A dominant antigenic epitope on SARS-CoV spike protein identified by an avian single-chain variable fragment (scFv)-expressing phage
Identifieur interne : 003933 ( Main/Merge ); précédent : 003932; suivant : 003934A dominant antigenic epitope on SARS-CoV spike protein identified by an avian single-chain variable fragment (scFv)-expressing phage
Auteurs : Yu-Ching Lee [Taïwan] ; Sy-Jye C. Leu [Taïwan] ; Han-Chang Hung [Taïwan] ; Hsueh-Hsia Wu [Taïwan] ; I.-Jen Huang [Taïwan] ; Wen-Shyang Hsieh [Taïwan] ; Wen-Ta Chiu [Taïwan] ; Ming-Song Hsieh [Taïwan] ; Tsui-Fen Cheng [Taïwan] ; Yi-Yuan Yang [Taïwan]Source :
- Veterinary Immunology and Immunopathology [ 0165-2427 ] ; 2007.
Descripteurs français
- KwdFr :
- ARN viral (), ARN viral (génétique), Alignement de séquences, Animaux, Banque de peptides, Cartographie épitopique, Données de séquences moléculaires, Femelle, Fragments d'immunoglobuline (génétique), Fragments d'immunoglobuline (immunologie), Glycoprotéine de spicule des coronavirus, Glycoprotéines membranaires (génétique), Glycoprotéines membranaires (immunologie), Humains, Immunoglobulines (immunologie), Poulets, Protéines de l'enveloppe virale (génétique), Protéines de l'enveloppe virale (immunologie), Protéines recombinantes (immunologie), RT-PCR, Région variable d'immunoglobuline (génétique), Région variable d'immunoglobuline (immunologie), Syndrome respiratoire aigu sévère (immunologie), Séquence d'acides aminés, Séquence nucléotidique, Test ELISA, Virus du SRAS (génétique), Virus du SRAS (immunologie).
- MESH :
- génétique : ARN viral, Fragments d'immunoglobuline, Glycoprotéines membranaires, Protéines de l'enveloppe virale, Région variable d'immunoglobuline, Virus du SRAS.
- immunologie : Fragments d'immunoglobuline, Glycoprotéines membranaires, Immunoglobulines, Protéines de l'enveloppe virale, Protéines recombinantes, Région variable d'immunoglobuline, Syndrome respiratoire aigu sévère, Virus du SRAS.
- ARN viral, Alignement de séquences, Animaux, Banque de peptides, Cartographie épitopique, Données de séquences moléculaires, Femelle, Glycoprotéine de spicule des coronavirus, Humains, Poulets, RT-PCR, Séquence d'acides aminés, Séquence nucléotidique, Test ELISA.
English descriptors
- KwdEn :
- Amino Acid Sequence, Animals, Base Sequence, Chickens, Enzyme-Linked Immunosorbent Assay, Epitope Mapping, Female, Humans, Immunoglobulin Fragments (genetics), Immunoglobulin Fragments (immunology), Immunoglobulin Variable Region (genetics), Immunoglobulin Variable Region (immunology), Immunoglobulins (immunology), Membrane Glycoproteins (genetics), Membrane Glycoproteins (immunology), Molecular Sequence Data, Peptide Library, RNA, Viral (chemistry), RNA, Viral (genetics), Recombinant Proteins (immunology), Reverse Transcriptase Polymerase Chain Reaction, SARS Virus (genetics), SARS Virus (immunology), Sequence Alignment, Severe Acute Respiratory Syndrome (immunology), Spike Glycoprotein, Coronavirus, Viral Envelope Proteins (genetics), Viral Envelope Proteins (immunology).
- MESH :
- chemical , chemistry : RNA, Viral.
- chemical , genetics : Immunoglobulin Fragments, Immunoglobulin Variable Region, Membrane Glycoproteins, RNA, Viral, Viral Envelope Proteins.
- chemical , immunology : Immunoglobulin Fragments, Immunoglobulin Variable Region, Immunoglobulins, Membrane Glycoproteins, Recombinant Proteins, Viral Envelope Proteins.
- genetics : SARS Virus.
- immunology : SARS Virus, Severe Acute Respiratory Syndrome.
- Amino Acid Sequence, Animals, Base Sequence, Chickens, Enzyme-Linked Immunosorbent Assay, Epitope Mapping, Female, Humans, Molecular Sequence Data, Peptide Library, Reverse Transcriptase Polymerase Chain Reaction, Sequence Alignment, Spike Glycoprotein, Coronavirus.
