Chemoenzymatic synthesis of para-nitrophenol (pNP)-tagged α2-8-sialosides and high-throughput substrate specificity studies of α2-8-sialidases

para-Nitrophenol (pNP)-tagged alpha 2-8-linked sialosides containing different sialic acid forms were chemoenzymatically synthesized using an efficient one-pot three-enzyme alpha 2-8-sialylation system. The resulting compounds allowed high-throughput substrate specificity studies of the alpha 2-8-sialidase activity of a recombinant human cytosolic sialidase hNEU2 and various bacterial sialidases. The sialoside substrate profiles obtained can be used to guide the selection of suitable sialidases for sialylglycan analysis and for cell and tissue surface glycan modification. They can also be used to guide sialidase inhibitor design.