The nuclear Overhauser effect has been observed in the nuclear magnetic resonance spectra of 31p. The information content of the nuclear Overhauser effect has been applied to the structure and dynamic properties of phosphatidylchoiine vesicles. In the vesicles only % of the theoretical maximum nuclear Overhauser effect enhancement is observed. This result is accounted for by dipolar interactions between the N-methyl protons and the phosphate of phosphatidylcholine, and a correlation time for internal motion of 1.4 X 10-9 sec. Addition of up to 30% cholesterol does not change the nuclear Overhauser effect enhancement or spin-lattice relaxation time of the vesicles. It is argued that the OH group of cholesterol is hydrogen bonded to the ester carbonyl oxygen of the phosphatidylcholine molecules. Amphiphilic molecules such as phospholipids and sterols are major lipid constituents of many membranes of mammalian cells and subcellular organelles. In recent years the molecular interactions of phospholipids and steroids in the lamellar gel and liquid-crystalline phases have been studied in detail by a variety of physical techniques (1-5). Although the effects of cholesterol on the structural and motional properties of phospholipid bilayers are becoming better understood (6, 7), the emphasis of previous investigations has been placed primarily on the hydrocarbon region of the bilayer system. Information on the importance of the interactions between phospholipids and steroids, especially the involvement of 30-OH group of cholesterol, in the polar head region is accumulating (8, 9), but little is known about either the precise position of the hydrophilic hydroxyl group of cholesterol in the bilayer, or which of the groups, such as the phosphate, quaternary ammonium, and carbonyl groups of phosphatidylcholine, is participating in the phospholipid-cholesterol interaction in the polar head region of the bilayer system. In order to investigate the interactions involving the polar groups of phosphatidylcholine with cholesterol, we have used phosphorus nuclear magnetic resonance (31P NMR).In this communication, we report evidence for a choline N-methyl proton interaction with the phosphate group of phosphatidylcholine in a vesicle bilayer system. The results suggest an intermolecular association which provides a model for the surface of the bilayer, and for molecular motion in the headgroup. The effects of added cholesterol lead to the conclusion that cholesterol does not interact with the phosphate group and that high percentages of cholesterol change the conformation of the headgroup, or, alternatively, the motion of that region.
EXPERIMENTAL MATERIALS AND PROCEDURESEgg phosphatidylcholine (PC) was isolated and purified as described (10). PC in which the N-methyl groups were fully deuterated was prepared by Dr. Barry Lentz with phosphatidic acid hydrolyzed from egg PC by action of phospholipase D, and subsequently condensed with deuterated choline acetate (11). The lipid was purified by silicic acid chromatography. Cho...