Evaluation of surface tension and ion occupancy effects on gramicidin A channel lifetime.

摘要

The surface tension of glycerylmonooleate-hexadecane lipid bilayer membranes and the lifetime of gramicidin A channels were measured at various concentrations of the surrounding solutions. For HCl the surface tension is essentially constant at approximately 5 mN/m up to approximately 1 M, whereas the average lifetime increases approximately 40-fold. At higher concentrations the surface tension decreases markedly. For CsCl the surface tension is constant up to about 1 M then increases with salt level. The average lifetime in this case increases about sixfold. In both cases the lifetime levels off and even decreases at higher salt levels. The increase in lifetime observed with ion activity is therefore qualitatively different from, and not explained by, the established dependence of lifetime on membrane properties (Elliot, J.R., D. Needham, J.P. Dilger, and D.A. Haydon. 1983. Biochim. Biophys. Acta. 735:95-103). We have previously proposed that ion occupancy is a determinant of channel stability, and to test this hypothesis the voltage dependence of channel lifetime was measured in asymmetrical solutions. For the case of a potassium chloride solution on one side of the membrane and a hydrogen chloride solution, on the other, the voltage dependence of the lifetime is asymmetrical. The asymmetry is such that when the electrical field is applied in the direction of the chemical gradient for each of the ions, the channel lifetime approaches, at increasing field strengths, that of a symmetrical solution of the respective ion. The voltage dependence of the surface tension, on the other hand, is negligible for the range of voltages used. These results, and the earlier findings that the order of the lifetimes for the alkali cations generally agree with the order of the permeability selectivity of the gramicidin A channel, support the hypothesis that ion occupancy is a major factor determining the lifetime of gramicidin A channels. The effects of multivalent blockers and osmotic agents were also tested. Ba2", La3+,and Mg2" decrease the lifetime and conductance markedly. Sucrose and urea increase the lifetime and decrease the conductance. The voltage dependence of the lifetime in symmetrical solutions was examined. Contrary to previous reports it was found that the lifetimes for K+, Cs', and H+ are voltage dependent. For 0.5 M HCI the lifetime decreases monotonically by .60% at 150 mV, and for 0.5 M KCI the lifetime increases by -60% at 200 mV. Below 10 mM there is no effect of voltage for H+, K+, and Cs+. These effects of blockers, osmotic agents, and voltage on the lifetime, as well as the lack of effect of voltage at low salt levels, are consistent with the occupancy hypothesis.