Structure of a Human Astrovirus Capsid-Antibody Complex and Mechanistic Insights into Virus Neutralization

ABSTRACT Human astroviruses (HAstVs) are a leading cause of viral diarrhea in young children, the immunocompromised, and the elderly. There are no vaccines or antiviral therapies against HAstV disease. Several lines of evidence point to the presence of protective antibodies in healthy adults as a mechanism governing protection against reinfection by HAstV. However, development of anti-HAstV therapies is hampered by the gap in knowledge of protective antibody epitopes on the HAstV capsid surface. Here, we report the structure of the HAstV capsid spike domain bound to the neutralizing monoclonal antibody PL-2. The antibody uses all six complementarity-determining regions to bind to a quaternary epitope on each side of the dimeric capsid spike. We provide evidence that the HAstV capsid spike is a receptor-binding domain and that the antibody neutralizes HAstV by blocking virus attachment to cells. We identify patches of conserved amino acids that overlap the antibody epitope and may comprise a receptor-binding site. Our studies provide a foundation for the development of therapies to prevent and treat HAstV diarrheal disease. IMPORTANCE Human astroviruses (HAstVs) infect nearly every person in the world during childhood and cause diarrhea, vomiting, and fever. Despite the prevalence of this virus, little is known about how antibodies in healthy adults protect them against reinfection. Here, we determined the crystal structure of a complex of the HAstV capsid protein and a virus-neutralizing antibody. We show that the antibody binds to the outermost spike domain of the capsid, and we provide evidence that the antibody blocks virus attachment to human cells. Importantly, our findings suggest that a subunit-based vaccine focusing the immune system on the HAstV capsid spike domain could be effective in protecting children against HAstV disease.

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Structures of Two Human Astrovirus Capsid/Neutralizing Antibody Complexes Reveal Distinct Epitopes and Inhibition of Virus Attachment to Cells

Journal of Virology ◽

10.1128/jvi.01415-21 ◽

2021 ◽

Author(s):

Lena Ricemeyer ◽

Nayeli Aguilar-Hernández ◽

Tomás López ◽

Rafaela Espinosa ◽

Sarah Lanning ◽

...

Keyword(s):

Receptor Binding ◽

Neutralizing Antibodies ◽

Severe Disease ◽

Neutralizing Antibody ◽

Host Cells ◽

Viral Gastroenteritis ◽

Receptor Binding Site ◽

Virus Attachment ◽

Human Astroviruses ◽

Human Astrovirus

Human astrovirus is an important cause of viral gastroenteritis worldwide. Young children, the elderly, and the immunocompromised are especially at risk for contracting severe disease. However, no vaccines exist to combat human astrovirus infection. Evidence points to the importance of antibodies in enabling protection of healthy adults from reinfection. To develop an effective subunit vaccine that broadly protects against diverse astrovirus serotypes, we must understand how neutralizing antibodies target the capsid surface at the molecular level. Here, we report the structures of the human astrovirus capsid spike domain bound to two neutralizing monoclonal antibodies. These antibodies bind two distinct conformational epitopes on the spike surface. We add to existing evidence that the human astrovirus capsid spike contains a receptor-binding domain and demonstrate that both antibodies neutralize human astrovirus by blocking virus attachment to host cells. We identify patches of conserved amino acids that overlap or border the antibody epitopes and may constitute a receptor-binding site. Our findings provide a basis to develop therapies that prevent and treat human astrovirus gastroenteritis. Importance Human astroviruses infect nearly every person in the world during childhood and cause diarrhea, vomiting, and fever. Despite the prevalence of this virus, little is known about how antibodies block virus infection. Here, we determined crystal structures of the astrovirus capsid protein in complex with two virus-neutralizing antibodies. We show that the antibodies bind two distinct sites on the capsid spike domain; however, both antibodies block virus attachment to human cells. Importantly, our findings support the use of the human astrovirus capsid spike as an antigen in a subunit-based vaccine to prevent astrovirus disease.

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Epitope–Paratope Interaction of a Neutralizing Human Anti-Hepatitis B Virus PreS1 Antibody That Recognizes the Receptor-Binding Motif

Vaccines ◽

10.3390/vaccines9070754 ◽

2021 ◽

Vol 9 (7) ◽

pp. 754

Author(s):

Jisu Hong ◽

Youngjin Choi ◽

Yoonjoo Choi ◽

Jiwoo Lee ◽

Hyo Jeong Hong

Keyword(s):

