CTP:2,3-di-O-geranylgeranyl-sn-glycero-1-phosphate Cytidyltransferase in the Methanogenic ArchaeonMethanothermobacter thermoautotrophicus

Abstract

CDP-2,3-di-O-geranylgeranyl-sn-glycerol synthase (CDP-archaeol synthase) activity was discovered in the membrane fraction of the methanoarchaeon Methanothermobacter thermoautotrophicus cells. It catalyzed the formation of CDP-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 bombardment-mass spectroscopy, chemical analysis, and by UV spectroscopy. One mole of the product was formed from approximately 1 mol of each of the reactants. The enzyme showed maximal activity at pH 8.5 and 55°C in the presence of Mg 2؉ and K ؉ ions. By in vivo pulse labeling of phospholipids with 32 P i , CDP-archaeol was found to be an intracellular intermediate. A cell-free homogenate of M. thermoautotrophicus, when incubated with L-serine, converted the product of CDP-archaeol synthase reaction to a product with the same chromatographic mobility as archaetidylserine. It was concluded from these results that both CDP-archaeol and CDP-archaeol synthase were involved in cellular phospholipid biosynthesis. Among various synthetic substrate analogs, both enantiomers of unsaturated archaetidic acid possessing geranylgeranyl chains showed similar levels of activity, while archaetidic acid with saturated or monounsaturated isoprenoid or straight chains was a poor substrate, despite having the same stereostructure as the fully active substrate. The ester analogs with geranylgeranioyl chains showed significant activities. These results suggest that the enzyme dose not recognize ether or ester bonds between glycerophosphate and hydrocarbon chains nor the stereostructure of the glycerophosphate backbone but mainly targets substrates with geranylgeranyl chains.

Recent Advances in Structural Research on Ether Lipids from Archaea Including Comparative and Physiological Aspects

“…The biosynthetic intermediates of ether polar lipids in archaea include geranylgeranyl ether lipids, which are a kind of allyl ethers. [78][79][80][81] If a stable saturated ether bond is essential for survival at high temperatures and unstable allyl ether bonds are degraded at those temperatures, the organism cannot survive at high temperatures because a trace amount of the unstable allyl ether intermediates would be degraded and the stable lipid could not be synthesized at high temperatures. It can, therefore, be concluded that stable ether bonds are not a result of adaptation to hot environments.…”

(S)-2,3-Di-O-geranylgeranylglyceryl Phosphate Synthase from the Thermoacidophilic Archaeon Sulfolobus solfataricus

“…DGGGP, which has been shown to be the intermediate of archaeal membrane lipids, as evidenced by an incorporation experiment by Eguchi et al (15), would be expected to subsequently undergo saturation of the alkyl group, modification of the polar head group, and the formation of the cyclic tetraether structure (16). However, comprehensive information concerning these biosynthetic reactions is not available: the genes and proteins involved and even the order of the reactions are unknown at this time, expect for some enzymes that catalyze the modification of polar head groups (17,18).…”

CDP-2,3-Di -O- Geranylgeranyl- sn -Glycerol: l -Serine O -Archaetidyltransferase (Archaetidylserine Synthase) in the Methanogenic Archaeon Methanothermobacter thermautotrophicus

“…The GP backbone of phospholipid in the Archaea is sn-glycerol-1-phosphate (G-1-P), which is the enantiomer of its bacterial and eucaryotic counterparts (10). The mechanism by which these specific characteristics are formed is now partly elucidated through in vitro studies of polar lipid biosynthesis in Archaea, which have already revealed four enzymatic reactions and their substrate specificities (14,16,28,29) (Fig. 1).…”

“…CDP-diacylglycerol, which corresponds to CDP-unsaturated archaeol, plays a central role in the biosynthesis of a number of phospholipids in Bacteria (4). In order to prove the next reaction of polar lipid (14,16,28,29). Reaction 5 is described in the present work.…”

“…Genes encoding the first ether bond-forming enzyme are detected in seven species of archaeal genomes (23). The intermediate formed is then activated by CTP to form CDP-unsaturated archaeol by the action of CTP:2,3-di-O-geranylgeranyl-sn-glycerol-1-phosphate cytidyltransferase (14) (Fig. 1, reaction 4).…”