A novel saccharide was synthesized by incubating globe-N-tetraose, GalNAc beta 1-3Gal alpha 1-4Gal beta 1-4Glc, and UDP-[H-3]GlcNAc with hog gastric mucosal microsomes, known to contain beta 1,6-N-acetylglucosaminyltransferase activity of a broad acceptor specificity, Chromatography and MALDI-TOF mass spectrometry of the product, as well as the amount of incorporated radioactivity indicated that one [3H]GlcNAc residue was transferred to the acceptor saccharide, One- and two-dimensional H-1 NMR-spectroscopic analysis of the product and ESI-CID mass spectrometry of the pentasaccharide in permethylated form established its structure as GalNAc beta 1-3([H-3]GlcNAc beta 1-6)Gal alpha 1-4Gal beta 1-4Glc. The new enzyme activity possesses substrate specificity features common to a purified beta 1,6-GlcNac-transferase from bovine tracheal epithelium, which forms branches at the subterminal beta 1,3-substituted galactose and accepts both GlcNAc- and Gal-configuration at the terminal residue of the acceptor (Ropp ef al, (1991) J. Biol, Chem,, 266, 23863-23871), The new beta 1,6-GlcNac-branch was readily galactosylated by bovine milk beta 1,4-galactosyltransferase, revealing a pathway to novel hybrid type glycans with N-acetyllactosamine chains on globotype saccharides, This pathway may lead to the rare IP blood-group antigen and to globoside-like molecules mediating cell adhesion.
