Rotary coupling mechanism of two nano motors which constitute ATP Synthase

构成ATP合酶的两个纳米电机的旋转耦合机构

• 批准号: 13308036
• 负责人: YOSHIDA Masasuke
• 金额: $ 28.87万
• 依托单位: Tokyo Institute of Technology
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (A)
• 财政年份: 2001
• 资助国家: 日本
• 起止时间: 2001 至 2003
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13308036/
• 关键词: ATP Synthase; Molecular Motor; F_1-ATPase; F_0 channel; ATPase; 回転モーター; ATP; F0F1; ATP合成; F1-ATPアーゼ; 回転

Major results are as follows.1)A regulatory subunit, ε, binds ATP and it changes its conformation depending on the protonic electrochemical potential difference across the membrane.2)By introducing Trp residue into the vicinity of the catalytic site on β subunits, we could clarify the origin of apparent negative cooperativity.3)ATP binding per se drives 80 degree step motion before hydrolysis on the enzyme.4)By using fluorescent ATP analogue, we could directly observe ATP binding and rotation simultaneously We found that once ATP binds, it remains bound until γ subunit rotates 240 degree.5)By using magnetic beads and forcing F_1-ATPase to rotate in synthetic direction, we could show continuous ATP synthesis.

主要结果如下。1）一个调节亚基ε结合ATP，并根据整个膜的质子电化学势差而改变其构象。2）通过将TRP居住地引入催化位点附近β亚基的附近，我们可以在80上启用PRINGIND PRINGIND aTP iSS SE SE SE SE SE SE SE SE SE SE SE SE SERISS。使用荧光ATP类似物，我们可以直接观察ATP结合和旋转，简单地发现，一旦ATP结合，它一直结合，直到γ亚基旋转240度。5）通过使用磁珠并迫使F_1-ATPase沿合成方向旋转，我们可以表现出连续的ATP合成。

项目成果

Tsunoda, S.P., Rodgers, A.J.W., Ageller R, Wilce, M.C.J., Yoshida, M., Capaldi, R.A.: "Conformational changes of the ε subunit in the bacterial F_1F_0 ATP synthase provide a ratchet action to regulate this rotary motor enzyme."Proc.Natl.Acad.Sci.USA. 98.

Tsunoda, S.P.、Rodgers, A.J.W.、Ageller R、Wilce, M.C.J.、Yoshida, M.、Capaldi, R.A.：“细菌 F_1F_0 ATP 合酶中 ε 亚基的构象变化提供了棘轮作用来调节这种旋转马达酶。”Proc .美国国家科学院98。

Yokoyama, K., Nakano, M., Imamura, H., Yoshida, M., Tamakoshi, M.: "Subunit Arrangement in V-ATPase from Thermus thermophilus"J.Biol.Chem. 278. 24255-24258 (2003)

Yokoyama, K.、Nakano, M.、Imamura, H.、Yoshida, M.、Tamakoshi, M.：“嗜热栖热菌 V-ATP 酶中的亚基排列”J.Biol.Chem。

Bald D, Noji H, Yosbida M, Hirono-Hara Y, Hisabori T.: "Redox regulation of the rotation of F_1-ATP synthase."J Biol Chem.. 276. 39505-39507 (2001)

Bald D、Noji H、Yosbida M、Hirono-Hara Y、Hisabori T.：“F_1-ATP 合酶旋转的氧化还原调节。”J Biol Chem.. 276. 39505-39507 (2001)

Kato-Yamada, Y., Yoshida, M.: "Isolated ε Subunit of Thermophilic F_1-ATPase Binds ATP"J.Biol.Chem.. 278. 36013-36016 (2003)

Kato-Yamada, Y., Yoshida, M.：“嗜热 F_1-ATP 酶结合 ATP 的分离 ε 亚基”J.Biol.Chem.. 278. 36013-36016 (2003)

Shimabukuro, K., Yasuda, Y., Muneyuki, E,.Hara, K.Y., Kinosita.K.Jr., Yoshida, M.: "Catalysis and rotation of F_1 motor ; cleavage of ATP at the catalytic site occurs in 1 ms before 40 degree substep rotation."Proc Natl Acad Sci USA. 100. 14731-14736 (200

Shimabukuro, K., Yasuda, Y., Muneyuki, E,.Hara, K.Y., Kinosita.K.Jr., Yoshida, M.：“F_1 电机的催化和旋转；催化位点 ATP 的裂解发生在 1 ms 内