PURIFICATION AND CDNA CLONING OF A HUMAN UDP-N-ACETYL-ALPHA-D-GALACTOSAMINE-POLYPEPTIDE N-ACETYLGALACTOSAMINYLTRANSFERASE

A UDP-GalNAc:polypeptide N-acetylgalactosaminyl-transferase (GalNAc-tr ansferase) from human placenta was purified to apparent homogeneity us ing a synthetic acceptor peptide as affinity ligand. The purified GalN Ac-transferase migrated as a single band with an approximate molecular weight of 52,000 by reducing sodium dodecyl sulfate-polyacrylamide ge l electrophoresis. Based on a partial amino acid sequence, the cDNA en coding the transferase was cloned and sequenced from a cDNA library of a human cancer cell line. The cDNA sequence has a 571-amino acid codi ng region in indicating a protein of 64.7 kDa with a type II domain st ructure. The deduced protein sequence showed significant similarity to a recently cloned bovine polypeptide GalNAc-transferase (Homa, F. L., Hollanders, T., Lehman, D. J., Thomsen, D. R., and Elhammer, Angstrom P. (1993) J. Biol. Chem, 268, 12609-12616), A polymerase chain reacti on construct was expressed in insect cells using a baculovirus vector, Northern analysis of eight human tissues differed clearly from that o f the bovine GalNAc-transferase. Polymerase chain reaction cloning and sequencing of the human version of the bovine transferase are present ed, and 98% similarity at the amino acid level was found. The data sug gest that the purified human GalNAc-transferase is a novel member of a family of polypeptide GalNAc-transferases, and a nomenclature GalNAc- T1 and GalNAc-T2 is introduced to distinguish the members.