Biochemical characterization of a Kunitz type inhibitor similar to dendrotoxins produced by Rhipicephalus (Boophilus) microplus (Acari: Ixodidae) hemocytes

Página do item simplificado
dc.contributor.authorLima, Cassia A. [UNIFESP]
dc.contributor.authorTorquato, Ricardo J. S. [UNIFESP]
dc.contributor.authorSasaki, Sergio D. [UNIFESP]
dc.contributor.authorJusto, Giselle Z. [UNIFESP]
dc.contributor.authorTanaka, Aparecida S. [UNIFESP]
dc.contributor.institutionUniversidade Federal de São Paulo (UNIFESP)
dc.contributor.institutionUniversidade Estadual de Campinas (UNICAMP)
dc.date.accessioned2016-01-24T13:59:19Z
dc.date.available2016-01-24T13:59:19Z
dc.date.issued2010-02-10
dc.description.abstractA novel chymotrypsin inhibitor identified in fat body and hemocyte cDNA libraries of Boophilus microplus was named BmCI (B. microplus Chymotrypsin Inhibitor) (Genbank EU636772). the putative BmCI amino acid sequence presented a 22-residue-signal peptide and 58-residue-mature protein. BmCI amino acid sequence analysis allowed its classification as a Kunitz-BPTI inhibitor with six cysteine residues, a theoretical pI of 7.8, and the presence of Tyr at P1 position in the putative reactive site, suggesting inhibitory activity toward chymotrypsin. in this work, we reported the biochemical characterization of BmCI. the recombinant BmCI expressed in yeast Pichia pastoris was purified by size exclusion and reverse phase chromatographies. rBmCI expression yield was of I mg L-1 of culture. Purified rBmCI confirmed its chymotrypsin inhibitory activity with a low K-i (6.2 pM). the BmCI gene expression analysis by semi-quantitative RT-PCR indicated its transcription in the hemocytes, salivary gland and ovary. the cytotoxic activity of purified rBmCI was demonstrated in BALB/c 3T3 mouse fibroblasts. As assessed by the MTT reduction assay, rBMCI induced a dose-dependent decrease in 3T3 fibroblasts viability (IC50 = 8 mu M). Moreover, flow cytometry analysis revealed that rBmCI is able to induce apoptosis, whereas no effect was observed on cell cycle progression. in conclusion, we demonstrated that rBmCI is cytotoxic against mammalian cells and obtained evidence that this growth inhibition is caused by an apoptosis-inducing activity. (C) 2009 Published by Elsevier B.V.en
dc.description.affiliationUniversidade Federal de São Paulo, Dept Bioquim, EPM, BR-04044020 São Paulo, Brazil
dc.description.affiliationUniv Estadual Campinas, Dept Bioquim, Inst Biol, BR-13083970 Campinas, SP, Brazil
dc.description.affiliationUnifespUniversidade Federal de São Paulo, Dept Bioquim, EPM, BR-04044020 São Paulo, Brazil
dc.description.sourceWeb of Science
dc.description.sponsorshipFundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)
dc.description.sponsorshipConselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)
dc.description.sponsorshipCoordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)
dc.description.sponsorshipIDFAPESP: 05/03514-9
dc.description.sponsorshipIDFAPESP: 06/52206-8
dc.format.extent279-287
dc.identifierhttp://dx.doi.org/10.1016/j.vetpar.2009.09.030
dc.identifier.citationVeterinary Parasitology. Amsterdam: Elsevier B.V., v. 167, n. 2-4, p. 279-287, 2010.
dc.identifier.doi10.1016/j.vetpar.2009.09.030
dc.identifier.issn0304-4017
dc.identifier.urihttp://repositorio.unifesp.br/handle/11600/32267
dc.identifier.wosWOS:000274942600021
dc.language.isoeng
dc.publisherElsevier B.V.
dc.relation.ispartofVeterinary Parasitology
dc.rightsAcesso restrito
dc.rights.licensehttp://www.elsevier.com/about/open-access/open-access-policies/article-posting-policy
dc.subjectKunitz-BPTI domainen
dc.subjectChymotrypsin inhibitoren
dc.subjectHemocytesen
dc.subjectEctoparasiteen
dc.subjectHematophagousen
dc.subjectDendrotoxinsen
dc.subjectCytotoxicityen
dc.subjectApoptosisen
dc.titleBiochemical characterization of a Kunitz type inhibitor similar to dendrotoxins produced by Rhipicephalus (Boophilus) microplus (Acari: Ixodidae) hemocytesen
dc.typeArtigo
Arquivos
Coleções
Em verificação - Geral