Respiratory syncytial virus fusion glycoprotein: further characterization of a major epitope involved in virus neutralization

1987 ◽  
Vol 33 (10) ◽  
pp. 933-938 ◽  
Author(s):  
Michel Trudel ◽  
Francine Nadon ◽  
Cécile Séguin ◽  
Pierre Payment ◽  
Pierre J. Talbot
Keyword(s):  
Respiratory Syncytial Virus ◽  
Fusion Protein ◽  
Neutralizing Antibodies ◽  
Peptide Mapping ◽  
Antigenic Site ◽  
Virus Neutralization ◽  
Protein Cleavage ◽  
Biological Assays ◽  
Fusion Glycoprotein ◽  
Syncytial Virus

Competition experiments and biological assays with a panel of 15 monoclonal antibodies confirmed the presence of at least four antigenic sites on the fusion protein of human respiratory syncytial virus, three of which were involved in virus neutralization. One antigenic site, recognized by two strongly neutralizing antibodies, was conserved after reduction and denaturation and shown by immunoblotting to be localized on the F1 fragment of the fusion protein. Cleavage of this protein with staphylococcal protease V8 or papain produced a series of smaller peptides from 11 to 7 kilodaltons that retained this important neutralization determinant. Compared with the other neutralization sites, the epitope defined by monoclonal antibody 7C2 thus appears as the major neutralization epitope. Our peptide mapping results support the hypothesis that this major epitope is composed of a continuous sequence on the viral genome.

scholarly journals Structure of a Major Antigenic Site on the Respiratory Syncytial Virus Fusion Glycoprotein in Complex with Neutralizing Antibody 101F

2010 ◽  
Vol 84 (23) ◽  
pp. 12236-12244 ◽  
Author(s):  
Jason S. McLellan ◽  
Man Chen ◽  
Jung-San Chang ◽  
Yongping Yang ◽  
Albert Kim ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Neutralizing Antibodies ◽  
Antigenic Site ◽  
Neutralizing Antibody ◽  
Structural Basis ◽  
Effective Vaccine ◽  
Linear Epitope ◽  
Peptide Epitope ◽  
Fusion Glycoprotein ◽  
Syncytial Virus

ABSTRACT Respiratory syncytial virus (RSV) is a major cause of pneumonia and bronchiolitis in infants and elderly people. Currently there is no effective vaccine against RSV, but passive prophylaxis with neutralizing antibodies reduces hospitalizations. To investigate the mechanism of antibody-mediated RSV neutralization, we undertook structure-function studies of monoclonal antibody 101F, which binds a linear epitope in the RSV fusion glycoprotein. Crystal structures of the 101F antigen-binding fragment in complex with peptides from the fusion glycoprotein defined both the extent of the linear epitope and the interactions of residues that are mutated in antibody escape variants. The structure allowed for modeling of 101F in complex with trimers of the fusion glycoprotein, and the resulting models suggested that 101F may contact additional surfaces located outside the linear epitope. This hypothesis was supported by surface plasmon resonance experiments that demonstrated 101F bound the peptide epitope ∼16,000-fold more weakly than the fusion glycoprotein. The modeling also showed no substantial clashes between 101F and the fusion glycoprotein in either the pre- or postfusion state, and cell-based assays indicated that 101F neutralization was not associated with blocking virus attachment. Collectively, these results provide a structural basis for RSV neutralization by antibodies that target a major antigenic site on the fusion glycoprotein.

scholarly journals Characterization of Epitope-Specific Anti-Respiratory Syncytial Virus (Anti-RSV) Antibody Responses after Natural Infection and after Vaccination with Formalin-Inactivated RSV

2016 ◽  
Vol 90 (13) ◽  
pp. 5965-5977 ◽  
Author(s):  
Ivy Widjaja ◽  
Oliver Wicht ◽  
Willem Luytjes ◽  
Kees Leenhouts ◽  
Peter J. M. Rottier ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Neutralizing Antibodies ◽  
Antigenic Site ◽  
Virus Neutralization ◽  
Specific Antibodies ◽  
F Protein ◽  
Human Sera ◽  
Cotton Rats ◽  
Syncytial Virus ◽  
Antibody Levels

