Differential Effects of Membrane Order on Membrane Permeability

Phospholipid membranes segregate into lateral domains of liquid ordered (lo) and liquid disordered (ld) phases when cholesterol and mixed species of lipids with saturated and unsaturated acyl chains are present. To examine membrane permeability and rate of vesicle rupture by Triton in pure lo, pure ld, and mixed lo/ld phases, LUVs were prepared based on the ternary phase diagram of POPC, sphingomyelin, and cholesterol. These LUVs were loaded with 2mM carboxyfluorescein (CF) and formed by extrusion at 65oC. Using a stopped-flow fluorometer, changes in CF fluorescence were measured when LUVs were exposed to sudden osmotic gradients, pH gradients, or 0.1% Triton. Acyl chain and phospholipid headgroup packing were assessed in all compositions with time-resolved measurements of DPH fluorescence lifetime and anisotropy decay. Water permeability was highest in the pure ld phase, and a factor of more than 100 lower in the pure lo phase. Proton permeability was lowest in the pure ld phase, and approximately five fold higher in the lo phase. The rate of membrane rupture was higher in the pure ld phase than in the pure lo phase, with inconsistent results in the coexistence region. Water permeability was found to correlate with acyl chain packing, decreasing with increased membrane order. Proton permeability increased exponentially with increasing membrane order.