A sensitive assay for phosphatidate phosphohydrolase in mouse liver microsomes.
A technique is described for the assay of phosphatidate phosphohydrolase using 1,2-[9,10-3H]dioleoyl-sn-glycero-3-phosphate as a substrate. This substrate was prepared enzymatically using mouse liver microsomes washed with 0.5 M NaCl, which synthesize minimal amounts of neutral lipids at high enzyme concentrations. Measurement of the product, 1,2-[9,10-3H]dioleoylglycerol, was 10-fold more sensitive than the usual colorimetric assay for inorganic phosphate release. In addition, the assay provides information about the relative contribution of other activities which limit the availability of diacylglycerols for further esterification to triacylglycerols and/or phospholipids.
D000818:Animals; D002855:Chromatography, Thin Layer / Q000379:methods*; D008297:Male; D051379:Mice; D008862:Microsomes, Liver / Q000201:enzymology*; D010711:Phosphatidate Phosphatase / Q000032:analysis*; D010712:Phosphatidic Acids / Q000302:isolation & purification / Q000378:metabolism; D010744:Phosphoric Monoester Hydrolases / Q000032:analysis*
D010712:Phosphatidic Acids; D010744:Phosphoric Monoester Hydrolases; D010711:Phosphatidate Phosphatase