Identification and characterization of the phosphatidylinositol-(4,5)- bisphosphate 5-phosphatase in human platelets

M. Matzaris, S. P. Jackson, K. M. Laxminarayan, C. J. Speed, C. A. Mitchell

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    Phosphatidylinositol (4,5)-bisphosphate (PtdIns(4,5)-P2) is the precursor of several second messenger molecules. In unstimulated cells PtdIns(4,5)P2 is hydrolyzed by a PtdIns(4,5)P2 5-phosphatase to form phosphatidylinositol 4-phosphate (PtdIns(4)P) which is subsequently recycled to phosphatidylinositol. PtdIns(4,5)P2 5-phosphatase activity was detected in platelet cytosolic and particulate fractions. The platelet PtdIns(4,5)P25- phosphatase activity was magnesium but not calcium dependent. The elution profile of platelet cytosolic PtdIns(4,5)P2 5-phosphatase from anion exchange resins, exactly matched that of the 75-kDa inositol-polyphosphate 5- phosphatase (Ins(1,4,5)P3 5-phosphatase). The latter is a signal terminating enzyme responsible for the hydrolysis of inositol (1,4,5)-trisphosphate (Ins(1,4,5)P3) to inositol (1,4)-bisphosphate (Mitchell, C. A., Connolly, T. M., and Majerus, P. W. (1989) J. Biol. Chem. 264, 8873-8877). Polyclonal antibodies raised against recombinant 75-kDa Ins(1,4,5)P3 5-phosphatase specifically immunoprecipitated all PtdIns(4,5)P2 5-phosphatase activity from both the platelet membrane and cytosolic fractions. Purified 75-kDa Ins(1,4,5)P3 5-phosphatase hydrolyzed PtdIns(4,5)P2 forming PtdIns(4)P (K(m) = 250 μM). By contrast, purified membrane-associated 43-kDa Ins(1,4,5)P3 5-phosphatase did not hydrolyze PtdIns(4,5)P2. In the unstimulated platelet, recycling of PtdIns(4,5)P2 to PtdIns(4)P is mediated by the 75-kDa Ins(1,4,5)P3 5-phosphatase.

    Original languageEnglish
    Pages (from-to)3397-3402
    Number of pages6
    JournalJournal of Biological Chemistry
    Issue number5
    Publication statusPublished - 1994

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