Abstract
Severe acute respiratory syndrome (SARS) is a newly emergent human disease, which requires rapid diagnosis and effective therapy. Among antibody sources, immunoglobulin Y (IgY) is the major antibody found in chicken eggs and can be used as an alternative to mammalian antibodies normally used in research and immunotherapy. In this study, phage-expressing chicken monoclonal scFv antibody was chosen and characterized with phage display antibody technology. Truncated fragments of SARS-CoV spike protein were cloned in pET-21 vector and expressed in BL-21
Url:
DOI: 10.1016/j.vetimm.2007.02.001
PubMed: 17360045
PubMed Central: 7112517
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PMC:7112517Le document en format XML
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level="a" type="main">A dominant antigenic epitope on SARS-CoV spike protein identified by an avian single-chain variable fragment (scFv)-expressing phage</title><author><name sortKey="Lee, Yu Ching" sort="Lee, Yu Ching" uniqKey="Lee Y" first="Yu-Ching" last="Lee">Yu-Ching Lee</name><affiliation wicri:level="1"><nlm:aff id="aff1">Graduate Institute of Pharmaceutical Science, Taipei Medical University, Taipei, Taiwan</nlm:aff><country xml:lang="fr">Taïwan</country><wicri:regionArea>Graduate Institute of Pharmaceutical Science, Taipei Medical University, Taipei</wicri:regionArea><wicri:noRegion>Taipei</wicri:noRegion></affiliation></author><author><name sortKey="Leu, Sy Jye C" sort="Leu, Sy Jye C" uniqKey="Leu S" first="Sy-Jye C." last="Leu">Sy-Jye C. Leu</name><affiliation wicri:level="1"><nlm:aff id="aff2">Graduate Institute of Cell and Molecular Biology, Taipei Medical University, Taipei, Taiwan</nlm:aff><country xml:lang="fr">Taïwan</country><wicri:regionArea>Graduate Institute of Cell and Molecular Biology, Taipei Medical University, Taipei</wicri:regionArea><wicri:noRegion>Taipei</wicri:noRegion></affiliation></author><author><name sortKey="Hung, Han Chang" sort="Hung, Han Chang" uniqKey="Hung H" first="Han-Chang" last="Hung">Han-Chang Hung</name><affiliation wicri:level="1"><nlm:aff id="aff3">School of Medical Laboratory Science and Biotechnology, Taipei Medical University, Taipei, Taiwan</nlm:aff><country xml:lang="fr">Taïwan</country><wicri:regionArea>School of Medical Laboratory Science and Biotechnology, Taipei Medical University, Taipei</wicri:regionArea><wicri:noRegion>Taipei</wicri:noRegion></affiliation></author><author><name sortKey="Wu, Hsueh Hsia" sort="Wu, Hsueh Hsia" uniqKey="Wu H" first="Hsueh-Hsia" last="Wu">Hsueh-Hsia Wu</name><affiliation wicri:level="1"><nlm:aff id="aff3">School of Medical Laboratory Science and Biotechnology, Taipei Medical University, Taipei, Taiwan</nlm:aff><country xml:lang="fr">Taïwan</country><wicri:regionArea>School of Medical Laboratory Science and Biotechnology, Taipei Medical University, Taipei</wicri:regionArea><wicri:noRegion>Taipei</wicri:noRegion></affiliation></author><author><name sortKey="Huang, I Jen" sort="Huang, I Jen" uniqKey="Huang I" first="I.-Jen" last="Huang">I.-Jen Huang</name><affiliation wicri:level="1"><nlm:aff id="aff4">Department of Applied Bioscience Division, Taiwan Sugar Research Institute, Tainan, Taiwan</nlm:aff><country xml:lang="fr">Taïwan</country><wicri:regionArea>Department of Applied Bioscience Division, Taiwan Sugar Research Institute, Tainan</wicri:regionArea><wicri:noRegion>Tainan</wicri:noRegion></affiliation></author><author><name sortKey="Hsieh, Wen Shyang" sort="Hsieh, Wen Shyang" uniqKey="Hsieh W" first="Wen-Shyang" last="Hsieh">Wen-Shyang Hsieh</name><affiliation wicri:level="1"><nlm:aff id="aff5">Department of Laboratory Medicine, Mackay Memorial Hospital, Taipei, Taiwan</nlm:aff><country xml:lang="fr">Taïwan</country><wicri:regionArea>Department of Laboratory Medicine, Mackay Memorial Hospital, Taipei</wicri:regionArea><wicri:noRegion>Taipei</wicri:noRegion></affiliation></author><author><name sortKey="Chiu, Wen Ta" sort="Chiu, Wen Ta" uniqKey="Chiu W" first="Wen-Ta" last="Chiu">Wen-Ta Chiu</name><affiliation wicri:level="1"><nlm:aff id="aff6">Department of Neurosurgery, Taipei Municipal Wan Fang Hospital, Taipei, Taiwan</nlm:aff><country xml:lang="fr">Taïwan</country><wicri:regionArea>Department of Neurosurgery, Taipei Municipal Wan Fang Hospital, Taipei</wicri:regionArea><wicri:noRegion>Taipei</wicri:noRegion></affiliation></author><author><name