angiotensin Converting Enzyme 2 ◽

Protection Efficacy

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) emerged at the end of 2019 and has been responsible for the still ongoing coronavirus disease 2019 (COVID-19) pandemic. Prophylactic vaccines have been authorized by the United States (US) Food and Drug Administration (FDA) for the prevention of COVID-19. Identification of SARS-CoV-2 neutralizing antibodies (NAbs) is important to assess vaccine protection efficacy, including their ability to protect against emerging SARS-CoV-2 variants of concern (VoC). Here we report the generation and use of a recombinant (r)SARS-CoV-2 USA/WA1/2020 (WA-1) strain expressing Venus and a rSARS-CoV-2 expressing mCherry and containing mutations K417N, E484K, and N501Y found in the receptor binding domain (RBD) of the spike (S) glycoprotein of the South African (SA) B.1.351 (beta, β) VoC, in bifluorescent-based assays to rapidly and accurately identify human monoclonal antibodies (hMAbs) able to neutralize both viral infections in vitro and in vivo. Importantly, our bifluorescent-based system accurately recapitulated findings observed using individual viruses. Moreover, fluorescent-expressing rSARS-CoV-2 and the parental wild-type (WT) rSARS-CoV-2 WA-1 had similar viral fitness in vitro, as well as similar virulence and pathogenicity in vivo in the K18 human angiotensin converting enzyme 2 (hACE2) transgenic mouse model of SARS-CoV-2 infection. We demonstrate that these new fluorescent-expressing rSARS-CoV-2 can be used in vitro and in vivo to easily identify hMAbs that simultaneously neutralize different SARS-CoV-2 strains, including VoC, for the rapid assessment of vaccine efficacy or the identification of prophylactic and/or therapeutic broadly NAbs for the treatment of SARS-CoV-2 infection.

Download Full-text

Structural basis for bivalent binding and inhibition of SARS-CoV-2 infection by human potent neutralizing antibodies

10.1101/2020.10.13.336800 ◽

2020 ◽

Author(s):

Renhong Yan ◽

Ruoke Wang ◽

Bin Ju ◽

Jinfang Yu ◽

Yuanyuan Zhang ◽

...

Keyword(s):

Neutralizing Antibodies ◽

Clinical Intervention ◽

Structural Features ◽

Full Length ◽

Structural Basis ◽

Virus Infections ◽

Virus Diseases ◽

S Protein ◽

Angiotensin Converting Enzyme 2 ◽

Potent Neutralization

AbstractNeutralizing monoclonal antibodies (nAbs) to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) represent promising candidates for clinical intervention against coronavirus virus diseases 2019 (COVID-19). We isolated a large number of nAbs from SARS-CoV-2 infected individuals capable of disrupting proper interaction between the receptor binding domain (RBD) of the viral spike (S) protein and the receptor angiotensin converting enzyme 2 (ACE2). In order to understand the mechanism of these nAbs on neutralizing SARS-CoV-2 virus infections, we have performed cryo-EM analysis and here report cryo-EM structures of the ten most potent nAbs in their native full-length IgG or Fab forms bound to the trimeric S protein of SARS-CoV-2. The bivalent binding of the full-length IgG is found to associate with more RBD in the “up” conformation than the monovalent binding of Fab, perhaps contributing to the enhanced neutralizing activity of IgG and triggering more shedding of the S1 subunit from the S protein. Comparison of large number of nAbs identified common and unique structural features associated with their potent neutralizing activities. This work provides structural basis for further understanding the mechanism of nAbs, especially through revealing the bivalent binding and their correlation with more potent neutralization and the shedding of S1 subunit.

Download Full-text

The Host Interactome of Spike Expands the Tropism of SARS-CoV-2

10.1101/2021.02.16.431318 ◽

2021 ◽

Author(s):

Casimir Bamberger ◽

Sandra Pankow ◽

Salvador Martínez-Bartolomé ◽

Jolene Diedrich ◽

Robin Park ◽

...

