In this manuscript we address an important gap in our current carbohydrate active enzyme toolbox, by developing a highly a2,6-selective (over a2,3-selective) de facto sialidase that is necessary both for glycan analysis and glycoconjugate remodeling. Both glycosidic linkages are commonly found in animal biology and each has been shown to have distinct biological function.
Our approach is novel in that it harnesses the high selectivity of known glycosyltransferases ‘in reverse’ for effective hydrolysis, converting transferases to hydrolases by reaction engineering.
More specifically, we demonstrate that the a2,6-specific pseudosialidase activity of Photobacterium sp. JT-ISH-224 a2,6-sialyltransferase can be used effectively for highly a2,6 selective hydrolysis on a broad range of analytes: small synthetic probes, isolated complex glycans and complex mixtures of glycoproteins.