2002 Fiscal Year Final Research Report Summary
Developing an E. coli-expression-function platform for study of 3-D structure-function relationships of channel molecules
| Project/Area Number |
13680737
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Biophysics
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| Research Institution | Fukui Medical University |
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Principal Investigator |
OIKI Shigetoshi Fukui Medical University, Faculty of Medicine, Professor, 医学部, 教授 (10185176)
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| Co-Investigator(Kenkyū-buntansha) |
YAMADA Katsuyuki Fukui Medical University, Faculty of Medicine, Research Associate, 医学部, 助手 (20313744)
SHIMIZU Horofumi Fukui Medical University, Faculty of Medicine, Research Associate, 医学部, 助手 (50324158)
KONNO Takashi Fukui Medical University, Faculty of Medicine, Associate Professor, 医学部, 助教授 (50225637)
YABE Isao Tokyo University, Institute of Molecular and Cellular Biosciences, Assistant Professor, 分子細胞生物学研究所, 助手 (90111575)
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| Project Period (FY) |
2001 – 2002
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| Keywords | Ion channel / KcsA channel / patch clamp technique / planar lipid bilayer method / single-channel recording / E. coli / whole-cell recording / liposome |
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| Research Abstract |
To elucidate structure-function relationships of ion channels a novel experimental platform is developed. So far only ion channels originated from prokaryotes were successfully crystallized. To study such targets more effectively, electrophysiological recording system for ion channels from prokaryotes is critically important. Based on Yabe's method for current recordings from giant E. coli, channel recording techniques were explored. Many technical problems arose for recording whole-cell current from giant bacteria. These problems have been mostly solved, although a few problems still remain.
KcsA channel was expressed and recorded by inside-out excised patch methods from giant E. coli. Reconstituted KcsA channel into liposome was also recorded by a modified tip-dip method. On the other hand, whole-cell current of KcsA channel could not be observed from giant bacteria. The discrepancy between successful single channel recordings and unsuccessful whole-cell recordings poses fundamental issue on functional role of ion channels on bacteria.
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[Publications] Horie, T., Yoshida, K., Nakayama, H., Yamada, K., Oiki, S. and Shinmyo, A.: "Two types of HKT transporters with different properties of Na^+ and K^+ transport in Oryza sativa"Plant J.. 27. 129-138 (2001)
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[Publications] Maeser, P., Hosoo, Y., Goshima, S., Horie, T., Eckelman, B., Yamada, K., Yoshida, K., Bakker, E.P., Shinmyo, A., Oiki, S., Schroeder, J.I., and Uozumi, N.: "Glycine residues in potassium-channel-like selectivity filters determine potassium selectivity in four-loop-per-subunit HKT transporters from plants"Proc.Natl.Acad.Sci.USA. 99. 6428-6433 (2002)
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「研究成果報告書概要(欧文)」より
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[Publications] Berthomieu, P., Nublat, A., Brackenbury, W.J., Lambert, C., Savio, C., Uozumi, N., Oiki, S., Yamada, K., Conejero, G., Cellier, F., Gosti, F., Simonneau, T., Very, A.-A., Sentenac, H. and Casse, F.: "Functional analysis of AtHKT1 in Arabidopsis shows that Na^+ recirculation by the phloem is crucial for salt tolerance"EMBO J.. 22. 2004-2014 (2003)
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[Publications] Futaki, S., Zhang, Y., Kiwada, T., Yagami, T., Oiki, S. and Sugiura, Y.: "Gramicidin-based Channel Systems for the Detection of Protein-ligand Interaction"Bioorg.Med.Chem.. In press.
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「研究成果報告書概要(欧文)」より
