A pyridylamination method aimed at automatic oligosaccharide analysis of N-linked sugar chains.
The procedure for preparation of pyridylaminated sugar chains from glycoproteins was improved with a view to its eventual automation. Following on the coupling reaction improvement already reported [N. Kuraya and S. Hase (1992) J. Biochem. 112, 122-126], two further aspects were improved in this study. Instead of sodium bicarbonate-acetic anhydride, volatile reagents were adopted for the re-N-acetylation of hexosamine residues after hydrazinolysis to give rapid removal of excess reagents. Subsequent to the pyridylamination reaction, excess reagents were removed by cation-exchange to isolate the pyridylaminated oligosaccharides in place of gel filtration. These alterations rendered a one-pot reaction possible and resulted in a large reduction in the amount of time needed compared with other methods so far reported. The procedure was successfully applied to the detection of sugar chains from Taka-amylase A and human erythrocyte membranes.
Yanagida
K
K
Graduate School of Science, Osaka University, Machikaneyama 1-1, Toyonaka, Osaka, 560-0043, Japan.
Natsuka
S
S
Hase
S
S
D000631:Aminopyridines; D002236:Carbohydrate Conformation; D002851:Chromatography, High Pressure Liquid; D004910:Erythrocyte Membrane / Q000737:chemistry; D006801:Humans; D009439:Neuraminidase / Q000378:metabolism; D009844:Oligosaccharides / Q000032:analysis*; D000516:alpha-Amylases / Q000378:metabolism
D000631:Aminopyridines; D009844:Oligosaccharides; D000516:alpha-Amylases; D009439:Neuraminidase