Separation of radiolabelled glycosaminoglycan oligosaccharides by polyacrylamide-gel electrophoresis

Abstract

Glycosaminoglycan oligosaccharides generated by treatment of biosynthetically radiolabeled dermatan sulfate and hyaluronic acid with chondroitin AC lyase or testicular hyaluronidase may be resolved into a series of discrete bands by polyacrylamide-gel electrophoresis. Bands were identified by fixation in glacial acetic acid containing 20% (wt/vol) 2,5-diphenyloxazole followed by fluorography. The bands represented glycans which differed in size by 1 disaccharide unit. For the larger oligosaccharides (decasaccharides and above) of similar charge:mass ratio, there was a linear relationship between electrophoretic mobility and log MW. However, the smaller species showed anomalous migration patterns. Consideration of the structures of the fragments produced by the different enzyme treatments suggests that copolymeric and homopolymeric oligosaccharides may be separated by polyacrylamide-gel electrophoresis. There are many potential applications of this technique, foremost amongst them being studies on the molecular size heterogeneity and patterns of enzyme-mediated depolymerization of native glycosaminoglycan chains and investigations into rates of polymer chain elongation and post-polymerization modification reactions so essential to glycosaminoglycan function.