ABSTRACTAntibodies against the fusion (F) protein of respiratory syncytial virus (RSV) play an important role in the protective immune response to this important respiratory virus. Little is known, however, about antibody levels against multiple F-specific epitopes induced by infection or after vaccination against RSV, while this is important to guide the evaluation of (novel) vaccines. In this study, we analyzed antibody levels against RSV proteins and F-specific epitopes in human sera and in sera of vaccinated and experimentally infected cotton rats and the correlation thereof with virus neutralization. Analysis of human sera revealed substantial diversity in antibody levels against F-, G (attachment)-, and F-specific epitopes between individuals. The highest correlation with virus neutralization was observed for antibodies recognizing prefusion-specific antigenic site Ø. Nevertheless, our results indicate that high levels of antibodies targeting other parts of the F protein can also mediate a potent antiviral antibody response. In agreement, sera of experimentally infected cotton rats contained high neutralizing activity despite lacking antigenic site Ø-specific antibodies. Strikingly, vaccination with formalin-inactivated RSV (FI-RSV) exclusively resulted in the induction of poorly neutralizing antibodies against postfusion-specific antigenic site I, although antigenic sites I, II, and IV were efficiently displayed in FI-RSV. The apparent immunodominance of antigenic site I in FI-RSV likely explains the low levels of neutralizing antibodies upon vaccination and challenge and may play a role in the vaccination-induced enhancement of disease observed with such preparations.IMPORTANCERSV is an importance cause of hospitalization of infants. The development of a vaccine against RSV has been hampered by the disastrous results obtained with FI-RSV vaccine preparations in the 1960s that resulted in vaccination-induced enhancement of disease. To get a better understanding of the antibody repertoire induced after infection or after vaccination against RSV, we investigated antibody levels against fusion (F) protein, attachment (G) protein, and F-specific epitopes in human and animal sera. The results indicate the importance of prefusion-specific antigenic site Ø antibodies as well as of antibodies targeting other epitopes in virus neutralization. However, vaccination of cotton rats with FI-RSV specifically resulted in the induction of weakly neutralizing, antigenic site I-specific antibodies, which may play a role in the enhancement of disease observed after vaccination with such preparations.

The Design of Vaccines Based on the Shielding of Antigenic Site Ø of a Respiratory Syncytial Virus Fusion Protein Immunogen

2020 ◽  
pp. 2000714
Author(s):  
Steven J. Frey ◽  
Chad Varner ◽  
Ammar Arsiwala ◽  
Michael G. Currier ◽  
Martin L. Moore ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Fusion Protein ◽  
Antigenic Site ◽  
Virus Fusion ◽  
Syncytial Virus

scholarly journals Trivalency of a Nanobody Specific for the Human Respiratory Syncytial Virus Fusion Glycoprotein Drastically Enhances Virus Neutralization and Impacts Escape Mutant Selection

2016 ◽  
Vol 60 (11) ◽  
pp. 6498-6509 ◽  
Author(s):  
Concepción Palomo ◽  
Vicente Mas ◽  
Laurent Detalle ◽  
Erik Depla ◽  
Olga Cano ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Antigenic Site ◽  
Serial Passage ◽  
Human Respiratory Syncytial Virus ◽  
Escape Mutant ◽  
Comprehensive Description ◽  
Mutant Selection ◽  
Fusion Glycoprotein ◽  
Escape Mutants ◽  
Syncytial Virus

ABSTRACTALX-0171 is a trivalent Nanobody derived from monovalent Nb017 that binds to antigenic site II of the human respiratory syncytial virus (hRSV) fusion (F) glycoprotein. ALX-0171 is about 6,000 to 10,000 times more potent than Nb017 in neutralization tests with strains of hRSV antigenic groups A and B. To explore the effect of this enhanced neutralization on escape mutant selection, viruses resistant to either ALX-0171 or Nb017 were isolated after serial passage of the hRSV Long strain in the presence of suboptimal concentrations of the respective Nanobodies. Resistant viruses emerged notably faster with Nb017 than with ALX-0171 and in both cases contained amino acid changes in antigenic site II of hRSV F. Detailed binding and neutralization analyses of these escape mutants as well as previously described mutants resistant to certain monoclonal antibodies (MAbs) offered a comprehensive description of site II mutations which are relevant for neutralization by MAbs and Nanobodies. Notably, ALX-0171 showed a sizeable neutralization potency with most escape mutants, even with some of those selected with the Nanobody, and these findings make ALX-0171 an attractive antiviral for treatment of hRSV infections.

