Differential antigenicity of recombinant polyepitope-antigens based on loop- and helix-forming B and T cell epitopes
Identifieur interne : 003661 ( Main/Curation ); précédent : 003660; suivant : 003662Differential antigenicity of recombinant polyepitope-antigens based on loop- and helix-forming B and T cell epitopes
Auteurs : D. M Theisen [Luxembourg (pays)] ; F. B Bouche [Luxembourg (pays)] ; K. C El Kasmi [Luxembourg (pays)] ; I. Von Der Ahe [Luxembourg (pays)] ; W. Ammerlaan [Luxembourg (pays)] ; S. Demotz [Suisse] ; C. P Muller [Luxembourg (pays)]Source :
- Journal of Immunological Methods [ 0022-1759 ] ; 2000.
Descripteurs français
- KwdFr :
- Animaux, Antigènes viraux (), Antigènes viraux (biosynthèse), Antigènes viraux (génétique), Antigènes viraux (immunologie), Conformation des protéines, Cricetinae, Données de séquences moléculaires, Déterminants antigéniques des lymphocytes B (), Déterminants antigéniques des lymphocytes B (biosynthèse), Déterminants antigéniques des lymphocytes B (génétique), Déterminants antigéniques des lymphocytes B (immunologie), Déterminants antigéniques des lymphocytes T (), Déterminants antigéniques des lymphocytes T (biosynthèse), Déterminants antigéniques des lymphocytes T (génétique), Déterminants antigéniques des lymphocytes T (immunologie), Expression des gènes, Hémagglutinines virales (), Hémagglutinines virales (biosynthèse), Hémagglutinines virales (génétique), Hémagglutinines virales (immunologie), Lignée cellulaire, Peptides (), Peptides (génétique), Peptides (immunologie), Protéines de fusion recombinantes (), Protéines de fusion recombinantes (génétique), Protéines de fusion recombinantes (immunologie), Séquence d'acides aminés.
- MESH :
- biosynthèse : Antigènes viraux, Déterminants antigéniques des lymphocytes B, Déterminants antigéniques des lymphocytes T, Hémagglutinines virales.
- génétique : Antigènes viraux, Déterminants antigéniques des lymphocytes B, Déterminants antigéniques des lymphocytes T, Hémagglutinines virales, Peptides, Protéines de fusion recombinantes.
- immunologie : Antigènes viraux, Déterminants antigéniques des lymphocytes B, Déterminants antigéniques des lymphocytes T, Hémagglutinines virales, Peptides, Protéines de fusion recombinantes.
- Pascal (Inist)
- Animaux, Antigène, Antigènes viraux, Antigénicité, Chimère, Conformation des protéines, Cricetinae, Données de séquences moléculaires, Déterminant antigénique, Déterminants antigéniques des lymphocytes B, Déterminants antigéniques des lymphocytes T, Expression des gènes, Hémagglutinine, Hémagglutinines virales, Lignée cellulaire, Lymphocyte B, Lymphocyte T, Peptides, Protéine, Protéines de fusion recombinantes, Structure hélice, Séquence d'acides aminés, Virus rougeole.
English descriptors
- KwdEn :
- Amino Acid Sequence, Animals, Antigen, Antigenic determinant, Antigenicity, Antigens, Viral (biosynthesis), Antigens, Viral (chemistry), Antigens, Viral (genetics), Antigens, Viral (immunology), B-Lymphocyte, Cell Line, Chimera, Cricetinae, Epitopes, B-Lymphocyte (biosynthesis), Epitopes, B-Lymphocyte (chemistry), Epitopes, B-Lymphocyte (genetics), Epitopes, B-Lymphocyte (immunology), Epitopes, T-Lymphocyte (biosynthesis), Epitopes, T-Lymphocyte (chemistry), Epitopes, T-Lymphocyte (genetics), Epitopes, T-Lymphocyte (immunology), Gene Expression, Helical structure, Hemagglutinin, Hemagglutinins, Viral (biosynthesis), Hemagglutinins, Viral (chemistry), Hemagglutinins, Viral (genetics), Hemagglutinins, Viral (immunology), Measles virus, Molecular Sequence Data, Peptides (chemistry), Peptides (genetics), Peptides (immunology), Protein, Protein Conformation, Recombinant Fusion Proteins (chemistry), Recombinant Fusion Proteins (genetics), Recombinant Fusion Proteins (immunology), T-Lymphocyte.
- MESH :
- chemical , biosynthesis : Antigens, Viral, Epitopes, B-Lymphocyte, Epitopes, T-Lymphocyte, Hemagglutinins, Viral.
- chemical , chemistry : Antigens, Viral, Epitopes, B-Lymphocyte, Epitopes, T-Lymphocyte, Hemagglutinins, Viral, Peptides, Recombinant Fusion Proteins.
- chemical , genetics : Antigens, Viral, Epitopes, B-Lymphocyte, Epitopes, T-Lymphocyte, Hemagglutinins, Viral, Peptides, Recombinant Fusion Proteins.
- chemical , immunology : Antigens, Viral, Epitopes, B-Lymphocyte, Epitopes, T-Lymphocyte, Hemagglutinins, Viral, Peptides, Recombinant Fusion Proteins.
- Amino Acid Sequence, Animals, Cell Line, Cricetinae, Gene Expression, Molecular Sequence Data, Protein Conformation.
- Teeft :
- Amino acids, Anking sequences, Antigenic, Apc, Bces, Brons, Cassette, Cell epitope, Cell epitopes, Cell line, Cell proliferation assay, Chimeric, Class peptides, Conformation, Different permutational, Epitope, Fournier, General rules, Helical, Helical conformation, Helix, Helix conformation, Hemagglutinin, Hemagglutinin protein, Immune, Immunogenicity, Immunol, Immunological, Immunological methods, Jung, Kasmi, Mabs, Mammalian cells, Maternal antibodies, Measles, Measles virus, Measles virus hemagglutinin protein, Molecular environment, Molecular weight, More antigenic, Multiple copies, Negative control, Neutralizing, Neutralizing antibodies, Peptide, Permutational, Plasmid, Polyepitope, Polyepitopes, Protective antibodies, Recombinant, Rotzschke, Schneider, Semliki forest virus replicon, Sequential, Sequential bces, Sequential epitopes, Stimulation index, Synthetic peptides, Tandem, Tandem tces, Tce, Theisen, Transfected cells, Transmembrane signal sequence, Vaccine, Virol, Western blot, Wiesmuller.
