Formation of 1-alkyl-2-acetyl-sn-glycerols via the de novo biosynthetic pathway for platelet-activating factor. Characterization of 1-alkyl-2-acetyl-sn-glycero-3-phosphate phosphohydrolase in rat spleens.

Abstract 1-Alkyl-2-acetyl-sn-glycerol (alkylacetyl-G) is an important intermediate in the biosynthesis of 1-alkyl-2-acetyl-sn-glycero-3-phosphocholine (platelet-activating factor) from 1-alkyl-2-lyso-sn-glycero-3-phosphate (alkyllyso-GP) via the de novo pathway. In the present investigation, we have characterized a 1-alkyl-2-acetyl-sn-glycero-3-phosphate (alkylacetyl-GP) phosphohydrolase in rat spleens that catalyzes the conversion of alkylacetyl-GP to alkylacetyl-G. The bulk of the enzymatic activity (53%) is located in the microsomal fraction, whereas 28% of the activity is present in mitochondria. The microsomal enzyme has an optimal pH of 7.0-7.4, an "apparent" Km of 31.8 microM for alkylacetyl-GP, and is widely distributed in various rat tissues. Studies of alkylacetyl-GP phosphohydrolase with respect to substrate specificity, pH profiles, sensitivities to temperature, and effects of detergent, ethanol, or cations indicate the activity of this enzyme can be distinguished from the activities of a nonspecific phosphomonoesterase or phosphatidate phosphohydrolase. Like alkyllyso-GP:acetyl-CoA acetyltransferase, the alkylacetyl-GP phosphohydrolase shows no notable substrate selectivities with regard to variations in alkyl chain length (C16:0 versus C18:0) at the sn-1 position or short chain acyl groups (C2:0 to C6:0, with the exception of C3:0) at the sn-2 position of the glycerol moiety. The enzymatic activity of alkylacetyl-GP phosphohydrolase is 30-90-fold higher than alkyllyso-GP:acetyl-CoA acetyltransferase in most tissues examined. Even though alkyllyso-GP is a substrate for alkyllyso-GP:acetyl-CoA acetyltransferase, it can also be degraded by alkylacetyl-GP phosphohydrolase. Thus, our findings coupled with earlier results imply that specificities of the molecular species of platelet-activating factor synthesized de novo are determined by the enzyme involved in the final step of this pathway, the dithiothreitol-insensitive alkylacetyl-G:CDP-choline cholinephosphotransferase. Furthermore, alkyl-lyso-GP:acetyl-CoA acetyltransferase appears to be the rate-limiting step in the de novo synthesis of alkylacetyl-G.