Volume 11, Number 1, p.p. 11–17
Quasi-continuous illumination of bacteriorhodopsins: absorbance and electric current
R. Tóth-Boconádi,1 S.G. Taneva2 and L. Keszthelyi1,3,*
1 Institute of Biophysics, Biological Research Centre, Hungarian Academy of Sciences, P.O. B. 521, H-6701 Szeged
2 Institute of Biophysics, Bulgarian Academy of Sciences, Sofia
3 KFKI Research Institute for Particle and Nuclear Physics, Hungarian Academy of Sciences, P.O.B. 49, H-1525 Budapest
The study of the mutant D96N plays an important rôle in understanding light-driven proton translocation in bacteriorhodopsin. Here we describe absorbance changes at 575, 450 and 400 nm and photoelectric currents for quasicontinuous illumination (i.e., illumination with continuous light for a few s) and found the optical and electric data very different. The difference is due to the increased lifetime of this mutant and is explained with simple equations. We found a long-living negative current of small amplitude (opposite in direction to the main proton translocation) when the quasicontinuous illumination was switched off. Pyranine, used as a proton indicator in solution, influenced the electric current: increasing during illumination and reversing the negative current to positive after illumination. For comparison, similar measurements were carried out on wild-type bacteriorhodopsin and its mutants E204Q and R227Q. Data from absorption changes indicated proton release connected to the P and/or Q substates, products of the excitation of the O intermediate of wild-type bacteriorhodopsin.
Keywords: buffer effect, orientation, proton emission, proton pump, purple membrane, quasicontinuous current