Biochemical and biophysical properties of a highly active recombinant arginase from Leishmania (Leishmania) amazonensis and subcellular localization of native enzyme

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dc.contributor.authorSilva, Edson Roberto da
dc.contributor.authorLaranjeira da Silva, Maria Fernanda
dc.contributor.authorFischer, Hannes
dc.contributor.authorMortara, Renato A. [UNIFESP]
dc.contributor.authorMayer, Mario Gustavo
dc.contributor.authorFramesqui, Karine
dc.contributor.authorSilber, Ariel Mariano
dc.contributor.authorFloeter-Winter, Lucile Maria
dc.contributor.institutionUniversidade de São Paulo (USP)
dc.contributor.institutionUniv Luterana Brasil
dc.contributor.institutionUniversidade Federal de São Paulo (UNIFESP)
dc.contributor.institutionInst Butantan Serv Genet
dc.date.accessioned2016-01-24T13:51:26Z
dc.date.available2016-01-24T13:51:26Z
dc.date.issued2008-06-01
dc.description.abstractArginase (L-arginine amidinohydrolase, E.C. 3.5.3.1) is a metalloenzyme that catalyses the hydrolysis of L-arginine to L-ornithine and urea. in Leishmania spp., the biological role of the enzyme may be involved in modulating NO production upon macrophage infection. Previously, we cloned and characterized the arginase gene from Leishmania (Leishmania) amazonensis. in the present work, we successfully expressed the recombinant enzyme in E. coli and performed biochemical and biophysical characterization of both the native and recombinant enzymes. We obtained K-M and V-max. values of 23.9(+/- 0.96) mM and 192.3 mu mol/min mg protein (+/- 14.3), respectively, for the native enzyme. for the recombinant counterpart, K-M was 21.5(+/- 0.90) mM and V-max was 144.9(+/- 8.9) mu mol/min mg. Antibody against the recombinant protein confirmed a glycosomal cellular localization of the enzyme in promastigotes. Data from light scattering and small angle X-ray scattering showed that a trimeric state is the active form of the protein. We determined empirically that a manganese wash at room temperature is the best condition to purify active enzyme. the interaction of the recombinant protein with the immobilized nickel also allowed us to confirm the structural disposition of histidine at positions 3 and 324. the determined structural parameters provide substantial data to facilitate the search for selective inhibitors of parasitic sources of arginase, which could subsequently point to a candidate for leishmaniasis therapy. (c) 2008 Elsevier B.V. All rights reserved.en
dc.description.affiliationUniv São Paulo, Dept Fisiol IB, Inst Biociencias, BR-05508900 São Paulo, Brazil
dc.description.affiliationUniv Luterana Brasil, Ctr Univ Luterano Palmas, CEULP ULBRA Palmas, São Paulo, TO, Brazil
dc.description.affiliationUniv São Paulo, Inst Fis, BR-01498 São Paulo, Brazil
dc.description.affiliationUniversidade Federal de São Paulo, Dept Microbiol Imunol & Parasitol, UNIFESP, São Paulo, Brazil
dc.description.affiliationInst Butantan Serv Genet, São Paulo, Brazil
dc.description.affiliationUniv São Paulo, Inst Ciencias Biomed, BR-05508 São Paulo, Brazil
dc.description.affiliationUnifespUniversidade Federal de São Paulo, Dept Microbiol Imunol & Parasitol, UNIFESP, São Paulo, Brazil
dc.description.sourceWeb of Science
dc.format.extent104-111
dc.identifierhttp://dx.doi.org/10.1016/j.molbiopara.2008.02.011
dc.identifier.citationMolecular and Biochemical Parasitology. Amsterdam: Elsevier B.V., v. 159, n. 2, p. 104-111, 2008.
dc.identifier.doi10.1016/j.molbiopara.2008.02.011
dc.identifier.issn0166-6851
dc.identifier.urihttp://repositorio.unifesp.br/handle/11600/30704
dc.identifier.wosWOS:000256198000004
dc.language.isoeng
dc.publisherElsevier B.V.
dc.relation.ispartofMolecular and Biochemical Parasitology
dc.rightsinfo:eu-repo/semantics/restrictedAccess
dc.rights.licensehttp://www.elsevier.com/about/open-access/open-access-policies/article-posting-policy
dc.subjectprotein structureen
dc.subjectmanganese activationen
dc.subjectimmunolocalizationen
dc.subjectdrug designen
dc.titleBiochemical and biophysical properties of a highly active recombinant arginase from Leishmania (Leishmania) amazonensis and subcellular localization of native enzymeen
dc.typeinfo:eu-repo/semantics/article
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