NMR study of the conformation of galactocerebroside in bilayers and solution: galactose reorientation during the metastable-stable gel transition.

Conformations of two types of bovine brain cerebroside containing normal and alpha-hydroxy-fatty acids (NFA-CER and HFA-CER, respectively) in solution and in bilayers were investigated using 1H and 13C NMR in solution and in the solid state. The analysis of vicinal 1H-1H coupling constants and NOE measurements in solution indicated that in both cerebrosides the predominant conformation about the O1-C1, C1-C2, and C2-C3 bonds is ap/-sc/ap, respectively. The remarkable similarity in the 13C NMR chemical shifts in solution and in hydrated liquid-crystalline bilayers indicated that both cerebrosides in bilayers assume conformation essentially identical to those in solution. The obtained 13C NMR spectra in solution were used as a reference for comparison with the variable-temperature 13C CP-MAS NMR spectra in the metastable and stable gel phases. The lack of chemical shift changes of polar carbon atoms upon cooling the HFA-CER bilayers below the Tm strongly suggests that the liquid-crystalline-metastable gel transition is not associated with a conformational change of the head group. The observed line broadening can be interpreted in terms of the hydrocarbon chain crystallization and slow dynamics of the head group in the metastable phase. On the other hand, the relaxation of the metastable gel phase of HFA-CER caused profound changes in the 13C spectra, primarily of the signals of the galactose C1, the ceramide C2, C4, and C5, and the carbonyl group. These changes are interpreted using the known dependence of the chemical shifts of anomeric carbon on the conformation about the O1-C1 bond to suggest that the gel phase relaxation involves a significant reorientation of the galactose moiety caused by a change in the rotation of the O1-C1 bond from the ap to -sc conformer. Similar changes of chemical shifts were observed in the case of NFA-CER during the transition from the liquid-crystalline phase to the stable gel phase.