Escherichia coli-based expression system for the heterologous expression and purification of the elicitin β-cinnamomin from Phytophthora cinnamomi.
Identifieur interne : 001332 ( Main/Exploration ); précédent : 001331; suivant : 001333Escherichia coli-based expression system for the heterologous expression and purification of the elicitin β-cinnamomin from Phytophthora cinnamomi.
Auteurs : Sebastian Hofzumahaus [Allemagne] ; Anett SchallmeySource :
- Protein expression and purification [ 1096-0279 ] ; 2013.
Descripteurs français
- KwdFr :
- Escherichia coli (génétique), Escherichia coli (métabolisme), Phytophthora (composition chimique), Phytophthora (métabolisme), Pichia (génétique), Pichia (métabolisme), Protéines inactivant les ribosomes de type 2 (génétique), Protéines inactivant les ribosomes de type 2 (isolement et purification), Protéines inactivant les ribosomes de type 2 (métabolisme), Protéines recombinantes (génétique), Protéines recombinantes (isolement et purification), Protéines recombinantes (métabolisme), Séquence d'acides aminés (MeSH), Séquence nucléotidique (MeSH).
- MESH :
- composition chimique : Phytophthora.
- génétique : Escherichia coli, Pichia, Protéines inactivant les ribosomes de type 2, Protéines recombinantes.
- isolement et purification : Protéines inactivant les ribosomes de type 2, Protéines recombinantes.
- métabolisme : Escherichia coli, Phytophthora, Pichia, Protéines inactivant les ribosomes de type 2, Protéines recombinantes.
- Séquence d'acides aminés, Séquence nucléotidique.
English descriptors
- KwdEn :
- Amino Acid Sequence (MeSH), Base Sequence (MeSH), Escherichia coli (genetics), Escherichia coli (metabolism), Phytophthora (chemistry), Phytophthora (metabolism), Pichia (genetics), Pichia (metabolism), Recombinant Proteins (genetics), Recombinant Proteins (isolation & purification), Recombinant Proteins (metabolism), Ribosome Inactivating Proteins, Type 2 (genetics), Ribosome Inactivating Proteins, Type 2 (isolation & purification), Ribosome Inactivating Proteins, Type 2 (metabolism).
- MESH :
- chemical , genetics : Recombinant Proteins, Ribosome Inactivating Proteins, Type 2.
- chemistry : Phytophthora.
- genetics : Escherichia coli, Pichia.
- chemical , isolation & purification : Recombinant Proteins, Ribosome Inactivating Proteins, Type 2.
- metabolism : Escherichia coli, Phytophthora, Pichia, Recombinant Proteins, Ribosome Inactivating Proteins, Type 2.
- Amino Acid Sequence, Base Sequence.
Abstract
Elicitins are sterol carrier proteins from the Oomycete genera Phytophthora and Phytium and elicit a hypersensitive response in many economically important plants, in some cases causing a systemic acquired resistance. Their recombinant expression in bacteria is complicated by the presence of three disulfide bonds in the elicitin structure. In consequence, elicitins have so far only been produced in soluble form by isolation from native Phytophthora or Phytium strains or by recombinant expression in the yeast Pichia pastoris. Here, for the first time, we report the soluble expression of the elicitin β-cinnamomin from Phytophthora cinnamomi in Escherichia coli by secretion of the protein into the periplasm. β-Cinnamomin yields have been significantly improved after careful selection of the optimum secretion signal sequence. In total, 17.6 mg β-cinnamomin per liter cell culture have been obtained in shake flasks with the secretion signal sequence of the maltose-binding protein MalE from E. coli. Furthermore, by making use of a C-terminal His-tag, β-cinnamomin purification has been significantly simplified with only one step of immobilized metal ion affinity chromatography yielding protein of high purity (>90%). The established protocol has further been successfully applied to the soluble expression of another elicitin.
