SOLUTION STRUCTURE OF A LEWIS(X) ANALOG BY OFF-RESONANCE H-1-NMR SPECTROSCOPY WITHOUT USE OF AN INTERNAL DISTANCE REFERENCE

A recent H-1 NMR method has been applied to the determination of the s olution structure and internal dynamics of a synthetic mixed C/O trisa ccharide related to sialyl Lewis(x). Varying the rf field offset in RO ESY-type experiments enabled the measurement of longitudinal and trans verse dipolar cross-relaxation rates with high accuracy. Assuming that for each proton pair the motion could be represented by a single expo nential autocorrelation function, it was possible to derive geometrica l parameters (r) and dynamic parameters (tau(cp)). With this assumptio n, 224 cross-relaxation rates have been transformed into 30 interproto n distance constraints and 30 dipolar correlation times. The distance constraints have been used in a simulated-annealing procedure. This tr isaccharide exhibits a structure close to the O-glycosidic analogue, b ut its flexibility seems highly reduced. On the basis of the determine d structure and dy namics, it is shown that no conformational exchange occurs, the molecule existing in the form of a unique family in aqueo us solution. In order to assess the quality of the resulting structure s and to validate this new experimental procedure of distance extracti on, we finally compare these solution structures to the ones obtained using three different sets of distances deduced from three choices of internal reference. It appears that this procedure allows the determin ation of the most precise and accurate solution.