期刊
GLYCOBIOLOGY
卷 19, 期 7, 页码 797-807出版社
OXFORD UNIV PRESS INC
DOI: 10.1093/glycob/cwp051
关键词
endogalactosylceramidase; fluorescence derivatization; transglycosylation reaction; 6-gala series glycosphingolipids
资金
- New Energy and Industrial Technology Development Organization
- Ministry of Education, Culture, Sports, Science and Technology of the Japanese Government
- Japan Society for the Promotion of Science for Young Scientists
Although 6-gala series glycosphingolipids possessing R-Gal (alpha/beta) 1-6Gal beta 1-1'Cer have been found in some mollusks, pathogenic parasites, and fungi, their physiological functions and metabolic pathway are not fully understood. We described a novel method of detecting 6-gala series glyco- sphingolipids utilizing the specificity of endogalactosylceramidase (EGALC), which is capable of hydrolyzing 6-gala series glycosphingolipids to produce intact oligosaccharides and ceramides. EGALC catalyzes not only hydrolysis but also a transglycosylation reaction. In the latter reaction, EGALC transfers oligosaccharides from the glycosphingolipids to acceptors such as fluorescent 1-alkanols. Based on the transglycosylation reaction of EGALC, a specific, easy, fast, sensitive, and reproducible method of detecting 6-gala series glycosphingolipids was developed using NBD-pentanol as an acceptor. The fluorescent products, NBD-pentanol-conjugated 6-gala oligosaccharides, were separated and detected by TLC or HPLC with a fluorescent detector. Moreover, it was revealed that as well as glycosphingolipids, a glycoglycerolipid, digalactosyldiacylglycerol, was utilized by EGALC as a donor substrate. This method was successfully applied to detect 6-gala series glycosphingolipids in a fungus, Rhizopus oryzae, and a parasite, Taenia crassiceps. The method would be useful for studying glycosphingolipids and galactosyl glycerolipids which share the Gal (alpha/beta) 1-6Gal structure.
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