DOI: 10.1016/j.pep.2013.05.010
PubMed: 23747816
Affiliations:
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(génétique)</term><term>Protéines inactivant les ribosomes de type 2 (isolement et purification)</term><term>Protéines inactivant les ribosomes de type 2 (métabolisme)</term><term>Protéines recombinantes (génétique)</term><term>Protéines recombinantes (isolement et purification)</term><term>Protéines recombinantes (métabolisme)</term><term>Séquence d'acides aminés (MeSH)</term><term>Séquence nucléotidique (MeSH)</term></keywords><keywords scheme="MESH" type="chemical" qualifier="genetics" xml:lang="en"><term>Recombinant Proteins</term><term>Ribosome Inactivating Proteins, Type 2</term></keywords><keywords scheme="MESH" qualifier="chemistry" xml:lang="en"><term>Phytophthora</term></keywords><keywords scheme="MESH" qualifier="composition chimique" xml:lang="fr"><term>Phytophthora</term></keywords><keywords scheme="MESH" qualifier="genetics" xml:lang="en"><term>Escherichia coli</term><term>Pichia</term></keywords><keywords scheme="MESH" qualifier="génétique" xml:lang="fr"><term>Escherichia coli</term><term>Pichia</term><term>Protéines inactivant les ribosomes de type 2</term><term>Protéines recombinantes</term></keywords><keywords scheme="MESH" type="chemical" qualifier="isolation & purification" xml:lang="en"><term>Recombinant Proteins</term><term>Ribosome Inactivating Proteins, Type 2</term></keywords><keywords scheme="MESH" qualifier="isolement et purification" xml:lang="fr"><term>Protéines inactivant les ribosomes de type 2</term><term>Protéines recombinantes</term></keywords><keywords scheme="MESH" qualifier="metabolism" xml:lang="en"><term>Escherichia coli</term><term>Phytophthora</term><term>Pichia</term><term>Recombinant Proteins</term><term>Ribosome Inactivating Proteins, Type 2</term></keywords><keywords scheme="MESH" qualifier="métabolisme" xml:lang="fr"><term>Escherichia coli</term><term>Phytophthora</term><term>Pichia</term><term>Protéines inactivant les ribosomes de type 2</term><term>Protéines recombinantes</term></keywords><keywords scheme="MESH" xml:lang="en"><term>Amino Acid Sequence</term><term>Base Sequence</term></keywords><keywords scheme="MESH" xml:lang="fr"><term>Séquence d'acides aminés</term><term>Séquence nucléotidique</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">Elicitins are sterol carrier proteins from the Oomycete genera Phytophthora and Phytium and elicit a hypersensitive response in many economically important plants, in some cases causing a systemic acquired resistance. Their recombinant expression in bacteria is complicated by the presence of three disulfide bonds in the elicitin structure. In consequence, elicitins have so far only been produced in soluble form by isolation from native Phytophthora or Phytium strains or by recombinant expression in the yeast Pichia pastoris. Here, for the first time, we report the soluble expression of the elicitin β-cinnamomin from Phytophthora cinnamomi in Escherichia coli by secretion of the protein into the periplasm. β-Cinnamomin yields have been significantly improved after careful selection of the optimum secretion signal sequence. In total, 17.6 mg β-cinnamomin per liter cell culture have been obtained in shake flasks with the secretion signal sequence of the maltose-binding protein MalE from E. coli. Furthermore, by making use of a C-terminal His-tag, β-cinnamomin purification has been significantly simplified with only one step of immobilized metal ion affinity chromatography yielding protein of high purity (>90%). The established protocol has further been successfully applied to the soluble expression of another elicitin.</div></front></TEI><pubmed><MedlineCitation Status="MEDLINE" Owner="NLM"><PMID Version="1">23747816</PMID><DateCompleted><Year>2014</Year><Month>01</Month><Day>30</Day></DateCompleted><DateRevised><Year>2013</Year><Month>07</Month><Day>22</Day></DateRevised><Article PubModel="Print-Electronic"><Journal><ISSN IssnType="Electronic">1096-0279</ISSN><JournalIssue CitedMedium="Internet"><Volume>90</Volume><Issue>2</Issue><PubDate><Year>2013</Year><Month>Aug</Month></PubDate></JournalIssue><Title>Protein expression and purification</Title><ISOAbbreviation>Protein Expr Purif</ISOAbbreviation></Journal><ArticleTitle>Escherichia coli-based expression system for the heterologous expression and purification of the elicitin β-cinnamomin from Phytophthora cinnamomi.