Surface charge control of electropermeabilization and glycophorin electroinsertion with 1,2-diacyl-sn-glycero-3-phosphocholine (lecithin) liposomes.

Back insertion of a solubilized membrane protein, glycophorin A, has been obtained in lipid multilamellar vesicles by applying calibrated electric field pulses on a lipid/protein mixture. Experimental evidence for insertion is given by means of immunofluorescence. Insertion was obtained only under field conditions that induced the leakage of a soluble hydrophilic molecule, calcein, which was trapped between the lipid layers. Studies were performed on mixed liposomes where charged species were present. The critical permeabilizing field is the same whatever the composition, but with overcritical fields the associated calcein transmembraneous flow is higher with positively charged lipids. Field conditions that where prone to trigger glycophorin insertion were similar to those that induced electropermeabilization. No electroinsertion has been obtained with stearylamine (SteNH2)/1,2-dipalmitoyl-sn-glycero-3-phosphocholine (Pam2GroPCho) liposomes under the same conditions. Calcein efflux as well as glycophorin insertion are controlled by the electric surface charge of the host liposome. These observations confirm our previous conclusions that spontaneous membrane protein insertion is obtained when the host membrane is brought to its electropermeabilized state, but show that a strong control due to the surface charges is nevertheless present.