P-31 2-DIMENSIONAL SOLID-STATE EXCHANGE NMR - APPLICATION TO MODEL MEMBRANE AND BIOLOGICAL-SYSTEMS

Two-dimensional solid-state P-31 NMR has been used to investigate the orientational exchange of phospholipids in gel and liquid-crystalline aqueous multilamellar dispersions and oriented multibilayers, and in biological membranes. In liquid-crystalline L-alpha multilamellar dispersions, orientational exchange originates from the lateral diffusion of phospholipid molecules over the curved surface of the liposomes and is manifest by an increase in off-diagonal intensity, which correlates the 90 and 0-degrees orientations of the membrane normal with respect to the magnetic field when the system is fully exchanged. Spectral simulations of the time evolution of exchange allowed determination of the correlation times tau-d lateral diffusion. For DMPC and DPPC at comparable reduced temperatures, tau-d values of 44 and 8 ms were obtained, respectively. The nature and rate of exchange observed for POPE at 30-degrees-C is similar to the of DMPC at the same temperature. The measured correlation times are consistent with diffusion rates obtained by FRAP for liposomes with radii in the 1 mu-m range. In the gel phase of DPPC (30-degrees), little orientational exchange is observed at mixing times up to 200 ms, demonstrating that the lateral diffusion is very slow. The correlation time for orientational exchange obtained from spectral simulations was approximately 900 ms; thus, exchange in the gel state is at least two orders of magnitude slower than in the liquid-crystalline state. In the P-beta' (ripple) phase, at temperatures between 34 and 39-degrees-C, significant exchange is observed for mixing times between 50 and 200 ms. Exchange is also observed in oriented samples of DPPC in the P-beta' phase for mixing times for 50 ms, but not for oriented liquid-crystalline samples for mixing times up to 100 ms. The exchange observed in the ripple phase cloud originate from rapid lateral diffusion of "fast" diffusing phospholipid within defect structures, and/or from "slow" lateral diffusion of ordered phospholipid over the ripples. 2D experiments were also performed on pig erythrocyte ghosts and on intact pig spinal cord. Significant orientational exchanges was observed with the erythrocyte ghosts at a mixing time of 200 ms, but almost no exchange was observed with the spinal cord at the same mixing time. Spectral simulations suggest tau-d values of approximately 400 ms and 1.3 s for the erythrocyte ghosts and spinal cord at 30-degrees-C. The results demonstrate that exchange in the biological membranes is significantly slower than in the model membrane systems, which suggests that the cell surfaces are relatively "smooth," i.e., any local surface perturbations are either present in small number or have little effect on the mean orientation of the phospholipids with respect to the membrane normal.