Cloning, expression, purification and characterization of recombinant glutathione-S-transferase from Xylella fastidiosa

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dc.contributor.authorTravensolo, Regiane F.
dc.contributor.authorGarcia, Wanius
dc.contributor.authorMuniz, Joao R. C.
dc.contributor.authorCaruso, Celia S.
dc.contributor.authorLemos, Eliana G. M. [UNESP]
dc.contributor.authorCarrilho, Emanuel
dc.contributor.authorAraujo, Ana P. U.
dc.contributor.institutionUniversidade de São Paulo (USP)
dc.contributor.institutionUniversidade Estadual Paulista (Unesp)
dc.date.accessioned2014-05-20T15:33:27Z
dc.date.available2014-05-20T15:33:27Z
dc.date.issued2008-05-01
dc.description.abstractXylella fastidiosa is an important pathogen bacterium transmitted by xylem-feedings leafhoppers that colonizes the xylem of plants and causes diseases on several important crops including citrus variegated chlorosis (CVC) in orange and lime trees. Glutathione-S-transferases (GST) form a group of multifunctional isoenzymes that catalyzes both glutathione (GSH)-dependent conjugation and reduction reactions involved in the cellular detoxification of xenobiotic and endobiotic compounds. GSTs are the major detoxification enzymes found in the intracellular space and mainly in the cytosol from prokaryotes to mammals, and may be involved in the regulation of stress-activated signals by suppressing apoptosis signal-regulating kinase 1. In this study, we describe the cloning of the glutathione-S-transferase from X. fastidiosa into pET-28a(+) vector, its expression in Escherichia coli, purification and initial structural characterization. The purification of recombinant xfGST (rxfGST) to near homogeneity was achieved using affinity chromatography and size-exclusion chromatography (SEC). SEC demonstrated that rxfGST is a homodimer in solution. The secondary and tertiary structures of recombinant protein were analyzed by circular dichroism and fluorescence spectroscopy, respectively. The enzyme was assayed for activity and the results taken together indicated that rxfGST is a stable molecule, correctly folded, and highly active. Several members of the GST family have been extensively studied. However, xfGST is part of a less-studied subfamily which yet has not been structurally and biochemically characterized. In addition, these studies should provide a useful basis for future studies and biotechnological approaches of rxfGST. (C) 2008 Elsevier B.V. All rights reserved.en
dc.description.affiliationUniv São Paulo, Lab Biofis Mol Sergio Mascarenhas, Inst Fis São Carlos, BR-13560970 São Carlos, SP, Brazil
dc.description.affiliationUSP, Lab Bioanalit Microfabricacao & Separacoes, Inst Quim São Carlos, São Carlos, Brazil
dc.description.affiliationUniv Estadual Paulista UNESP, Lab Bioquim Microrganismos & Plantas, Jaboticabal, Brazil
dc.description.affiliationUnespUniv Estadual Paulista UNESP, Lab Bioquim Microrganismos & Plantas, Jaboticabal, Brazil
dc.format.extent153-160
dc.identifierhttp://dx.doi.org/10.1016/j.pep.2008.01.017
dc.identifier.citationProtein Expression and Purification. San Diego: Academic Press Inc. Elsevier B.V., v. 59, n. 1, p. 153-160, 2008.
dc.identifier.doi10.1016/j.pep.2008.01.017
dc.identifier.issn1046-5928
dc.identifier.urihttp://hdl.handle.net/11449/42074
dc.identifier.wosWOS:000255312300021
dc.language.isoeng
dc.publisherAcademic Press Inc. Elsevier B.V.
dc.relation.ispartofProtein Expression and Purification
dc.relation.ispartofjcr1.338
dc.relation.ispartofsjr0,648
dc.rights.accessRightsAcesso restrito
dc.sourceWeb of Science
dc.subjectXylella fastidiosaen
dc.subjectglutathione-S-transferase (GST)en
dc.subjectdetoxification enzymesen
dc.subjectcircular dichroism spectroscopy (CD)en
dc.subjectfluorescence spectroscopyen
dc.titleCloning, expression, purification and characterization of recombinant glutathione-S-transferase from Xylella fastidiosaen
dc.typeArtigo
dcterms.licensehttp://www.elsevier.com/about/open-access/open-access-policies/article-posting-policy
dcterms.rightsHolderAcademic Press Inc. Elsevier B.V.
unesp.author.orcid0000-0001-7351-8220[6]
unesp.author.orcid0000-0001-5455-084X[7]
unesp.author.orcid0000-0003-2731-3966[3]
unesp.campusUniversidade Estadual Paulista (Unesp), Faculdade de Ciências Agrárias e Veterinárias, Jaboticabalpt

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