Hydrolysis of short acyl chain inositol lipids by phospholipase C-delta 1.
We investigated the relationship between substrate aggregation and activation of phosphoinositide-specific phospholipase C-delta 1 (PLC-delta 1), isolated from bovine brain cytosol. The inositol lipids 1,2-dibutyryl-sn-glycero-3-phosphoinositol (di-C4-PI), 1,2-dihexanoyl-sn-glycero-3-phosphoinositol (di-C6-PI), and 1,2-dioctanoyl-sn-glycero-3-phosphoinositol (di-C8-PI) were prepared from synthetic cytidine diphosphate diglyceride analogs in a reaction with myo-inositol catalyzed by yeast phosphatidylinositol synthase. All three lipids served as substrates for PLC-delta 1 at concentrations significantly below their critical micelle concentration (cmc). Under these conditions, steps that might limit the reaction rate, such as membrane adsorption or penetration into the phospholipid surface, were eliminated. Below the cmc, the concentration of lipid substrate required to produce hydrolysis followed the order: di-C8-PI < di-C6-PI << di-C4-PI. Calcium was essential for hydrolysis of the short chain substrates at all lipid concentrations tested. The dependence of the reaction on calcium suggests that this ion activates PLC-delta 1 at a step other than adsorption to or penetration of the membrane surface. As the concentration of di-C8-PI was raised above the cmc, the reaction velocity increased 2-3-fold. These results are consistent with the idea that micellar or bilayer aggregates of phosphoinositol are not required for PLC-catalyzed hydrolysis, although the reaction rate is enhanced by micelle formation.
Rebecchi
M J
MJ
Department of Physiology and Biophysics, State University of New York, Stony Brook 11794.
Eberhardt
R
R
Delaney
T
T
Ali
S
S
Bittman
R
R
D000818:Animals; D001921:Brain / Q000201:enzymology; D051607:CDP-Diacylglycerol-Inositol 3-Phosphatidyltransferase; D002118:Calcium / Q000494:pharmacology; D002417:Cattle; D003600:Cytosol / Q000201:enzymology; D004789:Enzyme Activation / Q000187:drug effects; D006868:Hydrolysis; D007700:Kinetics; D010716:Phosphatidylinositols / Q000378:metabolism*; D010770:Phosphotransferases / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000201:enzymology; D013379:Substrate Specificity; D017855:Transferases (Other Substituted Phosphate Groups); D010738:Type C Phospholipases / Q000378:metabolism*
D010716:Phosphatidylinositols; C079262:1,2-dioctanoyl-sn-glycero-3-phosphoinositol; C079261:1,2-dihexanoyl-sn-glycero-3-phosphoinositol; C079260:1,2-dibutyryl-sn-glycero-3-phosphoinositol; D010770:Phosphotransferases; D017855:Transferases (Other Substituted Phosphate Groups); D051607:CDP-Diacylglycerol-Inositol 3-Phosphatidyltransferase; D010738:Type C Phospholipases; D002118:Calcium