Structural determination of the polar glycoglycerolipids from thermophilic bacteria Meiothermus taiwanensis

Abstract

The polar glycolipids were isolated from the thermophilic bacteria Meiothermus taiwanensis ATCC BAA-400 by ethanol extraction and purified by Sephadex LH-20 and silica gel column chromatography. The fatty acid composition of O-acyl groups in the glycolipids was obtained by gas chromatography mass spectroscopy analysis on their methyl esters derived from methanolysis and was made mainly of C 15:0 (34.0%) and C 17:0 (42.3%) fatty acids, with the majority as branched fatty acids (over 80%). Removal of O-acyl groups under mild basic conditions provided two glycolipids, which differ only in N-acyl substitution on a hexosamine. Electrospray mass spectroscopy analysis revealed that one has a C 17:0 N-acyl group and the other hydroxy C 17:0 in a ratio of about 1 : 3.5. Furthermore, complete de-lipidation with strong base followed by selective N-acetylation resulted in a homogeneous tetraglycosyl glycerol. The linkages and configurations of the carbohydrate moiety were then elucidated by MS and various NMR analyses. Thus, the major glycolipid from M. taiwanensis ATCC BAA-400 was determined with the following structure:where N-acyl is C 17:0 or hydroxy C 17:0 fatty acid and the glycerol esters were mainly iso-and anteisobranched C 15:0 and C 17:0 .Keywords: glycolipid; Meiothermus taiwanensis; MS; NMR; thermophilic bacteria.The thermophilic bacteria such as Aquifex pyrophilus, Thermodesulfotobacterium commune, Thermus scotoductus, Thermomicrobium roseum and Thermodesulfatator indicus contain unique polar lipids as major membrane components [1][2][3][4][5][6][7][8]. Those lipids are essential for the thermal stability and biological functions of the bacteria in extreme environments [9][10][11]. The polar lipids found in Thermus aquaticus, Thermus filiformis, Thermus scotoductus, and Thermus oshimai were mostly phospholipids and glycolipids [12], and the glycolipids from Thermus species examined thus far usually contain three hexoses, one N-hexosamine, and one glycerol [7,10,[12][13][14][15]. Although the sequences of those carbohydrate moieties have been studied by chemical and mass spectroscopic analysis, no complete structure is available as yet due to the lack of information on the linkages and configurations of the carbohydrate moiety. We have been working on a newly discovered species of thermophilic bacteria, Meiothermus taiwanensis, recently isolated from the Wu-rai hot spring in Taiwan [16], as part of our program to investigate the immunomodulation activity of the glycolipids and the structure-activity relationship. In this study, we determined the structure of a major glycolipid from the thermophilic bacteria M. taiwanensis ATCC BAA-400. The fatty acids were examined by gas chromatography mass spectroscopy (GC-MS) analysis on their methyl esters derived from methanolysis, whereas, the structure of the carbohydrate moiety was elucidated by MS/MS and NMR spectroscopic analyses. To the best of our knowledge this is the first complete glycolipid structure from thermophilic bacteria. Materials and methodsIsolation and...

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