Isolation and structural characterization of adhesin polysaccharide receptors

The procedure for the purification of the adhesin polysaccharide receptor and its hexasaccharide repeating unit from whole S. oralis ATCC 55229 by chemical, enzymatic, and chromatographic techniques has been described. Chemical, chromatographic, and mass spectrometric procedures allow preliminary structural characterization of the hexasaccharide repeating unit and polysaccharide. The structural characterizations of the hexasaccharide and polysaccharide are completed using several 1D and 2D NMR techniques. Identification of the anomeric 1H and 13C signals of the glycosyl residues permits, by virtue of their chemical shifts and coupling constants (3JHH and 1JCH), the determination of the configurations of the glycosidic linkages. The HMBC connectivities permit the establishment of the hexasaccharide sequence as Rhap alpha(1-->2)Rhap alpha(1-->3)Galp alpha(1-->3)Galp beta(1-->4)Glcp beta(1-->3)Gal. The 1H NMR chemical shifts of the polysaccharide, as determined by the combination of COSY and TOCSY experiments, and the observed interglycosidic NOESY cross-peaks reveal the structure of the polysaccharide to be [formula: see text] where the position of the glycerol (Gro) phosphate moiety has been determined by [1H, 31P] NMR spectroscopy.