PROTON TRANSLOCATION ACROSS BACTERIORHODOPSIN CONTAINING SOLID SUPPORTED LIPID BILAYERS

Bacteriorhodopsin (BR) was incorporated in solid supported lipid bilay ers by fusion of reverse phase vesicles on chemisorbed monolayers of 1 ,2-dimyristoyl-sn-glycero-3-phosphothioethanol (DMPTE) on gold substra tes. The passive electrical behavior of the artificial membranes was m onitored by impedance spectroscopy in order to determine both the memb rane resistances and capacitances and to guarantee reproducibility of the bilayer formation. Illumination of the BR containing solid support ed lipid bilayers resulted in a transient photocurrent as expected fro m earlier experiments with black lipid membranes. The present preparat ion technique however is advantageous because of its long term stabili ty up to 1 day without loss of BR activity and its easy handling. We i nvestigated the dependence of the photocurrent on the BR content, lipi d environment, pH, and a proton carrier using a common current amplifi er. Maximum current densities were obtained in the presence of negativ ely charged lipids like 1,2-dimyristoyl-sn-glycero-3-phosphatidic acid (DMPA) or 1-palmitoyl-2-sn-glycero-3-phosphoglycerol (POPG) at a pH o f 6.4. Moreover it could be shown that the pump activity of reconstitu ted BR is insignificantly influenced by the capacitance of the first s elf-assembled DMPTE-monolayer on the gold electrodes. This may be expl ained by an incomplete fusion of BR containing vesicles on the hydroph obic surface. Carbonylcyanid-4-trifluoromethoxy-phenylhydrazone (FCCP) , a membrane soluble proton translocator, increases the membrane condu ctance as well as the capacitance of the lipid bilayer that was derive d either from impedance spectroscopy or evaluation of the time constan ts of the transient photocurrents. (C) 1997 Elsevier Science Ireland L td.