A necessary and sufficient determinant for protein-selective glycosylation in vivo

A limited number of glycoproteins including luteinizing hormone and carbonic anhydrase-VI(CA6) bearN-linked oligosaccharides that are modified with beta 1,4-linked N-acetylgalactosamine ( GalNAc). The selective addition of GalNAc to these glycoproteins requires that the beta 1,4-N-acetylgalactosaminyl-transferase (beta GT) recognize both the oligosaccharide acceptor and a peptide recognition determinant on the substrate glycoprotein. We report here that two recently cloned beta GTs, beta GT3 and beta GT4, that are able to transfer GalNAc to GlcNAc in beta 1,4-linkage display the necessary glycoprotein specificity in vivo. Both beta GTs transfer GalNAc to N-linked oligosaccharides on the luteinizing hormone alpha subunit and CA6 but not to those on transferrin ( Trf). A single peptide recognition determinant encoded in the carboxyl-terminal 19-amino acid sequence of bovine CA6 mediates transfer of GalNAc to each of its two N-linked oligosaccharides. The addition of this 19-amino acid sequence to the carboxyl terminus of Trf confers full acceptor activity onto Trf for both beta GT3 and beta GT4 in vivo. The complete 19-amino acid sequence is required for optimal GalNAc addition in vivo, indicating that the peptide sequence is both necessary and sufficient for recognition by beta GT3 and beta GT4.