sortKey="Hsieh, Ming Song" sort="Hsieh, Ming Song" uniqKey="Hsieh M" first="Ming-Song" last="Hsieh">Ming-Song Hsieh</name><affiliation wicri:level="1"><nlm:aff id="aff7">Department of Laboratory Medicine, Taipei Municipal Wan Fang Hospital, Taipei, Taiwan</nlm:aff><country xml:lang="fr">Taïwan</country><wicri:regionArea>Department of Laboratory Medicine, Taipei Municipal Wan Fang Hospital, Taipei</wicri:regionArea><wicri:noRegion>Taipei</wicri:noRegion></affiliation></author><author><name sortKey="Cheng, Tsui Fen" sort="Cheng, Tsui Fen" uniqKey="Cheng T" first="Tsui-Fen" last="Cheng">Tsui-Fen Cheng</name><affiliation wicri:level="1"><nlm:aff id="aff8">Department of General Surgery, Shin Kong Wu Ho-Su Memorial Hospital, Taipei, Taiwan</nlm:aff><country xml:lang="fr">Taïwan</country><wicri:regionArea>Department of General Surgery, Shin Kong Wu Ho-Su Memorial Hospital, Taipei</wicri:regionArea><wicri:noRegion>Taipei</wicri:noRegion></affiliation></author><author><name sortKey="Yang, Yi Yuan" sort="Yang, Yi Yuan" uniqKey="Yang Y" first="Yi-Yuan" last="Yang">Yi-Yuan Yang</name><affiliation wicri:level="1"><nlm:aff id="aff3">School of Medical Laboratory Science and Biotechnology, Taipei Medical University, Taipei, Taiwan</nlm:aff><country xml:lang="fr">Taïwan</country><wicri:regionArea>School of Medical Laboratory Science and Biotechnology, Taipei Medical University, Taipei</wicri:regionArea><wicri:noRegion>Taipei</wicri:noRegion></affiliation></author></analytic><series><title level="j">Veterinary Immunology and Immunopathology</title><idno type="ISSN">0165-2427</idno><idno type="eISSN">1873-2534</idno><imprint><date when="2007">2007</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Amino Acid Sequence</term><term>Animals</term><term>Base Sequence</term><term>Chickens</term><term>Enzyme-Linked Immunosorbent Assay</term><term>Epitope Mapping</term><term>Female</term><term>Humans</term><term>Immunoglobulin Fragments (genetics)</term><term>Immunoglobulin Fragments (immunology)</term><term>Immunoglobulin Variable Region (genetics)</term><term>Immunoglobulin Variable Region (immunology)</term><term>Immunoglobulins (immunology)</term><term>Membrane Glycoproteins (genetics)</term><term>Membrane Glycoproteins (immunology)</term><term>Molecular Sequence Data</term><term>Peptide Library</term><term>RNA, Viral (chemistry)</term><term>RNA, Viral (genetics)</term><term>Recombinant Proteins (immunology)</term><term>Reverse Transcriptase Polymerase Chain Reaction</term><term>SARS Virus (genetics)</term><term>SARS Virus (immunology)</term><term>Sequence Alignment</term><term>Severe Acute Respiratory Syndrome (immunology)</term><term>Spike Glycoprotein, Coronavirus</term><term>Viral Envelope Proteins (genetics)</term><term>Viral Envelope Proteins (immunology)</term></keywords><keywords scheme="KwdFr" xml:lang="fr"><term>ARN viral ()</term><term>ARN viral (génétique)</term><term>Alignement de séquences</term><term>Animaux</term><term>Banque de peptides</term><term>Cartographie épitopique</term><term>Données de séquences moléculaires</term><term>Femelle</term><term>Fragments d'immunoglobuline (génétique)</term><term>Fragments d'immunoglobuline (immunologie)</term><term>Glycoprotéine de spicule des coronavirus</term><term>Glycoprotéines membranaires (génétique)</term><term>Glycoprotéines membranaires (immunologie)</term><term>Humains</term><term>Immunoglobulines (immunologie)</term><term>Poulets</term><term>Protéines de l'enveloppe virale (génétique)</term><term>Protéines de l'enveloppe virale (immunologie)</term><term>Protéines recombinantes (immunologie)</term><term>RT-PCR</term><term>Région variable d'immunoglobuline (génétique)</term><term>Région variable d'immunoglobuline (immunologie)</term><term>Syndrome respiratoire aigu sévère (immunologie)</term><term>Séquence d'acides aminés</term><term>Séquence nucléotidique</term><term>Test ELISA</term><term>Virus du SRAS (génétique)</term><term>Virus du SRAS (immunologie)</term></keywords><keywords scheme="MESH" type="chemical" qualifier="chemistry" xml:lang="en"><term>RNA, Viral</term></keywords><keywords scheme="MESH" type="chemical" qualifier="genetics" xml:lang="en"><term>Immunoglobulin Fragments</term><term>Immunoglobulin Variable Region</term><term>Membrane