Efficient expression of the anti‐AahI' scorpion toxin nanobody under a new functional form in a Pichia pastoris system
Identifieur interne : 000241 ( Main/Exploration ); précédent : 000240; suivant : 000242Efficient expression of the anti‐AahI' scorpion toxin nanobody under a new functional form in a Pichia pastoris system
Auteurs : Aymen Ezzine [Tunisie] ; Sonia M'Hirsi El Adab [Tunisie] ; Balkiss Bouhaouala-Zahar [Tunisie] ; Issam Hmila [Tunisie] ; Laura Baciou [France] ; Mohamed Najib Marzouki [Tunisie]Source :
- Biotechnology and Applied Biochemistry [ 0885-4513 ] ; 2012-01.
Descripteurs français
- Pascal (Inist)
English descriptors
- KwdEn :
- AahI′ toxin, Animals, Antibodies (genetics), Antibodies (isolation & purification), Antibodies (metabolism), Gene Expression, Glycosylation, His‐Tag, N‐glycosylation, Pichia (genetics), Pichia (metabolism), Pichia pastoris, Recombinant Proteins (genetics), Recombinant Proteins (isolation & purification), Recombinant Proteins (metabolism), Recombinant protein, Scorpions (genetics), Toxin, heterologous protein, nanobody.
- MESH :
- chemical , genetics : Antibodies, Recombinant Proteins.
- chemical , isolation & purification : Antibodies, Recombinant Proteins.
- chemical , metabolism : Antibodies, Recombinant Proteins.
- genetics : Pichia, Scorpions.
- metabolism : Pichia.
- Animals, Gene Expression.
Abstract
Most large‐scale microbial production of recombinant proteins are based on Escherichia coli, yeasts, or filamentous fungi systems. Using eukaryotic hosts, antibody fragments are generally expressed by targeting to the secretory pathway. This enables not only efficient disulfide bond formation but also secretion of soluble and correctly folded product. For this goal, a recombinant vector was constructed to produce a single‐domain antibody (NbAahI′22) directed against AahI′ scorpion toxin using the methylotrophic yeast Pichia pastoris. The corresponding complementary DNA was cloned under control of the alcohol oxidase promoter in frame with the Saccharomyces α‐factor secretion signal and then transferred to P. pastoris cell strain X‐33. Using Western blot, we detected the expression of the recombinant NbAahI′22 exclusively in the culture medium. Targeting to the histidine label, the secreted nanobody was easily purified on nickel–nitrilotriacetic acid resin and then tested in enzyme‐linked immunosorbent assay. Interestingly, the production level of the NbAahI′22 in its new glycosylated form reached more than sixfold that obtained in E. coli. These findings give more evidence for the utilization of P. pastoris as a heterologous expression system.
Url:
DOI: 10.1002/bab.67
Affiliations:
Links toward previous steps (curation, corpus...)
- to stream Istex, to step Corpus: 001053
- to stream Istex, to step Curation: 000C21
- to stream Istex, to step Checkpoint: 000065
- to stream PubMed, to step Corpus: 000081
- to stream PubMed, to step Curation: 000081
- to stream PubMed, to step Checkpoint: 000081
- to stream Ncbi, to step Merge: 000066
- to stream Ncbi, to step Curation: 000066
- to stream Ncbi, to step Checkpoint: 000066
- to stream Main, to step Merge: 000246
- to stream PascalFrancis, to step Corpus: 000103
- to stream PascalFrancis, to step Curation: 000338
- to stream PascalFrancis, to step Checkpoint: 000096
- to stream Main, to step Merge: 000293
- to stream Main, to step Curation: 000241
Le document en format XML