Keyword(s):

Epithelial Cells ◽

Enrichment Analysis ◽

Cell Types ◽

Extracellular Proteins ◽

Spike Protein ◽

Angiotensin Converting Enzyme 2 ◽

Global Pandemic ◽

Additional Cell ◽

Progression Of Disease ◽

Different Cell Types

AbstractThe SARS-CoV-2 virus causes severe acute respiratory syndrome (COVID-19) and has rapidly created a global pandemic. Patients that survive may face a slow recovery with long lasting side effects that can afflict different organs. SARS-CoV-2 primarily infects epithelial airway cells that express the host entry receptor Angiotensin Converting Enzyme 2 (ACE2) which binds to spike protein trimers on the surface of SARS-CoV-2 virions. However, SARS-CoV-2 can spread to other tissues even though they are negative for ACE2. To gain insight into the molecular constituents that might influence SARS-CoV-2 tropism, we determined which additional host factors engage with the viral spike protein in disease-relevant human bronchial epithelial cells (16HBEo−). We found that spike recruited the extracellular proteins laminin and thrombospondin and was retained in the endoplasmatic reticulum (ER) by the proteins DJB11 and FBX2 which support re-folding or degradation of nascent proteins in the ER. Because emerging mutations of the spike protein potentially impact the virus tropism, we compared the interactome of D614 spike with that of the rapidly spreading G614 mutated spike. More D614 than G614 spike associated with the proteins UGGT1, calnexin, HSP7A and GRP78/BiP which ensure glycosylation and folding of proteins in the ER. In contrast to G614 spike, D614 spike was endoproteolytically cleaved, and the N-terminal S1 domain was degraded in the ER even though C-terminal ‘S2 only’ proteoforms remained present. D614 spike also bound more laminin than G614 spike, which suggested that extracellular laminins may function as co-factor for an alternative, ‘S2 only’ dependent virus entry. Because the host interactome determines whether an infection is productive, we developed a novel proteome-based cell type set enrichment analysis (pCtSEA). With pCtSEA we determined that the host interactome of the spike protein may extend the tropism of SARS-CoV-2 beyond mucous epithelia to several different cell types, including macrophages and epithelial cells in the nephron. An ‘S2 only’ dependent, alternative infection of additional cell types with SARS-CoV-2 may impact vaccination strategies and may provide a molecular explanation for a severe or prolonged progression of disease in select COVID-19 patients.

Download Full-text

Structural basis for neutralization of SARS-CoV-2 and SARS-CoV by a potent therapeutic antibody

Science ◽

10.1126/science.abc5881 ◽

2020 ◽

Vol 369 (6510) ◽

pp. 1505-1509 ◽

Cited By ~ 35

Author(s):

Zhe Lv ◽

Yong-Qiang Deng ◽

Qing Ye ◽

Lei Cao ◽

Chun-Yun Sun ◽

...

Keyword(s):

Neutralizing Antibodies ◽

Conformational Epitope ◽

Therapeutic Interventions ◽

Structural Basis ◽

Fab Fragment ◽

Health Crisis ◽

Angiotensin Converting Enzyme 2 ◽

Humanized Monoclonal Antibody ◽

Microscopy Characterization

The coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has resulted in an unprecedented public health crisis. There are no approved vaccines or therapeutics for treating COVID-19. Here we report a humanized monoclonal antibody, H014, that efficiently neutralizes SARS-CoV-2 and SARS-CoV pseudoviruses as well as authentic SARS-CoV-2 at nanomolar concentrations by engaging the spike (S) receptor binding domain (RBD). H014 administration reduced SARS-CoV-2 titers in infected lungs and prevented pulmonary pathology in a human angiotensin-converting enzyme 2 mouse model. Cryo–electron microscopy characterization of the SARS-CoV-2 S trimer in complex with the H014 Fab fragment unveiled a previously uncharacterized conformational epitope, which was only accessible when the RBD was in an open conformation. Biochemical, cellular, virological, and structural studies demonstrated that H014 prevents attachment of SARS-CoV-2 to its host cell receptors. Epitope analysis of available neutralizing antibodies against SARS-CoV and SARS-CoV-2 uncovered broad cross-protective epitopes. Our results highlight a key role for antibody-based therapeutic interventions in the treatment of COVID-19.

Download Full-text