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Updated: Aug 15, 2026

Identification and Characterization of Protein Glycosylation using Specific Endo- and Exoglycosidases
Published on: December 26, 2011
Reducing-end modification of N-linked oligosaccharides with tyrosine
T Tamura1, M S Wadhwa, K G Rice
1Division of Pharmaceutics and Pharmaceutical Chemistry, College of Pharmacy, Ohio State University, Columbus 43210.
Abstract:
The N-linked oligosaccharides from bovine fetuin were purified using newly developed preparative purification methodology. N-linked oligosaccharides were released from tryptic glycopeptides utilizing N-glycosidase F on the 5-g scale. Selective desialylation with neuraminidase from Clostridium perfringens resulted in the formation of a mono-sialyl-oligosaccharide and asialo-oligosaccharides. The reducing ends of the oligosaccharides were converted to the glycosylamine and reacted with the N-hydroxysuccinimide ester of Boctyrosine. The tyrosinated oligosaccharides were resolved into individual peaks on RP-HPLC and then characterized by proton NMR and FAB-MS. A single asialo-triantennary, an asialo-biantennary, and a mono-sialyl-triantennary oligosaccharide were recovered in good yield. Each product contained a single Boc-Tyr residue attached to the reducing-end GlcNAc residue through a beta-glycosylamide linkage. The procedure was utilized to isolate multi-micromole quantities of oligosaccharides from gram quantities of glycoprotein, thus providing a new route to purify large quantities of N-linked oligosaccharides which contain a terminal tyrosine residue. The tyrosinated oligosaccharides are valuable glycoconjugate ligands which contain a chromophore that absorbs at 280 nm and has sufficient hydrophobicity to facilitate RP-HPLC separations. Furthermore, this group can be deprotected by removal of Boc to reveal a primary amine suitable for further derivatization and can also be radioiodinated for tracking during biological experiments.
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