OSSEOUS PLATE ALKALINE-PHOSPHATASE IS ANCHORED BY GPI

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dc.contributor.authorPizauro, J. M.
dc.contributor.authorCiancaglini, P.
dc.contributor.authorLeone, F. A.
dc.contributor.institutionUniversidade de São Paulo (USP)
dc.contributor.institutionUniversidade Estadual Paulista (Unesp)
dc.date.accessioned2014-05-20T15:26:30Z
dc.date.available2014-05-20T15:26:30Z
dc.date.issued1994-02-01
dc.description.abstractAlkaline phosphatase activity was released up to 100% from the membrane by using 0.1 U of phosphatidylinositol-specific phospholipase C from B. thuringiensis. The Mr of solubilized enzyme was 145,000 by Sephacryl S-300 gel filtration and 66,000 by SDS-PAGE, suggesting a dimeric structure. Solubilization of the membrane-bound enzyme with phospholipase C did not destroy its ability to hydrolyze p-nitrophenyl phosphate (PNPP) (264.3 mu mol min(-1) mg(-1)), ATP (42.0 mu mol min(-1) mg(-1)) and pyrophosphate (28.4 mu mol min(-1) mg(-1)). The hydrolysis of ATP and PNPP by solubilized enzyme exhibited ''Michaelian'' kinetics with K-0.5 = 70 and 979 mu M, respectively. For pyrophosphate, K-0.5 was 128 mu M and site-site interactions were observed (n = 1.4). Magnesium ions were stimulatory (K-d = 1.5 mM) but zinc ions were powerful non-competitive inhibitors (K-d = 6.2 mu M) of solubilized enzyme. Treatment of solubilized alkaline phosphatase with Chellex 100 reduced the original PNPPase activity to 5%. Cobalt (K-0.5 = 10.1 mu M), magnesium (K-0.5 = 29.5 mu M) and manganese ions (K-0.5 = 5 mu M) restored the activity of the apoenzyme with positive cooperativity, suggesting that phosphatidylinositol-specific phospholipase C-solubilized alkaline phosphatase is a metalloenzyme. The stimulation of the apoenzyme by calcium ions (K-0.5 = 653 mu M) was lower than that observed for the other ions (26%) and exhibited site-site interactions (n = 0.7). Zinc ions had no effect on the apoenzyme of the solubilized enzyme.en
dc.description.affiliationUNIV SAO PAULO,FFCLRP,DEPT QUIM,BR-14040901 RIBEIRAO PRET,SP,BRAZIL
dc.description.affiliationUNESP,FAC CIENCIAS AGR & VET JABOTICABAL,DEPT TECNOL,BR-14870000 JABOTICABAL,SP,BRAZIL
dc.description.affiliationUnespUNESP,FAC CIENCIAS AGR & VET JABOTICABAL,DEPT TECNOL,BR-14870000 JABOTICABAL,SP,BRAZIL
dc.format.extent453-456
dc.identifierhttp://www.scielo.br/scielo.php?script=sci_issues&pid=0100-879X&lng=en&nrm=iso
dc.identifier.citationBrazilian Journal of Medical and Biological Research. São Paulo: Associação Bras Divulg Cientifica, v. 27, n. 2, p. 453-456, 1994.
dc.identifier.issn0100-879X
dc.identifier.urihttp://hdl.handle.net/11449/36667
dc.identifier.wosWOS:A1994MX60900052
dc.language.isoeng
dc.publisherAssociação Brasileira de Divulgação Científica (ABRADIC)
dc.relation.ispartofBrazilian Journal of Medical and Biological Research
dc.relation.ispartofjcr1.492
dc.rights.accessRightsAcesso restrito
dc.sourceWeb of Science
dc.subjectPHOSPHATIDYLINOSITOLpt
dc.subjectANCHORpt
dc.subjectAlkaline phosphatasept
dc.subjectOsseous platept
dc.subjectP-NITROPHENYL PHOSPHATEpt
dc.titleOSSEOUS PLATE ALKALINE-PHOSPHATASE IS ANCHORED BY GPIen
dc.typeArtigo
dcterms.licensehttp://www.scielo.br/revistas/bjmbr/iaboutj.htm
dcterms.rightsHolderAssociação Bras Divulg Cientifica
unesp.author.orcid0000-0002-2785-1345[2]
unesp.campusUniversidade Estadual Paulista (Unesp), Faculdade de Ciências Agrárias e Veterinárias, Jaboticabalpt

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