Biophys J, October 2001, p. 2059-2068, Vol. 81, No. 4

The Voltage-Dependent Proton Pumping in Bacteriorhodopsin Is Characterized by Optoelectric Behavior

Sven Geibel,^* Thomas Friedrich,^* Pal Ormos, Phillip G. Wood,^* Georg Nagel,^* and Ernst Bamberg^*

 ^*Max Planck Institut für Biophysik, D-60596 Frankfurt am Main, Germany; and  Institute of Biophysics, Biological Research Centre, Hungarian Academy of Sciences, H-6701 Szeged, Hungary

The light-driven proton pump bacteriorhodopsin (bR) was functionally expressed in Xenopus laevis oocytes and in HEK-293 cells. The latter expression system allowed high time resolution of light-induced current signals. A detailed voltage clamp and patch clamp study was performed to investigate the pH versus dependence of the pump current. The following results were obtained. The current voltage behavior of bR is linear in the measurable range between 160 mV and +60 mV. The pH dependence is less than expected from thermodynamic principles, i.e., one pH unit produces a shift of the apparent reversal potential of 34 mV (and not 58 mV). The M₂-BR decay shows a significant voltage dependence with time constants changing from 20 ms at +60 mV to 80 ms at 160 mV. The linear I-V curve can be reconstructed by this behavior. However, the slope of the decay rate shows a weaker voltage dependence than the stationary photocurrent, indicating that an additional process must be involved in the voltage dependence of the pump. A slowly decaying M intermediate (decay time > 100 ms) could already be detected at zero voltage by electrical and spectroscopic means. In effect, bR shows optoelectric behavior. The long-lived M can be transferred into the active photocycle by depolarizing voltage pulses. This is experimentally demonstrated by a distinct charge displacement. From the results we conclude that the transport cycle of bR branches via a long-lived M₁* in a voltage-dependent manner into a nontransporting cycle, where the proton release and uptake occur on the extracellular side.

Biophys J, October 2001, p. 2059-2068, Vol. 81, No. 4
© 2001 by the Biophysical Society   0006-3495/01/10/2059/10  $2.00

This article has been cited by other articles:

				R. Hagedorn, D. Gradmann, and P. Hegemann Dynamics of Voltage Profile in Enzymatic Ion Transporters, Demonstrated in Electrokinetics of Proton Pumping Rhodopsin Biophys. J., December 1, 2008; 95(11): 5005 - 5013. [Abstract] [Full Text] [PDF]

				S. P. Tsunoda, D. Ewers, S. Gazzarrini, A. Moroni, D. Gradmann, and P. Hegemann H+-Pumping Rhodopsin from the Marine Alga Acetabularia Biophys. J., August 15, 2006; 91(4): 1471 - 1479. [Abstract] [Full Text] [PDF]

				P. Hegemann, S. Ehlenbeck, and D. Gradmann Multiple Photocycles of Channelrhodopsin Biophys. J., December 1, 2005; 89(6): 3911 - 3918. [Abstract] [Full Text] [PDF]

				L. Rivas, S. Hippler-Mreyen, M. Engelhard, and P. Hildebrandt Electric-Field Dependent Decays of Two Spectroscopically Different M-States of Photosensory Rhodopsin II from Natronobacterium pharaonis Biophys. J., June 1, 2003; 84(6): 3864 - 3873. [Abstract] [Full Text] [PDF]

				G. Nagel, D. Ollig, M. Fuhrmann, S. Kateriya, A. M. Musti, E. Bamberg, and P. Hegemann Channelrhodopsin-1: A Light-Gated Proton Channel in Green Algae Science, June 28, 2002; 296(5577): 2395 - 2398. [Abstract] [Full Text] [PDF]