Dynamic exchange between stabilized conformations predicted for hyaluronan tetrasaccharides: comparison of molecular dynamics simulations with available NMR data

Studies of the hyaluronan (HA) tetrasaccharides are important forunderstanding hydrogen-bonding in the HA polymer, as they are probably thesmallest oligomers in which characteristics of the constituentmonosaccharides and the polymer are simultaneously exhibited. Here wepresent extensive molecular dynamics simulations of the twotetrasaccharides of HA in dilute aqueous solution. These simulations haveconfirmed the existence of intramolecular hydrogen-bonds between theneighboring sugar residues of HA in solution, as proposed by Scott (1989).However, our simulations predict that these intramolecular hydrogen-bondsare not static as previously proposed, but are in constant dynamic exchangeon the sub-nanosecond time-scale. This process results in discrete internalmotion of the HA tetrasaccharides where they rapidly move between lowenergy conformations. Specific interactions between water andintramolecular hydrogen-bonds involving the hydroxymethyl group were foundto result in differing conformations and dynamics for the two alternativetetrasaccharides of HA. This new observation suggests that this residue mayplay a key role in the entropy and stability of HA in solution, allowing itto stay soluble up to high concentration. The vicinal coupling constants3 JNHCH of the acetamido groups have been calculated from our aqueoussimulations of HA. We found that high values of 3J NHCH approximately 8 Hz,as experimentally measured for HA, are consistent with mixtures of bothtrans and cis conformations, and thus3 J NHCH cannot be used to imply apurely trans conformation of the acetamido. The rapid exchange ofintramolecular hydrogen-bonds indicates that although the structure is atany moment stabilized by these hydrogen-bonds, no one hydrogen-bond existsfor an extended period of time. This could explain why NMR often fails toprovide evidence for intramolecular hydrogen-bonds in HA and other aqueouscarbohydrate structures.