Abstract
The proton pump of bacteriorhodopsin in an aqueous solution at varied pH upon pulsed excitation was monitored using a solution-based electrochemical module. The photocurrent action spectrum agreed with the absorption contour at 495-645nm. Diminishing the photocurrent amplitude by adding a protonophore, carbonyl cyanide m-chlorophenyl hydrazone, revealed that protons were the charge carriers of the photocurrent. The evolution of the conventional proton pump is proposed to occur in three elementary steps consecutively: first, the proton relay from the protonated Schiff base to the purple membrane (PM) surface (k 1 ), then the proton exchange between PM surface and bulk (k 2 ), and finally, the proton uptake (k 3 ). The fitted temporal profiles of the photocurrent agreed with observations in the pH range 5.8-9.5. At pH7.3, k 1 , k 2 , and k 3 were 2098s -1 , 412s -1 , and 44s -1 , respectively. The rate coefficients at pH9.5 were smaller than those at pH6.3 by a factor of approximately 2, consistent with the differences in the intrinsic mobilities of the charge carriers proton and hydroxide ion. The combination of the electrochemical detection module and the concomitant model provides a promising tool for quantitative and qualitative characterization of the light-driven ion pumps. © 2014 Elsevier B.V.