<record><TEI wicri:istexFullTextTei="biblStruct"><teiHeader><fileDesc><titleStmt><title xml:lang="en">Efficient expression of the anti‐AahI' scorpion toxin nanobody under a new functional form in a Pichia pastoris system</title><author><name sortKey="Ezzine, Aymen" sort="Ezzine, Aymen" uniqKey="Ezzine A" first="Aymen" last="Ezzine">Aymen Ezzine</name></author><author><name sortKey="M Hirsi El Adab, Sonia" sort="M Hirsi El Adab, Sonia" uniqKey="M Hirsi El Adab S" first="Sonia" last="M'Hirsi El Adab">Sonia M'Hirsi El Adab</name></author><author><name sortKey="Bouhaouala Ahar, Balkiss" sort="Bouhaouala Ahar, Balkiss" uniqKey="Bouhaouala Ahar B" first="Balkiss" last="Bouhaouala-Zahar">Balkiss Bouhaouala-Zahar</name></author><author><name sortKey="Hmila, Issam" sort="Hmila, Issam" uniqKey="Hmila I" first="Issam" last="Hmila">Issam Hmila</name></author><author><name sortKey="Baciou, Laura" sort="Baciou, Laura" uniqKey="Baciou L" first="Laura" last="Baciou">Laura Baciou</name></author><author><name sortKey="Marzouki, Mohamed Najib" sort="Marzouki, Mohamed Najib" uniqKey="Marzouki M" first="Mohamed Najib" last="Marzouki">Mohamed Najib Marzouki</name></author></titleStmt><publicationStmt><idno type="wicri:source">ISTEX</idno><idno type="RBID">ISTEX:CC3886A98B196938DC8D47BB1065C97D18160EA1</idno><date when="2012" year="2012">2012</date><idno type="doi">10.1002/bab.67</idno><idno type="url">https://api.istex.fr/document/CC3886A98B196938DC8D47BB1065C97D18160EA1/fulltext/pdf</idno><idno type="wicri:Area/Istex/Corpus">001053</idno><idno type="wicri:explorRef" wicri:stream="Istex" wicri:step="Corpus" wicri:corpus="ISTEX">001053</idno><idno type="wicri:Area/Istex/Curation">000C21</idno><idno type="wicri:Area/Istex/Checkpoint">000065</idno><idno type="wicri:explorRef" wicri:stream="Istex" wicri:step="Checkpoint">000065</idno><idno type="wicri:doubleKey">0885-4513:2012:Ezzine A:efficient:expression:of</idno><idno type="wicri:source">PubMed</idno><idno type="RBID">pubmed:22332740</idno><idno type="wicri:Area/PubMed/Corpus">000081</idno><idno type="wicri:explorRef" wicri:stream="PubMed" wicri:step="Corpus" wicri:corpus="PubMed">000081</idno><idno type="wicri:Area/PubMed/Curation">000081</idno><idno type="wicri:explorRef" wicri:stream="PubMed" wicri:step="Curation">000081</idno><idno type="wicri:Area/PubMed/Checkpoint">000081</idno><idno type="wicri:explorRef" wicri:stream="Checkpoint" wicri:step="PubMed">000081</idno><idno type="wicri:Area/Ncbi/Merge">000066</idno><idno type="wicri:Area/Ncbi/Curation">000066</idno><idno type="wicri:Area/Ncbi/Checkpoint">000066</idno><idno type="wicri:Area/Main/Merge">000246</idno><idno type="wicri:source">INIST</idno><idno type="RBID">Pascal:12-0135969</idno><idno type="wicri:Area/PascalFrancis/Corpus">000103</idno><idno type="wicri:Area/PascalFrancis/Curation">000338</idno><idno type="wicri:Area/PascalFrancis/Checkpoint">000096</idno><idno type="wicri:explorRef" wicri:stream="PascalFrancis" wicri:step="Checkpoint">000096</idno><idno type="wicri:doubleKey">0885-4513:2012:Ezzine A:efficient:expression:of</idno><idno type="wicri:Area/Main/Merge">000293</idno><idno type="wicri:Area/Main/Curation">000241</idno><idno type="wicri:Area/Main/Exploration">000241</idno></publicationStmt><sourceDesc><biblStruct><analytic><title level="a" type="main" xml:lang="en">Efficient expression of the anti‐AahI' scorpion toxin nanobody under a new functional form in a Pichia pastoris system</title><author><name sortKey="Ezzine, Aymen" sort="Ezzine, Aymen" uniqKey="Ezzine A" first="Aymen" last="Ezzine">Aymen Ezzine</name><affiliation wicri:level="3"><country xml:lang="fr">Tunisie</country><wicri:regionArea>Laboratoire d'Ingénierie des Protéines et des Molécules Bioactives (LIP‐MB), Institut National des Sciences Appliquées et de Technologie, Université de Carthage, Tunis</wicri:regionArea><placeName><settlement type="city">Tunis</settlement><region nuts="2">Gouvernorat de Tunis</region></placeName></affiliation><affiliation></affiliation></author><author><name sortKey="M Hirsi El Adab, Sonia" sort="M Hirsi El Adab, Sonia" uniqKey="M Hirsi El Adab S" first="Sonia" last="M'Hirsi El Adab">Sonia M'Hirsi El Adab</name><affiliation wicri:level="3"><country xml:lang="fr">Tunisie</country><wicri:regionArea>Laboratoire d'Ingénierie des Protéines et des Molécules Bioactives (LIP‐MB), Institut National des