Abstract
Abstract: To investigate a strategy for the design of chimeric antigens based on B cell epitopes (BCEs) we have genetically recombined multiple copies of loop- (L) and helix-forming (H) sequential and protective BCEs of the measles virus hemagglutinin protein (MVH) in a number of high-molecular-weight polyepitope constructs (24.5–45.5 kDa). The BCE cassettes were combined semi-randomly together with a promiscuous T cell epitope (TCE; tt830–844) to yield 13 different permutational constructs. When expressed in mammalian cells, all constructs were detectable by Western blot as distinct bands of predicted molecular weight. Flow cytometry with conformation-specific antibodies revealed the Cys-loop in two [(L4T4)2 and (L2T2)4] and the helix conformation in one [(H2T2)4] of the different permutational constructs. The larger constructs, containing 16 epitope cassettes, seemed more likely to express the BCEs in their native conformation than the 8-mers. In the T cell proliferation assay, constructs with a higher copy number of TCEs, such as (L2T2)4, were more antigenic, as long as tandem repeats were separated by spacers. Since the conformation of even sequential BCEs and the processing of TCEs are both sensitive to their molecular environment it is difficult to predict the antigenic properties of polyepitopes. However, with the permutational approach we have developed several polyepitope constructs [(L4T4)2, (L2T2)4, (H2T2)4] based on complex sequential BCEs that are antigenic for both T and B cells. Several constructs induced sera that reacted with reporter peptides, demonstrating that the sequential nature of the viral epitopes was conserved in the polyepitopes. Although several sera contained antibodies directed against amino acids critical for neutralization, only one construct induced antibodies that cross-reacted with the virus. Our results show the difficulty of designing chimeric antigens based on B cell epitopes mimicking their antigenic and immunologic properties even when these are sequential in nature.
Url:
DOI: 10.1016/S0022-1759(00)00197-6
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<profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Amino Acid Sequence</term>
<term>Animals</term>
<term>Antigen</term>
<term>Antigenic determinant</term>
<term>Antigenicity</term>
<term>Antigens, Viral (biosynthesis)</term>
<term>Antigens, Viral (chemistry)</term>
<term>Antigens, Viral (genetics)</term>
<term>Antigens, Viral (immunology)</term>
<term>B-Lymphocyte</term>
<term>Cell Line</term>
<term>Chimera</term>
<term>Cricetinae</term>
<term>Epitopes, B-Lymphocyte (biosynthesis)</term>
<term>Epitopes, B-Lymphocyte (chemistry)</term>
<term>Epitopes, B-Lymphocyte (genetics)</term>
<term>Epitopes, B-Lymphocyte (immunology)</term>
<term>Epitopes, T-Lymphocyte (biosynthesis)</term>
<term>Epitopes, T-Lymphocyte (chemistry)</term>
<term>Epitopes, T-Lymphocyte (genetics)</term>
<term>Epitopes, T-Lymphocyte (immunology)</term>
<term>Gene Expression</term>
<term>Helical structure</term>
<term>Hemagglutinin</term>
<term>Hemagglutinins, Viral (biosynthesis)</term>
<term>Hemagglutinins, Viral (chemistry)</term>
<term>Hemagglutinins, Viral (genetics)</term>
<term>Hemagglutinins, Viral (immunology)</term>
<term>Measles virus</term>
<term>Molecular Sequence Data</term>
<term>Peptides (chemistry)</term>
<term>Peptides (genetics)</term>
<term>Peptides (immunology)</term>
<term>Protein</term>
<term>Protein Conformation</term>
<term>Recombinant Fusion Proteins (chemistry)</term>
<term>Recombinant Fusion Proteins (genetics)</term>
<term>Recombinant Fusion Proteins (immunology)</term>
<term>T-Lymphocyte</term>
</keywords>
<keywords scheme="KwdFr" xml:lang="fr"><term>Animaux</term>
<term>Antigènes viraux ()</term>
<term>Antigènes viraux (biosynthèse)</term>
<term>Antigènes viraux (génétique)</term>
<term>Antigènes viraux (immunologie)</term>
<term>Conformation des protéines</term>
<term>Cricetinae</term>
<term>Données de séquences moléculaires</term>
<term>Déterminants antigéniques des lymphocytes B ()</term>
<term>Déterminants antigéniques des lymphocytes B (biosynthèse)</term>
<term>Déterminants antigéniques des lymphocytes B (génétique)</term>
<term>Déterminants antigéniques des lymphocytes B (immunologie)</term>
<term>Déterminants antigéniques des lymphocytes T ()</term>
<term>Déterminants antigéniques des lymphocytes T (biosynthèse)</term>
<term>Déterminants antigéniques des lymphocytes T (génétique)</term>
<term>Déterminants antigéniques des lymphocytes T (immunologie)</term>
<term>Expression des gènes</term>
<term>Hémagglutinines virales ()</term>
<term>Hémagglutinines virales (biosynthèse)</term>
<term>Hémagglutinines virales (génétique)</term>
<term>Hémagglutinines virales (immunologie)</term>
<term>Lignée cellulaire</term>
<term>Peptides ()</term>
<term>Peptides (génétique)</term>
<term>Peptides (immunologie)</term>
<term>Protéines de fusion recombinantes ()</term>
<term>Protéines de fusion recombinantes (génétique)</term>
<term>Protéines de fusion recombinantes (immunologie)</term>
<term>Séquence d'acides aminés</term>
</keywords>
<keywords scheme="MESH" type="chemical" qualifier="biosynthesis" xml:lang="en"><term>Antigens, Viral</term>
<term>Epitopes, B-Lymphocyte</term>
<term>Epitopes, T-Lymphocyte</term>
<term>Hemagglutinins, Viral</term>
</keywords>
<keywords scheme="MESH" type="chemical" qualifier="chemistry" xml:lang="en"><term>Antigens, Viral</term>
<term>Epitopes, B-Lymphocyte</term>
<term>Epitopes, T-Lymphocyte</term>
<term>Hemagglutinins, Viral</term>
<term>Peptides</term>
<term>Recombinant Fusion Proteins</term>
</keywords>
<keywords scheme="MESH" type="chemical" qualifier="genetics" xml:lang="en"><term>Antigens, Viral</term>
