Location of cholesterol in model membranes by magic-angle-sample-spinning NMR

High-resolution magic-angle-sample-spinning C-13-NMR was applied to determine the specific location of cholesterol in non-perturbed multilamellar model membranes formed by egg yolk phosphatidylcholine. C-13 spin-lattice relaxation times of both the phospholipid and cholesterol molecules were measured in the absence and in the presence of Gd3+, a paramagnetic agent, in order to obtain information on molecular distances. The effect of Gd3+ on the spin-lattice relaxation times of the lipid resonances has an explicit distance dependence, allowing it to be used to evaluate relative distances on a molecular scale. It has been found that cholesterol is placed in such a position that it is not readily exposed to the solvent: the hydrophobic steroid ring is oriented parallel to the membrane phospholipids, the hydroxyl group is in close vicinity to the phospholipid ester carbonyl groups and the isooctyl side chain is deeply buried in the center of the membrane. These data are consistent with an organization such that mixtures of cholesterol and phospholipids present a packing similar to that found in interdigitated lipid bilayer systems.