Immunovirological studies on human respiratory syncytial virus structural proteins

1986 ◽  
Vol 32 (1) ◽  
pp. 15-21 ◽  
Author(s):  
Michel Trudel ◽  
Francine Nadon ◽  
Cécile Séguin ◽  
Simone Ghoubril ◽  
Pierre Payment ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Fusion Protein ◽  
Neutralizing Antibodies ◽  
Sodium Dodecyl ◽  
Human Respiratory Syncytial Virus ◽  
Structural Proteins ◽  
Membrane Glycoproteins ◽  
Protein Dimer ◽  
Syncytial Virus ◽  
Nucleoprotein N

Immunovirological studies suggest that human respiratory syncytial virus may well be composed of five structural proteins as are other members of the Paramyxoviridae family: the two external membrane glycoproteins H (90 000) and Fo (F1, 49 000; F2, 20 000; disulfide linked), the internal membrane protein M (34 000), the nucleoprotein N (42 000), and a protein (78 000) designated P that could be the equivalent of the polymerase of the morbillivirus and paramyxovirus genus. Neutralizing monoclonal antibodies showed, by immunoprecipitation and immunoblotting, that the fusion protein carries neutralizing epitopes. One monoclonal antibody, which shows a high neutralizing titer, immunoblotted directly with the F1 fragment (49 000) of the fusion protein. Analysis in mice of the immunogenicity of the structural proteins separated on sodium dodecyl sulphate gels indicated that, under our conditions, only the fusion protein dimer Fo and its F1 fragment were capable of inducing neutralizing antibodies.

scholarly journals Replication-Competent or Attenuated, Nonpropagating Vesicular Stomatitis Viruses Expressing Respiratory Syncytial Virus (RSV) Antigens Protect Mice against RSV Challenge

2001 ◽  
Vol 75 (22) ◽  
pp. 11079-11087 ◽  
Author(s):  
Jeffrey S. Kahn ◽  
Anjeanette Roberts ◽  
Carla Weibel ◽  
Linda Buonocore ◽  
John K. Rose
Keyword(s):  
Respiratory Syncytial Virus ◽  
Neutralizing Antibodies ◽  
Protective Immunity ◽  
Enzyme Linked Immunosorbent Assay ◽  
Complete Protection ◽  
Attractive Candidate ◽  
Fusion Glycoprotein ◽  
Vesicular Stomatitis ◽  
Syncytial Virus ◽  
Detectable Serum

ABSTRACT Foreign glycoproteins expressed in recombinant vesicular stomatitis virus (VSV) can elicit specific and protective immunity in the mouse model. We have previously demonstrated the expression of respiratory syncytial virus (RSV) G (attachment) and F (fusion) glycoprotein genes in recombinant VSV. In this study, we demonstrate the expression of RSV F and G glycoproteins in attenuated, nonpropagating VSVs which lack the VSV G gene (VSVΔG) and the incorporation of these RSV proteins into recombinant virions. We also show that intranasal vaccination of mice with nondefective VSV recombinants expressing RSV G (VSV-RSV G) or RSV F (VSV-RSV F) elicited RSV-specific antibodies in serum (by enzyme-linked immunosorbent assay [ELISA]) as well as neutralizing antibodies to RSV and afford complete protection against RSV challenge. In contrast, VSVΔG-RSV F induced detectable serum antibodies to RSV by ELISA, but no detectable neutralizing antibodies, yet it still protected from RSV challenge. VSVΔG-RSV G failed to induce any detectable serum (by ELISA) or neutralizing antibodies and failed to protect from RSV challenge. The attenuated, nonpropagating VSVΔG-RSV F is a particularly attractive candidate for a live attenuated recombinant RSV vaccine.

scholarly journals Antigenic Structure of Human Respiratory Syncytial Virus Fusion Glycoprotein

1998 ◽  
Vol 72 (8) ◽  
pp. 6922-6928 ◽  
Author(s):  
Juan A. López ◽  
Regla Bustos ◽  
Claes Örvell ◽  
Mabel Berois ◽  
Juan Arbiza ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Protein Sequence ◽  
Neutralizing Antibodies ◽  
Human Respiratory Syncytial Virus ◽  
Antigenic Structure ◽  
Antigenic Sites ◽  
Fusion Glycoprotein ◽  
Escape Mutants ◽  
Syncytial Virus ◽  
Prediction Of Secondary Structure

ABSTRACT New series of escape mutants of human respiratory syncytial virus were prepared with monoclonal antibodies specific for the fusion (F) protein. Sequence changes selected in the escape mutants identified two new antigenic sites (V and VI) recognized by neutralizing antibodies and a group-specific site (I) in the F1 chain of the F molecule. The new epitopes, and previously identified antigenic sites, were incorporated into a refined prediction of secondary-structure motifs to generate a detailed antigenic map of the F glycoprotein.

