Amphotericin B Channel-Forming Activity Depends on Membrane Dipole Potential

Abstract

Amphotericin B (AmB) is the most effective polyene antibiotic used in treatment of systemic fungal infections. The activity of AmB results from its interaction with sterol-containing membranes and consequent pore formation. As some sterols influence membrane dipole potential (Vd) one can assume that Vd may regulate the ability of AmB to form ion channels. The aim of the present work was to study the channel-forming activity of AmB as well as the properties of its single pores in planar lipid bilayers of different dipole potentials. AmB (0.1-0.2 µM) was added on both sides of a membrane prepared from diphytanoylphosphocholine (67 mol %) and cholesterol (33 mol %) in 2 M KCl (5 mM Hepes, pH 7.0) solutions. A reduction of Vd by addition of 20 µM phloretin to the bilayer bathing solutions is accompanied by ∼3-fold decrease of a single channel conductance and ∼20-fold increase of a steady-state AmB-induced membrane conductance. Increasing the membrane dipole potential by adding 5 µM RH421 produces ∼2-fold increase of the conductance of AmB channels and practically does not affect the steady-state bilayer conductance. The observed changes in the channel-forming activity of AmB in the presence of dipole modifying agents correspond to ∼60-fold increase and ∼2 fold decreases in steady-state open channel number in the presence of phloretin and RH421, respectively. The obtained results allow us to speculate that AmB molecule possess a certain dipole moment. Therefore, the decrease of membrane dipole potential produces a reduction of the energy barrier for interfacial accumulation of AmB dipoles. These results may be instructive in understanding of the membrane activity of amphotericin B. The study was supported in part by RFBR (# 09-04-00883), SS-3796.2010.4, the Program “Molecular and Cell Biology”, RAS, and the State contract # P1372 (MES, FTP“SSEPIR”).