Structural characterization of a novel mono-sulfated gangliotriaosylceramide containing a 3-O-sulfated N-acetylgalactosamine from rat kidney

Abstract

A novel mono-sulfated glycosphingolipid based on the gangliotriaose core structure was isolated from rat kidney. The isolation procedure involved extraction of lipids with chloroform/methanol, mild alkaline methanolysis, column chromatographies with anion exchangers and silica beads. The structure was characterized by compositional analysis, FTIR spectroscopy, methylation analysis, 1H-NMR spectroscopy and negative-ion liquid secondary ion mass spectrometry (LSMIS) using the intact glycolipid and its desulfation product. The two dimensional chemical shift correlated spectroscopy provided information on the sugar sequence as well as anomeric configurations, and indicated the presence of a 3-O-sulfated N-acetylgalactosamine within the molecule. Negative-ion LSIMS with high- and low-energy collision-induced dissociation defined the sugar sequence and ceramide composition, confirming the presence of a sulfated N-acetylgalactosamine at the non-reducing terminus. From these results, the complete structure was proposed to be HSO3-3GalNAc beta 1-4Gal beta 1-4Glc beta 1-1Cer (Gg3Cer III3-sulfate, SM2b).