CTP : 2,3-di-O-geranylgeranyl-sn-glycero-1-phosphate cytidyltransferase inthe methanogenic archaeon Methanothermobacter thermoautotrophicus

CDP-S, 3-di-O-geranylgeranyl-sn-glycerol synthase (CDP-archaeol synthase) a ctivity was discovered in the membrane fraction of the methanoarchaeon Meth ano-thennobacter thermoautotrophicus cells. It catalyzed the formation of C DP-2,3-di-O-geranylgeranyl-sn-glycerol from CTP and 2,3-di-O-geranylgeranyl -sn-glycero-1-phosphate (unsaturated archaetidic acid). The identity of the reaction product was confirmed by thin layer chromatography, fast atom bom bardment-mass spectroscopy, chemical analysis, and by UV spectroscopy. One mole of the product was formed hom approximately 1 mol of each of the react ants. The enzyme showed maximal. activity at pH 8.5 and 55 degreesC in the presence of Mg2+ and K+ ions. By in vivo pulse labeling of phospholipids wi th P-32(i), CDP-archaeol was found to be an intracellular intermediate. A c ell-free homogenate of Bf. thermoautotrophicus, when incubated with L-serin e, converted the product of CDP-archaeol synthase reaction to a product wit h the same chromatographic mobility as archaetidylserine. It was concluded from these results that both CDP-archaeol and CDP-archaeol synthase were in volved in cellular phospholipid biosynthesis. Among various synthetic subst rate analogs, both enantiomers of unsaturated archaetidic acid possessing g eranylgeranyl chains showed similar levels of activity, while archaetidic a cid with saturated or monounsaturated isoprenoid or straight chains was a p oor substrate, despite having the same stereostructure as the fully active substrate. The ester analogs with geranylgeranioyl chains showed significan t activities. These results suggest that the enzyme dose not recognize ethe r or ester bonds between glycerophosphate and hydrocarbon chains nor the st ereostructure of the glycerophosphate backbone but mainly targets substrate s with geranylgeranyl chains.