THE ESCHERICHIA-COLI PGPB GENE ENCODES FOR A DIACYLGLYCEROL PYROPHOSPHATE PHOSPHATASE-ACTIVITY

We provided genetic and biochemical evidence that supported the conclu sion that the product of pgpB gene of Escherichia coil exhibited diacy lglycerol pyrophosphate (DGPP) phosphatase activity. DGPP phosphatase activity was absent in pgpB mutant cells and was expressed at high lev els in cells carrying the wild-type pgpB gene on a runaway replication plasmid. The pgpB mutant has been primarily characterized by a defect in phosphatidate (PA) phosphatase activity and also exhibits defects in lyse-PA phosphatase and phosphatidylglycerophosphate phosphatase ac tivities. The defective PA phosphatase in the pgpB mutant was shown to be a Mg2+-independent PA phosphatase activity of the DGPP phosphatase enzyme. We characterized DG:PP phosphatase activity in membranes from cells overproducing the pgpB gene product. DGPP phosphatase catalyzed the dephosphorylation of the beta phosphate of DGPP to form PA follow ed by the dephosphorylation of PA to form diacylglycerol. The specific ity constant (V-max/K-m) for DG:PP was 9.3-fold greater than that for PA. The pH optimum for the DGPP phosphatase reaction was 6.5. Activity was independent of a divalent cation requirement, was potently inhibi ted by Mn2+ ions, and was insensitive to inhibition by N-ethylmaleimid e. Pure DGPP phosphatase from Saccharomyces cerevisiae was shown to be similar to the E. coil DGPP phosphatase in its ability to utilize lys o-PA and phosphatidylglycerophosphate as substrates in vitro.