PORIN PORE FORMATION : KINETICS IN DIFFERENT PLANAR LIPID MEMBRANES

The porin, a protein of outer membrane of mitochondria, changes the membrane ion permeability forming channels that are voltage-dependent. In this study we present comparative results on porin incorporation kinetics in black lipid membranes (BLM) made with different lipids: phosphatidylinositol (PI) and oxidized cholesterol (OxCh). The experimental methods have been previously reported (Proc. X School Biophys. Memb. Transp. 1: 328, Poland 1990). PI and OxCh were extracted from ox brain following respectively the methods of Folk J. (J. Biol. Chem. 146: 35, 1942) and of Tien H.T. et al. (Nature 212: 718, 1966). Porin was purified from heart beef mitochondria. Bathing solutions were KC1 1M. The capacitance and the conductance of OxCh membranes were 422.0 ± 23.5 nF/cm2 (n=30) and 4.77 ± 0.53 μS/cm2 (n=30) and those of PI membranes were 251.0 ± 9.3 nF/cm2 (n=61) and 2.27 ± 0.09 μS/cm2 (n=61). Porin addition to the bathing solutions determines an increase of the membrane conductance indicating a progressive insertion of new protein forming channels into BLM. The incorporation kinetics follows an hyperbolic function. As for PI, in the OxCh BLM the incorporation is a function of the porin concentration and of the applied voltage. The increase and the behaviour were different, in that PI membranes needed an higher porin concentration to determine a large increase of the conductance as compared to OxCh membranes confirming the observation of Benz R. et al. (J. Membrane Biol. 56: 19, 1980). The difference between the two types of kinetics is confirmed by the experiments in which porin was present before the membrane formation. The former continues to display a S-shaped kinetics, shifted on the right, the latter starts with an elevated conductance and continues to increase through an hyperbolic curve. It could be assumed that the protein incorporation into the two different BLMs is governed by the different lipid matrix