Recombinant (2→3)-α-sialyltransferase immobilized on nickel-Agarose for preparative synthesis of sialyl Lewisx and Lewisa precursor oligosaccharides Original Research Article

The specificity of recombinant (2→3)-α-sialyltransferase (ST3Gal-III), expressed in baculovirus-infected insect cells, has been determined with various oligosaccharide acceptors and sugar-nucleotide donors using a fluorescence based assay. Recombinant ST3Gal-III tagged with a polyhistidine tail was immobilized on Ni2+-NTA-Agarose as an active enzyme for use in the synthesis of three sialylated oligosaccharides: (i) the divalent molecule [α-Neu5Ac-(2→3)-d-Galp-(1→4)-β-d-GlcpNAc-O-CH2]2-C-(CH2OBn)2 (12); (ii) the dansylated derivative, α-Neu5Ac-(2→3)-d-Galp-(1→3)-β-d-GlcpNAc-O-(CH2)6-NH-dansyl and; (iii) the tetrasacharide α-Neu5Ac-(2→3)-β-d-Galp-(1→4)-β-d-GlcpNAc-(1→2)-α-d-Manp-O-CH3. Compound 12 was itself prepared from the divalent N-acetyllactosamine molecule built on pentaerythritol by a chemo-enzymatic route.