Sciences Appliquées et de Technologie, Université de Carthage, Tunis</wicri:regionArea><placeName><settlement type="city">Tunis</settlement><region nuts="2">Gouvernorat de Tunis</region></placeName></affiliation></author><author><name sortKey="Bouhaouala Ahar, Balkiss" sort="Bouhaouala Ahar, Balkiss" uniqKey="Bouhaouala Ahar B" first="Balkiss" last="Bouhaouala-Zahar">Balkiss Bouhaouala-Zahar</name><affiliation wicri:level="3"><country xml:lang="fr">Tunisie</country><wicri:regionArea>Laboratoire des Venins et Toxines, Institut Pasteur de Tunis, Tunis</wicri:regionArea><placeName><settlement type="city">Tunis</settlement><region nuts="2">Gouvernorat de Tunis</region></placeName></affiliation></author><author><name sortKey="Hmila, Issam" sort="Hmila, Issam" uniqKey="Hmila I" first="Issam" last="Hmila">Issam Hmila</name><affiliation wicri:level="3"><country xml:lang="fr">Tunisie</country><wicri:regionArea>Laboratoire des Venins et Toxines, Institut Pasteur de Tunis, Tunis</wicri:regionArea><placeName><settlement type="city">Tunis</settlement><region nuts="2">Gouvernorat de Tunis</region></placeName></affiliation></author><author><name sortKey="Baciou, Laura" sort="Baciou, Laura" uniqKey="Baciou L" first="Laura" last="Baciou">Laura Baciou</name><affiliation wicri:level="3"><country xml:lang="fr">France</country><wicri:regionArea>Laboratoire de Chimie Physique, Université Paris Sud 11, Orsay</wicri:regionArea><placeName><settlement type="city">Orsay</settlement></placeName></affiliation></author><author><name sortKey="Marzouki, Mohamed Najib" sort="Marzouki, Mohamed Najib" uniqKey="Marzouki M" first="Mohamed Najib" last="Marzouki">Mohamed Najib Marzouki</name><affiliation wicri:level="3"><country xml:lang="fr">Tunisie</country><wicri:regionArea>Laboratoire d'Ingénierie des Protéines et des Molécules Bioactives (LIP‐MB), Institut National des Sciences Appliquées et de Technologie, Université de Carthage, Tunis</wicri:regionArea><placeName><settlement type="city">Tunis</settlement><region nuts="2">Gouvernorat de Tunis</region></placeName></affiliation></author></analytic><monogr></monogr><series><title level="j">Biotechnology and Applied Biochemistry</title><title level="j" type="abbrev">Biotechnology and Applied Biochemistry</title><idno type="ISSN">0885-4513</idno><idno type="eISSN">1470-8744</idno><imprint><publisher>Wiley Subscription Services, Inc., A Wiley Company</publisher><pubPlace>Hoboken</pubPlace><date type="published" when="2012-01">2012-01</date><biblScope unit="volume">59</biblScope><biblScope unit="issue">1</biblScope><biblScope unit="page" from="15">15</biblScope><biblScope unit="page" to="21">21</biblScope></imprint><idno type="ISSN">0885-4513</idno></series><idno type="istex">CC3886A98B196938DC8D47BB1065C97D18160EA1</idno><idno type="DOI">10.1002/bab.67</idno><idno type="ArticleID">BAB67</idno></biblStruct></sourceDesc><seriesStmt><idno type="ISSN">0885-4513</idno></seriesStmt></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>AahI′ toxin</term><term>Animals</term><term>Antibodies (genetics)</term><term>Antibodies (isolation & purification)</term><term>Antibodies (metabolism)</term><term>Gene Expression</term><term>Glycosylation</term><term>His‐Tag</term><term>N‐glycosylation</term><term>Pichia (genetics)</term><term>Pichia (metabolism)</term><term>Pichia pastoris</term><term>Recombinant Proteins (genetics)</term><term>Recombinant Proteins (isolation & purification)</term><term>Recombinant Proteins (metabolism)</term><term>Recombinant protein</term><term>Scorpions (genetics)</term><term>Toxin</term><term>heterologous protein</term><term>nanobody</term></keywords><keywords scheme="MESH" type="chemical" qualifier="genetics" xml:lang="en"><term>Antibodies</term><term>Recombinant Proteins</term></keywords><keywords scheme="MESH" type="chemical" qualifier="isolation & purification" xml:lang="en"><term>Antibodies</term><term>Recombinant