1. Formation of M-Like Intermediates in Proteorhodopsin in Alkali Solutions (pH ≥ ∼8.5) Where the Proton Release Occurs First in Contrast to the Sequence at Lower pH.
- Author
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Tamogami J, Sato K, Kurokawa S, Yamada T, Nara T, Demura M, Miyauchi S, Kikukawa T, Muneyuki E, and Kamo N
- Subjects
- Algorithms, Amino Acid Substitution, Aquatic Organisms metabolism, Aquatic Organisms radiation effects, Bacterial Proteins chemistry, Bacterial Proteins genetics, Biocatalysis radiation effects, Biological Transport radiation effects, Eubacterium metabolism, Eubacterium radiation effects, Gammaproteobacteria metabolism, Gammaproteobacteria radiation effects, Halobacterium salinarum metabolism, Halobacterium salinarum radiation effects, Hydrogen-Ion Concentration, Immobilized Proteins chemistry, Immobilized Proteins genetics, Immobilized Proteins metabolism, Lipid Bilayers chemistry, Membranes, Artificial, Mutation, Phosphatidylcholines chemistry, Photochemical Processes, Proton Pumps chemistry, Proton Pumps genetics, Recombinant Proteins chemistry, Recombinant Proteins metabolism, Rhodopsins, Microbial chemistry, Rhodopsins, Microbial genetics, Bacterial Proteins metabolism, Models, Molecular, Proton Pumps metabolism, Rhodopsins, Microbial metabolism
- Abstract
Proteorhodopsin (PR) is an outward light-driven proton pump observed in marine eubacteria. Despite many structural and functional similarities to bacteriorhodopsin (BR) in archaea, which also acts as an outward proton pump, the mechanism of the photoinduced proton release and uptake is different between two H(+)-pumps. In this study, we investigated the pH dependence of the photocycle and proton transfer in PR reconstituted with the phospholipid membrane under alkaline conditions. Under these conditions, as the medium pH increased, a blue-shifted photoproduct (defined as Ma), which is different from M, with a pKa of ca. 9.2 was produced. The sequence of the photoinduced proton uptake and release during the photocycle was inverted with the increase in pH. A pKa value of ca. 9.5 was estimated for this inversion and was in good agreement with the pKa value of the formation of Ma (∼ 9.2). In addition, we measured the photoelectric current generated by PRs attached to a thin polymer film at varying pH. Interestingly, increases in the medium pH evoked bidirectional photocurrents, which may imply a possible reversal of the direction of the proton movement at alkaline pH. On the basis of these findings, a putative photocycle and proton transfer scheme in PR under alkaline pH conditions was proposed.
- Published
- 2016
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