Bovine pituitary membrane glycoproteins contain beta-N-acetylgalactosaminylated N-linked sugar chains

Bovine pituitary glycoprotein hormones contain unique N-linked sugar chains with GalNAc beta 1-->4GlcNAc 4GlcNAc structure in their outer chain moieties. In the present study, whether bovine pituitary membrane glycoproteins contain the sugar chains with the disaccharide structure was investigated. Western blot analysis of the membrane glycoproteins using Wistaria floribunda agglutinin (WFA), which binds oligosaccharides terminating with beta-N-acetylgalactosamine residue(s), showed that most protein bands detected with Coomassie Brilliant Blue staining bind to WFA. However, no WFA binding was observed for the bands after treatment of the blotted filter with jack bean beta-N-acetylhexosaminidase or N-Glycanase. The WFA-positive bands were also detected in membrane glycoprotein samples from bovine cerebrum, cerebellum, and medulla oblongata, although their expression levels were low. Structural analysis of the oligosaccharides released by hydrazinolysis from the pituitary membrane glycoproteins by serial lectin column chromatography and sequential exoglycosidase digestion revealed that the major oligosaccharides, which bound to a WFA-agarose column, are of biantennary complex type with one and two GalNAc beta 1-->4GlcNAc groups in their outer chain moieties. These results indicate that the beta-N-acetylgalactosaminylation is not unique to the glycoprotein hormones but occurs to most bovine pituitary glycoproteins.