ATP synthases play a crucial role in energy production by utilizing the proton motive force (pmf) across the membrane to rotate their membrane-embedded rotor c-ring, and thus driving ATP synthesis in the hydrophilic catalytic hexamer. However, the mechanism of how pmf converts into c-ring rotation remains unclear. This study presents a 2.8 Å cryo-EM structure of the V_o domain of V/A-ATPase from Thermus thermophilus, revealing precise orientations of glutamate (Glu) residues in the c₁₂-ring. Three Glu residues face a water channel, with one forming a salt bridge with the Arginine in the stator (a/Arg). Molecular dynamics (MD) simulations show that protonation of specific Glu residues triggers unidirectional Brownian motion of the c₁₂-ring towards ATP synthesis. When the key Glu remains unprotonated, the salt bridge persists, blocking rotation. These findings suggest that asymmetry in the protonation of c/Glu residues biases c₁₂-ring movement, facilitating rotation and ATP synthesis.

ATP 合酶在能量产生中起着至关重要的作用，它利用跨膜的质子动力 （pmf） 来旋转其膜嵌入的转子 C 环，从而在亲水催化六聚体中驱动 ATP 合成。然而，pmf 如何转化为 c 环旋转的机制仍不清楚。本研究提出了来自嗜热热藻的 V/A-ATP 酶 V_o 结构域的 2.8 Å 冷冻电镜结构，揭示了 c₁₂ 环中谷氨酸 （Glu） 残基的精确方向。三个 Glu 残基面向水通道，其中一个残基与定子中的精氨酸 （a/Arg） 形成盐桥。分子动力学 （MD） 模拟表明，特定 Glu 残基的质子化触发 c₁₂ 环向 ATP 合成的单向布朗运动。当键 Glu 保持未质子化时，盐桥会持续存在，从而阻止旋转。这些发现表明，c/Glu 残基质子化的不对称性偏向于 c₁₂ 环运动，促进旋转和 ATP 合成。