AustralieFrV1, Main, Exploration, bibRecord, 00A823

Purification and Functional Reconstitution of N- and C-Halves of the MscL Channel

Identifieur interne : 00A823 ( Main/Exploration ); précédent : 00A822; suivant : 00A824

Purification and Functional Reconstitution of N- and C-Halves of the MscL Channel

Auteurs : Kyu-Ho Park [France] ; Catherine Berrier [France] ; Boris Martinac [Australie] ; Alexandre Ghazi [France]

Source :

Biophysical Journal [ 0006-3495 ] ; 2004.

RBID : PMC:1304064

Abstract

MscL is a mechanosensitive channel gated by membrane tension in the lipid bilayer alone. Its structure, known from x-ray crystallography, indicates that it is a homopentamer. Each subunit comprises two transmembrane segments TM1 and TM2 connected by a periplasmic loop. The closed pore is lined by five TM1 helices. We expressed in Escherichia coli and purified two halves of the protein, each containing one of the transmembrane segments. Their electrophysiological activity was studied by the patch-clamp recording upon reconstitution in artificial liposomes. The TM2 moiety had no electrophysiological activity, whereas the TM1 half formed channels, which were not affected by membrane tension and varied in conductance between 50 and 350 pS in 100 mM KCl. Coreconstitution of the two halves of MscL however, yielded mechanosensitive channels having the same conductance as the native MscL (1500 pS), but exhibiting increased sensitivity to pressure. Our results confirm the current view on the functional role of TM1 and TM2 helices in the MscL gating and emphasize the importance of helix-helix interactions for the assembly and functional properties of the channel protein. In addition, the results indicate a crucial role of the periplasmic loop for the channel mechanosensitivity.

Url:

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1304064

PubMed: 15041653
PubMed Central: 1304064

Affiliations:

