The tetrameric MotA complex as the core of the flagellar motor stator from hyperthermophilic bacterium
Norihiro Takekawa, Naoya Terahara, Takayuki Kato, Mizuki Gohara, Kouta Mayanagi, Atsushi Hijikata, Yasuhiro Onoue, Seiji Kojima, Tsuyoshi Shirai, Keiichi Namba & Michio Homma
Scientific Reports 6, Article number: 31526 (2016)
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Biochemistry Biophysics
Received: 09 June 2016 Accepted: 20 July 2016 Published online: 17 August 2016
Source/Fonte: Michio Homma
Abstract
Rotation of bacterial flagellar motor is driven by the interaction between the stator and rotor, and the driving energy is supplied by ion influx through the stator channel. The stator is composed of the MotA and MotB proteins, which form a hetero-hexameric complex with a stoichiometry of four MotA and two MotB molecules. MotA and MotB are four- and single-transmembrane proteins, respectively. To generate torque, the MotA/MotB stator unit changes its conformation in response to the ion influx, and interacts with the rotor protein FliG. Here, we overproduced and purified MotA of the hyperthermophilic bacterium Aquifex aeolicus. A chemical crosslinking experiment revealed that MotA formed a multimeric complex, most likely a tetramer. The three-dimensional structure of the purified MotA, reconstructed by electron microscopy single particle imaging, consisted of a slightly elongated globular domain and a pair of arch-like domains with spiky projections, likely to correspond to the transmembrane and cytoplasmic domains, respectively. We show that MotA molecules can form a stable tetrameric complex without MotB, and for the first time, demonstrate the cytoplasmic structure of the stator.
Acknowledgements
We thank Dr. Chiaki Kato for providing chromosomal DNA of P. profundum, S. benthica and S. violacea. We thank Akio Kitao for providing the model of the transmembrane region of the MotA tetramer. This work was supported in part by Grants-in-Aid for scientific research from the Japan Society for the Promotion of Science (24117004 and 23247024 to M.H., 26840035 to N.Te., 26650021, 24117001, 24117004 to T.K., and 25000013 to K.N.), by Platform for Drug Discovery, Informatics, and Structural Life Science from Japan Agency for Medical Research and Development (to A.H. and T.S.), and from the Japan Society for the Promotion of Science (13J02161 to N.Ta.). N.Ta. was partly supported by the Integrative Graduate Education and Research Program in Green Natural Sciences of Nagoya University.
Author information
Author notes
Norihiro Takekawa, Naoya Terahara & Takayuki Kato
These authors contributed equally to this work.
Affiliations
Division of Biological Science, Graduate School of Science, Nagoya University, Chikusa-ku, Nagoya 464-8602, Japan
Norihiro Takekawa, Mizuki Gohara, Yasuhiro Onoue, Seiji Kojima & Michio Homma
Graduate School of Frontier Biosciences, Osaka University, 1-3 Yamadaoka, Suita, Osaka 565-0871, Japan
Naoya Terahara, Takayuki Kato & Keiichi Namba
Medical Institute of Bioregulation, Kyushu University, Higashi-ku, Fukuoka 812-8581, Japan
Kouta Mayanagi
JST, PRESTO, Fukuoka 812-8582, Japan
Kouta Mayanagi
Department of Bioscience, Nagahama Institute of BioScience and Technology, 1266 Tamura, Nagahama, 526-0829, Japan
Atsushi Hijikata & Tsuyoshi Shirai
Contributions
N. Takekawa, T.K., K.N. and M.H. designed the experiments; N. Takekawa, N. Terahara, M.G., K.M., A.H., Y.O., T.S. and T.K. performed the experiments; N. Takekawa, N. Terahara, T.K., S.K., T.S, K.N. and M.H. analyzed the data; and N. Takekawa, N. Terahara, T.K., K.N. and M.H. wrote the manuscript.
Competing interests
The authors declare no competing financial interests.
Corresponding authors
Correspondence to Keiichi Namba or Michio Homma.
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