Biosynthesis of phosphatidylinositol in Crithidia fasciculata.
Microsomal preparations from the protozoan (Crithidia fasciculata were shown to incorporate myo-[2-3H]inositol into phosphatidylinositol by both the CDPdiacylglycerol:myo-inositol phosphatidyltransferase reaction and by a myo-inositol exchange reaction. Non-ionic detergent and Mg2+ were necessary for the measurement of transferase activity. Untreated preparations could not be saturated with Mg2+, even at very high concentrations (50-75 mM). However, low concentrations of EGTA (75 micro M) both stimulated the activity 3-fold and reduced the Mg2+ required for saturation to 15-20 mM. EGTA also increased the apparent Km for CDPdiacylglycerol while increasing the sensitivity to substrate inhibition above 1 mM. The transferase activity was inhibited by relatively low concentrations of Ca2+ (50 micro M). This and the EGTA effect suggest a possible role for Ca2+ in the modulation of phosphatidylinositol synthesis. The myo-inositol exchange activity required Mn2+, was insensitive to Ca2+ inhibition and was only slightly stimulated by detergents and EGTA. This activity was preferentially inactivated by heating at 50 degrees C in the presence of Triton X-100. In a detergent solubilized preparation the exchange activity but not the transferase exhibited a non-specific requirement for phospholipid. The differences in properties of the two activities suggest the presence of a separate exchange enzyme.
D000818:Animals; D051607:CDP-Diacylglycerol-Inositol 3-Phosphatidyltransferase; D002118:Calcium / Q000494:pharmacology; D003566:Cytidine Diphosphate Choline / Q000494:pharmacology; D003567:Cytidine Diphosphate Diglycerides / Q000378:metabolism; D003902:Detergents / Q000494:pharmacology; D004533:Egtazic Acid / Q000494:pharmacology; D056890:Eukaryota / Q000378:metabolism*; D006358:Hot Temperature; D008345:Manganese / Q000494:pharmacology; D008861:Microsomes / Q000378:metabolism; D010716:Phosphatidylinositols / Q000096:biosynthesis*; D010770:Phosphotransferases / Q000378:metabolism; D012995:Solubility; D013268:Stimulation, Chemical; D017855:Transferases (Other Substituted Phosphate Groups)
D003567:Cytidine Diphosphate Diglycerides; D003902:Detergents; D010716:Phosphatidylinositols; D008345:Manganese; D004533:Egtazic Acid; D003566:Cytidine Diphosphate Choline; D010770:Phosphotransferases; D017855:Transferases (Other Substituted Phosphate Groups); D051607:CDP-Diacylglycerol-Inositol 3-Phosphatidyltransferase; D002118:Calcium