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

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-Neup5Ac-(2→8)-α-D-Neup5Ac-(2→3)-β-D-Galp-(1→4)-β-D-Glcp-), GT3 (α-D-Neup5Ac-(2→8)-α-D-Neup5Ac-(2→8)-α-D-Neup5Ac-(2→3)-β-D-Galp-(1→4)-β-D-Glcp-), GM2 (β-D-GalpNAc-(1→4)-[α-D-Neup5Ac-(2→3)]-β-D-Galp-(1→4)-β-D-Glcp-), GD2 (β-D-GalpNAc-(1→4)-[α-D-Neup5Ac-(2→8)-α-D-Neup5Ac-(2→3)]-β-D-Galp-(1→4)-β-D-Glcp-), GT2 (β-D-GalpNAc-(1→4)-[α-D-Neup5Ac-(2→8)-α-D-Neup5Ac-(2→8)-α-D-Neup5Ac-(2→3)]-β-D-Galp-(1→4)-β-D-Glcp-), and GM1 (β-D-Galp-(1→3)-β-D-GalpNAc-(1→4)-[α-D-Neup5Ac-(2→3)]-β-D-Galp-(1→4)-β-D-Glcp-) 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-Neup5Ac-(2→3)-β-D-Galp-(1→3)-β-D-GalpNAc-(1→4)-[α-D-Neup5Ac-(2→3)]-β-D-Galp-(1→4)-β-D-Glcp-) 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.