Membrane-bound alkaline phosphatase from ectopic mineralization and rat bone marrow cell culture

dc.contributor.authorSimao, Ana Maria. S.
dc.contributor.authorBeloti, Marcio M.
dc.contributor.authorCezarino, Rodrigo M.
dc.contributor.authorRosa, Adalberto Luiz
dc.contributor.authorPizauro, Joao M.
dc.contributor.authorCiancaglini, Pietro
dc.contributor.institutionUniversidade de São Paulo (USP)
dc.contributor.institutionUniversidade Estadual Paulista (Unesp)
dc.date.accessioned2014-05-20T13:17:31Z
dc.date.available2014-05-20T13:17:31Z
dc.date.issued2007-04-01
dc.description.abstractCells from rat bone marrow exhibit the proliferation-differentiation sequence of osteoblasts, form mineralized extracellular matrix in vitro and release alkaline phosphatase into the medium. Membrane-bound alkaline phosphatase was obtained by method that is easy to reproduce, simpler and fast when compared with the method used to obtain the enzyme from rat osseous plate. The membrane-bound alkaline phosphatase from cultures of rat bone marrow cells has a MWr of about 120 kDa and specific PNPP activity of 1200 U/tng. The ecto-enzyme is anchored to the plasma membrane by the GPI anchor and can be released by PIPLC (selective treatment) or polidocanol (0.2 mg/mL protein and 1% (w/v) detergent). The apparent optimum pH for PNPP hydrolysis by the enzyme was pH 10. This fraction hydrolyzes ATP (240 U/mg), ADP (350 U/ mg), glucose 1-phosphate (1100 U/mg), glucose 6-phosphate (340 Wing), fructose 6-phosphate (460 U/mg), pyrophosphate (330 U/mg) and (3glycerophosphate (600 U/mg). Cooperative effects were observed for the hydrolysis of PPi and beta-glycerophosphate. PNPPase activity was inhibited by 0.1 mM vanadate (46%), 0.1 mM ZnCl2 (68%), 1 mM levamisole (66%), 1 mM arsenate (44%), 10 mM phosphate (21%) and 1 mM theophylline (72%). We report the biochemical characterization of membrane-bound alkaline phosphatase obtained from rat bone marrow cells cultures, using a method that is simple, rapid and easy to reproduce. Its properties are compared with those of rat osseous plate enzyme and revealed that the alkaline phosphatase obtained has some kinetics and structural behaviors with higher levels of enzymatic activity, facilitating the comprehension of the mineralization process and its function. (c) 2006 Elsevier B.V. All rights reserved.en
dc.description.affiliationUSP, FFCLRP, Dept Quim, BR-14040901 Ribeirao Preto, Brazil
dc.description.affiliationUSP, FORP, Dept Cirugia TBMF & Periodontia, BR-14040901 Ribeirao Preto, Brazil
dc.description.affiliationUNESP, FCAVJ, Dept Tecnol, BR-14884900 Jaboticabal, SP, Brazil
dc.description.affiliationUnespUNESP, FCAVJ, Dept Tecnol, BR-14884900 Jaboticabal, SP, Brazil
dc.format.extent679-687
dc.identifierhttp://dx.doi.org/10.1016/j.cbpa.2006.05.008
dc.identifier.citationComparative Biochemistry and Physiology A-molecular & Integrative Physiology. New York: Elsevier B.V., v. 146, n. 4, p. 679-687, 2007.
dc.identifier.doi10.1016/j.cbpa.2006.05.008
dc.identifier.issn1095-6433
dc.identifier.urihttp://hdl.handle.net/11449/3952
dc.identifier.wosWOS:000245735500025
dc.language.isoeng
dc.publisherElsevier B.V.
dc.relation.ispartofComparative Biochemistry and Physiology A-molecular & Integrative Physiology
dc.relation.ispartofjcr2.258
dc.relation.ispartofsjr0,836
dc.rights.accessRightsAcesso restrito
dc.sourceWeb of Science
dc.subjectAlkaline phosphatasept
dc.subjectbone marrowpt
dc.subjectOsseous platept
dc.subjectcell culturept
dc.subjectglycosyl phosphatidylinositol anchor (GPI)pt
dc.subjectmembrane solubilizationpt
dc.subjectkinetic datapt
dc.subjectinhibitionpt
dc.titleMembrane-bound alkaline phosphatase from ectopic mineralization and rat bone marrow cell cultureen
dc.typeArtigo
dcterms.licensehttp://www.elsevier.com/about/open-access/open-access-policies/article-posting-policy
dcterms.rightsHolderElsevier B.V.
unesp.author.orcid0000-0003-0149-7189[2]
unesp.author.orcid0000-0002-6495-2778[4]
unesp.author.orcid0000-0002-2785-1345[6]
unesp.author.orcid0000-0002-0911-5053[5]
unesp.campusUniversidade Estadual Paulista (Unesp), Faculdade de Ciências Agrárias e Veterinárias, Jaboticabalpt
unesp.departmentTecnologia - FCAVpt

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