Structure of the O-linked oligosaccharides from a major thyroid cell surface glycoprotein

A major glycoprotein at the surface of calf thyroid cells, GP-3 (M(r) 20,000), contains the I-antigenic activity of calf thyroid which has been attributed to its poly-N-acetyllactosamine N-linked saccharide chains (Edge, A. S. B., and Spiro, R. G., J. Biol. Chem. 260, 15332-15338, 1985). The present study demonstrated that alkaline borohydride treatment of GP-3 results in the release of five neutral and five acidic saccharides that were found to represent over 30% of the saccharides of this carbohydrate-rich glycoprotein. Three of the oligosaccharides contained terminal alpha1 --> 3-linked galactose residues which accounted for their affinity toward Bandeiraea simplicifolia I lectin. The saccharides could be grouped into several distinct categories on the basis of their internal sequence. A novel tetrasaccharide in GP-3 was shown to have the structure: Gal alpha1 --> 3Gal beta1 --> 6(Gal beta1 --> 3)GalNAcH2. An unsubstituted N-acetylgalactosamine unit and a Gal beta1 --> 3GalNAc disaccharide were prominent O-linked constituents, with the disaccharide serving as a core unit for the attachment of sialic acid residues to form tetra- and trisaccharides. A branched core structure, Gal beta1 --> 3(GlcNAcbeta1 --> 6)GalNAcH2 was shared by 4 of the 10 saccharides, the most complete of which was assigned the sequence NeuAc alpha2 --> 3Gal beta1 --> 3(Gal alpha1 --> 3Gal beta1 --> 4GlcNAc beta1 --> 6)GalNAcH2.