Solid state nuclear magnetic resonance studies of select electrolyte interactions with phospholipid bilayer membranes in various model membrane systems

Several select interactions between electrolytes and phospholipid bilayer membranes were investigated by a variety of nuclear magnetic resonance (NMR) techniques. The incorporation of charged amphiphiles into bilayered micelles (bicelles), a new model membrane system, was investigated by a combination of phosphorus-31 (31P) and deuterium (2H) solid state NMR spectroscopy. 31P NMR results confirmed that, in the presence of sufficient ionic strength, charged amphiphiles can routinely be incorporated into bicelles and that they retain the ability to spontaneously align within strong magnetic fields. 2H NMR spectroscopy revealed that the changes in quadrupole splittings of head group deuterons in response to changes in membrane surface charge were consistent with the phosphocholine head group functioning as a molecular voltmeter. The interaction of the paramagnetic lanthanide ion Eu3+ with bicelles of various lipid compositions was investigated by 2H NMR spectroscopy and several different effects were observed. First, there was a progressive change in α- and β-deuteron quadrupole splittings in response to Eu3+ binding. Second was the observation of paramagnetic shifts in the isotropic chemical shifts of the observed deuterium signals. Third was the observation of a reorientation of the bicelles in the magnetic field in response to Eu3+ binding. Fluorescence experiments quantified the binding of Eu3+ so that the relative proportion of the two differently oriented phases could be correlated directly with the amount of Eu3+ bound. Additional experiments involving bicelles explored temperature effects on the 2H NMR spectra, in addition to 2H longitudinal and transverse relaxation measurements. A separate line of investigation probed the binding of Al3+ to phospholipids by employing several different NMR techniques. The amount of Al3+ that bound to phospholipid membranes was determined by means of an aluminum-27 (27Al) NMR difference assay. It was found that anionic membrane surfaces significantly increased the degree of Al3+ binding. Fast magic angle spinning (MAS) 31P NMR experiments quantified the amount of POPC and POPG that were bound by Al3+ and a distinct preference for POPG over POPC was observed. Slow MAS 31P NMR experiments indicated the potential to determine the relative proximity of Al3+ to the phosphate center of the phospholipid head group. 31P NMR two-dimensional (2D) exchange experiments allowed for a calculation of the lifetime of the Al3+-phospholipid complexes, with the Al 3+-POPG complex having a lifetime approximately 2.5 times greater than that for the Al3+-POPC complex. One final avenue of experimentation focused on observing and quantifying the rate of lipid lateral diffusion through the application of 2D 31P 2D exchange experiments under static or slow MAS conditions. Results from the latter experiments, coupled with a new spectral simulation program, identified lipid diffusion constants on the order of 5 * 10−8 cm2/sec.