Reversal of tyrosinamide-oligosaccharide derivatization by Edman degradation

In a previous report (Tamura, T., Wadhwa, M.S., and Rice, K.G. (1994) Anal. Biochem. 216, 335-344) we described the derivatization of N-linked oligosaccharides with Boc-tyrosine resulting in the formation of tyrosinamide-oligosaccharides. Attachment of Boc-tyrosine to the reducing-end of an oligosaccharide through a glycosylamide linkage provides a hydrophobic chromophore that facilitates the purification of individual N-linked oligosaccharides derived from glycoproteins on preparative reverse-phase HPLC. In the present report we extend the utility of tyrosinamide-oligosaccharides by demonstrating two additional attributes of these glycoconjugates. Edman degradation was used to reverse both sialyl and asialyl tyrosinamide-oligosaccharide derivatives resulting in the formation of reducing oligosaccharides. These can be used as analytical standards for chromatography or can be further derivatized with other probes that react with the reducing-end of an oligosaccharide. Second, we report that the fluorescence of tyrosinamide-oligosaccharides allows their sensitive detection (5 pmol) on HPLC. These two attributes expand the versatility of tyrosinamide-oligosaccharides as glycoconjugates suitable for both analytical and biological studies.