The diversity of the liquid ordered (Lo) phase of phosphatidylcholine/cholesterol membranes:: A variable temperature multinuclear solid-state NMR and X-ray diffraction study

To investigate the properties of a pure liquid ordered (L-o) phase in a model membrane system, a series of saturated phosphatidylcholines combined with cholesterol were examined by variable temperature multinuclear (H-1, H-2, C-13, P-31) solid-state NMR spectroscopy and x-ray scattering. Compositions with cholesterol concentrations >= 40 mol %, well within the L-o phase region, are shown to exhibit changes in properties as a function of temperature and cholesterol content. The H-2-NMR data of both cholesterol and phospholipids were used to more accurately map the L-o phase boundary. It has been established that the gel-L-o phase coexistence extends to 60 mol % cholesterol and a modified phase diagram is presented. Combined H-1-, H-2-, (1)3C-NMR, and x-ray scattering data indicate that there are large changes within the L-o phase region, in particular, H-1-magic angle spinning NMR and wide-angle x-ray scattering were used to examine the in-plane intermolecular spacing, which approaches that of a fluid L-alpha phase at high temperature and high cholesterol concentrations. Although it is well known for cholesterol to broaden the gel-to-fluid transition temperature, we have observed, from the C-13 magic angle spinning NMR data, that the glycerol region can still undergo a "melting", though this is broadened with increasing cholesterol content and changes with phospholipid chain length. Also from H-2-NMR order parameter data it was observed that the effect of temperature on chain length became smaller with increasing cholesterol content. Finally, from the cholesterol order parameter, it has been previously suggested that it is possible to determine the degree to which cholesterol associates with different phospholipids. However, we have found that by taking into account the relative temperature above the phase boundary this relationship may not be correct.