A direct nonchromatographic assay for 1-acyl-sn-glycerol-3-phosphate acyltransferase

1-Acyl-sn-glycerol-3-phosphate acyltransferase (also called lysophosphatidic acid acyltransferase) which catalyzes the acylation of 1-acyl-sn-glycerol-3-phosphate to phosphatidic acid is generally assayed by the use of a radioactive substrate followed by a time-consuming chromatographic separation of substrate and product. We report a direct and highly sensitive nonchromatographic assay for this enzyme based on the ability of Escherichia coli alkaline phosphatase to dephosphorylate 1-acyl-sn-glycerol-3-phosphate but not phosphatidic acid. This selective hydrolysis coupled with the use of ³²P-labeled 1-acyl-sn-glycerol-3-phosphate as substrate permits measurement of the product, ³²P-labeled phosphatidic acid by solvent extraction or precipitation. We also report a series of enzymatic reactions for the efficient conversion of ³²Pi to ³²P-labeled 1-acyl-sn-glycerol-3-phosphate.