Bacterial sialidases cleave terminal sialic acid from a variety of host glycoproteins, and contribute to survival and growth of many human-dwelling bacterial species, including various pathogens. Tannerella forsythia, an oral, Gram-negative, fastidious anaerobe, is a key organism in periodontal disease and possesses a dedicated sialic acid utilisation and scavenging (nan) operon, including NanH sialidase. Here, we describe biochemical characterisation of recombinant NanH, including its action on host-relevant sialoglycans such as sialyl Lewis A and sialyl Lewis X (SLeA/X), and on human cell-attached sialic acids directly, uncovering that it is a highly active broad specificity sialidase. Furthermore, the N-terminal domain of NanH was hypothesised and proved to be capable of binding to a range of sialoglycans and non-sialylated derivatives with Kd in the micromolar range, as determined by steady-state tryptophan fluorescence spectroscopy, but it has no catalytic activity in isolation from the active site. We consider this domain to represent the founding member of a novel subfamily of carbohydrate-binding module (CBM), involved in glycosidase-ligand binding. In addition, we created a catalytically inactive version of the NanH enzyme (FRIP → YMAP) that retained its ability to bind sialic acid-containing ligands and revealed for the first time that binding activity of a CBM is enhanced by association with the catalytic domain. Finally, we investigated the importance of Lewis-type sialoglycans on T. forsythia–host interactions, showing that nanomolar amounts of SLeA/X were capable of reducing invasion of oral epithelial cells by T. forsythia, suggesting that these are key ligands for bacterial–cellular interactions during periodontal disease.

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