</ArticleTitle><Pagination><MedlinePgn>117-23</MedlinePgn></Pagination><ELocationID EIdType="doi" ValidYN="Y">10.1016/j.pep.2013.05.010</ELocationID><ELocationID EIdType="pii" ValidYN="Y">S1046-5928(13)00105-8</ELocationID><Abstract><AbstractText>Elicitins are sterol carrier proteins from the Oomycete genera Phytophthora and Phytium and elicit a hypersensitive response in many economically important plants, in some cases causing a systemic acquired resistance. Their recombinant expression in bacteria is complicated by the presence of three disulfide bonds in the elicitin structure. In consequence, elicitins have so far only been produced in soluble form by isolation from native Phytophthora or Phytium strains or by recombinant expression in the yeast Pichia pastoris. Here, for the first time, we report the soluble expression of the elicitin β-cinnamomin from Phytophthora cinnamomi in Escherichia coli by secretion of the protein into the periplasm. β-Cinnamomin yields have been significantly improved after careful selection of the optimum secretion signal sequence. In total, 17.6 mg β-cinnamomin per liter cell culture have been obtained in shake flasks with the secretion signal sequence of the maltose-binding protein MalE from E. coli. Furthermore, by making use of a C-terminal His-tag, β-cinnamomin purification has been significantly simplified with only one step of immobilized metal ion affinity chromatography yielding protein of high purity (>90%). The established protocol has further been successfully applied to the soluble expression of another elicitin.</AbstractText><CopyrightInformation>Copyright © 2013 Elsevier Inc. All rights reserved.</CopyrightInformation></Abstract><AuthorList CompleteYN="Y"><Author ValidYN="Y"><LastName>Hofzumahaus</LastName><ForeName>Sebastian</ForeName><Initials>S</Initials><AffiliationInfo><Affiliation>Institute of Biotechnology, RWTH Aachen University, Aachen, Germany. s.hofzumahaus@biotec.rwth-aachen.de</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Schallmey</LastName><ForeName>Anett</ForeName><Initials>A</Initials></Author></AuthorList><Language>eng</Language><PublicationTypeList><PublicationType UI="D016428">Journal Article</PublicationType><PublicationType UI="D013485">Research Support, Non-U.S. Gov't</PublicationType></PublicationTypeList><ArticleDate DateType="Electronic"><Year>2013</Year><Month>06</Month><Day>04</Day></ArticleDate></Article><MedlineJournalInfo><Country>United States</Country><MedlineTA>Protein Expr 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MajorTopicYN="N">metabolism</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D010838" MajorTopicYN="N">Phytophthora</DescriptorName><QualifierName UI="Q000737" MajorTopicYN="N">chemistry</QualifierName><QualifierName UI="Q000378" MajorTopicYN="Y">metabolism</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D010843" MajorTopicYN="N">Pichia</DescriptorName><QualifierName UI="Q000235" MajorTopicYN="N">genetics</QualifierName><QualifierName UI="Q000378" MajorTopicYN="N">metabolism</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D011994" MajorTopicYN="N">Recombinant Proteins</DescriptorName><QualifierName UI="Q000235" MajorTopicYN="N">genetics</QualifierName><QualifierName UI="Q000302" MajorTopicYN="N">isolation & purification</QualifierName><QualifierName UI="Q000378" MajorTopicYN="N">metabolism</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D054790" MajorTopicYN="N">Ribosome Inactivating Proteins, Type 2</DescriptorName><QualifierName UI="Q000235" MajorTopicYN="Y">genetics</QualifierName><QualifierName UI="Q000302" MajorTopicYN="N">isolation & purification</QualifierName><QualifierName UI="Q000378" MajorTopicYN="N">metabolism</QualifierName></MeshHeading></MeshHeadingList><KeywordList Owner="NOTNLM"><Keyword MajorTopicYN="N">Affinity chromatography</Keyword><Keyword MajorTopicYN="N">Disulfide bonds</Keyword><Keyword MajorTopicYN="N">Periplasmic expression</Keyword><Keyword MajorTopicYN="N">Secretion signal peptide</Keyword><Keyword MajorTopicYN="N">Sterol carrier protein</Keyword></KeywordList></MedlineCitation><PubmedData><History><PubMedPubDate PubStatus="received"><Year>2013</Year><Month>04</Month><Day>04</Day></PubMedPubDate><PubMedPubDate PubStatus="revised"><Year>2013</Year><Month>05</Month><Day>17</Day></PubMedPubDate><PubMedPubDate PubStatus="accepted"><Year>2013</Year><Month>05</Month><Day>18</Day></PubMedPubDate><PubMedPubDate PubStatus="entrez"><Year>2013</Year><Month>6</Month><Day>11</Day><Hour>6</Hour><Minute>0</Minute></PubMedPubDate><PubMedPubDate PubStatus="pubmed"><Year>2013</Year><Month>6</Month><Day>12</Day><Hour>6</Hour><Minute>0</Minute></PubMedPubDate><PubMedPubDate PubStatus="medline"><Year>2014</Year><Month>1</Month><Day>31</Day><Hour>6</Hour><Minute>0</Minute></PubMedPubDate></History><PublicationStatus>ppublish</PublicationStatus><ArticleIdList><ArticleId IdType="pubmed">23747816</ArticleId><ArticleId IdType="pii">S1046-5928(13)00105-8</ArticleId><ArticleId IdType="doi">10.1016/j.pep.2013.05.010</ArticleId></ArticleIdList></PubmedData></pubmed><affiliations><list><country><li>Allemagne</li></country><region><li>District de Cologne</li><li>Rhénanie-du-Nord-Westphalie</li></region><settlement><li>Aix-la-Chapelle</li></settlement></list><tree><noCountry><name sortKey="Schallmey, Anett" sort="Schallmey, Anett" uniqKey="Schallmey A" first="Anett" last="Schallmey">Anett Schallmey</name></noCountry><country name="Allemagne"><region name="Rhénanie-du-Nord-Westphalie"><name sortKey="Hofzumahaus, Sebastian" sort="Hofzumahaus, Sebastian" uniqKey="Hofzumahaus S" first="Sebastian" last="Hofzumahaus">Sebastian Hofzumahaus</name></region></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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