Interactions of thiophosphatidic acid with enzymes which metabolize phosphatidic acid. Inhibition of phosphatidic acid phosphatase and utilization by CDP-diacylglycerol synthase.
Thiophosphatidic acid (1,2-diacyl-sn-glycero-3-phosphorothioate; thioPA) was chemically synthesized from egg phosphatidylcholine-derived 1,2-diacylglycerol and PSCl3 and tested for its effects on enzymes which utilize phosphatidic acid (PA) in phospholipid biosynthesis. The compound was not a substrate for rat liver cytosolic PA phosphatase and strongly inhibited this enzyme activity. ThioPA was also a potent inhibitor of purified membrane-associated PA phosphatase from Saccharomyces cerevisiae in a competitive manner and exhibited an apparent Ki = 60 microM. In contrast, purified CDPdiacylglycerol synthase (PA:CTP cytidylyltransferase) from this organism was able to convert thioPA to CDP-diacylglycerol. The apparent Vmax for thioPA was 7-fold lower than that for PA, whereas the apparent Km for thioPA (70 microM) was 4-fold lower than that for PA. Calculation of the specificity constant (Vmax/Km) demonstrated that PA was the preferred substrate. These properties of thioPA indicate that this substance may prove useful in studies of phospholipid metabolism and function.
Bonnel
S I
SI
Department of Biochemical and Biophysical Sciences, University of Houston 77204-5500.
Lin
Y P
YP
Kelley
M J
MJ
Carman
G M
GM
Eichberg
J
J
D000818:Animals; D003600:Cytosol / Q000201:enzymology; D007202:Indicators and Reagents; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D009713:Nucleotidyltransferases / Q000302:isolation & purification / Q000378:metabolism*; D010711:Phosphatidate Phosphatase / Q000037:antagonists & inhibitors* / Q000302:isolation & purification / Q000378:metabolism; D010712:Phosphatidic Acids / Q000138:chemical synthesis* / Q000378:metabolism / Q000494:pharmacology; D010713:Phosphatidylcholines; D010744:Phosphoric Monoester Hydrolases / Q000037:antagonists & inhibitors*; D051381:Rats; D012441:Saccharomyces cerevisiae / Q000201:enzymology; D013379:Substrate Specificity; D014316:Tritium
D007202:Indicators and Reagents; D010712:Phosphatidic Acids; D010713:Phosphatidylcholines; D014316:Tritium; D009713:Nucleotidyltransferases; C022348:phosphatidate cytidylyltransferase; D010744:Phosphoric Monoester Hydrolases; D010711:Phosphatidate Phosphatase