Cholesterol orientation and dynamics in dimyristoylphosphatidylcholine bilayers: A solid state deuterium NMR analysis

Proton decoupled deuterium NMR spectra of oriented bilayers made of DMPC and 30 mot % deuterated cholesterol acquired at 76.8 MHz (30 degrees C) have provided a set of very accurate quadrupolar splitting for eight C-D bonds of cholesterol. Due to the new precision of the experimental data, the original analysis by Dufourc et al. (1984. Biochemistry. 23:6062-6071) had to be reconsidered. We performed a systematic study of the influence on the precision and uniqueness of the data-fitting procedure of: (i) the coordinates derived from x-ray, neutron scattering, or force field-minimized structures, (ii) internal mobility, (iii) the axial symmetry hypothesis, and (iv) the knowledge of some quadrupolar splitting assignments. Good agreement between experiment and theory could be obtained only with the neutron scattering structure, for which both axial symmetry hypothesis and full order parameter matrix analysis gave satisfactory results. Finally, this work revealed an average orientation of cholesterol slightly different from those previously published and, most importantly, a molecular order parameter equal to 0.95 +/- 0.01, instead of 0.79 +/- 0.03 previously found for the same system at 30 degrees C. Temperature dependence in the 20-50 degrees C range shows a constant average orientation and a monotonous decrease of cholesterol S-mol, with a slope of -0.0016 K-1. A molecular order parameter of 0.89 +/- 0.01 at 30 degrees C was determined for a DMPC/16 mol % of cholesterol.