Conformational analysis of a dermatan sulfate-derived tetrasaccharide by NMR, molecular modeling, and residual dipolar couplings

The solution conformation behavior of a dermatan-derived tetrasaccharide - ΔHexA-(1→3)-GalNAc4S-β-(1-4)-IdoA-α-(1→3) -red-GalNAc4S (S is a sulfate group) - has been explored by means of NMR spectroscopy, especially by NOE-based conformational analysis. The tetrasaccharide was present as four species, two of which are chemically different in the anomeric orientation of the reducing 2-deoxy-2-acetamido- galactose (red-GalNAc) residue, while the other two are the result of different conformations of the iduronic acid (IdoA) unit. The two α-β- interconverting anomers were present in a 0.6:1 ratio. Ring conformations have been defined by analysis of ³J_HH coupling constants and interresidual NOE contacts. Both 2-deoxy-2-acetamido-galactose (GalNAc) residues were found in the ⁴C₁ chair conformation, the unsaturated uronic acid (Δ-Hex A) adopts a strongly predominant half-chair ¹H₂ conformation, while the IdoA residue exists either in the ¹C₄ chair or in the ²S₀ skewed boat geometries, in a 4:1 ratio. There is a moderate flexibility of Φ and Ψ torsions as suggested by nuclear Overhauser effects (NOEs), molecular modeling (MM), and molecular dynamics (MD) studies. This was further investigated by residual dipolar couplings (RDCs). One-bond C-H RDCs ( ¹D_CH) and long-range H-H (³D_H,H) RDCs were measured for the tetrasaccharide in a phage solution and interpreted in combination with restrained MD simulation. The RDC-derived data substantially confirmed the validity of the conformer distribution resulting from the NOE-derived simulations, but allowed an improved definition of the conformational behavior of the oligosaccharides in solution. In summary, the data show a moderate flexibility of the four tetrasaccharide species at the central glycosidic linkage. Differences in the shapes of species with the IdoA in skew and in chair conformations and in the distribution of the sulfate groups have also been highlighted.