Oligosaccharides display both rigidity and high flexibility in water as determined by 13C NMR relaxation and 1H,1H NOE spectroscopy: evidence of anti-phi and anti-psi torsions in the same glycosidic linkage.

The trisaccharide beta-D-Glcp-(1-->2)-beta-D-Glcp-(1-->3)-alpha-D-Glcp-OMe has been investigated by molecular dynamics (MD) simulations and NMR experiments in water. 13C spin-lattice (T1) and spin-spin (T2) relaxation times, together with 1H,13C NOE data were measured at two magnetic field strengths (9.4 and 14.1 T) in a 277 K D2O solution. Relaxation data interpreted by means of the model-free formalism revealed a rigid (S2 approximately 0.9) oligosaccharide tumbling in solution. 1H,1H Cross-relaxation rates were determined at 600 MHz by 1D DPFGSE NOESY and T-ROESY experiments, which provided high quality data and subsequently proton-proton distances within the trisaccharide. The presence of anti conformers at both torsions of a glycosidic linkage is demonstrated for the first time. MD simulations were carried out to facilitate analysis of the NOE data. In total, 15 simulations-starting from five different conformational states--were performed, with production runs of up to 10 ns, resulting in 83 ns of oligosaccharide dynamics in water. anti Conformers were populated to different degrees in the simulations, especially at the phi2 torsion angle. By combining the results from the NOE experiments and the MD simulations, the anti conformers at the (1-->2)-linkage were quantified as 7% anti-phi2 and 2% anti-psi2, revealing a highly flexible trisaccharide in which large conformational changes occur. From the MD simulations, interresidue hydrogen bonding, from HO2" to O2 or O3, was significantly populated (approximately 40%) in both of the anti conformational states. The contentious issue over rigidity versus flexibility in oligosaccharides has thus been thoroughly examined, showing that the dynamics should be taken into account for a relevant description of the molecular system.

Structural analysis of the O-antigen polysaccharide from the Shiga toxin-producing Escherichia coli O172.

The structure of the O-antigen polysaccharide from Escherichia coli O172 has been determined. In combination with sugar analysis, NMR spectroscopy shows that the polysaccharide is composed of pentasaccharide repeating units. Sequential information was obtained by mass spectrometry and two-dimensional NMR techniques. An O-acetyl group was present as 0.7 equivalent per repeating unit. Treatment of the O-deacetylated polysaccharide with aqueous 48% hydrofluoric acid rendered cleavage of the phosphodiester in the backbone of the polymer and the pentasaccharide isolated after gel permeation chromatography was structurally characterized. Subsequent NMR experiments on polymeric materials revealed the structure of the repeating unit of the O-polysaccharide from E. coli O172 as:-->P-4)-alpha-D-Glcp-(1-->3)-alpha-L-FucpNAc-(1-->3)-alpha-D- GlcpNAc-(1-->3)-alpha-L-FucpNAc-(1-->4)-alpha-D-Glcp6Ac-(1-->

Structural analysis of the O-antigenic polysaccharide from Vibrio mimicus N-1990.

The O-antigenic polysaccharide part of the lipopolysaccharide from Vibrio mimicus N-1990 has been investigated. Sugar and methylation analysis of native and dephosphorylated polysaccharide together with NMR spectroscopy show that the polysaccharide is composed of tetrasaccharide repeating units. The structure of the repeating unit of the polysaccharide from V. mimicus N-1990 could be determined as: -->4)-alpha-D-GalpNAc-(1-->3)-beta-D-GalpNAc-(1-->2)-4,6-P-beta-D-Galp-( 1-->3)-alpha-D-GalpNAc-(1-->. The V. mimicus N-1990 strain cross-reacts with antibodies elicited against the Vibrio cholerae O139. The nature of this cross-reactivity resides in the partial structure comprising the galactosyl residue substituted with a cyclic phosphate. This element is present in the cell-wall-associated polysaccharides of both strains.

Structural analysis of the O-antigenic polysaccharide from the enteropathogenic Escherichia coli O142.

The polysaccharide part of the lipopolysaccharide obtained from the enteropathogenic Escherichia coli O142 has been isolated, and its structure determined. Together with 1H-NMR and 13C-NMR spectroscopy, sugar and methylation analyses show that the polysaccharide is composed of repeating pentasaccharide units. Sequential information on the O-polysaccharide was obtained by two-dimensional NMR techniques, namely heteronuclear-multiple-bond-connectivity and NOESY experiments. The repeating unit of the O-polysaccharide of E. coli strain O142 has the following structure: [structure: see text].