著者
Yu Nakajima Takashi Tsukamoto Yohei Kumagai Yoshitoshi Ogura Tetsuya Hayashi Jaeho Song Takashi Kikukawa Makoto Demura Kazuhiro Kogure Yuki Sudo Susumu Yoshizawa
出版者
Japanese Society of Microbial Ecology · The Japanese Society of Soil Microbiology
雑誌
Microbes and Environments (ISSN:13426311)
巻号頁・発行日
pp.ME17197, (Released:2018-03-16)
被引用文献数
1

Light-driven ion-pumping rhodopsins are widely distributed among bacteria, archaea, and eukaryotes in the euphotic zone of the aquatic environment. H+-pumping rhodopsin (proteorhodopsin: PR), Na+-pumping rhodopsin (NaR), and Cl–-pumping rhodopsin (ClR) have been found in marine bacteria, which suggests that these genes evolved independently in the ocean. Putative microbial rhodopsin genes were identified in the genome sequences of marine Cytophagia. In the present study, one of these genes was heterologously expressed in Escherichia coli cells and the rhodopsin protein named Rubricoccus marinus halorhodopsin (RmHR) was identified as a light-driven inward Cl– pump. Spectroscopic assays showed that the estimated dissociation constant (Kd,int.) of this rhodopsin was similar to that of haloarchaeal halorhodopsin (HR), while the Cl–-transporting photoreaction mechanism of this rhodopsin was similar to that of HR, but different to that of the already-known marine bacterial ClR. This amino acid sequence similarity also suggested that this rhodopsin is similar to haloarchaeal HR and cyanobacterial HRs (e.g., SyHR and MrHR). Additionally, a phylogenetic analysis revealed that retinal biosynthesis pathway genes (blh and crtY) belong to a phylogenetic lineage of haloarchaea, indicating that these marine Cytophagia acquired rhodopsin-related genes from haloarchaea by lateral gene transfer. Based on these results, we concluded that inward Cl–-pumping rhodopsin is present in genera of the class Cytophagia and may have the same evolutionary origins as haloarchaeal HR.
著者
Yu Nakajima Takashi Tsukamoto Yohei Kumagai Yoshitoshi Ogura Tetsuya Hayashi Jaeho Song Takashi Kikukawa Makoto Demura Kazuhiro Kogure Yuki Sudo Susumu Yoshizawa
出版者
Japanese Society of Microbial Ecology · The Japanese Society of Soil Microbiology
雑誌
Microbes and Environments (ISSN:13426311)
巻号頁・発行日
vol.33, no.1, pp.89-97, 2018 (Released:2018-03-29)
参考文献数
43
被引用文献数
1

Light-driven ion-pumping rhodopsins are widely distributed among bacteria, archaea, and eukaryotes in the euphotic zone of the aquatic environment. H+-pumping rhodopsin (proteorhodopsin: PR), Na+-pumping rhodopsin (NaR), and Cl−-pumping rhodopsin (ClR) have been found in marine bacteria, which suggests that these genes evolved independently in the ocean. Putative microbial rhodopsin genes were identified in the genome sequences of marine Cytophagia. In the present study, one of these genes was heterologously expressed in Escherichia coli cells and the rhodopsin protein named Rubricoccus marinus halorhodopsin (RmHR) was identified as a light-driven inward Cl− pump. Spectroscopic assays showed that the estimated dissociation constant (Kd,int.) of this rhodopsin was similar to that of haloarchaeal halorhodopsin (HR), while the Cl−-transporting photoreaction mechanism of this rhodopsin was similar to that of HR, but different to that of the already-known marine bacterial ClR. This amino acid sequence similarity also suggested that this rhodopsin is similar to haloarchaeal HR and cyanobacterial HRs (e.g., SyHR and MrHR). Additionally, a phylogenetic analysis revealed that retinal biosynthesis pathway genes (blh and crtY) belong to a phylogenetic lineage of haloarchaea, indicating that these marine Cytophagia acquired rhodopsin-related genes from haloarchaea by lateral gene transfer. Based on these results, we concluded that inward Cl−-pumping rhodopsin is present in genera of the class Cytophagia and may have the same evolutionary origins as haloarchaeal HR.
著者
Keiichi Kojima Hiroshi C. Watanabe Satoko Doi Natsuki Miyoshi Misaki Kato Hiroshi Ishikita Yuki Sudo
出版者
The Biophysical Society of Japan
雑誌
Biophysics and Physicobiology (ISSN:21894779)
巻号頁・発行日
vol.15, pp.179-188, 2018 (Released:2018-09-07)
参考文献数
42

Anion channelrhodopsin-2 (ACR2), a light-gated channel recently identified from the cryptophyte alga Guillardia theta, exhibits anion channel activity with exclusive selectivity. In addition to its novel function, ACR2 has become a focus of interest as a powerful tool for optogenetics. Here we combined experimental and computational approaches to investigate the roles of conserved carboxylates on the anion transport activity of ACR2 in Escherichia coli membrane. First, we replaced six conserved carboxylates with a neutral residue (i.e. E9Q, E56Q, E64Q, E159Q, E219Q and D230N), and measured anion transport activity using E. coli expression system. E159Q and D230N exhibited significantly lower anion transport activity compared with wild-type ACR2 (1/12~1/3.4), which suggests that E159 and D230 play important roles in the anion transport. Second, to explain its molecular aspects, we constructed a homology model of ACR2 based on the crystal structure of a cation channelrhodopsin (ChR). The model structure showed a cavity formed by four transmembrane helices (TM1, TM2, TM3 and TM7) similar to ChRs, as a putative anion conducting pathway. Although E159 is not located in the putative pathway, the model structure showed hydrogen bonds between E159 and R129 with a water molecule. D230 is located in the pathway near the protonated Schiff base (PSB) of the chromophore retinal, which suggests that there is an interaction between D230 and the PSB. Thus, we demonstrated the functional importance and the hypothetical roles of two conserved carboxylates, E159 and D230, in the anion transport activity of ACR2 in E. coli membrane.
著者
Marie Kurihara Yuki Sudo
出版者
一般社団法人 日本生物物理学会
雑誌
Biophysics and Physicobiology (ISSN:21894779)
巻号頁・発行日
vol.12, pp.121-129, 2015 (Released:2015-12-11)
参考文献数
71
被引用文献数
11

One of the major topics in biophysics and physicobiology is to understand and utilize biological functions using various advanced techniques. Taking advantage of the photoreactivity of the seven-transmembrane rhodopsin protein family has been actively investigated by a variety of methods. Rhodopsins serve as models for membrane-embedded proteins, for photoactive proteins and as a fundamental tool for optogenetics, a new technology to control biological activity with light. In this review, we summarize progress of microbial rhodopsin research from the viewpoint of distribution, diversity and potential.