<term>Epitopes, B-Lymphocyte</term>
<term>Epitopes, T-Lymphocyte</term>
<term>Hemagglutinins, Viral</term>
<term>Peptides</term>
<term>Recombinant Fusion Proteins</term>
</keywords>
<keywords scheme="MESH" type="chemical" qualifier="immunology" xml:lang="en"><term>Antigens, Viral</term>
<term>Epitopes, B-Lymphocyte</term>
<term>Epitopes, T-Lymphocyte</term>
<term>Hemagglutinins, Viral</term>
<term>Peptides</term>
<term>Recombinant Fusion Proteins</term>
</keywords>
<keywords scheme="MESH" qualifier="biosynthèse" xml:lang="fr"><term>Antigènes viraux</term>
<term>Déterminants antigéniques des lymphocytes B</term>
<term>Déterminants antigéniques des lymphocytes T</term>
<term>Hémagglutinines virales</term>
</keywords>
<keywords scheme="MESH" qualifier="génétique" xml:lang="fr"><term>Antigènes viraux</term>
<term>Déterminants antigéniques des lymphocytes B</term>
<term>Déterminants antigéniques des lymphocytes T</term>
<term>Hémagglutinines virales</term>
<term>Peptides</term>
<term>Protéines de fusion recombinantes</term>
</keywords>
<keywords scheme="MESH" qualifier="immunologie" xml:lang="fr"><term>Antigènes viraux</term>
<term>Déterminants antigéniques des lymphocytes B</term>
<term>Déterminants antigéniques des lymphocytes T</term>
<term>Hémagglutinines virales</term>
<term>Peptides</term>
<term>Protéines de fusion recombinantes</term>
</keywords>
<keywords scheme="MESH" xml:lang="en"><term>Amino Acid Sequence</term>
<term>Animals</term>
<term>Cell Line</term>
<term>Cricetinae</term>
<term>Gene Expression</term>
<term>Molecular Sequence Data</term>
<term>Protein Conformation</term>
</keywords>
<keywords scheme="Pascal" xml:lang="fr"><term>Animaux</term>
<term>Antigène</term>
<term>Antigènes viraux</term>
<term>Antigénicité</term>
<term>Chimère</term>
<term>Conformation des protéines</term>
<term>Cricetinae</term>
<term>Données de séquences moléculaires</term>
<term>Déterminant antigénique</term>
<term>Déterminants antigéniques des lymphocytes B</term>
<term>Déterminants antigéniques des lymphocytes T</term>
<term>Expression des gènes</term>
<term>Hémagglutinine</term>
<term>Hémagglutinines virales</term>
<term>Lignée cellulaire</term>
<term>Lymphocyte B</term>
<term>Lymphocyte T</term>
<term>Peptides</term>
<term>Protéine</term>
<term>Protéines de fusion recombinantes</term>
<term>Structure hélice</term>
<term>Séquence d'acides aminés</term>
<term>Virus rougeole</term>
</keywords>
<keywords scheme="Teeft" xml:lang="en"><term>Amino acids</term>
<term>Anking sequences</term>
<term>Antigenic</term>
<term>Apc</term>
<term>Bces</term>
<term>Brons</term>
<term>Cassette</term>
<term>Cell epitope</term>
<term>Cell epitopes</term>
<term>Cell line</term>
<term>Cell proliferation assay</term>
<term>Chimeric</term>
<term>Class peptides</term>
<term>Conformation</term>
<term>Different permutational</term>
<term>Epitope</term>
<term>Fournier</term>
<term>General rules</term>
<term>Helical</term>
<term>Helical conformation</term>
<term>Helix</term>
<term>Helix conformation</term>
<term>Hemagglutinin</term>
<term>Hemagglutinin protein</term>
<term>Immune</term>
<term>Immunogenicity</term>
<term>Immunol</term>
<term>Immunological</term>
<term>Immunological methods</term>
<term>Jung</term>
<term>Kasmi</term>
<term>Mabs</term>
<term>Mammalian cells</term>
<term>Maternal antibodies</term>
<term>Measles</term>
<term>Measles virus</term>
<term>Measles virus hemagglutinin protein</term>
<term>Molecular environment</term>
<term>Molecular weight</term>
<term>More antigenic</term>
<term>Multiple copies</term>
<term>Negative control</term>
<term>Neutralizing</term>
<term>Neutralizing antibodies</term>
<term>Peptide</term>
<term>Permutational</term>
<term>Plasmid</term>
<term>Polyepitope</term>
<term>Polyepitopes</term>
<term>Protective antibodies</term>
<term>Recombinant</term>
<term>Rotzschke</term>
<term>Schneider</term>
<term>Semliki forest virus replicon</term>
<term>Sequential</term>
<term>Sequential bces</term>
<term>Sequential epitopes</term>
<term>Stimulation index</term>
<term>Synthetic peptides</term>
<term>Tandem</term>
<term>Tandem tces</term>
<term>Tce</term>
<term>Theisen</term>
<term>Transfected cells</term>
<term>Transmembrane signal sequence</term>
<term>Vaccine</term>
<term>Virol</term>
<term>Western blot</term>
<term>Wiesmuller</term>
</keywords>
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</langUsage>
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<front><div type="abstract" xml:lang="en">Abstract: To investigate a strategy for the design of chimeric antigens based on B cell epitopes (BCEs) we have genetically recombined multiple copies of loop- (L) and helix-forming (H) sequential and protective BCEs of the measles virus hemagglutinin protein (MVH) in a number of high-molecular-weight polyepitope constructs (24.5–45.5 kDa). The BCE cassettes were combined semi-randomly together with a promiscuous T cell epitope (TCE; tt830–844) to yield 13 different permutational constructs. When expressed in mammalian cells, all constructs were detectable by Western blot as distinct bands of predicted molecular weight. Flow cytometry with conformation-specific antibodies revealed the Cys-loop in two [(L4T4)2 and (L2T2)4] and the helix conformation in one [(H2T2)4] of the different permutational constructs. The larger constructs, containing 16 epitope cassettes, seemed more likely to express the BCEs in their native conformation than the 8-mers. In the T cell proliferation assay, constructs with a higher copy number of TCEs, such as (L2T2)4, were more antigenic, as long as tandem repeats were separated by spacers. Since the conformation of even sequential BCEs and the processing of TCEs are both sensitive to their molecular environment it is difficult to predict the antigenic properties of polyepitopes. However, with the permutational approach we have developed several polyepitope constructs [(L4T4)2, (L2T2)4, (H2T2)4] based on complex sequential BCEs that are antigenic for both T and B cells. Several constructs induced sera that reacted with reporter peptides, demonstrating that the sequential nature of the viral epitopes was conserved in the polyepitopes. Although several sera contained antibodies directed against amino acids critical for neutralization, only one construct induced antibodies that cross-reacted with the virus. Our results show the difficulty of designing chimeric antigens based on B cell epitopes mimicking their antigenic and immunologic properties even when these are sequential in nature.</div>
