title: Platelet membrane phosphatidylinositol kinase activity. Triton X-100 effects provide evidence for intramicellar reaction.
abstract: Phosphatidylinositol (PI) kinase activity of platelet membranes was solubilized and partially purified by anion-exchange chromatography to measure the initial enzymatic rates. Kinetic studies were performed in the presence of Triton X-100 to obtain mixed micelles. The partially purified enzyme exhibited a Michaelian behaviour towards ATP, with a Km of 58 microM. The enzymatic rates were dependent upon Triton concentrations. Upon increasing its concentration, this detergent exhibited an activating effect followed by an inhibitory one. The optimal micellar Triton concentration was proportionnal to the PI concentration used in the assay. Conversely, the behaviour of the enzyme towards PI was dependent upon the Triton concentration. However, when PI concentration was expressed as its surfacic concentration within the micelles, the activity became independent of the detergent concentration, and a Km value of 0.09 mol/mol was estimated. Therefore, in vitro phosphorylation of phosphatidylinositol by PI kinase is rate-limited by an intramicellar reaction, and provides a study model for the in vivo reaction.
authors.affiliation: Institut de Biochimie Cellulaire et Neurochimie, Centre National de la Recherche Scientifique, Bordeaux, France.
authors.forename: D
authors.initials: D
authors.lastname: Boué
authors.forename: O M
authors.initials: OM
authors.lastname: Viratelle
mesh_terms: D019870:1-Phosphatidylinositol 4-Kinase; D001792:Blood Platelets / Q000201:enzymology*; D002462:Cell Membrane / Q000201:enzymology; D002852:Chromatography, Ion Exchange; D003902:Detergents; D006801:Humans; D007700:Kinetics; D008823:Micelles; D017830:Octoxynol; D010770:Phosphotransferases / Q000302:isolation & purification / Q000378:metabolism*; D011092:Polyethylene Glycols
chemical_list: D003902:Detergents; D008823:Micelles; D011092:Polyethylene Glycols; D017830:Octoxynol; D010770:Phosphotransferases; D019870:1-Phosphatidylinositol 4-Kinase