Le document en format XML

<record><TEI><teiHeader><fileDesc><titleStmt><title xml:lang="en">Purification and Functional Reconstitution of N- and C-Halves of the MscL Channel</title>
<author><name sortKey="Park, Kyu Ho" sort="Park, Kyu Ho" uniqKey="Park K" first="Kyu-Ho" last="Park">Kyu-Ho Park</name>
<affiliation wicri:level="4"><nlm:aff wicri:cut="; and" id="N0x9cacbb8.0x9b48048">Unité Mixte de Recherche Centre National de la Recherche Scientifique 8619, Université Paris-Sud, 91405, Orsay Cedex, France</nlm:aff>
<country xml:lang="fr">France</country>
<wicri:regionArea>Unité Mixte de Recherche Centre National de la Recherche Scientifique 8619, Université Paris-Sud, 91405, Orsay Cedex</wicri:regionArea>
<wicri:noRegion>91405, Orsay Cedex</wicri:noRegion>
<orgName type="university">Université Paris-Sud</orgName>
<placeName><settlement type="city">Orsay</settlement>
<region type="region" nuts="2">Île-de-France</region>
</placeName>
</affiliation>
</author>
<author><name sortKey="Berrier, Catherine" sort="Berrier, Catherine" uniqKey="Berrier C" first="Catherine" last="Berrier">Catherine Berrier</name>
<affiliation wicri:level="4"><nlm:aff wicri:cut="; and" id="N0x9cacbb8.0x9b48048">Unité Mixte de Recherche Centre National de la Recherche Scientifique 8619, Université Paris-Sud, 91405, Orsay Cedex, France</nlm:aff>
<country xml:lang="fr">France</country>
<wicri:regionArea>Unité Mixte de Recherche Centre National de la Recherche Scientifique 8619, Université Paris-Sud, 91405, Orsay Cedex</wicri:regionArea>
<wicri:noRegion>91405, Orsay Cedex</wicri:noRegion>
<orgName type="university">Université Paris-Sud</orgName>
<placeName><settlement type="city">Orsay</settlement>
<region type="region" nuts="2">Île-de-France</region>
</placeName>
</affiliation>
</author>
<author><name sortKey="Martinac, Boris" sort="Martinac, Boris" uniqKey="Martinac B" first="Boris" last="Martinac">Boris Martinac</name>
<affiliation wicri:level="1"><nlm:aff id="N0x9cacbb8.0x9b48048">School of Medicine and Pharmacology M510, Queen Elizabeth II Medical Center, University of Western Australia, Crawley, Western Australia 6009, Australia</nlm:aff>
<country xml:lang="fr">Australie</country>
<wicri:regionArea>School of Medicine and Pharmacology M510, Queen Elizabeth II Medical Center, University of Western Australia, Crawley, Western Australia 6009</wicri:regionArea>
<wicri:noRegion>Western Australia 6009</wicri:noRegion>
</affiliation>
</author>
<author><name sortKey="Ghazi, Alexandre" sort="Ghazi, Alexandre" uniqKey="Ghazi A" first="Alexandre" last="Ghazi">Alexandre Ghazi</name>
<affiliation wicri:level="4"><nlm:aff wicri:cut="; and" id="N0x9cacbb8.0x9b48048">Unité Mixte de Recherche Centre National de la Recherche Scientifique 8619, Université Paris-Sud, 91405, Orsay Cedex, France</nlm:aff>
<country xml:lang="fr">France</country>
<wicri:regionArea>Unité Mixte de Recherche Centre National de la Recherche Scientifique 8619, Université Paris-Sud, 91405, Orsay Cedex</wicri:regionArea>
<wicri:noRegion>91405, Orsay Cedex</wicri:noRegion>
<orgName type="university">Université Paris-Sud</orgName>
<placeName><settlement type="city">Orsay</settlement>
<region type="region" nuts="2">Île-de-France</region>
</placeName>
</affiliation>
</author>
</titleStmt>
<publicationStmt><idno type="wicri:source">PMC</idno>
<idno type="pmid">15041653</idno>
<idno type="pmc">1304064</idno>
<idno type="url">http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1304064</idno>
<idno type="RBID">PMC:1304064</idno>
<date when="2004">2004</date>
<idno type="wicri:Area/Pmc/Corpus">000F64</idno>
<idno type="wicri:explorRef" wicri:stream="Pmc" wicri:step="Corpus" wicri:corpus="PMC">000F64</idno>
<idno type="wicri:Area/Pmc/Curation">000E25</idno>
<idno type="wicri:explorRef" wicri:stream="Pmc" wicri:step="Curation">000E25</idno>
<idno type="wicri:Area/Pmc/Checkpoint">002C77</idno>
<idno type="wicri:explorRef" wicri:stream="Pmc" wicri:step="Checkpoint">002C77</idno>
<idno type="wicri:Area/Ncbi/Merge">000104</idno>
<idno type="wicri:Area/Ncbi/Curation">000104</idno>
<idno type="wicri:Area/Ncbi/Checkpoint">000104</idno>
<idno type="wicri:doubleKey">0006-3495:2004:Park K:purification:and:functional</idno>
<idno type="wicri:Area/Main/Merge">00B474</idno>
<idno type="wicri:Area/Main/Curation">00A823</idno>
<idno type="wicri:Area/Main/Exploration">00A823</idno>
</publicationStmt>
<sourceDesc><biblStruct><analytic><title xml:lang="en" level="a" type="main">Purification and Functional Reconstitution of N- and C-Halves of the MscL Channel</title>
<author><name sortKey="Park, Kyu Ho" sort="Park, Kyu Ho" uniqKey="Park K" first="Kyu-Ho" last="Park">Kyu-Ho Park</name>
<affiliation wicri:level="4"><nlm:aff wicri:cut="; and" id="N0x9cacbb8.0x9b48048">Unité Mixte de Recherche Centre National de la Recherche Scientifique 8619, Université Paris-Sud, 91405, Orsay Cedex, France</nlm:aff>
<country xml:lang="fr">France</country>
<wicri:regionArea>Unité Mixte de Recherche Centre National de la Recherche Scientifique 8619, Université Paris-Sud, 91405, Orsay Cedex</wicri:regionArea>
