a1>3-GALACTOSYLTRANSFERASE : THE USE OF RECOMBINANT ENZYME FOR THE SYNTHESIS OF a-GALACTOSYLATED GLYCOCONJUGATES

Abstract

We have reported the isolation and characterization of a bovine cDNA clone containing the complete coding sequence for UDPGal: Galß1>4G1cNAc «1>3-galactosyltransferase (Joziasse, D.H. et al., (1989) J. Biol. Chem. 264, 14290-14297). Insertion of this cDNA clone into the genome of Autographa californica nuclear polyhedrosis virus (AcNPV) and subsequent infection of Sf9 insect cells with recombinant virus, resulted in high-level expression of enzymatically active al>3-galactosyltransferase. The recombinant &1>3-galactosyltransferase could be readily detergent solubilized and subsequently purified by affinity-chromatography on UDP-hexanolamine-Sepharose. The recombinant «1>3-galactosyltransferase showed the expected preference for the acceptor substrate N-acetyllactosamine (GalB1>4G1cNAc), and demonstrated enzyme kinetics identical to those previously reported for affinity-purified calf thymus a1 >3- galactosyltransferase.