Analysis of glycosphingolipids by fluorophore-assisted carbohydrate electrophoresis using ceramide glycanase from Mercenaria mercenaria

Anal Biochem. 1994 Oct;222(1):270-4. doi: 10.1006/abio.1994.1484.

Abstract

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 order to react with the amine group of the fluorophore (8-aminonaphthalene-1,3,6-trisulfonic acid or 2-aminoacridone). We have used exoglycosidase-free ceramide glycanase from hard-shelled clam to expose the reducing terminal of the glycan in glycosphingolipid by cleaving the linkage between the glycan and the ceramide. The released glycan was used for fluorophore-assisted carbohydrate electrophoresis, in order to qualitatively determine its monosaccharide composition, chain length, and glycosidic linkages using linkage-specific glycosidases, including a beta 1-3,4 galactosidase from clam.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Bivalvia / enzymology*
  • Carbohydrate Sequence
  • Cattle
  • Electrophoresis, Polyacrylamide Gel / methods*
  • Fluorescent Dyes
  • Glycoside Hydrolases / chemistry*
  • Glycosphingolipids / analysis*
  • Molecular Sequence Data
  • Rabbits

Substances

  • Fluorescent Dyes
  • Glycosphingolipids
  • Glycoside Hydrolases
  • ceramide glycanase