<placeName><region type="region" nuts="2">Île-de-France</region>
<settlement type="city">Orsay</settlement>
<settlement type="city">Orsay</settlement>
</placeName>
<orgName type="university">Université Paris-Sud</orgName>
</affiliation>
</author>
<author><name sortKey="Berrier, Catherine" sort="Berrier, Catherine" uniqKey="Berrier C" first="Catherine" last="Berrier">Catherine Berrier</name>
<affiliation wicri:level="4"><nlm:aff wicri:cut="; and" id="N0x9cacbb8.0x9b48048">Unité Mixte de Recherche Centre National de la Recherche Scientifique 8619, Université Paris-Sud, 91405, Orsay Cedex, France</nlm:aff>
<country xml:lang="fr">France</country>
<wicri:regionArea>Unité Mixte de Recherche Centre National de la Recherche Scientifique 8619, Université Paris-Sud, 91405, Orsay Cedex</wicri:regionArea>
<wicri:noRegion>91405, Orsay Cedex</wicri:noRegion>
<orgName type="university">Université Paris-Sud</orgName>
<placeName><settlement type="city">Orsay</settlement>
<region type="region" nuts="2">Île-de-France</region>
</placeName>
</affiliation>
</author>
<author><name sortKey="Martinac, Boris" sort="Martinac, Boris" uniqKey="Martinac B" first="Boris" last="Martinac">Boris Martinac</name>
<affiliation wicri:level="1"><nlm:aff id="N0x9cacbb8.0x9b48048">School of Medicine and Pharmacology M510, Queen Elizabeth II Medical Center, University of Western Australia, Crawley, Western Australia 6009, Australia</nlm:aff>
<country xml:lang="fr">Australie</country>
<wicri:regionArea>School of Medicine and Pharmacology M510, Queen Elizabeth II Medical Center, University of Western Australia, Crawley, Western Australia 6009</wicri:regionArea>
<wicri:noRegion>Western Australia 6009</wicri:noRegion>
</affiliation>
</author>
<author><name sortKey="Ghazi, Alexandre" sort="Ghazi, Alexandre" uniqKey="Ghazi A" first="Alexandre" last="Ghazi">Alexandre Ghazi</name>
<affiliation wicri:level="4"><nlm:aff wicri:cut="; and" id="N0x9cacbb8.0x9b48048">Unité Mixte de Recherche Centre National de la Recherche Scientifique 8619, Université Paris-Sud, 91405, Orsay Cedex, France</nlm:aff>
<country xml:lang="fr">France</country>
<wicri:regionArea>Unité Mixte de Recherche Centre National de la Recherche Scientifique 8619, Université Paris-Sud, 91405, Orsay Cedex</wicri:regionArea>
<wicri:noRegion>91405, Orsay Cedex</wicri:noRegion>
<orgName type="university">Université Paris-Sud</orgName>
<placeName><settlement type="city">Orsay</settlement>
<region type="region" nuts="2">Île-de-France</region>
</placeName>
</affiliation>
</author>
</analytic>
<series><title level="j">Biophysical Journal</title>
<idno type="ISSN">0006-3495</idno>
<idno type="eISSN">1542-0086</idno>
<imprint><date when="2004">2004</date>
</imprint>
</series>
</biblStruct>
</sourceDesc>
</fileDesc>
<profileDesc><textClass></textClass>
</profileDesc>
</teiHeader>
<front><div type="abstract" xml:lang="en"><p>MscL is a mechanosensitive channel gated by membrane tension in the lipid bilayer alone. Its structure, known from x-ray crystallography, indicates that it is a homopentamer. Each subunit comprises two transmembrane segments TM1 and TM2 connected by a periplasmic loop. The closed pore is lined by five TM1 helices. We expressed in <italic>Escherichia coli</italic>
 and purified two halves of the protein, each containing one of the transmembrane segments. Their electrophysiological activity was studied by the patch-clamp recording upon reconstitution in artificial liposomes. The TM2 moiety had no electrophysiological activity, whereas the TM1 half formed channels, which were not affected by membrane tension and varied in conductance between 50 and 350 pS in 100 mM KCl. Coreconstitution of the two halves of MscL however, yielded mechanosensitive channels having the same conductance as the native MscL (1500 pS), but exhibiting increased sensitivity to pressure. Our results confirm the current view on the functional role of TM1 and TM2 helices in the MscL gating and emphasize the importance of helix-helix interactions for the assembly and functional properties of the channel protein. In addition, the results indicate a crucial role of the periplasmic loop for the channel mechanosensitivity.</p>
</div>
</front>
</TEI>
<affiliations><list><country><li>Australie</li>
<li>France</li>
</country>
<region><li>Île-de-France</li>
</region>
<settlement><li>Orsay</li>
</settlement>
<orgName><li>Université Paris-Sud</li>
</orgName>
</list>
<tree><country name="France"><region name="Île-de-France"><name sortKey="Park, Kyu Ho" sort="Park, Kyu Ho" uniqKey="Park K" first="Kyu-Ho" last="Park">Kyu-Ho Park</name>
</region>
<name sortKey="Berrier, Catherine" sort="Berrier, Catherine" uniqKey="Berrier C" first="Catherine" last="Berrier">Catherine Berrier</name>
<name sortKey="Ghazi, Alexandre" sort="Ghazi, Alexandre" uniqKey="Ghazi A" first="Alexandre" last="Ghazi">Alexandre Ghazi</name>
</country>
<country name="Australie"><noRegion><name sortKey="Martinac, Boris" sort="Martinac, Boris" uniqKey="Martinac B" first="Boris" last="Martinac">Boris Martinac</name>
</noRegion>
</country>
</tree>
</affiliations>
</record>

