SOLUBILIZATION AND BIOCHEMICAL-PROPERTIES OF PHOSPHATIDATE PHOSPHATASE FROM SPINACH CHLOROPLAST ENVELOPE MEMBRANES

Phosphatidate phosphatase activity was first solubilized from spinach chloroplast envelope membranes using a zwitterionic detergent, CHAPS. We have set up a method to assay the solubilized enzyme, using [P-32]p hosphatidic acid as a substrate. Addition of phosphatidylglycerol to t he incubation medium was essential for optimal enzyme activity, probab ly because it was responsible for a better solubilization of the subst rate by CHAPS. However, the possibility of a specific role for this ph ospholipid as a physiological activator cannot be ruled out since the inner envelope membrane from spinach chloroplasts contains significant amounts of phosphatidylglycerol. The biochemical properties of the so lubilized phosphatidate phosphatase from chloroplast envelope membrane s were investigated. As in the native membranes, the solubilized phosp hatidate phosphatase was inhibited by Mg2+ and also by a wide range of metal ions such as Mn2+ and Zn2+. A partial purification of the enzym e was obtained, using hydroxyapatite chromatography. Our results demon strate that the biochemical properties of the envelope phosphatidate p hosphatase are rather different from those of phosphatidate phosphatas e described in other systems, and especially in extraplastidial compar tments from plant tissues. Finally, the question of whether the two en velope enzymes which catalyze the conversion of phosphatidic acid into monogalactosyldiacylglycerol, i.e. phosphatidate phosphatase and 1,2- diacylglycerol galactosyltransferase (or MGDG synthase), could be asso ciated together within the inner envelope membrane is discussed.