</front>
</TEI>
<double idat="0022-1759:2000:Theisen D:differential:antigenicity:of"><INIST><TEI><teiHeader><fileDesc><titleStmt><title xml:lang="en" level="a">Differential antigenicity of recombinant polyepitope-antigens based on loop- and helix-forming B and T cell epitopes</title>
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<sourceDesc><biblStruct><analytic><title xml:lang="en" level="a">Differential antigenicity of recombinant polyepitope-antigens based on loop- and helix-forming B and T cell epitopes</title>
<author><name sortKey="Theisen, D M" sort="Theisen, D M" uniqKey="Theisen D" first="D. M." last="Theisen">D. M. Theisen</name>
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<sZ>1 aut.</sZ>
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</inist:fA14>
<country>Luxembourg (pays)</country>
<wicri:noRegion>1011 Luxembourg</wicri:noRegion>
</affiliation>
<affiliation wicri:level="3"><inist:fA14 i1="02"><s1>Fakultät für Biologie, University of Tübingen</s1>
<s2>Tübingen</s2>
<s3>DEU</s3>
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<country>Allemagne</country>
<placeName><region type="land" nuts="1">Bade-Wurtemberg</region>
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<s2>1011 Luxembourg</s2>
<s3>LUX</s3>
<sZ>1 aut.</sZ>
<sZ>2 aut.</sZ>
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</affiliation>
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</placeName>
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<author><name sortKey="El Kasmi, K C" sort="El Kasmi, K C" uniqKey="El Kasmi K" first="K. C." last="El Kasmi">K. C. El Kasmi</name>
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<sZ>1 aut.</sZ>
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<settlement type="city">Tübingen</settlement>
</placeName>
</affiliation>
</author>
<author><name sortKey="Von Der Ahe, I" sort="Von Der Ahe, I" uniqKey="Von Der Ahe I" first="I." last="Von Der Ahe">I. Von Der Ahe</name>
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<wicri:noRegion>1011 Luxembourg</wicri:noRegion>
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<author><name sortKey="Ammerlaan, W" sort="Ammerlaan, W" uniqKey="Ammerlaan W" first="W." last="Ammerlaan">W. Ammerlaan</name>
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<wicri:noRegion>1011 Luxembourg</wicri:noRegion>
</affiliation>
</author>
<author><name sortKey="Demotz, S" sort="Demotz, S" uniqKey="Demotz S" first="S." last="Demotz">S. Demotz</name>
<affiliation wicri:level="1"><inist:fA14 i1="05"><s1>Institut de Biochemie, University of Lausanne</s1>
<s2>Epalinges</s2>
<s3>CHE</s3>
<sZ>6 aut.</sZ>
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<country>Suisse</country>
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<author><name sortKey="Muller, C P" sort="Muller, C P" uniqKey="Muller C" first="C. P." last="Muller">C. P. Muller</name>
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<wicri:noRegion>1011 Luxembourg</wicri:noRegion>
</affiliation>
<affiliation wicri:level="3"><inist:fA14 i1="04"><s1>Medizinische Fakultät, University of Tübingen</s1>
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<s3>DEU</s3>
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<sZ>4 aut.</sZ>
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<series><title level="j" type="main">Journal of immunological methods</title>
<title level="j" type="abbreviated">J. immunol. methods</title>
<idno type="ISSN">0022-1759</idno>
<imprint><date when="2000">2000</date>
</imprint>
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</biblStruct>
</sourceDesc>
<seriesStmt><title level="j" type="main">Journal of immunological methods</title>
<title level="j" type="abbreviated">J. immunol. methods</title>
<idno type="ISSN">0022-1759</idno>
</seriesStmt>
</fileDesc>
<profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Antigen</term>
<term>Antigenic determinant</term>
<term>Antigenicity</term>
<term>B-Lymphocyte</term>
<term>Chimera</term>
<term>Helical structure</term>
<term>Hemagglutinin</term>
<term>Measles virus</term>
<term>Protein</term>
<term>T-Lymphocyte</term>
</keywords>
<keywords scheme="Pascal" xml:lang="fr"><term>Lymphocyte T</term>
<term>Lymphocyte B</term>
<term>Déterminant antigénique</term>
<term>Virus rougeole</term>
<term>Antigène</term>
<term>Chimère</term>
<term>Protéine</term>
<term>Hémagglutinine</term>
<term>Structure hélice</term>
<term>Antigénicité</term>
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<front><div type="abstract" xml:lang="en">To investigate a strategy for the design of chimeric antigens based on B cell epitopes (BCEs) we have genetically recombined multiple copies of loop- (L) and helix-forming (H) sequential and protective BCEs of the measles virus hemagglutinin protein (MVH) in a number of high-molecular-weight polyepitope constructs (24.5-45.5 kDa). The BCE cassettes were combined semi-randomly together with a promiscuous T cell epitope (TCE; tt830-844) to yield 13 different permutational constructs. When expressed in mammalian cells, all constructs were detectable by Western blot as distinct bands of predicted molecular weight. Flow cytometry with conformation-specific antibodies revealed the Cys-loop in two [(L<sub>4</sub>