Pour manipuler ce document sous Unix (Dilib)

EXPLOR_STEP=$WICRI_ROOT/Wicri/Asie/explor/AustralieFrV1/Data/Main/Exploration

HfdSelect -h $EXPLOR_STEP/biblio.hfd -nk 00A823 | SxmlIndent | more

HfdSelect -h $EXPLOR_AREA/Data/Main/Exploration/biblio.hfd -nk 00A823 | SxmlIndent | more

Pour mettre un lien sur cette page dans le réseau Wicri

{{Explor lien
   |wiki=    Wicri/Asie
   |area=    AustralieFrV1
   |flux=    Main
   |étape=   Exploration
   |type=    RBID
   |clé=     PMC:1304064
   |texte=   Purification and Functional Reconstitution of N- and C-Halves of the MscL Channel
}}

Pour générer des pages wiki

HfdIndexSelect -h $EXPLOR_AREA/Data/Main/Exploration/RBID.i   -Sk "pubmed:15041653" \
       | HfdSelect -Kh $EXPLOR_AREA/Data/Main/Exploration/biblio.hfd   \
       | NlmPubMed2Wicri -a AustralieFrV1

This area was generated with Dilib version V0.6.33.
Data generation: Tue Dec 5 10:43:12 2017. Site generation: Tue Mar 5 14:07:20 2024

Serveur d'exploration sur les relations entre la France et l'Australie

Purification and Functional Reconstitution of N- and C-Halves of the MscL Channel

Purification and Functional Reconstitution of N- and C-Halves of the MscL Channel

Source :

Abstract

Links toward previous steps (curation, corpus...)

Le document en format XML

Pour manipuler ce document sous Unix (Dilib)

Pour mettre un lien sur cette page dans le réseau Wicri

Pour générer des pages wiki

	Serveur d'exploration sur les relations entre la France et l'Australie
	Attention, ce site est en cours de développement ! Attention, site généré par des moyens informatiques à partir de corpus bruts. Les informations ne sont donc pas validées.