ANALYSIS OF GLYCOSPHINGOLIPIDS BY FLUOROPHORE-ASSISTED CARBOHYDRATE ELECTROPHORESIS USING CERAMIDE GLYCANASE FROM MERCENARIA-MERCENARIA

Polyacrylamide gel electrophoresis of fluorophore-labeled saccharides is a simple and sensitive separation method for glycan analysis. This method requires an aldehydic reducing carbon on the saccharide in orde r to react with the amine group of the fluorophore (8-aminonaphthalene - 1,3,6-trisulfonic acid or 2-aminoacridone). We have used exoglycosid ase-free ceramide glycanase from hard-shelled clam to expose the reduc ing terminal of the glycan in glycosphingolipid by cleaving the linkag e between the glycan and the ceramide. The released glycan was used fo r fluorophore-assisted carbohydrate electrophoresis, in order to quali tatively determine its monosaccharide composition, chain length, and g lycosidic linkages using linkage-specific glycosidases, including a be ta 1-3,4 galactosidase from clam. (C) 1994 Academic Press, Inc.