Metabolism of phospholipids IX. Phosphatidate phosphohydrolase in rat liver

Abstract

1. 1. Rat-liver homogenates were fractionated into mitochondrial, lysosomal, microsomal, and particle-free supernatant fractions which were characterised by specific marker enzymes, and the intracellular distribution of phosphatidate phosphohydrolase ( l-α-phosphatidate phosphohydrolase, EC 3.1.3.4) was studied. 2. 2. The lysosomal fraction had the highest specific activity with respect to phosphatidate phosphohydrolase activity but the mitochondrial and microsomal fractions contained the relatively highest proportions of this enzyme. It was concluded that phosphatidate phosphohydrolase is a true constituent of all three particulate sub-cellular fractions, with some activity occurring in the supernatant. 3. 3. Repeated extraction of the three particulate subcellular fractions by freezing and thawing in 0.3 M sucrose solubilised 60, 25 and 16% of the mitochondrial, lysosomal, and microsomal phosphatidate phosphohydrolase activity, respectively. 4. 4. Investigation of Michaelis constants, pH optima, and the effect of detergents and cations revealed significant differences between the mitochondrial enzyme which had been extracted by freezing and thawing and that which remained insoluble. 5. 5. Examination of an extract obtained from a mitochondrial preparation by repeated freezing and thawing showed that of a number of important enzymes in glyceride and phospholipid biosynthesis only phosphatidate phosphohydrolase was extracted.