ANALYSIS OF THE DC PHOTOELECTRIC SIGNAL FROM MODEL BACTERIORHODOPSIN MEMBRANES - DC PHOTOCONDUCTIVITY DETERMINATION BY THE NULL CURRENT METHOD AND THE EFFECT OF PROTON IONOPHORES

On illumination by continuous light, a reconstituted bacteriorhodopsin membrane exhibits a stationary photocurrent under short-circuit condi tions. It has been widely reported that this photocurrent is linearly dependent on the applied transmembrane potential, and that the photocu rrent reverses its polarity at a critical potential. It is also well k nown that the stationary photosignal of a bacteriorhodopsin membrane i s linearly dependent on the light intensity and eventually reaches sat uration. In this paper, the null current method (F.T. Hong and D. Mauz erall, Biochim. Biophys. Acta, 275 (1972) 479) is applied to decompose the photocurrent into a photovoltaic part (photoemf) and a photocondu ctive part (photoconductance). It is found that the photoconductance i s zero in the dark, and is activated by illumination to reach a fixed magnitude which is independent of a further increase in the light inte nsity (''step-function'' photoswitching). Furthermore, the photoconduc tance is ohmic (i.e. independent of the applied potential). The linear voltage dependence of the photocurrent can be explained in terms of t he photoswitching by assuming that the light-activated proton conducta nce channel is also available for a transmembrane potential to drive a proton current through in either direction. With this assumption, the photoemf is shown to be voltage independent. The photoemf is initiall y linearly light dependent at low light intensities, but eventually re aches a saturation level. We confirm the reported enhancement effect o f the proton ionophores carbonyl cyanide-p-trifluoromethoxyphenylhydra zone (FCCP) and carbonyl cyanide-m-chlorophenylhydrazone (CCCP), which is caused by increases in both the photoconductance and the ionic con ductance. The action of the Cl- ionophore, nystatin, is quite differen t. Nystatin inhibits the photocurrent and increases the ionic conducta nce, but does not affect the photoconductance. The enhancement effect of proton ionophores cannot be explained by the shunting effect alone, even if the sandwich model postulated by Bamberg et al. (Biophys. Str uct. Mech., 5 (1979) 277) is invoked. We suspect that the incorporatio n of bacteriorhodopsin into the artificial black lipid membrane may be more complete than initially believed.