ID | 65948 |
FullText URL | |
Author |
Kurihara, Marie
Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama University
Thiel, Vera
Department of Biological Sciences, Tokyo Metropolitan University
Takahashi, Hirona
Department of Chemistry, Graduate School of Science, Okayama University of Science
Kojima, Keiichi
Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama University
Ward, David M.
Department of Land Resources and Environmental Sciences, Montana State University
Bryant, Donald A.
Department of Biochemistry and Molecular Biology, The Pennsylvania State University
Sakai, Makoto
Department of Chemistry, Graduate School of Science, Okayama University of Science
Yoshizawa, Susumu
Atmosphere and Ocean Research Institute, The University of Tokyo
Sudo, Yuki
Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama University
ORCID
Kaken ID
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Abstract | Rhodopsins are transmembrane proteins with retinal chromophores that are involved in photo-energy conversion and photo-signal transduction in diverse organisms. In this study, we newly identified and characterized a rhodopsin from a thermophilic bacterium, Bellilinea sp. Recombinant Escherichia coli cells expressing the rhodopsin showed light-induced alkalization of the medium only in the presence of sodium ions (Na+), and the alkalization signal was enhanced by addition of a protonophore, indicating an outward Na+ pump function across the cellular membrane. Thus, we named the protein Bellilinea Na+-pumping rhodopsin, BeNaR. Of note, its Na+-pumping activity is significantly greater than that of the known Na+-pumping rhodopsin, KR2. We further characterized its photochemical properties as follows: (i) Visible spectroscopy and HPLC revealed that BeNaR has an absorption maximum at 524 nm with predominantly (>96%) the all-trans retinal conformer. (ii) Time-dependent thermal denaturation experiments revealed that BeNaR showed high thermal stability. (iii) The time-resolved flash-photolysis in the nanosecond to millisecond time domains revealed the presence of four kinetically distinctive photointermediates, K, L, M and O. (iv) Mutational analysis revealed that Asp101, which acts as a counterion, and Asp230 around the retinal were essential for the Na+-pumping activity. From the results, we propose a model for the outward Na+-pumping mechanism of BeNaR. The efficient Na+-pumping activity of BeNaR and its high stability make it a useful model both for ion transporters and optogenetics tools.
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Keywords | rhodopsin
ion transport
retinal
isomerization
optogenetics
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Published Date | 2023-02-01
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Publication Title |
Chemical and Pharmaceutical Bulletin
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Volume | volume71
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Issue | issue2
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Publisher | Pharmaceutical Society of Japan
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Start Page | 154
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End Page | 164
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ISSN | 0009-2363
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NCID | AA00602100
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Content Type |
Journal Article
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language |
English
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OAI-PMH Set |
岡山大学
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Copyright Holders | © 2023 The Pharmaceutical Society of Japan
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File Version | publisher
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PubMed ID | |
DOI | |
Web of Science KeyUT | |
Related Url | isVersionOf https://doi.org/10.1248/cpb.c22-00774
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Funder Name |
Japan Society for the Promotion of Science
Japan Science and Technology Agency
Japan Agency for Medical Research and Development
Department of Energy
NASA Exobiology program
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助成番号 | JP19K16090
JP21K15054
JP18H02411
JP19H04727
JP19H05396
JP20K21482
JP21H0040413
JP21H0244613
JPMJCR1656
20dm0207060h0004
DE-FG02-94ER20137
NX09AM87G
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