[Studies of phosphorylation in rat mast cells (supplement 2). Diphosphoinositide kinase in rat mast cell granules].

Rat mast cells were purified by a Percoll gradient, and the granules were isolated from the sonicated mast cells on a Percoll gradient. The granules were shown to contain a diphosphoinositide kinase which catalyzes the formation of triphosphoinositide (TPI) from diphosphoinositide (DPI). The enzyme requires ATP and Mg2+ for the activity. TPI formation is almost completely dependent on MgCl2 or MnCl2, and maximal response is observed at 20 mM or 1 mM, respectively. The Km for ATP is 3 microM. TPI formation in the granules is dependent on the reaction time. The maximal response is seen at 23 degrees C. NaCl, KCl, Na2HPO4 and KH2PO4 have no effect on the activity. One hundred microM adenosine, AMP, ADP, and 10 microM cyclic AMP inhibit the kinase activity.