Chemoenzymatic synthesis of 2-azidoethyl-ganglio-oligosaccharides GD3, GT3, GM2, GD2, GT2, GM1, and GD1a

We have synthesized several ganglio-oligosaccharide structures using glycosyltransferases from Campylobacter jejuni. The enzymes, α-(2→3/8)-sialyltransferase (Cst-II), β-(1→4)- N-acetylgalactosaminyltransferase (CgtA), and β-(1→3)-galactosyltransferase (CgtB), were produced in large-scale fermentation from Escherichia coli and further characterized based on their acceptor specificities. 2-Azidoethyl-glycosides corresponding to the oligosaccharides of GD3 (α- D-Neu p5Ac-(2→8)-α- D-Neu p5Ac-(2→3)-β- D-Gal p-(1→4)-β- D-Glc p-), GT3 (α- D-Neu p5Ac-(2→8)-α- D-Neu p5Ac-(2→8)-α- D-Neu p5Ac-(2→3)-β- D-Gal p-(1→4)-β- D-Glc p-), GM2 (β- D-Gal pNAc-(1→4)-[α- D-Neu p5Ac-(2→3)]-β- D-Gal p-(1→4)-β- D-Glc p-), GD2 (β- D-Gal pNAc-(1→4)-[α- D-Neu p5Ac-(2→8)-α- D-Neu p5Ac-(2→3)]-β- D-Gal p-(1→4)-β- D-Glc p-), GT2 (β- D-Gal pNAc-(1→4)-[α- D-Neu p5Ac-(2→8)-α- D-Neu p5Ac-(2→8)-α- D-Neu p5Ac-(2→3)]-β- D-Gal p-(1→4)-β- D-Glc p-), and GM1 (β- D-Gal p-(1→3)-β- D-Gal pNAc-(1→4)-[α- D-Neu p5Ac-(2→3)]-β- D-Gal p-(1→4)-β- D-Glc p-) were synthesized in high yields (gram-scale). In addition, a mammalian α-(2→3)-sialyltransferase (ST3Gal I) was used to sialylate GM1 and generate GD1a (α- D-Neu p5Ac-(2→3)-β- D-Gal p-(1→3)-β- D-Gal pNAc-(1→4)-[α- D-Neu p5Ac-(2→3)]-β- D-Gal p-(1→4)-β- D-Glc p-) oligosaccharide. We also cloned and expressed a rat UDP- N-acetylglucosamine-4′epimerase (GalNAcE) in E. coli AD202 cells for cost saving in situ conversion of less expensive UDP-GlcNAc to UDP-GalNAc.