Torque generation mechanism of F1-ATPase probed by site-specificand reaction-specific single-molecule manipulation

通过位点特异性和反应特异性单分子操作探测 F1-ATPase 扭矩产生机制

• 批准号: 22247025
• 负责人: NOJI Hiroyuki
• 金额: $ 27.71万
• 依托单位: The University of Tokyo
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (A)
• 财政年份: 2010
• 资助国家: 日本
• 起止时间: 2010 至 2012
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-22247025/
• 关键词: 1 分子操作; 1 分子計測; 分子モーター; 1分子操作; 1分子計測

We determined the rate constants and equilibrium constants of catalytic reactions on F1-ATPase as a function of rotary angle by manipulating individual F1 molecules. The results show ATP-binding process is the major torque-generating step. A novel single-molecule imaging system using polarized scattering light from a gold nanorod that has high spatiotemporal resolution was established to visualize the conformational change of the subunit, the torque-generating unit of F1. It was revealed that the subunit makes a power-stroke type conformational change. High-speed AFM was applied to visualize the conformational change of the subunit in the isolated stator ring with three subunits. subunits showed highly cooperative conformational transition, propagating the conformational state in the anticlockwise direction as the same as the rotor rotation.This finding means that the structural basis is programmed in the stator ring.

我们通过操纵单个F1分子，测定了F1-ATPase上催化反应的速率常数和平衡常数随旋转角的变化。结果表明，ATP结合过程是扭矩产生的主要步骤。建立了一种新的单分子成像系统，利用具有高时空分辨率的金纳米棒的偏振散射光来可视化F1的扭矩产生单元亚基的构象变化。结果表明，该亚基使一个动力冲程型构象变化。利用高速原子力显微镜观察了定子环中三个亚基的构象变化。亚基表现出高度协同的构象转变，构象状态沿与转子旋转相同的反时针方向传播，这一发现意味着结构基础在定子环中编程。

项目成果

A single-molecule enzymatic assay in a directly accessible femtoliter droplet array

• DOI: 10.1039/c0lc00062k
• 发表时间: 2010-01-01
• 期刊: LAB ON A CHIP
• 影响因子: 6.1
• 作者: Sakakihara, Shouichi;Araki, Suguru;Noji, Hiroyuki
• 通讯作者: Noji, Hiroyuki

Winding single-molecule double-stranded DNA on a nanometer-sized reel.

• DOI: 10.1093/nar/gks651
• 发表时间: 2012-10
• 期刊: Nucleic acids research
• 影响因子: 14.9
• 作者: You H;Iino R;Watanabe R;Noji H
• 通讯作者: Noji H

Mechanical Modulation of Catalytic Power of F1-ATPase

F1-ATP酶催化能力的机械调节

• 发表时间: 2012

細胞機能再構成のための2つのナノバイオ技術 ; マイクロドロプレットとFRET型Arpセンサー

两种用于重建细胞功能的纳米生物技术；微滴和FRET型Arp传感器

• 发表时间: 2010
• 作者: 河村正二、松本圭史、平松千尋、松下裕香、小澤範宏、蘆野龍一、中田真紀子、笠木聡、Di Fiore;A.、Aureli;F.、Schaffner;C. M.、Melin;A.;野地博行
• 通讯作者: 野地博行

Stiffness of γ subunit of F1-ATPase

• DOI: 10.1007/s00249-010-0616-9
• 发表时间: 2010-11-01
• 期刊: EUROPEAN BIOPHYSICS JOURNAL WITH BIOPHYSICS LETTERS
• 影响因子: 2
• 作者: Okuno, Daichi;Iino, Ryota;Noji, Hiroyuki
• 通讯作者: Noji, Hiroyuki