A precise structural analysis of a fertilization-associated carbohydrate-rich glycopeptide isolated from the fertilized eggs of euryhaline killi fish (Fundulus heteroclitus). Novel penta-antennary N-glycan chains with a bisecting N-acetylglucosaminyl residue

A novel carbohydrate-rich sialoglycopeptide of apparent molecular mass $$$6 kDa was isolated from the fertilized eggs of Fundulus heteroclitus (euryhaline killi fish). Thisglycopeptide is a member of the L-hyosophorin family, characterized by its high content of carbohydrate (8090% by weight) and formed by depolymerization of the precursor glycopolyprotein (H-hyosophorin) upon fertilization. The structures of the N-glycan chains were unambiguously established by a combination of compositional analysis, methylation analysis, selective chemical degradation (periodate oxidation-Smith degradation and hydrazinolysis-nitrous acid deamination), enzymatic (peptide:N-g]ycosidase F, several β-galactosidases, (β-hexosaminidase and α-galactosidase) digestions and instrumental analyses (¹H-NMR and fast atom bombardment mass spectrometry) tohave the novel and unique carbohydrate sequences, Galα1→3(Galβ1→4)Ga1β1࢐4GlcNAcβ1→ and Galα1→3(±GalNAcβ1→4GlcNAcβ1→3Galβ1→4)Gal-β1→4GlcNAcβ1→. This study representsthe first detailed investigation of the nature of bulky complex asparagine-linked penta-antennary glycans with a bisecting GlcNAc residue in glycoproteins. Expression of such bulky multiantennaryglycanunits on proteins may be essential during early embryogenesis.