I. The use of deuterated phospholipids to elucidate lipid-lipid interactions in bilayer vesicles. II. The effect of chain length on the secondary structure of oligoadenylates

Abstract

Part I: The Use of Deuterated Phospholipids to Elucidate Lipid-Lipid Interactions in Bilayer Vesicles. The motional state of lecithin molecules in single walled bilayer vesicles has been studied by nuclear magnetic resonance spectroscopy. Inter- and intramolecular relaxation rates have been determined for the hydrocarbon chain methylene protons. The intermolecular contribution is much smaller than previously estimated. The data suggest a model in which hydrocarbon chain motion is described by rapid kink formation and diffusion along the chains in addition to slower C-C trans-gauche rotations. A lateral diffusion coefficient of 2.5 - 5.5 x 10^(-8) cm^2 sec^(-1) is estimated. Both linewidths and spin-lattice relaxation rates have been measured for lecithins with different hydrocarbon chain lengths. Above the crystal to liquid crystal phase transition temperature, the linewidths are independent of chain length. This is taken to indicate that the hydrocarbon chain mobility and order are also independent of chain length. The motional state of cholesterol in bilayer vesicles has been studied. Cholesterol dispersed in bilayers consisting of dipalmitoyl lecithin with perdeuterated hydrocarbon chains has an NMR spectrum with well defined features. Two of the five methyl resonances are sharp and clearly resolved. These are assigned to the isopropyl methyls of the hydrocarbon tail. The other methyl resonances are much broader. This is taken to indicate that the hydrocarbon tail is much more mobile and disordered than the steroid nucleus. These conclusions are in agreement with and provide experimental support for the model proposed by Rothman and Engelman, but contrast with recent NMR studies which conclude that both the steroid nucleus and hydrocarbon tail are highly immobilized. Part II: The Effect of Chain Length on the Secondary Structure of Oligoadenylates. The oligoadenylates (Ap)_(2_4)A have been studied by proton magnetic resonance (pmr) spectroscopy. All the exterior base protons and a number of the interior base proton resonances have been assigned. The results of this work showed that the adenine bases in these oligoadenylates are intramolecularly stacked at 20°C with their bases oriented preferentially in the anti conformation about their respective glycosidic bonds. The oligomers were found to associate extensively even at concentrations of 0.02 M, primarily via to end stacking. With increasing temperature the oligomer bases destack, but it is argued that this unfolding process cannot be described in terms of a stacked versus unstacked model. Instead, the temperature dependences of the base proton chemical shifts support a base-oscillation model. The relationship between this model and the two-state model is discussed. Finally, on the basis of the chain-length dependence of the proton chemical shifts of the various adenine bases, it was concluded that subtle variations in the secondary structure of these oligomers exist with increasing chain length. Evidence is presented to show that the effects of distant base shielding are considerably smaller than what was previously estimated. The observed departures from the extended dimer model are attributed to differences in the relative orientations of the bases with respect to their neighbors in the oligomer.