Temperature Dependence of Single Molecule Rotation of the Escherichia coli ATP Synthase F1 Sector Reveals the Importance of γ-β Subunit Interactions in the Catalytic Dwell*
Temperature Dependence of Single Molecule Rotation of the Escherichia coli ATP Synthase F1 Sector Reveals the Importance of γ-β Subunit Interactions in the Catalytic Dwell*
复制标题
DOI：
10.1074/jbc.m109.009019
复制
发表时间：
2009-06
期刊：
影响因子：
--
通讯作者：
Mizuki Sekiya;R. Nakamoto;M. Al-Shawi;M. Nakanishi‐Matsui;M. Futai
中科院分区：
文献类型：
--
作者：
Mizuki Sekiya;R. Nakamoto;M. Al-Shawi;M. Nakanishi‐Matsui;M. Futai
使用60-nm金珠在Vmax条件下在低粘滞阻力下观察到F1-ATP酶γ亚基的温度依赖性旋转（Nakanishi-Matsui，M.，Kashiwagi，S.，美国，细川，H.，西普里亚诺，D. J.，Dunn，S. D、韦达，Y.，和Futai，M.（2006）J.Biol.Chem.281，4126-4131）。各个120°步骤的速度的阿耳忒弥斯斜率和旋转步骤之间的停顿长度的倒数非常相似，表明旋转耦合催化循环遵循平坦的能量路径。相反，γ M23 K突变体F1的倒数停顿长度的Arrhenius斜率显著增加，而旋转速率的Arrhenius斜率与野生型相似。相互作用的定子亚基中转子γ M23 K取代的效应和β E381 D突变的抵消效应表明转子-定子相互作用在储存弹性能的利用中起着关键作用。γ M23 K酶必须克服一个突然的活化能障碍，迫使它进入一个不太有利的途径，导致从旋转解偶联催化。
The temperature-dependent rotation of F1-ATPase γ subunit was observed in Vmax conditions at low viscous drag using a 60-nm gold bead (Nakanishi-Matsui, M., Kashiwagi, S., Hosokawa, H., Cipriano, D. J., Dunn, S. D., Wada, Y., and Futai, M. (2006) J. Biol. Chem. 281, 4126–4131). The Arrhenius slopes of the speed of the individual 120° steps and reciprocal of the pause length between rotation steps were very similar, indicating a flat energy pathway followed by the rotationally coupled catalytic cycle. In contrast, the Arrhenius slope of the reciprocal pause length of the γM23K mutant F1 was significantly increased, whereas that of the rotation rate was similar to wild type. The effects of the rotor γM23K substitution and the counteracting effects of βE381D mutation in the interacting stator subunits demonstrate that the rotor-stator interactions play critical roles in the utilization of stored elastic energy. The γM23K enzyme must overcome an abrupt activation energy barrier, forcing it onto a less favored pathway that results in uncoupling catalysis from rotation.