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J. Bacteriol. doi:10.1128/JB.01875-06
Copyright (c) 2007, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

In vitro Biosynthesis of Ether-type Glycolipids in the Methanoarchaeon Methanothermobacter thermautotrophicus

Department of Chemistry, School of Medicine, University of Occupational and Environmental Health, Yahatanishi-ku, Kitakyushu 807-8555, and Department of Chemistry and Materials Science, Tokyo Institute of Technology, Meguro-ku Tokyo 152-8551, Japan

^* To whom correspondence should be addressed. Email: h-morii{at}health.uoeh-u.ac.jp.

	Abstract

The biosynthesis of archaeal ether-type glycolipids was investigated in vitro using Methanothermobacter thermautotrophicus cell-free homogenates. The sole sugar moiety of glycolipids and phosphoglycolipids of the organism is the -D-glucosyl-(1->6)-D-glucosyl (gentiobiosyl) unit. The enzyme activities of archaeol:UDP-glucose -glucosyltransferase (monoglucosylarchaeol synthase, MGA synthase) and monoglucosylarchaeol:UDP-glucose -1,6-glucosyltransferase (diglucosylarchaeol synthase, DGA synthase) were found in the methanoarchaeon. The synthesis of DGA is probably a two-step glucosylation: (i) archaeol + UDP-glucose -> MGA + UDP; and (ii) MGA + UDP-glucose -> DGA + UDP. Both enzymes required the addition of K⁺ ions and archaetidylinositol for their activities. Ten mM MgCl₂ stimulated DGA synthase, in contrast to MGA synthase, which did not require Mg²⁺. It was likely that the activities of MGA synthesis and DGA synthesis were carried out by different proteins because of the Mg²⁺ requirement and their cellular localization. MGA synthase and DGA synthase can be distinguished in cell extracts greatly enriched for each activity by demonstrating differing Mg²⁺ requirements of each enzyme. MGA synthase preferred a lipid-substrate with the sn-2,3 stereostructure of the glycerol backbone on which two saturated isoprenoid chains are bound at the sn-2 and sn-3 positions. A lipid-substrate with unsaturated isoprenoid chains or sn-1,2-dialkylglycerol configuration exhibited low activity. Tetraether type caldarchaetidylinositol was also actively glucosylated by the homogenates to form monoglucosyl caldarchaetidylinositol and small amount of diglucosyl caldarchaetidylinositol. The addition of Mg²⁺ increased the formation of diglucosyl caldarchaetidylinositol. This suggested that the same enzyme set synthesized the sole sugar moiety of diether type glycolipids and tetraether type phosphoglycolipids.