Characterization of the phosphatidylinositol-specific phospholipase C-released form of rat osseous plate alkaline phosphatase and its possible significance on endochondral ossification

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MetadadosDescriçãoIdioma
Autor(es): dc.contributorUniversidade Estadual Paulista (UNESP)-
Autor(es): dc.contributorUniversidade de São Paulo (USP)-
Autor(es): dc.creatorPizauro, João M.-
Autor(es): dc.creatorCiancaglini, Pietro-
Autor(es): dc.creatorLeone, Francisco A.-
Data de aceite: dc.date.accessioned2025-08-21T20:57:38Z-
Data de disponibilização: dc.date.available2025-08-21T20:57:38Z-
Data de envio: dc.date.issued2022-04-28-
Data de envio: dc.date.issued2022-04-28-
Data de envio: dc.date.issued1995-11-01-
Fonte completa do material: dc.identifierhttp://dx.doi.org/10.1007/BF01076074-
Fonte completa do material: dc.identifierhttp://hdl.handle.net/11449/224010-
Fonte: dc.identifier.urihttp://educapes.capes.gov.br/handle/11449/224010-
Descrição: dc.descriptionAlkaline phosphatase activity was released up to 100% from the membrane by incubating the rat osseous plate membrane-bound enzyme with phosphatidylinositol-specific phospholipase C. The molecular weight of the released enzyme was 145,000 on Sephacryl S-300 gel filtration and 66,000 on PAGE-SDS, suggesting a dimeric structure. Solubilization of the membrane-bound enzyme with phospholipase C did not destroy its ability to hydrolyse PNPP, ATP and pyrophosphate. The hydrolysis of ATP and PNPP by phosphatidylinositol-specific phospholipase C-released enzyme exhibited 'Michaelian' kinetics with K0.5=70 and 979 μM, respectively. For pyrophosphate, K0.5 was 128 μM and site-site interactions were observed (n=1.4). Magnesium ions were stimulatory (K0.5=1.5 mM) and zinc ions were a powerful noncompetitive inhibitor (Ki=6.2 μM) of phosphatidylinositol-specific phospholipase C-released enzyme. Phosphatidylinositol-specific phospholipase C-released alkaline phosphatase was relatively stable at 40°C. However, with increasing temperature from 40-60°C, the enzyme was inactivated rapidly following first order kinetics and thermal inactivation constants varied from 5.08×10-4 min-1 to 0.684 min-1. Treatment of phosphatydilinositol-specific phospholipase C-released alkaline phosphatase with Chellex 100 depleted to 5% its original PNPPase activity. Magnesium (K0.5=29.5 μM), manganese (K0.5=5 μM) and cobalt ions (K0.5=10.1 μM) restored the activity of Chelex-treated enzyme, demonstrating its metalloenzyme nature. The stimulation of Chelex-treated enzyme by calcium ions (K0.5=653 μM) was less effective (only 26%) and occurred with site-site interactions (n=0.7). Zinc ions had no stimulatory effects. The possibility that the soluble form of the enzyme, detected during endochondral ossification, would arise by the hydrolysis of the P1-anchored form of osseous plate alkaline phosphatase is discussed. © 1995 Kluwer Academic Publishers.-
Descrição: dc.descriptionDepartamento de Tecnologia Faculdade de Ciências Agrárias e Veterinárias de Jaboticabal-UNESP, Av. Bandeirantes 3900, Ribeirão Preto, 14040-901, SP-
Descrição: dc.descriptionDepartamento de Química, Faculdade de Filosofia Ciências e Letras de Ribeirão Preto-USP, Av. Bandeirantes 3900, Ribeirão Preto, 14040-901, SP-
Descrição: dc.descriptionDepartamento de Tecnologia Faculdade de Ciências Agrárias e Veterinárias de Jaboticabal-UNESP, Av. Bandeirantes 3900, Ribeirão Preto, 14040-901, SP-
Formato: dc.format121-129-
Idioma: dc.languageen-
Relação: dc.relationMolecular and Cellular Biochemistry-
???dc.source???: dc.sourceScopus-
Palavras-chave: dc.subjectalkaline phosphatase-
Palavras-chave: dc.subjecthydrophobic chromatography-
Palavras-chave: dc.subjectosseous plate-
Palavras-chave: dc.subjectp-nitrophenyl phosphate-
Palavras-chave: dc.subjectphosphatidylinositol-specific phospholipase C-
Palavras-chave: dc.subjectphospholipase C-
Título: dc.titleCharacterization of the phosphatidylinositol-specific phospholipase C-released form of rat osseous plate alkaline phosphatase and its possible significance on endochondral ossification-
Tipo de arquivo: dc.typelivro digital-
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