T<sub>4</sub>
)<sub>2</sub>
and (L<sub>2</sub>
T<sub>2</sub>
)<sub>4</sub>
] and the helix conformation in one [(H<sub>2</sub>
T<sub>2</sub>
)<sub>4</sub>
] of the different permutational constructs. The larger constructs, containing 16 epitope cassettes, seemed more likely to express the BCEs in their native conformation than the 8-mers. In the T cell proliferation assay, constructs with a higher copy number of TCEs, such as (L<sub>2</sub>
T<sub>2</sub>
)<sub>4</sub>
were more antigenic, as long as tandem repeats were separated by spacers. Since the conformation of even sequential BCEs and the processing of TCEs are both sensitive to their molecular environment it is difficult to predict the antigenic properties of polyepitopes. However, with the permutational approach we have developed several polyepitope constructs [(L<sub>4</sub>
T<sub>4</sub>
)<sub>2</sub>
, (L<sub>2</sub>
T<sub>2</sub>
)<sub>4</sub>
, (H<sub>2</sub>
T<sub>2</sub>
)<sub>4</sub>
] based on complex sequential BCEs that are antigenic for both T and B cells. Several constructs induced sera that reacted with reporter peptides, demonstrating that the sequential nature of the viral epitopes was conserved in the polyepitopes. Although several sera contained antibodies directed against amino acids critical for neutralization, only one construct induced antibodies that cross-reacted with the virus. Our results show the difficulty of designing chimeric antigens based on B cell epitopes mimicking their antigenic and immunologic properties even when these are sequential in nature.</div>
</front>
</TEI>
</INIST>
<ISTEX><TEI wicri:istexFullTextTei="biblStruct"><teiHeader><fileDesc><titleStmt><title>Differential antigenicity of recombinant polyepitope-antigens based on loop- and helix-forming B and T cell epitopes</title>
<author><name sortKey="Theisen, D M" sort="Theisen, D M" uniqKey="Theisen D" first="D. M" last="Theisen">D. M Theisen</name>
</author>
<author><name sortKey="Bouche, F B" sort="Bouche, F B" uniqKey="Bouche F" first="F. B" last="Bouche">F. B Bouche</name>
</author>
<author><name sortKey="El Kasmi, K C" sort="El Kasmi, K C" uniqKey="El Kasmi K" first="K. C" last="El Kasmi">K. C El Kasmi</name>
</author>
<author><name sortKey="Von Der Ahe, I" sort="Von Der Ahe, I" uniqKey="Von Der Ahe I" first="I" last="Von Der Ahe">I. Von Der Ahe</name>
</author>
<author><name sortKey="Ammerlaan, W" sort="Ammerlaan, W" uniqKey="Ammerlaan W" first="W" last="Ammerlaan">W. Ammerlaan</name>
</author>
<author><name sortKey="Demotz, S" sort="Demotz, S" uniqKey="Demotz S" first="S" last="Demotz">S. Demotz</name>
</author>
<author><name sortKey="Muller, C P" sort="Muller, C P" uniqKey="Muller C" first="C. P" last="Muller">C. P Muller</name>
</author>
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<sourceDesc><biblStruct><analytic><title level="a">Differential antigenicity of recombinant polyepitope-antigens based on loop- and helix-forming B and T cell epitopes</title>
<author><name sortKey="Theisen, D M" sort="Theisen, D M" uniqKey="Theisen D" first="D. M" last="Theisen">D. M Theisen</name>
<affiliation wicri:level="1"><country xml:lang="fr">Luxembourg (pays)</country>
<wicri:regionArea>Department of Immunology and WHO Collaborating Center for Measles, Laboratoire National de Santé, B.P. 1102, L-1011 Luxembourg</wicri:regionArea>
<wicri:noRegion>L-1011 Luxembourg</wicri:noRegion>
</affiliation>
</author>
<author><name sortKey="Bouche, F B" sort="Bouche, F B" uniqKey="Bouche F" first="F. B" last="Bouche">F. B Bouche</name>
<affiliation wicri:level="1"><country xml:lang="fr">Luxembourg (pays)</country>
<wicri:regionArea>Department of Immunology and WHO Collaborating Center for Measles, Laboratoire National de Santé, B.P. 1102, L-1011 Luxembourg</wicri:regionArea>
<wicri:noRegion>L-1011 Luxembourg</wicri:noRegion>
</affiliation>
</author>
<author><name sortKey="El Kasmi, K C" sort="El Kasmi, K C" uniqKey="El Kasmi K" first="K. C" last="El Kasmi">K. C El Kasmi</name>
<affiliation wicri:level="1"><country xml:lang="fr">Luxembourg (pays)</country>
<wicri:regionArea>Department of Immunology and WHO Collaborating Center for Measles, Laboratoire National de Santé, B.P. 1102, L-1011 Luxembourg</wicri:regionArea>
<wicri:noRegion>L-1011 Luxembourg</wicri:noRegion>
</affiliation>
</author>
<author><name sortKey="Von Der Ahe, I" sort="Von Der Ahe, I" uniqKey="Von Der Ahe I" first="I" last="Von Der Ahe">I. Von Der Ahe</name>
<affiliation wicri:level="1"><country xml:lang="fr">Luxembourg (pays)</country>
<wicri:regionArea>Department of Immunology and WHO Collaborating Center for Measles, Laboratoire National de Santé, B.P. 1102, L-1011 Luxembourg</wicri:regionArea>
<wicri:noRegion>L-1011 Luxembourg</wicri:noRegion>
</affiliation>
</author>
<author><name sortKey="Ammerlaan, W" sort="Ammerlaan, W" uniqKey="Ammerlaan W" first="W" last="Ammerlaan">W. Ammerlaan</name>
<affiliation wicri:level="1"><country xml:lang="fr">Luxembourg (pays)</country>
<wicri:regionArea>Department of Immunology and WHO Collaborating Center for Measles, Laboratoire National de Santé, B.P. 1102, L-1011 Luxembourg</wicri:regionArea>
<wicri:noRegion>L-1011 Luxembourg</wicri:noRegion>
</affiliation>
</author>
<author><name sortKey="Demotz, S" sort="Demotz, S" uniqKey="Demotz S" first="S" last="Demotz">S. Demotz</name>
<affiliation wicri:level="1"><country xml:lang="fr">Suisse</country>
