Phosphorylation of phosphatidylinositol in rat liver Golgi.
The phosphorylation of phosphatidylinositol in subcellular fractions from rat liver has been examined. Fractions enriched in Golgi showed by far the highest specific activity while a plasma membrane fraction depleted in Golgi, as well as rough microsomes, mitochondria, and particle-free supernatant had much lower activity. The product formed from [gamma-32P]ATP and endogenous or exogenously added phosphatidylinositol was predominantly diphosphoinositide with no more than 5% triphosphoinositide. The phosphatidylinositol kinase showed a broad pH optimum with peak activity around pH 7.8. The kinase reaction was not inhibited by the detergent Triton X-100, except at very high concentration, while it was severely inhibited by digitonin. Exogenous phosphatidylinositol did not serve as substrate for the kinase when added in the form of sonicated vesicles, but did so in the presence of Triton. In the latter form it also restored kinase activity after enzymatic depletion of endogenous substrate. The diphosphoinositide formed from endogenous phosphatidylinositol remained fairly stable in the intact membrane, while its degradation was enhanced significantly in the presence of detergent. This study indicates that the phosphatidylinositol kinase in rat liver is highly enriched in the Golgi and suggests that it can be solubilized and assayed by the use of a nonionic detergent.
D000818:Animals; D002458:Cell Fractionation; D003902:Detergents / Q000494:pharmacology; D006056:Golgi Apparatus / Q000378:metabolism*; D007700:Kinetics; D008099:Liver / Q000378:metabolism*; D008297:Male; D017830:Octoxynol; D010716:Phosphatidylinositols / Q000378:metabolism*; D010766:Phosphorylation; D011092:Polyethylene Glycols / Q000494:pharmacology; D051381:Rats; D011919:Rats, Inbred Strains; D013347:Subcellular Fractions / Q000378:metabolism
D003902:Detergents; D010716:Phosphatidylinositols; D011092:Polyethylene Glycols; D017830:Octoxynol