| Project/Area Number |
13680689
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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 |
Structural biochemistry
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| Research Institution | Department of Biological Sciences, Graduate School of Science, Osaka University |
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Principal Investigator |
KANAZAWA Hiroshi Osaka Univ. Dept. Biological Sciences, Professor, 大学院・理学研究科, 教授 (50116448)
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| Co-Investigator(Kenkyū-buntansha) |
NAKAMURA Norihiro Osaka Univ. Dept. Biological Sciences, Lecturer, 大学院・理学研究科, 助手 (90324748)
INOUE Hiroki Osaka Univ. Dept. Biological Sciences, Lecturer, 大学院・理学研究科, 助手 (10294448)
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| Project Period (FY) |
2001 – 2002
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| Project Status |
Completed (Fiscal Year 2002)
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| Budget Amount *help |
¥3,500,000 (Direct Cost: ¥3,500,000)
Fiscal Year 2002: ¥2,000,000 (Direct Cost: ¥2,000,000)
Fiscal Year 2001: ¥1,500,000 (Direct Cost: ¥1,500,000)
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| Keywords | Na^+ / H^+ antiporter / Ion transport / Salt tolerance / Active transport / Genetic approach / Intracellular localization / Regulatory protein / 膜タンパク質 / 膜タンパクの細胞内局在化 / 膜蛋白質の精製と再構成 / イオン輸送蛋白質の構想と機能 / 能動輸送の分子機構 / 新規カルシューム結合性蛋白質 / キメラ蛋白質の合成 / イオン輸送分子機構 / H^+交換輸送 / アポトーシスキナーゼ / カルシューム結合蛋白 |
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| Research Abstract |
Maintaining homeostasis of intracellular ion concentrations is the most basic requirement for every living cells. This mechanism is supported by various ion transpoting proteins located in cellular membranes. Na^+/H^+ antiporters among the ion-transporting proteins play a central role to maintain intracellular Na^+ and H^+ concentrations and named Nha or NHE, for bacteria or mammalian cells, respectively. In the present study, we set a final goal of this project to clarify the molecular basis of adaptation mechanism of living cells to various ion environments, especially Na^+ and H^+, by analyzing structure, function and regulation of Na^2/H^+ antiporters from bacteria, yeast and mammalian cells. During 2 years project term, we successfully obtained several new information of Na^+/H^2 antiporters as follows: (1) We clarified transmembrane domains essential for ion transport as well as domains required for pH sensing. We showed that H. pylori antiporter have a specific structure for pH
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sensing which is different from E. coli antiproter. We also successfully purified H. pylon antiporters and reconstituted the antiporter activity in vitro. (2) We showed that various different yeast species have similar Na^+/H^+ (Nhalp) by cloning the genes. We also found that Nhalp has a two-domain structure, comprised of hydrophobic membrane region, and hydrophilic cytoplasmic region which overall structure is close to mammalian NHE. Within this cytoplasmic region, we revealed that domains important for antiport and also intracellular localization exist. Further we identified that a novel membrane protein binds to the domain and enhance the antiporter activity. (3) We identified a novel Ca^<2+> protein (CHP) capable of binding to NHE previously. In this project we identified a new protein kinase and a new kinesin as the binding target of CHP. We studied functional significance of CHP in vitro and in vivo and discussed the multifunction of CHP. Further study is required for understanding intracellular regulatory function of CHP in the future study.
The several different lines of result in terms of Na^+/H^+ antiporters in this project opened a new future study. The results were published as 6 independent full papers. Less
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