<wicri:regionArea>Institut de Biochemie, University of Lausanne, Epalinges</wicri:regionArea>
<wicri:noRegion>Epalinges</wicri:noRegion>
</affiliation>
</author>
<author><name sortKey="Muller, C P" sort="Muller, C P" uniqKey="Muller C" first="C. P" last="Muller">C. P Muller</name>
<affiliation wicri:level="1"><country wicri:rule="url">Luxembourg (pays)</country>
</affiliation>
<affiliation wicri:level="1"><country xml:lang="fr">Luxembourg (pays)</country>
<wicri:regionArea>Department of Immunology and WHO Collaborating Center for Measles, Laboratoire National de Santé, B.P. 1102, L-1011 Luxembourg</wicri:regionArea>
<wicri:noRegion>L-1011 Luxembourg</wicri:noRegion>
</affiliation>
</author>
</analytic>
<monogr></monogr>
<series><title level="j">Journal of Immunological Methods</title>
<title level="j" type="abbrev">JIM</title>
<idno type="ISSN">0022-1759</idno>
<imprint><publisher>ELSEVIER</publisher>
<date type="published" when="2000">2000</date>
<biblScope unit="volume">242</biblScope>
<biblScope unit="issue">1–2</biblScope>
<biblScope unit="page" from="145">145</biblScope>
<biblScope unit="page" to="157">157</biblScope>
</imprint>
<idno type="ISSN">0022-1759</idno>
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<term>Anking sequences</term>
<term>Antigenic</term>
<term>Apc</term>
<term>Bces</term>
<term>Brons</term>
<term>Cassette</term>
<term>Cell epitope</term>
<term>Cell epitopes</term>
<term>Cell line</term>
<term>Cell proliferation assay</term>
<term>Chimeric</term>
<term>Class peptides</term>
<term>Conformation</term>
<term>Different permutational</term>
<term>Epitope</term>
<term>Fournier</term>
<term>General rules</term>
<term>Helical</term>
<term>Helical conformation</term>
<term>Helix</term>
<term>Helix conformation</term>
<term>Hemagglutinin</term>
<term>Hemagglutinin protein</term>
<term>Immune</term>
<term>Immunogenicity</term>
<term>Immunol</term>
<term>Immunological</term>
<term>Immunological methods</term>
<term>Jung</term>
<term>Kasmi</term>
<term>Mabs</term>
<term>Mammalian cells</term>
<term>Maternal antibodies</term>
<term>Measles</term>
<term>Measles virus</term>
<term>Measles virus hemagglutinin protein</term>
<term>Molecular environment</term>
<term>Molecular weight</term>
<term>More antigenic</term>
<term>Multiple copies</term>
<term>Negative control</term>
<term>Neutralizing</term>
<term>Neutralizing antibodies</term>
<term>Peptide</term>
<term>Permutational</term>
<term>Plasmid</term>
<term>Polyepitope</term>
<term>Polyepitopes</term>
<term>Protective antibodies</term>
<term>Recombinant</term>
<term>Rotzschke</term>
<term>Schneider</term>
<term>Semliki forest virus replicon</term>
<term>Sequential</term>
<term>Sequential bces</term>
<term>Sequential epitopes</term>
<term>Stimulation index</term>
<term>Synthetic peptides</term>
<term>Tandem</term>
<term>Tandem tces</term>
<term>Tce</term>
<term>Theisen</term>
<term>Transfected cells</term>
<term>Transmembrane signal sequence</term>
<term>Vaccine</term>
<term>Virol</term>
<term>Western blot</term>
<term>Wiesmuller</term>
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<front><div type="abstract" xml:lang="en">Abstract: To investigate a strategy for the design of chimeric antigens based on B cell epitopes (BCEs) we have genetically recombined multiple copies of loop- (L) and helix-forming (H) sequential and protective BCEs of the measles virus hemagglutinin protein (MVH) in a number of high-molecular-weight polyepitope constructs (24.5–45.5 kDa). The BCE cassettes were combined semi-randomly together with a promiscuous T cell epitope (TCE; tt830–844) to yield 13 different permutational constructs. When expressed in mammalian cells, all constructs were detectable by Western blot as distinct bands of predicted molecular weight. Flow cytometry with conformation-specific antibodies revealed the Cys-loop in two [(L4T4)2 and (L2T2)4] and the helix conformation in one [(H2T2)4] of the different permutational constructs. The larger constructs, containing 16 epitope cassettes, seemed more likely to express the BCEs in their native conformation than the 8-mers. In the T cell proliferation assay, constructs with a higher copy number of TCEs, such as (L2T2)4, were more antigenic, as long as tandem repeats were separated by spacers. Since the conformation of even sequential BCEs and the processing of TCEs are both sensitive to their molecular environment it is difficult to predict the antigenic properties of polyepitopes. However, with the permutational approach we have developed several polyepitope constructs [(L4T4)2, (L2T2)4, (H2T2)4] based on complex sequential BCEs that are antigenic for both T and B cells. Several constructs induced sera that reacted with reporter peptides, demonstrating that the sequential nature of the viral epitopes was conserved in the polyepitopes. Although several sera contained antibodies directed against amino acids critical for neutralization, only one construct induced antibodies that cross-reacted with the virus. Our results show the difficulty of designing chimeric antigens based on B cell epitopes mimicking their antigenic and immunologic properties even when these are sequential in nature.</div>
</front>
</TEI>
</ISTEX>
<PubMed><TEI><teiHeader><fileDesc><titleStmt><title xml:lang="en">Differential antigenicity of recombinant polyepitope-antigens based on loop- and helix-forming B and T cell epitopes.</title>
<author><name sortKey="Theisen, D M" sort="Theisen, D M" uniqKey="Theisen D" first="D M" last="Theisen">D M Theisen</name>
<affiliation wicri:level="1"><nlm:affiliation>Department of Immunology and WHO Collaborating Center for Measles, Laboratoire National de Santé, B.P. 1102, L-1011 Luxembourg, Luxembourg.</nlm:affiliation>