Hepatitis B Virus ◽

Hepatitis B ◽

Receptor Binding ◽

Neutralizing Antibody ◽

Hbv Infection ◽

Binding Motif ◽

Alanine Scanning Mutagenesis ◽

B Virus ◽

Complementarity Determining Regions

Hepatitis B virus (HBV) is a global health burden that causes acute and chronic hepatitis. To develop an HBV-neutralizing antibody that effectively prevents HBV infection, we previously generated a human anti-preS1 monoclonal antibody (1A8) that binds to genotypes A–D and validated its HBV-neutralizing activity in vitro. In the present study, we aimed to determine the fine epitope and paratope of 1A8 to understand the mechanism of HBV neutralization. We performed alanine-scanning mutagenesis on the preS1 (aa 19–34, genotype C) and the heavy (HCDR) and light (LCDR) chain complementarity-determining regions. The 1A8 recognized the three residues (Leu22, Gly23, and Phe25) within the highly conserved receptor-binding motif (NPLGFFP) of the preS1, while four CDR residues of 1A8 were critical in antigen binding. Structural analysis of the epitope–paratope interaction by molecular modeling revealed that Leu100 in the HCDR3, Ala50 in the HCDR2, and Tyr96 in the LCDR3 closely interacted with Leu22, Gly23, and Phe25 of the preS1. Additionally, we found that 1A8 also binds to the receptor-binding motif (NPLGFLP) of infrequently occurring HBV. The results suggest that 1A8 may broadly and effectively block HBV entry and thus have potential as a promising candidate for the prevention and treatment of HBV infection.

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SARS-CoV-2 spike protein arrested in the closed state induces potent neutralizing responses

10.1101/2021.01.14.426695 ◽

2021 ◽

Author(s):

George W. Carnell ◽

Katarzyna A. Ciazynska ◽

David A. Wells ◽

Xiaoli Xiong ◽

Ernest T. Aguinam ◽

...

Keyword(s):

Receptor Binding ◽

Neutralizing Antibodies ◽

Neutralizing Antibody ◽

Protein S ◽

Spike Protein ◽

Receptor Binding Site ◽

S Protein ◽

Closed State ◽

S Proteins ◽

Protein Constructs

AbstractThe majority of SARS-CoV-2 vaccines in use or in advanced clinical development are based on the viral spike protein (S) as their immunogen. S is present on virions as pre-fusion trimers in which the receptor binding domain (RBD) is stochastically open or closed. Neutralizing antibodies have been described that act against both open and closed conformations. The long-term success of vaccination strategies will depend upon inducing antibodies that provide long-lasting broad immunity against evolving, circulating SARS-CoV-2 strains, while avoiding the risk of antibody dependent enhancement as observed with other Coronavirus vaccines. Here we have assessed the results of immunization in a mouse model using an S protein trimer that is arrested in the closed state to prevent exposure of the receptor binding site and therefore interaction with the receptor. We compared this with a range of other modified S protein constructs, including representatives used in current vaccines. We found that all trimeric S proteins induce a long-lived, strongly neutralizing antibody response as well as T-cell responses. Notably, the protein binding properties of sera induced by the closed spike differed from those induced by standard S protein constructs. Closed S proteins induced more potent neutralising responses than expected based on the degree to which they inhibit interactions between the RBD and ACE2. These observations suggest that closed spikes recruit different, but equally potent, virus-inhibiting immune responses than open spikes, and that this is likely to include neutralizing antibodies against conformational epitopes present in the closed conformation. Together with their improved stability and storage properties we suggest that closed spikes may be a valuable component of refined, next-generation vaccines.

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Structural basis of Chikungunya virus inhibition by monoclonal antibodies

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2008051117 ◽

2020 ◽

Vol 117 (44) ◽

pp. 27637-27645

Author(s):

Qun Fei Zhou ◽

Julie M. Fox ◽

James T. Earnest ◽

Thiam-Seng Ng ◽

Arthur S. Kim ◽

...

Keyword(s):

Monoclonal Antibodies ◽

Chikungunya Virus ◽

Neutralizing Antibody ◽

Structural Basis ◽

Chikv Infection ◽

Receptor Binding Site ◽

Viral Pathogen ◽

Virus Attachment

Chikungunya virus (CHIKV) is an emerging viral pathogen that causes both acute and chronic debilitating arthritis. Here, we describe the functional and structural basis as to how two anti-CHIKV monoclonal antibodies, CHK-124 and CHK-263, potently inhibit CHIKV infection in vitro and in vivo. Our in vitro studies show that CHK-124 and CHK-263 block CHIKV at multiple stages of viral infection. CHK-124 aggregates virus particles and blocks attachment. Also, due to antibody-induced virus aggregation, fusion with endosomes and egress are inhibited. CHK-263 neutralizes CHIKV infection mainly by blocking virus attachment and fusion. To determine the structural basis of neutralization, we generated cryogenic electron microscopy reconstructions of Fab:CHIKV complexes at 4- to 5-Å resolution. CHK-124 binds to the E2 domain B and overlaps with the Mxra8 receptor-binding site. CHK-263 blocks fusion by binding an epitope that spans across E1 and E2 and locks the heterodimer together, likely preventing structural rearrangements required for fusion. These results provide structural insight as to how neutralizing antibody engagement of CHIKV inhibits different stages of the viral life cycle, which could inform vaccine and therapeutic design.

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A human neutralizing antibody targets the receptor-binding site of SARS-CoV-2

Nature ◽

10.1038/s41586-020-2381-y ◽

2020 ◽

Vol 584 (7819) ◽

pp. 120-124 ◽

Cited By ~ 221

Author(s):

Rui Shi ◽

Chao Shan ◽

Xiaomin Duan ◽

Zhihai Chen ◽

Peipei Liu ◽

...

