METABOLISM OF DIACYLGLYCEROL PYROPHOSPHATE BY SUSPENSION-CULTURED CATHARANTHUS-ROSEUS CELLS - IDENTIFICATION AND CHARACTERIZATION OF DIACYLGLYCEROL PYROPHOSPHATE PHOSPHATASE IN PLANTS

Diacylglycerol pyrophosphate (DGPP) is a novel phospholipid found in p lants that is the product of the reaction catalyzed by phosphatidate k inase. In this study we examined the metabolism of DGPP in Catharanthu s roseus cells. Metabolic labeling studies showed that DGPP was rapidl y metabolized to phosphatidate and P-i. The enzyme catalyzing this rea ction was named DGPP phosphatase. DGPP phosphatase activity was 20 tim es more active than phosphatidate kinase activity. Microsomal and solu ble isoforms of DGPP phosphatase were found in C. roseus cells. The mi crosomal enzyme was primarily associated with intracellular membranes. The pH optima of the microsomal and soluble DGPP phosphatases were 4. 5 and 3.5, respectively. Both activities were, independent of a divale nt cation requirement and were insensitive to EDTA and N-ethylmaleimid e. The DGPP phosphatase activities were inhibited by NaF, MgCl2, MnCl2 , and sphingosine. The dependence of the microsomal and soluble DGPP p hosphatase activities on DGPP were dose-dependent with maximum activit ies obtained at 100 mu M (3 mol %) and 600 mu M (20 mol %) respectivel y, using a Triton X-100/DGPP mixed micellar substrate. Pyrophosphate, p -nitrophenylphosphate, and adenosine diphosphate (ADP) were not subs trates for the DGPP phosphatase enzymes. DGPP phosphatase activity was also found in a wide range of organisms including Escherichia coli, S accharomyces cerevisiae, rat, pig, and cattle. (C) 1997 Elsevier Scien ce Ireland Ltd.