<country xml:lang="fr">Luxembourg (pays)</country>
<wicri:regionArea>Department of Immunology and WHO Collaborating Center for Measles, Laboratoire National de Santé, B.P. 1102, L-1011 Luxembourg</wicri:regionArea>
<wicri:noRegion>L-1011 Luxembourg</wicri:noRegion>
</affiliation>
</author>
<author><name sortKey="Bouche, F B" sort="Bouche, F B" uniqKey="Bouche F" first="F B" last="Bouche">F B Bouche</name>
</author>
<author><name sortKey="El Kasmi, K C" sort="El Kasmi, K C" uniqKey="El Kasmi K" first="K C" last="El Kasmi">K C El Kasmi</name>
</author>
<author><name sortKey="Von Der Ahe, I" sort="Von Der Ahe, I" uniqKey="Von Der Ahe I" first="I" last="Von Der Ahe">I. Von Der Ahe</name>
</author>
<author><name sortKey="Ammerlaan, W" sort="Ammerlaan, W" uniqKey="Ammerlaan W" first="W" last="Ammerlaan">W. Ammerlaan</name>
</author>
<author><name sortKey="Demotz, S" sort="Demotz, S" uniqKey="Demotz S" first="S" last="Demotz">S. Demotz</name>
</author>
<author><name sortKey="Muller, C P" sort="Muller, C P" uniqKey="Muller C" first="C P" last="Muller">C P Muller</name>
</author>
</titleStmt>
<publicationStmt><idno type="wicri:source">PubMed</idno>
<date when="2000">2000</date>
<idno type="RBID">pubmed:10986397</idno>
<idno type="pmid">10986397</idno>
<idno type="doi">10.1016/s0022-1759(00)00197-6</idno>
<idno type="wicri:Area/PubMed/Corpus">002576</idno>
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<idno type="wicri:Area/PubMed/Checkpoint">002449</idno>
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<idno type="wicri:Area/Ncbi/Merge">000069</idno>
<idno type="wicri:Area/Ncbi/Curation">000069</idno>
<idno type="wicri:Area/Ncbi/Checkpoint">000069</idno>
<idno type="wicri:doubleKey">0022-1759:2000:Theisen D:differential:antigenicity:of</idno>
<idno type="wicri:Area/Main/Merge">003667</idno>
</publicationStmt>
<sourceDesc><biblStruct><analytic><title xml:lang="en">Differential antigenicity of recombinant polyepitope-antigens based on loop- and helix-forming B and T cell epitopes.</title>
<author><name sortKey="Theisen, D M" sort="Theisen, D M" uniqKey="Theisen D" first="D M" last="Theisen">D M Theisen</name>
<affiliation wicri:level="1"><nlm:affiliation>Department of Immunology and WHO Collaborating Center for Measles, Laboratoire National de Santé, B.P. 1102, L-1011 Luxembourg, Luxembourg.</nlm:affiliation>
<country xml:lang="fr">Luxembourg (pays)</country>
<wicri:regionArea>Department of Immunology and WHO Collaborating Center for Measles, Laboratoire National de Santé, B.P. 1102, L-1011 Luxembourg</wicri:regionArea>
<wicri:noRegion>L-1011 Luxembourg</wicri:noRegion>
</affiliation>
</author>
<author><name sortKey="Bouche, F B" sort="Bouche, F B" uniqKey="Bouche F" first="F B" last="Bouche">F B Bouche</name>
</author>
<author><name sortKey="El Kasmi, K C" sort="El Kasmi, K C" uniqKey="El Kasmi K" first="K C" last="El Kasmi">K C El Kasmi</name>
</author>
<author><name sortKey="Von Der Ahe, I" sort="Von Der Ahe, I" uniqKey="Von Der Ahe I" first="I" last="Von Der Ahe">I. Von Der Ahe</name>
</author>
<author><name sortKey="Ammerlaan, W" sort="Ammerlaan, W" uniqKey="Ammerlaan W" first="W" last="Ammerlaan">W. Ammerlaan</name>
</author>
<author><name sortKey="Demotz, S" sort="Demotz, S" uniqKey="Demotz S" first="S" last="Demotz">S. Demotz</name>
</author>
<author><name sortKey="Muller, C P" sort="Muller, C P" uniqKey="Muller C" first="C P" last="Muller">C P Muller</name>
</author>
</analytic>
<series><title level="j">Journal of immunological methods</title>
<idno type="ISSN">0022-1759</idno>
<imprint><date when="2000" type="published">2000</date>
</imprint>
</series>
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</sourceDesc>
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<term>Animals</term>
<term>Antigens, Viral (biosynthesis)</term>
<term>Antigens, Viral (chemistry)</term>
<term>Antigens, Viral (genetics)</term>
<term>Antigens, Viral (immunology)</term>
<term>Cell Line</term>
<term>Cricetinae</term>
<term>Epitopes, B-Lymphocyte (biosynthesis)</term>
<term>Epitopes, B-Lymphocyte (chemistry)</term>
<term>Epitopes, B-Lymphocyte (genetics)</term>
<term>Epitopes, B-Lymphocyte (immunology)</term>
<term>Epitopes, T-Lymphocyte (biosynthesis)</term>
<term>Epitopes, T-Lymphocyte (chemistry)</term>
<term>Epitopes, T-Lymphocyte (genetics)</term>
<term>Epitopes, T-Lymphocyte (immunology)</term>
<term>Gene Expression</term>
<term>Hemagglutinins, Viral (biosynthesis)</term>
<term>Hemagglutinins, Viral (chemistry)</term>
<term>Hemagglutinins, Viral (genetics)</term>
<term>Hemagglutinins, Viral (immunology)</term>
<term>Molecular Sequence Data</term>
<term>Peptides (chemistry)</term>
<term>Peptides (genetics)</term>
<term>Peptides (immunology)</term>
<term>Protein Conformation</term>
<term>Recombinant Fusion Proteins (chemistry)</term>
<term>Recombinant Fusion Proteins (genetics)</term>
<term>Recombinant Fusion Proteins (immunology)</term>
</keywords>
<keywords scheme="KwdFr" xml:lang="fr"><term>Animaux</term>
<term>Antigènes viraux ()</term>
<term>Antigènes viraux (biosynthèse)</term>
<term>Antigènes viraux (génétique)</term>
<term>Antigènes viraux (immunologie)</term>
<term>Conformation des protéines</term>
<term>Cricetinae</term>
<term>Données de séquences moléculaires</term>
<term>Déterminants antigéniques des lymphocytes B ()</term>
<term>Déterminants antigéniques des lymphocytes B (biosynthèse)</term>
<term>Déterminants antigéniques des lymphocytes B (génétique)</term>
<term>Déterminants antigéniques des lymphocytes B (immunologie)</term>
<term>Déterminants antigéniques des lymphocytes T ()</term>
<term>Déterminants antigéniques des lymphocytes T (biosynthèse)</term>
