RESUMO
In this study we substituted the retinal units in proteorhodopsin (PR) and bacteriorhodopsin (BR) with azo chromophores to investigate the mechanism of photoinduced proton pumping in rhodopsins and potentially develop new artificial molecular pumps. We used an indium tin oxide electrode to investigate the photoinduced proton transfer of the azo analogues of PR and BR. We also employed flash photolysis to determine the characteristic photocycles, comprising multiple transient intermediates, of the azo chromophore-bound PR and BR. Moreover, our studies of the photoinduced proton pumping functions of azo-proteoopsin and azo-bacterioopsin complexes revealed that they did not pump protons upon illumination, even though they underwent photoinduced proton transfer and the characteristic photocycle. Mutational analysis suggested that the proton pumping malfunction of the azo analogues of PR and BR resulted from the absence of proton transfer reactions through cytoplasmic channels, even though these reactions were evoked in extracellular channels. Based on our experimental findings, we propose herein a putative model of the proton transfer reaction mechanism for the azo analogues of PR and BR.
