Antigenic and cellular localisation analysis of the severe acute respiratory syndrome coronavirus nucleocapsid protein using monoclonal antibodies

A member of the family of coronaviruses has previously been identified as the cause of the severe acute respiratory syndrome (SARS). In this study, several monoclonal antibodies against the nucleocapsid protein have been generated to examine distribution of the nucleocapsid in virus-infected cells a...

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Veröffentlicht in:Virus research 2006-12, Vol.122 (1), p.119-126
Hauptverfasser: Bussmann, Bianca M., Reiche, Sven, Jacob, Lotta H., Braun, Jan Matthias, Jassoy, Christian
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creator Bussmann, Bianca M.
Reiche, Sven
Jacob, Lotta H.
Braun, Jan Matthias
Jassoy, Christian
description A member of the family of coronaviruses has previously been identified as the cause of the severe acute respiratory syndrome (SARS). In this study, several monoclonal antibodies against the nucleocapsid protein have been generated to examine distribution of the nucleocapsid in virus-infected cells and to study antigenic regions of the protein. Confocal microscopic analysis identified nucleocapsids packaged in vesicles in the perinuclear area indicating viral synthesis at the endoplasmic reticulum and Golgi apparatus. The monoclonal antibodies bound to the central and carboxyterminal half of the nucleocapsid protein indicating prominent exposure and immunogenicity of this part of the protein. Antibodies recognised both linear and conformational epitopes. Predictions of antigenicity using mathematical modelling based on hydrophobicity analysis of SARS nucleoprotein could not be confirmed fully. Antibody binding to discontinuous peptides provides evidence that amino acids 274–283 and 373–382 assemble to a structural unit particularly rich in basic amino acids. In addition, amino acids 286–295, 316–325 and 361–367 that represent the epitope recognised by monoclonal antibody 6D11C1 converge indicating a well-structured C-terminal region of the SARS virus nucleocapsid protein and functional relationship of the peptide regions involved. Alternatively, dimerisation of the nucleocapsid protein may result in juxtaposition of the amino acid sequences 316–325 and 361–367 on one nucleoprotein molecule to amino acid 286–295 on the second peptide. The monoclonal antibodies will be available to assess antigenicity and immunological variabilities between different SARS CoV strains.
doi_str_mv 10.1016/j.virusres.2006.07.005
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In this study, several monoclonal antibodies against the nucleocapsid protein have been generated to examine distribution of the nucleocapsid in virus-infected cells and to study antigenic regions of the protein. Confocal microscopic analysis identified nucleocapsids packaged in vesicles in the perinuclear area indicating viral synthesis at the endoplasmic reticulum and Golgi apparatus. The monoclonal antibodies bound to the central and carboxyterminal half of the nucleocapsid protein indicating prominent exposure and immunogenicity of this part of the protein. Antibodies recognised both linear and conformational epitopes. Predictions of antigenicity using mathematical modelling based on hydrophobicity analysis of SARS nucleoprotein could not be confirmed fully. Antibody binding to discontinuous peptides provides evidence that amino acids 274–283 and 373–382 assemble to a structural unit particularly rich in basic amino acids. In addition, amino acids 286–295, 316–325 and 361–367 that represent the epitope recognised by monoclonal antibody 6D11C1 converge indicating a well-structured C-terminal region of the SARS virus nucleocapsid protein and functional relationship of the peptide regions involved. Alternatively, dimerisation of the nucleocapsid protein may result in juxtaposition of the amino acid sequences 316–325 and 361–367 on one nucleoprotein molecule to amino acid 286–295 on the second peptide. 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In this study, several monoclonal antibodies against the nucleocapsid protein have been generated to examine distribution of the nucleocapsid in virus-infected cells and to study antigenic regions of the protein. Confocal microscopic analysis identified nucleocapsids packaged in vesicles in the perinuclear area indicating viral synthesis at the endoplasmic reticulum and Golgi apparatus. The monoclonal antibodies bound to the central and carboxyterminal half of the nucleocapsid protein indicating prominent exposure and immunogenicity of this part of the protein. Antibodies recognised both linear and conformational epitopes. Predictions of antigenicity using mathematical modelling based on hydrophobicity analysis of SARS nucleoprotein could not be confirmed fully. Antibody binding to discontinuous peptides provides evidence that amino acids 274–283 and 373–382 assemble to a structural unit particularly rich in basic amino acids. In addition, amino acids 286–295, 316–325 and 361–367 that represent the epitope recognised by monoclonal antibody 6D11C1 converge indicating a well-structured C-terminal region of the SARS virus nucleocapsid protein and functional relationship of the peptide regions involved. Alternatively, dimerisation of the nucleocapsid protein may result in juxtaposition of the amino acid sequences 316–325 and 361–367 on one nucleoprotein molecule to amino acid 286–295 on the second peptide. The monoclonal antibodies will be available to assess antigenicity and immunological variabilities between different SARS CoV strains.</abstract><cop>Netherlands</cop><pub>Elsevier B.V</pub><pmid>16920216</pmid><doi>10.1016/j.virusres.2006.07.005</doi><tpages>8</tpages><oa>free_for_read</oa></addata></record>
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subjects Animals
Antibodies, Monoclonal - immunology
Antibodies, Viral - immunology
Antigens, Viral - analysis
Antigens, Viral - immunology
Cell Nucleus - chemistry
Chlorocebus aethiops
Coronaviruses
Cytoplasm - chemistry
Cytoplasm - virology
Dimerization
Endoplasmic Reticulum - chemistry
Endoplasmic Reticulum - virology
Epitope Mapping
Golgi Apparatus - chemistry
Golgi Apparatus - virology
Microscopy, Confocal
Microscopy, Fluorescence
Nucleocapsid Proteins - analysis
Nucleocapsid Proteins - immunology
Nucleoprotein
RNA viruses
SARS coronavirus
SARS Virus - physiology
Vero Cells
Virus structure
title Antigenic and cellular localisation analysis of the severe acute respiratory syndrome coronavirus nucleocapsid protein using monoclonal antibodies
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