<term>Déterminants antigéniques des lymphocytes T (génétique)</term>
<term>Déterminants antigéniques des lymphocytes T (immunologie)</term>
<term>Expression des gènes</term>
<term>Hémagglutinines virales ()</term>
<term>Hémagglutinines virales (biosynthèse)</term>
<term>Hémagglutinines virales (génétique)</term>
<term>Hémagglutinines virales (immunologie)</term>
<term>Lignée cellulaire</term>
<term>Peptides ()</term>
<term>Peptides (génétique)</term>
<term>Peptides (immunologie)</term>
<term>Protéines de fusion recombinantes ()</term>
<term>Protéines de fusion recombinantes (génétique)</term>
<term>Protéines de fusion recombinantes (immunologie)</term>
<term>Séquence d'acides aminés</term>
</keywords>
<keywords scheme="MESH" type="chemical" qualifier="biosynthesis" xml:lang="en"><term>Antigens, Viral</term>
<term>Epitopes, B-Lymphocyte</term>
<term>Epitopes, T-Lymphocyte</term>
<term>Hemagglutinins, Viral</term>
</keywords>
<keywords scheme="MESH" type="chemical" qualifier="chemistry" xml:lang="en"><term>Antigens, Viral</term>
<term>Epitopes, B-Lymphocyte</term>
<term>Epitopes, T-Lymphocyte</term>
<term>Hemagglutinins, Viral</term>
<term>Peptides</term>
<term>Recombinant Fusion Proteins</term>
</keywords>
<keywords scheme="MESH" type="chemical" qualifier="genetics" xml:lang="en"><term>Antigens, Viral</term>
<term>Epitopes, B-Lymphocyte</term>
<term>Epitopes, T-Lymphocyte</term>
<term>Hemagglutinins, Viral</term>
<term>Peptides</term>
<term>Recombinant Fusion Proteins</term>
</keywords>
<keywords scheme="MESH" type="chemical" qualifier="immunology" xml:lang="en"><term>Antigens, Viral</term>
<term>Epitopes, B-Lymphocyte</term>
<term>Epitopes, T-Lymphocyte</term>
<term>Hemagglutinins, Viral</term>
<term>Peptides</term>
<term>Recombinant Fusion Proteins</term>
</keywords>
<keywords scheme="MESH" qualifier="biosynthèse" xml:lang="fr"><term>Antigènes viraux</term>
<term>Déterminants antigéniques des lymphocytes B</term>
<term>Déterminants antigéniques des lymphocytes T</term>
<term>Hémagglutinines virales</term>
</keywords>
<keywords scheme="MESH" qualifier="génétique" xml:lang="fr"><term>Antigènes viraux</term>
<term>Déterminants antigéniques des lymphocytes B</term>
<term>Déterminants antigéniques des lymphocytes T</term>
<term>Hémagglutinines virales</term>
<term>Peptides</term>
<term>Protéines de fusion recombinantes</term>
</keywords>
<keywords scheme="MESH" qualifier="immunologie" xml:lang="fr"><term>Antigènes viraux</term>
<term>Déterminants antigéniques des lymphocytes B</term>
<term>Déterminants antigéniques des lymphocytes T</term>
<term>Hémagglutinines virales</term>
<term>Peptides</term>
<term>Protéines de fusion recombinantes</term>
</keywords>
<keywords scheme="MESH" xml:lang="en"><term>Amino Acid Sequence</term>
<term>Animals</term>
<term>Cell Line</term>
<term>Cricetinae</term>
<term>Gene Expression</term>
<term>Molecular Sequence Data</term>
<term>Protein Conformation</term>
</keywords>
<keywords scheme="MESH" xml:lang="fr"><term>Animaux</term>
<term>Antigènes viraux</term>
<term>Conformation des protéines</term>
<term>Cricetinae</term>
<term>Données de séquences moléculaires</term>
<term>Déterminants antigéniques des lymphocytes B</term>
<term>Déterminants antigéniques des lymphocytes T</term>
<term>Expression des gènes</term>
<term>Hémagglutinines virales</term>
<term>Lignée cellulaire</term>
<term>Peptides</term>
<term>Protéines de fusion recombinantes</term>
<term>Séquence d'acides aminés</term>
</keywords>
</textClass>
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<front><div type="abstract" xml:lang="en">To investigate a strategy for the design of chimeric antigens based on B cell epitopes (BCEs) we have genetically recombined multiple copies of loop- (L) and helix-forming (H) sequential and protective BCEs of the measles virus hemagglutinin protein (MVH) in a number of high-molecular-weight polyepitope constructs (24.5-45.5 kDa). The BCE cassettes were combined semi-randomly together with a promiscuous T cell epitope (TCE; tt830-844) to yield 13 different permutational constructs. When expressed in mammalian cells, all constructs were detectable by Western blot as distinct bands of predicted molecular weight. Flow cytometry with conformation-specific antibodies revealed the Cys-loop in two [(L(4)T(4))(2) and (L(2)T(2))(4)] and the helix conformation in one [(H(2)T(2))(4)] of the different permutational constructs. The larger constructs, containing 16 epitope cassettes, seemed more likely to express the BCEs in their native conformation than the 8-mers. In the T cell proliferation assay, constructs with a higher copy number of TCEs, such as (L(2)T(2))(4), were more antigenic, as long as tandem repeats were separated by spacers. Since the conformation of even sequential BCEs and the processing of TCEs are both sensitive to their molecular environment it is difficult to predict the antigenic properties of polyepitopes. However, with the permutational approach we have developed several polyepitope constructs [(L(4)T(4))(2), (L(2)T(2))(4), (H(2)T(2))(4)] based on complex sequential BCEs that are antigenic for both T and B cells. Several constructs induced sera that reacted with reporter peptides, demonstrating that the sequential nature of the viral epitopes was conserved in the polyepitopes. Although several sera contained antibodies directed against amino acids critical for neutralization, only one construct induced antibodies that cross-reacted with the virus. Our results show the difficulty of designing chimeric antigens based on B cell epitopes mimicking their antigenic and immunologic properties even when these are sequential in nature.</div>
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