Phospholipase D activity in subcellular membranes of rat ventricular myocardium.
The phospholipase D (PL D), which catalyzes the formation of phosphatidic acid (PA), was studied in rat myocardium using 14C-labelled phosphatidylcholine (PC) as an exogenous substrate. Subcellular distribution experiments indicated the presence of PL D in particulate fractions only. Different procedures for the isolation of purified cardiac subcellular organelles showed the presence of PL D in sarcolemma (SL), sarcoplasmic reticulum (SR) and mitochondria with 14-, 11- and 5-fold enrichment when compared to the homogenate value, respectively. The activity of SL PL D was observed over a narrow acid pH range with an optimum at 6.5, and it showed a high specificity for PC while phosphatidylethanolamine and phosphatidylinositol showed a low rate of hydrolysis. Under optimal conditions, PA formation was linear for a 90-min period of incubation and the reaction rate was constant for 10 to 100 micrograms SL protein in the assay medium. The SR PL D displayed properties similar to those seen with the SL PL D. In membrane fractions PL D was also found to catalyze a transphosphatidylation reaction for the synthesis of phosphatidylglycerol. Assessment of the intramembranal levels of radioactive 1,2-diacylglycerol (DAG) in the absence or presence of KF suggested the presence of an active PA phosphohydrolase activity. This study indicates that a PC-specific PL D activity is localized in different membrane systems of the myocardium and may be associated with PA phosphohydrolase to act in a coordinated manner. The functional significance of PL D-dependent formation of PA in cardiac membranes is discussed.
Panagia
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V
Division of Cardiovascular Sciences, St. Boniface General Hospital Research Centre, Winnipeg, Manitoba, Canada.
Ou
C
C
Taira
Y
Y
Dai
J
J
Dhalla
N S
NS
D000818:Animals; D004075:Diglycerides / Q000378:metabolism; D006352:Heart Ventricles / Q000201:enzymology*; D006868:Hydrolysis; D008297:Male; D008928:Mitochondria / Q000201:enzymology; D009206:Myocardium / Q000201:enzymology*; D010715:Phosphatidylglycerols / Q000378:metabolism; D010739:Phospholipase D / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains; D012508:Sarcolemma / Q000201:enzymology; D012519:Sarcoplasmic Reticulum / Q000201:enzymology; D013379:Substrate Specificity
C011439:1,2-diacylglycerol; D004075:Diglycerides; D010715:Phosphatidylglycerols; D010739:Phospholipase D