Glycosyl transferases of baby hamster kidney cells and ricin‐resistant mutants

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Abstract
Extracts of baby hamster kidney (BHK) cells catalyzed the incorporation of N‐acetylgalactosamine from UDP‐N‐acetyl[14C]galactosamine into myelin basic protein and an acylated tetrapeptide, N‐acetylthreonyl‐triproline, based on the threonine residue 98, glycosylated in myelin basic protein. The incorporated N‐acetylgalactosamine residues were shown to be in α linkage to the peptide moieties. Several ricin‐resistant BHK cell lines contained enhanced (approximately twofold) levels of the transferase activity. Apomucins obtained from bovine submaxillary gland mucin by chemical or enzymic degradation were relatively poor acceptors. Using asialomucin as acceptor, galactosyl transferase activities and a weak sialyl transferase activity were detected in BHK cell extracts. Galactose transfer occurred at two sites: to peptide‐linked N‐acetylgalactosamine residues to form the linkage, galactosyl‐(β1→3)‐N‐acetylgalactosamine and to terminally linked N‐acetylglucosamine residues that exist as a minor constituent in bovine submaxillary mucin O‐glycans, to form a galactosyl N‐acetylglucosamine linkage. This reaction was not inhibited by ovalbumin, an efficient acceptor of the β1→4 galactosyl transferase involved in N‐glycan assembly. Incorporation of galactose and N‐acetylgalactosamine into endogenous proteins of BHK cell extracts was also detected. Sialic acid, fucose and N‐acetylglucosamine residues were not incorporated. The incorporated N‐acetylgalactosamine residues were shown to be in α linkage to polypeptide, and galactose incorporation represented synthesis of the galactosyl‐(β1→3)‐N‐acetylgalactosamine sequence linked to polypeptide. The major endogenous protein labelled by either sugar had a molecular weight of approximately 80000. A BHK‐cell‐associated glycoprotein, analogous to the urinary Tamm‐Horsfall glycoprotein of molecular weight similar to the major endogenous acceptor of glycosylation, was not glycosylated in the experiments in vitro.