OSMOTIC AND PH TRANSMEMBRANE GRADIENTS CONTROL THE LYTIC POWER OF MELITTIN

Transmembrane osmotic gradients applied on large unilamellar 1-palmito yl-2-oleoyl-phosphatidylcholine vesicles were used to modulate the pot ency of melittin to induce leakage. Melittin, an amphipathic peptide, changes the permeability of vesicles, as studied using the release of entrapped calcein, a fluorescent marker. A promotion of the ability of melittin to induce leakage was observed when a hyposomotic gradient ( i.e., internal salt concentration higher than the external one) was im posed on the vesicles. It is proposed that structural perturbations ca used by the osmotic pressure loosen the compactness of the outer leafl et, which facilitates the melittin-induced change in membrane permeabi lity. Additionally, we have shown that this phenomenon is not due to e nhanced binding of melittin to the vesicles using intrinsic fluorescen ce of the melittin tryptophan. Furthermore, we investigated the possib ility of using a transmembrane pH gradient to control the lytic activi ty of melittin. The potency of melittin in inducing release is known t o be inhibited by increased negative surface charge density. A transme mbrane pH gradient causing an asymmetric distribution of unprotonated palmitic acid in the bilayer is shown to be an efficient way to modula te the lytic activity of melittin, without changing the overall lipid composition of the membrane. We demonstrate that the protective effect of negatively charged lipids is preserved for asymmetric membranes.