Keyword(s):

Binding Site ◽

Receptor Binding ◽

Neutralizing Antibody ◽

Receptor Binding Site

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Broadly neutralizing antibodies to SARS-related viruses can be readily induced in rhesus macaques

10.1101/2021.07.05.451222 ◽

2021 ◽

Author(s):

Wan-ting He ◽

Meng Yuan ◽

Sean Callaghan ◽

Rami Musharrafieh ◽

Ge Song ◽

...

Keyword(s):

Receptor Binding ◽

Neutralizing Antibodies ◽

Rhesus Macaques ◽

Evaluation Studies ◽

Neutralizing Antibody ◽

Primate Species ◽

Vaccine Response ◽

Cell Repertoire ◽

Receptor Binding Site ◽

Human Antibodies

To prepare for future coronavirus (CoV) pandemics, it is desirable to generate vaccines capable of eliciting neutralizing antibody responses against multiple CoVs. Because of the phylogenetic similarity to humans, rhesus macaques are an animal model of choice for many virus-challenge and vaccine-evaluation studies, including SARS-CoV-2. Here, we show that immunization of macaques with SARS-CoV-2 spike (S) protein generates potent receptor binding domain cross-neutralizing antibody (nAb) responses to both SARS-CoV-2 and SARS-CoV-1, in contrast to human infection or vaccination where responses are typically SARS-CoV-2-specific. Furthermore, the macaque nAbs are equally effective against SARS-CoV-2 variants of concern. Structural studies show that different immunodominant sites are targeted by the two primate species. Human antibodies generally target epitopes strongly overlapping the ACE2 receptor binding site (RBS), whereas the macaque antibodies recognize a relatively conserved region proximal to the RBS that represents another potential pan-SARS-related virus site rarely targeted by human antibodies. B cell repertoire differences between the two primates appear to significantly influence the vaccine response and suggest care in the use of rhesus macaques in evaluation of vaccines to SARS-related viruses intended for human use.

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One-shot identification of SARS-CoV-2 S RBD escape mutants using yeast screening

10.1101/2021.03.15.435309 ◽

2021 ◽

Author(s):

Irene Francino Urdaniz ◽

Paul J. Steiner ◽

Monica B. Kirby ◽

Fangzhu Zhao ◽

Cyrus M. Haas ◽

...

Keyword(s):

Binding Site ◽

Rapid Method ◽

Receptor Binding ◽

Neutralizing Antibody ◽

Therapeutic Strategies ◽

Recognition Site ◽

Community Resource ◽

Receptor Binding Site ◽

The Public ◽

Escape Mutants

ABSTRACTThe potential emergence of SARS-CoV-2 Spike (S) escape mutants is a threat to reduce the efficacy of existing vaccines and neutralizing antibody (nAb) therapies. An understanding of the antibody/S escape mutations landscape is urgently needed to preemptively address this threat. Here we describe a rapid method to identify escape mutants for nAbs targeting the S receptor binding site. We identified escape mutants for five nAbs, including three from the public germline class VH3-53 elicited by natural COVID-19 infection. Escape mutations predominantly mapped to the periphery of the ACE2 recognition site on the RBD with K417, D420, Y421, F486, and Q493 as notable hotspots. We provide libraries, methods, and software as an openly available community resource to accelerate new therapeutic strategies against SARS-CoV-2.One Sentence SummaryWe present a facile method to identify antibody escape mutants on SARS-CoV-2 S RBD.

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A highly conserved cryptic epitope in the receptor binding domains of SARS-CoV-2 and SARS-CoV

Science ◽

10.1126/science.abb7269 ◽

2020 ◽

Vol 368 (6491) ◽

pp. 630-633 ◽

Cited By ~ 407

Author(s):

Meng Yuan ◽

Nicholas C. Wu ◽

Xueyong Zhu ◽

Chang-Chun D. Lee ◽

Ray T. Y. So ◽

...

Keyword(s):

Receptor Binding ◽

Neutralizing Antibody ◽

Receptor Binding Domain ◽

Receptor Binding Site ◽

S Protein ◽

Antibody Recognition ◽

Binding Domains ◽

Binding Epitope ◽

Cryptic Epitope ◽

Conserved Epitope

The outbreak of coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome–coronavirus 2 (SARS-CoV-2) has now become a pandemic, but there is currently very little understanding of the antigenicity of the virus. We therefore determined the crystal structure of CR3022, a neutralizing antibody previously isolated from a convalescent SARS patient, in complex with the receptor binding domain (RBD) of the SARS-CoV-2 spike (S) protein at 3.1-angstrom resolution. CR3022 targets a highly conserved epitope, distal from the receptor binding site, that enables cross-reactive binding between SARS-CoV-2 and SARS-CoV. Structural modeling further demonstrates that the binding epitope can only be accessed by CR3022 when at least two RBDs on the trimeric S protein are in the “up” conformation and slightly rotated. These results provide molecular insights into antibody recognition of SARS-CoV-2.

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