LacdiNAc (GalNAcβ1–4GlcNAc) is a major motif in N-glycan structures of the chicken eggshell protein ovocleidin-116

Abstract

The avian eggshell matrix protein ovocleidin-116 (OC-116) contains two N-glycosylation sites in its sequence. One of them, 293N-D-S, is modified only marginally while the second one, 62N-Q-T, is completely occupied by N-linked glycans. The glycopeptide bearing the modified site was isolated by size exclusion chromatography and reversed phase HPLC after cleavage of the protein with lysyl endopeptidase. The carbohydrate structures attached to Asn62 were determined by carbohydrate compositional analysis, methylation analysis and electrospray MS/MS. We identified 17 different oligosaccharide structures. Four of them were of the high-mannose type, eight were hybrid type and five were complex type structures. Both, hybrid and complex type glycans comprised core-fucosylated and peripherally fucosylated structures. Most of the antennae contained the relatively rare lacdiNAc (GalNAcbeta1-4GlcNAc) motif, which was fucosylated in 9 out of 15 structures. The lacNAc (Galbeta1-4GlcNAc) motif, which is the more frequent motif in mammals, only occurred in 3 of the 17 glycoforms. This is the first detailed study of N-glycan structures occurring in an avian shell-specific protein and, to our knowledge, the first description of fucosylated lacdiNAc structures present in avian glycoproteins.