Application of 1D and 2D NMR techniques to the structure elucidation of the O-polysaccharide from Proteus mirabilis O:57.
The LPS O-polysaccharide (O-PS) produced by Proteus mirabilis serotype O:57 (ATCC 49995) was shown by NMR spectroscopy and chemical analysis to be a high-molecular-weight acidic branched polymer of pentasaccharide repeating units, composed of D-glucose, D-galactose, 2-acetamido-2-deoxy-D-galactose and glycerophosphate residues (1:2:2:2:1). Application of one- and two-dimensional NMR methods allowed the complete assignment of notoriously crowded 1H and 13C spectra of the O-PS, leading to the determination of its structure. Several of the NMR techniques used were applied for the first time to the structure elucidation of polysaccharides. The resonances of galactose H5, H6 and H6' were identified by a 1D analog of 3D NOESY-TOCSY and 2D (1H,1H) triple-quantum experiments. The position and the nature of the phosphate group were determined from 2D 31P (omega 1)-half-filtered COSY and 2D 31P-relayed COSY spectra. 2D HMQC-TOCSY and 2D single-quantum proton-carbon long-range correlation techniques were used to overcome the difficulties of severe overlap and higher order effects in the 1H NMR spectrum of the O-PS. The latter technique, together with 2D NOESY, enabled us to identify the substitution positions, the anomeric configurations and the sequence of the component glycose residues in the O-PS.
Uhrín
D
D
Institute for Biological Sciences, National Research Council, Ottawa, ON, Canada.
Brisson
J R
JR
MacLean
L L
LL
Richards
J C
JC
Perry
M B
MB
D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D008070:Lipopolysaccharides / Q000737:chemistry*; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D019081:O Antigens; D011135:Polysaccharides, Bacterial / Q000737:chemistry*; D011513:Proteus mirabilis / Q000737:chemistry*
D008070:Lipopolysaccharides; D019081:O Antigens; D011135:Polysaccharides, Bacterial