Elucidation of molecular mechanism in ATP-depending translation activity in plant seed cells
| Project/Area Number |
16K14665
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| Research Category |
Grant-in-Aid for Challenging Exploratory Research
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| Allocation Type | Multi-year Fund |
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| Research Field |
Molecular biology
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| Research Institution | Saitama University |
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Principal Investigator |
Tozawa Yuzuru 埼玉大学, 理工学研究科, 教授 (90363267)
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| Research Collaborator |
Suzuki Kakeru
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| Project Period (FY) |
2016-04-01 – 2019-03-31
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| Project Status |
Completed (Fiscal Year 2018)
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| Budget Amount *help |
¥3,640,000 (Direct Cost: ¥2,800,000、Indirect Cost: ¥840,000)
Fiscal Year 2018: ¥1,040,000 (Direct Cost: ¥800,000、Indirect Cost: ¥240,000)
Fiscal Year 2017: ¥1,040,000 (Direct Cost: ¥800,000、Indirect Cost: ¥240,000)
Fiscal Year 2016: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000)
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| Keywords | 無細胞タンパク質合成 / イネ / GTP / カルス / 翻訳 / 翻訳因子 / 植物 / コムギ / 酵母 / 蛋白質 / 生体分子 / 遺伝子 / 酵素 |
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| Outline of Final Research Achievements |
In this study, we have established a new plant cell-free translation system based on rice callus extracts. We revealed that the rice cell-free system allows efficient protein expression in the absence of GTP. We confirmed generation of low level GTP from GMP, which is degradation product of mRNA by endogenous nuclease. On the other hand, By using polyuridylic acid (polyU), which contains only uridine but not guanine nucleotide, as a template, we demonstrated that translation elongation activity of the system is not affected by the lack of GTP in the reaction. This GTP-independence in the translation elongation activity is different from yeast cell-free system, and seems to be unique to plant seed cell system.
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| Academic Significance and Societal Importance of the Research Achievements |
新たに確立したイネカルス無細胞翻訳系は、1 mlあたり数百マイクログラム相当のタンパク質合成が可能で、精製用ヒスチジンタグの利用による合成タンパク質の精製も可能であり、生化学的機能解析用のタンパク質合成系に実用化できる生産性を有すると考える。また、薬害試験の対象となるヒト心筋カリウムチャネルタンパク質hERGの合成について検証したところ、脂質二分子膜ベシクル添加条件で、hERGとその再構成促進因子との共合成により、活性型hERGの効率的再構成系の構築が可能であることを確認した。このように、確立したシステムが、創薬関連タンパク質の機能型再構成系の構築にも資することを確認できた。
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Report
(4 results)
Research Products
(9 results)
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[Journal Article] Mechanically stable solvent-free lipid bilayers in nano- and micro-tapered apertures for reconstitution of cell-free synthesized hERG channels.2017
Author(s)
Daisuke Tadaki, Daichi Yamaura, Shun Araki, Miyu Yoshida, Kohei Arata, Takeshi Ohori, Ken-ichi Ishibashi, Miki Kato, Teng Ma, Ryusuke Miyata, Yuzuru Tozawa, Hideaki Yamamoto, Michio Niwano, and Ayumi Hirano-Iwata
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Journal Title
Scientific Report
Volume: 7
Issue: 1
Pages: 17736-17736
DOI
Related Report
Peer Reviewed / Open Access
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