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Purification and functional reconstitution of the human CHIP28 water channel expressed in Saccharomyces cerevisiae

1997-12Journal article Restricted access
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dc.contributor.authorLaizé, Vincent
dc.contributor.authorRipoche, P.
dc.contributor.authorTacnet, F.
dc.date.accessioned2014-06-05T14:40:48Z
dc.date.available2014-06-05T14:40:48Z
dc.date.issued1997-12
dc.date.updated2014-06-03T10:28:11Z
dc.description.abstractThe yeast Saccharomyces cerevisiae was used for these proteins are of primary importance to appreciate heterologous expression of the human CHIP28 water their physiological role in living organisms. Aquaporin-1 channel (Aquaporin-1). A nine-amino- Recently, CHIP28 was expressed functionally in acid epitope of the influenza hemagglutinin protein yeast secretory vesicles in our laboratory (3). This work (HA epitope), recognized by the monoclonal antibody indicated that wild-type or mutant aquaporins could 12CA5, was chosen to tag CHIP28 at its N-terminus. be produced easily in this new heterologous expression Epitope-tagged CHIP28 was purified from yeast ex- system for functional analyses. Such studies are fretracts by immunochromatography on protein A/ quently limited in the other known expression systems, 12CA5-coupled beads, after KI extraction and deter- mainly due to maturation or sorting defects (4). It begent solubilization, then concentrated by anion ex- came possible to produce large amounts of aquaporins change chromatography. Purified protein was recon- from yeast cultures for biochemical and biophysical stituted in proteoliposomes and was shown to function studies. Here, we describe a simple method to purify as a water channel by stopped-flow spectrophotome- try. This study demonstrates that the yeast has the yeast-expressed CHIP28, after addition of an epitope capacity to produce functional aquaporins at levels tag at the protein N-terminus. The purification procesufficient for biochemical and biophysical analyses.por
dc.identifier.citationLaizé V; Ripoche P; Tacnet F. Purification and functional reconstitution of the human CHIP28 water channel expressed in Saccharomyces cerevisiae, Protein Expression and Purification, 11, 3, 284-288, 1997.por
dc.identifier.issn1046-5928
dc.identifier.urihttp://hdl.handle.net/10400.1/4207
dc.language.isoengpor
dc.peerreviewedyespor
dc.publisherElsevierpor
dc.titlePurification and functional reconstitution of the human CHIP28 water channel expressed in Saccharomyces cerevisiaepor
dc.typejournal article
dspace.entity.typePublication
oaire.citation.endPage288por
oaire.citation.issue3por
oaire.citation.startPage284por
oaire.citation.titleProtein Expression and Purificationpor
oaire.citation.volume11por
person.familyNameLaizé
person.givenNameVincent
person.identifier.orcid0000-0001-9565-9198
person.identifier.ridB-4463-2008
person.identifier.scopus-author-id6602982778
rcaap.rightsrestrictedAccesspor
rcaap.typearticlepor
relation.isAuthorOfPublicationb51cd3d5-7ee4-47d8-9bc4-b3e297aa222f
relation.isAuthorOfPublication.latestForDiscoveryb51cd3d5-7ee4-47d8-9bc4-b3e297aa222f

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