Phosphate release in F1-ATPase catalytic cycle follows ADP release
Phosphate release in F1-ATPase catalytic cycle follows ADP release
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DOI：
10.1038/nchembio.443
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发表时间：
2010-11-01
影响因子：
14.8
通讯作者：
Noji, Hiroyuki
中科院分区：
文献类型：
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作者：
Watanabe, Rikiya;Iino, Ryota;Noji, Hiroyuki
F-1-ATP酶是一种ATP驱动的旋转马达蛋白，其中γ亚基逆着催化定子环旋转。虽然F-1的反应机制已基本被揭示，但无机磷酸盐（P-i）释放的时间仍然存在争议。在这里，我们通过用磁镊子验证ATP在被捕的F-1上水解的可逆性来解决这个问题。发现ATP水解基本上是可逆的，这意味着在γ旋转和ADP释放后释放P-i，尽管在ATP水解角下发现极慢的P-i释放作为解偶联副反应。在此基础上，我们推导了F-1的化学机械耦合方案。我们发现，对P-i的亲和力是强烈的角度依赖性的，这意味着P-i释放对扭矩产生有很大的贡献。这些发现意味着在ATP合成条件下，P-i结合到空的催化位点，阻止溶液ATP（尽管不是ADP）结合。因此，这支持了选择性ADP结合用于有效ATP合成的概念。
F-1-ATPase is an ATP-driven rotary motor protein in which the gamma-subunit rotates against the catalytic stator ring. Although the reaction scheme of F-1 has mostly been revealed, the timing of inorganic phosphate (P-i) release remains controversial. Here we addressed this issue by verifying the reversibility of ATP hydrolysis on arrested F-1 with magnetic tweezers. ATP hydrolysis was found to be essentially reversible, implying that P-i is released after the gamma rotation and ADP release, although extremely slow P-i release was found at the ATP hydrolysis angle as an uncoupling side reaction. On the basis of this finding, we deduced the chemomechanical coupling scheme of F-1. We found that the affinity for P-i was strongly angle dependent, implying a large contribution by P-i release to torque generation. These findings imply that under ATP synthesis conditions, P-i binds to an empty catalytic site, preventing solution ATP (though not ADP) from binding. Thus, this supports the concept of selective ADP binding for efficient ATP synthesis.