ANALYSIS OF GLYCAN STRUCTURES OF BARLEY (1-3,1-4)-BETA-D-GLUCAN 4-GLUCANOHYDROLASE ISOENZYME EII

Barley (1-3,1-4)-beta-glucanase (EC 3.2.1.73) isoenzyme EII has only one site for potential N-glycosylation. The carbohydrate moieties were released as oligosaccharides by hydrazinolysis followed by N-acetylation and reductive labelling with tritiated sodium borohydride. The tritium labelled oligosaccharide mixture was fractionated by Bio-Gel P-4 chromatography and high voltage electrophoresis of borate complexes. Partial exoglycosidase sequencing and co-electrophoresis with plant N-glycans of known structure made it possible to assign tentative structures to the five major oligosaccharide fractions obtained, representing about 85% of the labelled glycans. More than 50% of the N-glycan structures present are substituted with alpha-fucose, presumably in 3-linkage to the reducing terminal N-acetylglucosamine which is characteristic of plant modified-type N-glycans. Substitution of fucose in this position confers on the glycoprotein resistance to the enzyme most often used for deglycosylation, glycopeptidase F.