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Identifying epitopes responsible for neutralizing antibody and DC-SIGN binding on the spike glycoprotein of the severe acute respiratory syndrome coronavirus.

Identifieur interne : 001733 ( Ncbi/Checkpoint ); précédent : 001732; suivant : 001734

Identifying epitopes responsible for neutralizing antibody and DC-SIGN binding on the spike glycoprotein of the severe acute respiratory syndrome coronavirus.

Auteurs : Yi-Ping Shih [République populaire de Chine] ; Chia-Yen Chen ; Shih-Jen Liu ; Kuan-Hsuan Chen ; Yuan-Ming Lee ; Yu-Chan Chao ; Yi-Ming Arthur Chen

Source :

RBID : pubmed:17041212

Descripteurs français

English descriptors

Abstract

The severe acute respiratory syndrome-associated coronavirus (SARS-CoV) uses dendritic cell-specific ICAM-3 grabbing nonintegrin (DC-SIGN) to facilitate cell entry via cellular receptor-angiotensin-converting enzyme 2. For this project, we used recombinant baculoviruses expressing different lengths of SARS-CoV spike (S) protein in a capture assay to deduce the minimal DC-SIGN binding region. Our results identified the region location between amino acid (aa) residues 324 to 386 of the S protein. We then generated nine monoclonal antibodies (MAbs) against the S protein to map the DC-SIGN-binding domain using capture assays with pseudotyped viruses and observed that MAb SIa5 significantly blocked S protein-DC-SIGN interaction. An enhancement assay using the HKU39849 SARS-CoV strain and human immature dendritic cells confirmed our observation. Data from a pepscan analysis and M13 phage peptide display library system mapped the reactive MAb SIa5 epitope to aa residues 363 to 368 of the S protein. Results from a capture assay testing three pseudotyped viruses with mutated N-linked glycosylation sites of the S protein indicate that only two pseudotyped viruses (N330Q and N357Q, both of which lost glycosylation sites near the SIa5 epitope) had diminished DC-SIGN-binding capacity. We also noted that MAb SIb4 exerted a neutralizing effect against HKU39849; its reactive epitope was mapped to aa residues 435 to 439 of the S protein. We offer the data to facilitate the development of therapeutic agents and preventive vaccines against SARS-CoV infection.