scholarly journals A vulnerable, membrane-proximal site in human respiratory syncytial virus F revealed by a prefusion-specific single-domain antibody

2021 ◽  
Author(s):  
Iebe Rossey ◽  
Ching-Lin Hsieh ◽  
Koen Sedeyn ◽  
Marlies Ballegeer ◽  
Bert Schepens ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Neutralizing Antibodies ◽  
Antigenic Site ◽  
Human Respiratory Syncytial Virus ◽  
Single Domain ◽  
Respiratory Pathogen ◽  
Single Domain Antibody ◽  
Domain Antibody ◽  
Proximal Site ◽  
Syncytial Virus

Human respiratory syncytial virus (RSV) is a major cause of lower respiratory tract disease, especially in young children and the elderly. The fusion protein (F) exists in a pre- and postfusion conformation and is the main target of RSV-neutralizing antibodies. Highly potent RSV-neutralizing antibodies typically bind sites that are unique to the prefusion conformation of F. In this study we screened a single-domain antibody (VHH) library derived from a llama immunized with prefusion-stabilized F and identified a prefusion F-specific VHH that can neutralize RSV A at subnanomolar concentrations. Structural analysis revealed that this VHH primarily binds to antigenic site I while also making contacts with residues in antigenic site III and IV. This new VHH reveals a previously underappreciated membrane-proximal region sensitive for neutralization. Importance RSV is an important respiratory pathogen. This study describes a prefusion F-specific VHH that primarily binds to antigenic site I of RSV F. This is the first time that a prefusion F-specific antibody that binds this site is reported. In general, antibodies that bind to site I are poorly neutralizing, whereas the VHH described here neutralizes RSV A at subnanomolar concentrations. Our findings contribute to insights into the RSV F antigenic map.

scholarly journals In silico virtual screening of lead compounds for major antigenic sites in respiratory syncytial virus fusion protein

2021 ◽  
Author(s):  
Shilu Mathew ◽  
Sara Taleb ◽  
Ali Hussein Eid ◽  
Asmaa A. Althani ◽  
Hadi M. Yassine
Keyword(s):  
Respiratory Syncytial Virus ◽  
Virtual Screening ◽  
In Silico ◽  
Neutralizing Antibodies ◽  
Antigenic Site ◽  
Crystallographic Structure ◽  
F Protein ◽  
Antigenic Sites ◽  
Syncytial Virus ◽  
Neutralizing Epitopes

AbstractHuman respiratory syncytial virus (RSV) is a leading ubiquitous respiratory pathogen in newborn infants, young children, and the elderly, with no vaccine available to date. The viral fusion glycoprotein (RSV F) plays an essential role in the infection process, and it is a primary target of neutralizing antibodies, making it an attractive site for vaccine development. With this in view, there is a persistent need to identify selective antiviral drugs against RSV, targeting the major antigenic sites on the F protein. We aimed to conduct a robust in silico high-throughput drug screening of one million compounds to explore potential inhibitors that bind the major antigenic site Ø and site II on RSV F protein, which are the main target of neutralizing antibodies (NAb). We utilized the three-dimensional crystallographic structure of both antigenic site Ø on pre-F and antigenic II on post-F to screen for potential anti-RSV inhibitors. A library of one million small compounds was docked to explore lead binders in the major antigenic sites by using virtual lab bench CLC Drug Discovery. We also performed Quantitative Structure-Activity and Relationship (QSAR) for the lead best binders known for their antiviral activity. Among one million tested ligands, seven ligands (PubChem ID: 3714418, 24787350, 49828911, 24802036, 79824892, 49726463, and 3139884) were identified as the best binders to neutralizing epitopes site Ø and four ligands (PubChem ID: 865999, 17505357, 24802036, and 24285058) to neutralizing epitopes site II, respectively. These binders exhibited significant interactions with neutralizing epitopes on RSV F, with an average of six H bonds, docking energy of − 15.43 Kcal·mol−1, and minimum interaction energy of − 7.45 Kcal·mol−1. Using in silico virtual screening, we identified potential RSV inhibitors that bind two major antigenic sites on the RSV F protein. Using structure-based design and combination-based drug therapy, identified molecules could be modified to generate the next generation anti-RSV drugs.

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