Conformational Dynamics and Exchange Kinetics of <i>N</i>-Formyl and <i>N</i>-Acetyl Groups Substituting 3-Amino-3,6-dideoxy-α-<scp>d</scp>-galactopyranose, a Sugar Found in Bacterial O-Antigen Polysaccharides

Abstract

Three dimensional shape and conformation of carbohydrates are important factors in molecular recognition events and the N -acetyl group of a monosaccharide residue can function as a conformational gatekeeper whereby it influences the overall shape of the oligosaccharide. NMR spectroscopy and quantum mechanics (QM) calculations are used herein to investigate both the conformational preferences and the dynamic behavior of N -acetyl and N -formyl substituents of 3-amino-3,6-dideoxy-α- d -galactopyranose, a sugar and substitution pattern found in bacterial O-antigen polysaccharides. QM calculations suggest that the amide oxygen can be involved in hydrogen bonding with the axial OH4 group primarily but also with the equatorial OH2 group. However, an NMR J coupling analysis indicates that the θ 1 torsion angle, adjacent to the sugar ring, prefers an ap conformation where conformations <180° also are accessible, but does not allow for intramolecular hydrogen bonding. In the formyl-substituted compound 4 J HH coupling constants to the exo -cyclic group were detected and analyzed. A van’t Hoff analysis revealed that the trans conformation at the amide bond is favored by Δ G ° ≈ – 0.8 kcal·mol –1 in the formyl-containing compound and with Δ G ° ≈ – 2.5 kcal·mol –1 when the N -acetyl group is the substituent. In both cases the enthalpic term dominates to the free energy, irrespective of water or DMSO as solvent, with only a small contribution from the entropic term. The cis – trans isomerization of the θ 2 torsion angle, centered at the amide bond, was also investigated by employing 1 H NMR line shape analysis and 13 C NMR saturation transfer experiments. The extracted transition rate constants were utilized to calculate transition energy barriers that were found to be about 20 kcal·mol –1 in both DMSO- d 6 and D 2 O. Enthalpy had a higher contribution to the energy barriers in DMSO- d 6 compared to in D 2 O, where entropy compensated for the loss of enthalpy.