Detailed structural analysis of N-glycans released from glycoproteins in SDS-PAGE gel bands using HPLC combined with exoglycosidase array digestions

Methods Mol Biol. 2006:347:125-43. doi: 10.1385/1-59745-167-3:125.

Abstract

In contrast to the linear sequences of protein and DNA, oligosaccharides are branched structures. In addition, almost all glycoproteins consist of a heterogeneous collection of differently glycosylated variants. Glycan analysis therefore requires high-resolution separation techniques that can provide detailed structural analysis, including both monosaccharide sequence and linkage information. This chapter describes how a combination of high-performance liquid chromatography (HPLC) and exoglycosidase enzyme array digestions can deliver quantitative glycan analysis of sugars released from glycoproteins in sodium dodecyl sulfate-polyacrylamide gel electrophoresis gel bands by matching HPLC elution positions with a database of standard glycans.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Carbohydrate Conformation
  • Carbohydrate Sequence
  • Chromatography, High Pressure Liquid / methods*
  • Electrophoresis, Polyacrylamide Gel / methods*
  • Fluorescent Dyes / chemistry
  • Glycoproteins / chemistry
  • Glycoside Hydrolases / chemistry
  • Glycoside Hydrolases / metabolism*
  • Molecular Sequence Data
  • Polysaccharides / analysis*
  • Polysaccharides / chemistry*
  • Polysaccharides / metabolism

Substances

  • Fluorescent Dyes
  • Glycoproteins
  • Polysaccharides
  • Glycoside Hydrolases