Thin-layer Chromatographic Analysis of Asn-linked Oligosaccharides Derived from Glycoproteins by Specific Cleavage with Almond Glycopeptidase

Abstract

Almond glycopeptidase which cleaved specifically the β-aspartylglucosylamine linkages in glycopeptides proved to be useful for the simple and rapid analysis of Asn-linked oligosaccharides derived from glycoproteins. The oligosaccharide fraction in each glycopeptidase digest was desalted by paper chromatography with the solvent composed of 1-butanol/ethanol/water (4: 41) for 2 days, and analyzed by thin-layer chromatography on silica gel 60. The solvent system was 1-propanol/acetic acid/water (3: 3: 2). Only one kind of the high-mannose-type oligosaccharide was present in a Taka-amylase A molecule (Fig.3). The human fibrinogen molecule was found to have the identical four biantennary oligosaccharides of complex-type (Fig.4). Fig.6 shows that several kinds of high-mannose-type oligosaccharides and hybrid-type oligosaccharides were attached to the same position in a peptide sequence of hen ovalbumin. We extended the application of the enzyme to histochemical examination, i. e., the distribution of Asn-linked oligosaccharides in umbilical artery, vein and matrix (Fig.7).