Open AccessStructure of the O‐linked carbohydrate chains of porcine zona pellucida glycoproteins
Author(s) -
HOKKE Cornelis H.,
DAMM Jan B. L.,
PENNINKHOF Bea,
AITKEN R. John,
KAMERLING Johannis P.,
VLIEGENTHART Johannes F. G.
Publication year - 1994
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1994.tb18762.x
Subject(s) - chemistry , glycoprotein , gel permeation chromatography , oligosaccharide , exoglycosidase , ion chromatography , glycan , carbohydrate conformation , asparagine , chromatography , beta (programming language) , protein primary structure , peptide , nuclear magnetic resonance spectroscopy , stereochemistry , biochemistry , peptide sequence , amino acid , organic chemistry , polymer , computer science , gene , programming language
The N‐linked carbohydrate chains of procine zona pellucida glycoproteins were released by digestion with peptide‐ N 4 ‐( N ‐acetyl‐β‐glucosaminlyl)asparagine amidase F and subsequently separated from the O‐glycoprotein by gel‐permeation chromatography on Bio‐Gel P‐100. The O‐linked carbohydrate chains were relesed from the O‐glycoprotein by alakaline borohydride treatment. Fractitionation of the extremely heterogeneous mixture of O‐linked oligosaccharide alditols was achieved by a combination of chromatographic techniques comprising gel‐permeation chromatography on Bio‐Gel P‐4 and P‐6, anion‐exchange FPLC on Mono Q, and high‐pH anion‐exchange chromatography on CarboPac PA‐1. The primary structures of 32 O‐glycans were determined by one‐ and two‐dimensional 1 H‐NMR spectroscopy. The major part of the analyzed compounds contain a combination of the structural elements Galβ1‐4GlcNAcβ1‐3Galβ1‐3GalNAc‐ol, Galβ1‐4(6SO 4‐ )GlcNAc, and α2‐3‐linked Neu5Gc or Neu5Ac. This series of compounds has the following structure, where n = 0 to > 6 : [Neu5Gc/Acα2‐3] 0‐1 [Galβ1‐4(6SO 4‐ )GlcNAcβ1‐3] n Galβ1‐4GlcNAcβ1‐3Galβ1‐3GalNAc‐ol. In addition, smaller compounds were identified in which the Galβ1‐3GalNAc‐ol core is substituted by Neu5Gc/Ac α2‐6‐linked to GalNAc‐ol and/or Neu5Gc/Ac α2‐3‐linked to Gal. Furthermore, oligosaccharides were obtained in which the distribution of 6‐ O ‐sulfated GlcNAc residues differs from that in the above‐mentioned general structure, and a small portion of the oligosaccharides has the GlcNAcβ1‐3GalNAc‐ol core structure. Analysis of the endo‐β‐galactosidase digests of pools of N‐ and O‐glycans indicated that the two types of oligosaccharides contain qualitatively similar poly( N ‐acetyllactosamine) chains. In the case of the N‐linked carbohydrate chains, multiple branching of the core structures occurs, resulting in an even larger heterogeneity than observed for the O‐linked carbohydrate chains.