Conformational characterization of sphingomyelin and its interactions with water and cholesterol by infrared and nuclear magnetic resonance spectroscopies

Sphingomyelin (SM) is a major component of mammalian plasma membranes, particularly those in ocular lenses. Sphingomyelin exhibits a preferential affinity for cholesterol (CHol), another predominant lipid in lens membranes. Due to its flexibility, there is no published X-ray structure of SM. Although it has been recognized that H-bonds play a key role in the aggregation of SM and could facilitate the interactions between SM and CHol, the specific nature and impact of these bonds on the conformation of SM are unclear.; The first goal of this work was to establish the conformational preferences of SM in monomeric and aggregated forms in chloroform, a solvent significantly less polar than water that may provide a rational mimetic environment to CHol-rich membranes. The second goal was to characterize the interactions of SM with water and CHol.; Infrared and nuclear magnetic resonance spectral data indicate that intramolecular H-bonds are predominant in the monomeric form of SM and connect the OH and NH groups of the interface of SM to the ester oxygen and anionic oxygens of the phosphate group. Above the critical micelle concentration of SM in chloroform (3.2 +/- 0.3 mM at 32°C), some of the intramolecular H-bonds involving the NH moiety are disrupted to form new intermolecular H-bonds with the C=O group of adjacent molecules. Water is an integral part of the H-bonding network as it forms strong H-bonds connecting the amide groups of neighboring molecules.; The betaD-OH group of CHol is essential to the interaction of the steroid with SM. This OH group acts as an anchor that allows the docking of CHol between SM molecules by binding to bridging water molecules. CHol is thusly close to the interface region of SM and serves to disorder the hydrophobic tails of SM. As a result, the temperature and cooperativity of the gel to fluid phase transition for SM assemblies decrease. For CHol molar fractions exceeding 50%, CHol-CHol interactions prevail over those between CHol and SM, and the conformational features of SM resemble those seen for nearly monomeric arrangements. These results suggest the formation of CHol domains around SM molecules. Microscopic infrared studies suggest that the size of these microdomains is below 50mum.