DOI: 10.1128/JVI.01138-06
PubMed: 17041212


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pubmed:17041212

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<name sortKey="Chao, Yu Chan" sort="Chao, Yu Chan" uniqKey="Chao Y" first="Yu-Chan" last="Chao">Yu-Chan Chao</name>
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<term>Antibodies, Viral</term>
<term>Antigens, Viral (genetics)</term>
<term>Baculoviridae (genetics)</term>
<term>Base Sequence</term>
<term>Binding Sites (genetics)</term>
<term>Cell Adhesion Molecules (metabolism)</term>
<term>DNA, Viral (genetics)</term>
<term>Dendritic Cells (immunology)</term>
<term>Dendritic Cells (virology)</term>
<term>Epitope Mapping</term>
<term>Epitopes (chemistry)</term>
<term>Epitopes (genetics)</term>
<term>Glycosylation</term>
<term>Humans</term>
<term>In Vitro Techniques</term>
<term>Lectins, C-Type (metabolism)</term>
<term>Membrane Glycoproteins (chemistry)</term>
<term>Membrane Glycoproteins (genetics)</term>
<term>Membrane Glycoproteins (immunology)</term>
<term>Models, Molecular</term>
<term>Mutagenesis, Site-Directed</term>
<term>Neutralization Tests</term>
<term>Protein Binding</term>
<term>Protein Structure, Tertiary</term>
<term>Receptors, Cell Surface (metabolism)</term>
<term>Recombinant Proteins (chemistry)</term>
<term>Recombinant Proteins (genetics)</term>
<term>Recombinant Proteins (immunology)</term>
<term>SARS Virus (genetics)</term>
<term>SARS Virus (immunology)</term>
<term>Severe Acute Respiratory Syndrome (immunology)</term>
<term>Severe Acute Respiratory Syndrome (virology)</term>
<term>Spike Glycoprotein, Coronavirus</term>
<term>Viral Envelope Proteins (chemistry)</term>
<term>Viral Envelope Proteins (genetics)</term>
<term>Viral Envelope Proteins (immunology)</term>
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<term>Anticorps antiviraux</term>
<term>Anticorps monoclonaux</term>
<term>Antigènes viraux (génétique)</term>
<term>Baculoviridae (génétique)</term>
<term>Cartographie épitopique</term>
<term>Cellules dendritiques (immunologie)</term>
<term>Cellules dendritiques (virologie)</term>
<term>Glycoprotéine de spicule des coronavirus</term>
<term>Glycoprotéines membranaires ()</term>
<term>Glycoprotéines membranaires (génétique)</term>
<term>Glycoprotéines membranaires (immunologie)</term>
<term>Glycosylation</term>
<term>Humains</term>
<term>Lectines de type C (métabolisme)</term>
<term>Liaison aux protéines</term>
<term>Modèles moléculaires</term>
<term>Molécules d'adhérence cellulaire (métabolisme)</term>
<term>Mutagenèse dirigée</term>
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<term>Récepteurs de surface cellulaire (métabolisme)</term>
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<term>Structure tertiaire des protéines</term>
<term>Syndrome respiratoire aigu sévère (immunologie)</term>
<term>Syndrome respiratoire aigu sévère (virologie)</term>
<term>Séquence nucléotidique</term>
<term>Techniques in vitro</term>
<term>Tests de neutralisation</term>
<term>Virus du SRAS (génétique)</term>
<term>Virus du SRAS (immunologie)</term>
<term>Épitopes ()</term>
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<term>Recombinant Proteins</term>
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<term>Epitopes</term>
<term>Membrane Glycoproteins</term>
<term>Recombinant Proteins</term>
<term>Viral Envelope Proteins</term>
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<term>Membrane Glycoproteins</term>
<term>Recombinant Proteins</term>
<term>Viral Envelope Proteins</term>
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<term>Cell Adhesion Molecules</term>
<term>Lectins, C-Type</term>
<term>Receptors, Cell Surface</term>
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<term>Antibodies, Monoclonal</term>
<term>Antibodies, Viral</term>
<term>Spike Glycoprotein, Coronavirus</term>
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<term>Baculoviridae</term>
<term>Binding Sites</term>
<term>SARS Virus</term>
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<term>ADN viral</term>
<term>Antigènes viraux</term>
<term>Baculoviridae</term>
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<term>Protéines de l'enveloppe virale</term>
<term>Protéines recombinantes</term>
<term>Sites de fixation</term>
<term>Virus du SRAS</term>
<term>Épitopes</term>
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<term>Glycoprotéines membranaires</term>
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<term>Protéines recombinantes</term>
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<term>SARS Virus</term>
<term>Severe Acute Respiratory Syndrome</term>
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<term>Lectines de type C</term>
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<term>Récepteurs de surface cellulaire</term>
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<term>Syndrome respiratoire aigu sévère</term>
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<term>Dendritic Cells</term>
<term>Severe Acute Respiratory Syndrome</term>
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<keywords scheme="MESH" xml:lang="en">
<term>Base Sequence</term>
<term>Epitope Mapping</term>
<term>Glycosylation</term>
<term>Humans</term>
<term>In Vitro Techniques</term>
<term>Models, Molecular</term>
<term>Mutagenesis, Site-Directed</term>
<term>Neutralization Tests</term>
<term>Protein Binding</term>
<term>Protein Structure, Tertiary</term>
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<term>Anticorps antiviraux</term>
<term>Anticorps monoclonaux</term>
<term>Cartographie épitopique</term>
<term>Glycoprotéine de spicule des coronavirus</term>
<term>Glycoprotéines membranaires</term>
<term>Glycosylation</term>
<term>Humains</term>
<term>Liaison aux protéines</term>
<term>Modèles moléculaires</term>
<term>Mutagenèse dirigée</term>
<term>Protéines de l'enveloppe virale</term>
<term>Protéines recombinantes</term>
<term>Structure tertiaire des protéines</term>
<term>Séquence nucléotidique</term>
<term>Techniques in vitro</term>
<term>Tests de neutralisation</term>
<term>Épitopes</term>
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<front>
<div type="abstract" xml:lang="en">The severe acute respiratory syndrome-associated coronavirus (SARS-CoV) uses dendritic cell-specific ICAM-3 grabbing nonintegrin (DC-SIGN) to facilitate cell entry via cellular receptor-angiotensin-converting enzyme 2. For this project, we used recombinant baculoviruses expressing different lengths of SARS-CoV spike (S) protein in a capture assay to deduce the minimal DC-SIGN binding region. Our results identified the region location between amino acid (aa) residues 324 to 386 of the S protein. We then generated nine monoclonal antibodies (MAbs) against the S protein to map the DC-SIGN-binding domain using capture assays with pseudotyped viruses and observed that MAb SIa5 significantly blocked S protein-DC-SIGN interaction. An enhancement assay using the HKU39849 SARS-CoV strain and human immature dendritic cells confirmed our observation. Data from a pepscan analysis and M13 phage peptide display library system mapped the reactive MAb SIa5 epitope to aa residues 363 to 368 of the S protein. Results from a capture assay testing three pseudotyped viruses with mutated N-linked glycosylation sites of the S protein indicate that only two pseudotyped viruses (N330Q and N357Q, both of which lost glycosylation sites near the SIa5 epitope) had diminished DC-SIGN-binding capacity. We also noted that MAb SIb4 exerted a neutralizing effect against HKU39849; its reactive epitope was mapped to aa residues 435 to 439 of the S protein. We offer the data to facilitate the development of therapeutic agents and preventive vaccines against SARS-CoV infection.</div>
</front>
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