Proteins</term></keywords><keywords scheme="MESH" type="chemical" qualifier="metabolism" xml:lang="en"><term>Antibodies</term><term>Recombinant Proteins</term></keywords><keywords scheme="MESH" qualifier="genetics" xml:lang="en"><term>Pichia</term><term>Scorpions</term></keywords><keywords scheme="MESH" qualifier="metabolism" xml:lang="en"><term>Pichia</term></keywords><keywords scheme="MESH" xml:lang="en"><term>Animals</term><term>Gene Expression</term></keywords><keywords scheme="Pascal" xml:lang="fr"><term>Glycosylation</term><term>Pichia pastoris</term><term>Protéine recombinante</term><term>Toxine</term></keywords></textClass><langUsage><language ident="en">en</language></langUsage></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">Most large‐scale microbial production of recombinant proteins are based on Escherichia coli, yeasts, or filamentous fungi systems. Using eukaryotic hosts, antibody fragments are generally expressed by targeting to the secretory pathway. This enables not only efficient disulfide bond formation but also secretion of soluble and correctly folded product. For this goal, a recombinant vector was constructed to produce a single‐domain antibody (NbAahI′22) directed against AahI′ scorpion toxin using the methylotrophic yeast Pichia pastoris. The corresponding complementary DNA was cloned under control of the alcohol oxidase promoter in frame with the Saccharomyces α‐factor secretion signal and then transferred to P. pastoris cell strain X‐33. Using Western blot, we detected the expression of the recombinant NbAahI′22 exclusively in the culture medium. Targeting to the histidine label, the secreted nanobody was easily purified on nickel–nitrilotriacetic acid resin and then tested in enzyme‐linked immunosorbent assay. Interestingly, the production level of the NbAahI′22 in its new glycosylated form reached more than sixfold that obtained in E. coli. These findings give more evidence for the utilization of P. pastoris as a heterologous expression system.</div></front></TEI><affiliations><list><country><li>France</li><li>Tunisie</li></country><region><li>Gouvernorat de Tunis</li></region><settlement><li>Orsay</li><li>Tunis</li></settlement></list><tree><country name="Tunisie"><region name="Gouvernorat de Tunis"><name sortKey="Ezzine, Aymen" sort="Ezzine, Aymen" uniqKey="Ezzine A" first="Aymen" last="Ezzine">Aymen Ezzine</name></region><name sortKey="Bouhaouala Ahar, Balkiss" sort="Bouhaouala Ahar, Balkiss" uniqKey="Bouhaouala Ahar B" first="Balkiss" last="Bouhaouala-Zahar">Balkiss Bouhaouala-Zahar</name><name sortKey="Hmila, Issam" sort="Hmila, Issam" uniqKey="Hmila I" first="Issam" last="Hmila">Issam Hmila</name><name sortKey="M Hirsi El Adab, Sonia" sort="M Hirsi El Adab, Sonia" uniqKey="M Hirsi El Adab S" first="Sonia" last="M'Hirsi El Adab">Sonia M'Hirsi El Adab</name><name sortKey="Marzouki, Mohamed Najib" sort="Marzouki, Mohamed Najib" uniqKey="Marzouki M" first="Mohamed Najib" last="Marzouki">Mohamed Najib Marzouki</name></country><country name="France"><noRegion><name sortKey="Baciou, Laura" sort="Baciou, Laura" uniqKey="Baciou L" first="Laura" last="Baciou">Laura Baciou</name></noRegion></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
EXPLOR_STEP=$WICRI_ROOT/Wicri/Terre/explor/NickelMaghrebV1/Data/Main/Exploration
HfdSelect -h $EXPLOR_STEP/biblio.hfd -nk 000241 | SxmlIndent | more
Ou
HfdSelect -h $EXPLOR_AREA/Data/Main/Exploration/biblio.hfd -nk 000241 | SxmlIndent | more
Pour mettre un lien sur cette page dans le réseau Wicri
{{Explor lien
|wiki= Wicri/Terre
|area= NickelMaghrebV1
|flux= Main
|étape= Exploration
|type= RBID
|clé= ISTEX:CC3886A98B196938DC8D47BB1065C97D18160EA1
|texte= Efficient expression of the anti‐AahI' scorpion toxin nanobody under a new functional form in a Pichia pastoris system
}}
| This area was generated with Dilib version V0.6.27. |  |