High-resolution 1D- and 2D-correlation H-1 NMR and C-13 NMR, at 500 and 125 MHz, respectively, permitted assignment of the majority of the resonances in the per-N-acetylated, phosphorylated tetrasaccharide-ribitol repeat-unit, and in the complete polymer (n = 5-7) containing between five and seven repeating units attached to the deacylated lipid anchor, for the lipoteichoic acid from Streptococcus pneumoniae strain R6; the P-31 resonances were also assigned. Comparison of the P-31 spectra obtained for the per-N-acetylated oligosaccharide and for the oligosaccharide having the AATG 4-NH2 group still free, indicate a conformational difference brought about by interaction between the amino group and the neighboring phosphate group. Collectively these results prove the chain structure previously elucidated by chemical procedures and FAB-mass spectrometry, namely: -[ --> 6)-beta-Glc-(1 --> 3)-AATG-alpha-(1 --> 4)-[cholineP-6] -alpha-GalNAc-(1 --> 3)-[cholineP-6]-beta-GalNAc-(1 --> 1)-Ribitol-5P]- Proton-proton through-space connectivities were determined by NOESY experiments for both the oligomer (n = 5-7) and the isolated tetrasaccharide-ribitol repeat-unit. Quantitative estimation of the interproton distances from the NOE measurements allowed determination of the most probable 3D-conformations of the glycosidic linkages of the tetrasaccharide-ribitol repeat-unit in the teichoic acid polymer. Computer calculations by CHARMm molecular mechanics and dynamics methods, Monte Carlo simulation, and with experimental NOE-derived distances as constraints, indicate the most likely solution conformation for the tetrasaccharide-ribitol, taking account of charge, hydrogen-bonding, and solvent effects. The torsional mobility of the glycosidic linkages between the hexopyranose units is shown to be rather low, whereas the mobility of the final glycosidic linkage in the repeat-unit between the second choline-substituted galactopyranose unit and the ribitol residue is much higher. Two adjacent polymer chains, lying nest to one another, are capable of forming a relatively stable, pseudo-helical complex. The biological significance of these findings in terms of aggregation phenomena and the possible role of the side-chain phosphorylcholine moieties is discussed.