Glycoproteins</term><term>RNA, Viral</term><term>Viral Envelope Proteins</term></keywords><keywords scheme="MESH" type="chemical" qualifier="immunology" xml:lang="en"><term>Immunoglobulin Fragments</term><term>Immunoglobulin Variable Region</term><term>Immunoglobulins</term><term>Membrane Glycoproteins</term><term>Recombinant Proteins</term><term>Viral Envelope Proteins</term></keywords><keywords scheme="MESH" qualifier="genetics" xml:lang="en"><term>SARS Virus</term></keywords><keywords scheme="MESH" qualifier="génétique" xml:lang="fr"><term>ARN viral</term><term>Fragments d'immunoglobuline</term><term>Glycoprotéines membranaires</term><term>Protéines de l'enveloppe virale</term><term>Région variable d'immunoglobuline</term><term>Virus du SRAS</term></keywords><keywords scheme="MESH" qualifier="immunologie" xml:lang="fr"><term>Fragments d'immunoglobuline</term><term>Glycoprotéines membranaires</term><term>Immunoglobulines</term><term>Protéines de l'enveloppe virale</term><term>Protéines recombinantes</term><term>Région variable d'immunoglobuline</term><term>Syndrome respiratoire aigu sévère</term><term>Virus du SRAS</term></keywords><keywords scheme="MESH" qualifier="immunology" xml:lang="en"><term>SARS Virus</term><term>Severe Acute Respiratory Syndrome</term></keywords><keywords scheme="MESH" xml:lang="en"><term>Amino Acid Sequence</term><term>Animals</term><term>Base Sequence</term><term>Chickens</term><term>Enzyme-Linked Immunosorbent Assay</term><term>Epitope Mapping</term><term>Female</term><term>Humans</term><term>Molecular Sequence Data</term><term>Peptide Library</term><term>Reverse Transcriptase Polymerase Chain Reaction</term><term>Sequence Alignment</term><term>Spike Glycoprotein, Coronavirus</term></keywords><keywords scheme="MESH" xml:lang="fr"><term>ARN viral</term><term>Alignement de séquences</term><term>Animaux</term><term>Banque de peptides</term><term>Cartographie épitopique</term><term>Données de séquences moléculaires</term><term>Femelle</term><term>Glycoprotéine de spicule des coronavirus</term><term>Humains</term><term>Poulets</term><term>RT-PCR</term><term>Séquence d'acides aminés</term><term>Séquence nucléotidique</term><term>Test ELISA</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en"><p>Severe acute respiratory syndrome (SARS) is a newly emergent human disease, which requires rapid diagnosis and effective therapy. Among antibody sources, immunoglobulin Y (IgY) is the major antibody found in chicken eggs and can be used as an alternative to mammalian antibodies normally used in research and immunotherapy. In this study, phage-expressing chicken monoclonal scFv antibody was chosen and characterized with phage display antibody technology. Truncated fragments of SARS-CoV spike protein were cloned in pET-21 vector and expressed in BL-21 <italic>Escherichia coli</italic> (<italic>E. coli</italic>) cells. After purification, the purity of these recombinant spike proteins was examined on SDS–PAGE and their identity verified with Western blot analysis using anti-his antibodies and sera from convalescent stage SARS-CoV-infected patients. Using these bacteria-derived proteins to immunize chickens, it was found that polyclonal IgY antibodies in the egg yolk and sera were highly reactive to the immunogens, as shown by Western blot and immunocytochemical staining analysis. A phage displaying scFv library was also established from spleen B cells of immunized chicken with 5 × 10<sup>7</sup> clones. After four panning cycles, the eluted phage titer showed a 10-fold increase. In sequence analysis with chicken germline gene, five phage clones reacted, with large dissimilarities of between 31 and 62%, in the complementarity-determining regions, one dominant phage 4S1 had strong binding to fragment Se-e, located between amino acid residues 456–650 of the spike protein and this particular phage had significantly strong binding to SARS-CoV-infected Vero E6 cells. Based on the results, we conclude that generating specific scFv-expressing phage binders with the phage display system can be successfully achieved and that this knowledge can be applied in clinical or academic research.</p></div></front><back><div1 type="bibliography"><listBibl><biblStruct><analytic><author><name sortKey="Abouzid, K" uniqKey="Abouzid K">K. 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