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An improved non-denaturing method for the purification of spiralin, the main membrane lipoprotein of the pathogenic bacteria Spiroplasma melliferum

hal.structure.identifierInstitut de Génétique et Développement de Rennes [IGDR]
dc.contributor.authorDESFOUGERES, Yann
hal.structure.identifierBiologie du fruit et pathologie [BFP]
dc.contributor.authorPOITOU, Jean-Michel
hal.structure.identifierInstitut de Génétique et Développement de Rennes [IGDR]
dc.contributor.authorWRÓBLEWSKI, Henri
hal.structure.identifierInstitut de Génétique et Développement de Rennes [IGDR]
hal.structure.identifierBiologie du fruit et pathologie [BFP]
dc.contributor.authorBEVEN, Laure
dc.date.issued2016-11
dc.identifier.issn1570-0232
dc.description.abstractEnSpiralin is the most abundant protein of several species of spiroplasmas, helical, motile bacteria pathogenic for arthropods and plants. This amphiphilic protein is anchored to the outer face of the plasma membrane by a lipoylated N-terminal cysteine. Although spiroplasma pathogenicity in mammals is controversial, it was shown that spiralin is highly immunogenic and endowed with immunomodulatory activity. In this paper, we describe a high performance method for the purification of Spiroplasma melliferum spiralin under non-denaturing conditions. The protein was selectively extracted with 3-[(3-cholamidopropyl) dimethylammonio]-1-propyl sulfonate (CHAPS) from the membrane pre-treated with sodium dodecyl-N-sarcosinate (Sarkosyl), and purified to homogeneity by cation-exchange HPLC with an overall yield of ∼60%. Detergent-depleted, water-soluble micelles of spiralin displaying a mean diameter of 170Å, as evidenced by transmission electron microscopy, were obtained by dialysis detergent removal. Circular dichroism spectroscopy and cross immunoprecipitation assay of the purified spiralin strongly suggested that this purification method could retain the structural characteristics of the native spiralin. The strategy developed to purify spiralin (two successive selective extractions of membrane proteins with mild detergents followed by ion-exchange chromatography) should prove useful for the purification of membrane lipoproteins of other bacteria of the class Mollicutes including different pathogens for humans, animals and plants.
dc.language.isoen
dc.publisherElsevier
dc.subject.enMollicutes
dc.subject.enSpiralin
dc.subject.enCE-HPLC
dc.subject.enDetergents
dc.subject.enMembrane lipoprotein
dc.title.enAn improved non-denaturing method for the purification of spiralin, the main membrane lipoprotein of the pathogenic bacteria Spiroplasma melliferum
dc.typeArticle de revue
dc.identifier.doi10.1016/j.jchromb.2016.10.012
dc.subject.halSciences du Vivant [q-bio]
dc.subject.halSciences du Vivant [q-bio]/Microbiologie et Parasitologie/Bactériologie
bordeaux.journalJournal of Chromatography B - Analytical Technologies in the Biomedical and Life Sciences
bordeaux.page149-156
bordeaux.volume1036-1037
bordeaux.peerReviewedoui
hal.identifierhal-01558319
hal.version1
hal.popularnon
hal.audienceInternationale
hal.origin.linkhttps://hal.archives-ouvertes.fr//hal-01558319v1
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