ENZYMATIC-SYNTHESIS OF N-LINKED OLIGOSACCHARIDES TERMINATING IN MULTIPLE SIALYL-LEWIS(X) AND GALNAC-LEWIS(X) DETERMINANTS - CLUSTERED GLYCOSIDES FOR STUDYING SELECTIN INTERACTIONS

Galactosyltransferase, sialyltransferase, and fucosyltransferase were used to create a panel of complex oligosaccharides that possess multip le terminal sialyl-Le(x) (NeuAc alpha 2-3Gal[Fuc alpha 1-3]beta 1-4Glc NAc) and GalNAc-Le(x) (GalNAc[Fuc alpha 1-3]beta 1-4GlcNAc). The enzym atic synthesis of tyrosinamide biantennary, triantennary, and tetraant ennary N-linked oligosaccharides bearing multiple terminal sialyl-Le(x ) was accomplished on the 0.5 mu mol scale and the purified products w ere characterized by electrospray MS and H-1 NMR. Likewise, biantennar y and triantennary tyrosinamide oligosaccharides bearing multiple term inal GalNAc-Le(x) determinants were synthesized and similarly characte rized. The transfer kinetics of human milk alpha 3/4-fucosyltransferas e were compared for biantennary oligosaccharide acceptor substrates po ssessing Gal beta 1-4GlcNAc, GalNAc beta 1-4GlcNAc, and NeuAc alpha 2- 3Gal beta 1-4GlcNAc which established NeuAc alpha 2-3Gal beta 1-4GlcNA c as the most efficient acceptor substrate. The resulting complex olig osaccharides were chemically tethered through the tyrosinamide aglycon e to the surface of liposomes containing phosphatidylthioethanol, resu lting in the generation of glycoliposomes probe which will be useful t o study relationships between binding affinity and the micro- and macr o-clustering of selectin ligand. (C) 1998 Elsevier Science Ltd. All ri ghts reserved.