Cai Xiuhong, Haider Kamran, Lu Jianxun, Radic Slaven, Son Chang Yun, Cui Qiang, Gunner M R
Department of Physics, City College of New York, 160 Convent Avenue, New York, NY 10031, USA; Department of Physics, Graduate Center, City University of New York, 365 Fifth Avenue, New York, NY 10016, USA.
Department of Physics, City College of New York, 160 Convent Avenue, New York, NY 10031, USA.
Biochim Biophys Acta Bioenerg. 2018 Oct;1859(10):997-1005. doi: 10.1016/j.bbabio.2018.05.010. Epub 2018 May 18.
Cytochrome c Oxidase (CcO) reduces O, the terminal electron acceptor, to water in the aerobic, respiratory electron transport chain. The energy released by O reductions is stored by removing eight protons from the high pH, N-side, of the membrane with four used for chemistry in the active site and four pumped to the low pH, P-side. The proton transfers must occur along controllable proton pathways that prevent energy dissipating movement towards the N-side. The CcO N-side has well established D- and K-channels to deliver protons to the protein interior. The P-side has a buried core of hydrogen-bonded protonatable residues designated the Proton Loading Site cluster (PLS cluster) and many protonatable residues on the P-side surface, providing no obvious unique exit. Hydrogen bond pathways were identified in Molecular Dynamics (MD) trajectories of Rb. sphaeroides CcO prepared in the P state with the heme a propionate and Glu286 in different protonation states. Grand Canonical Monte Carlo sampling of water locations, polar proton positions and residue protonation states in trajectory snapshots identify a limited number of water mediated, proton paths from PLS cluster to the surface via a (P-exit) cluster of residues. Key P-exit residues include His93, Ser168, Thr100 and Asn96. The hydrogen bonds between PLS cluster and P-exit clusters are mediated by a water wire in a cavity centered near Thr100, whose hydration can be interrupted by a hydrophobic pair, Leu255B (near Cu) and Ile99. Connections between the D channel and PLS via Glu286 are controlled by a second, variably hydrated cavity. SIGNIFICANCE STATEMENT: Cytochrome C oxidase plays a crucial role in cellular respiration and energy generation. It reduces O to water and uses the released free energy to move protons across mitochondrial and bacterial cell membranes adding to the essential electrochemical gradient. Energy storage requires that protons are taken up from the high pH, N-side and released to the low pH, P-side of the membrane. We identify a potential proton exit from a buried cluster of polar residues (the proton loading site) to the P-side of CcO via paths made up of waters and conserved residues. Two water cavities connect the proton exit pathway to the surface only when hydrated. Changing the degree of hydration may control otherwise energetically favorable proton backflow from the P-side.
细胞色素c氧化酶（CcO）在需氧呼吸电子传递链中将末端电子受体O₂还原为水。O₂还原所释放的能量通过从膜的高pH值N侧移除八个质子来储存，其中四个用于活性位点的化学反应，另外四个被泵送到低pH值的P侧。质子转移必须沿着可控的质子通道进行，以防止能量耗散地向N侧移动。CcO的N侧有成熟的D通道和K通道，用于将质子输送到蛋白质内部。P侧有一个由氢键连接的可质子化残基组成的埋藏核心，称为质子负载位点簇（PLS簇），并且P侧表面有许多可质子化残基，没有明显的独特出口。在处于P状态且血红素a丙酸酯和Glu286处于不同质子化状态的球形红细菌CcO的分子动力学（MD）轨迹中确定了氢键通道。对轨迹快照中的水位置、极性质子位置和残基质子化状态进行巨正则蒙特卡罗采样，确定了从PLS簇通过一组（P出口）残基到表面的有限数量的水介导质子路径。关键的P出口残基包括His93、Ser168、Thr100和Asn96。PLS簇和P出口簇之间的氢键由位于Thr100附近中心的一个腔中的水线介导，其水合作用可被一对疏水残基Leu255B（靠近Cu）和Ile99中断。通过Glu286连接D通道和PLS的连接由第二个可变水合腔控制。
细胞色素C氧化酶在细胞呼吸和能量产生中起关键作用。它将O₂还原为水，并利用释放的自由能将质子跨线粒体和细菌细胞膜移动，增加基本的电化学梯度。能量储存要求质子从膜的高pH值N侧吸收并释放到低pH值的P侧。我们确定了一个潜在的质子出口，从一个埋藏的极性残基簇（质子负载位点）通过由水和保守残基组成的路径到达CcO的P侧。只有在水合时，两个水腔才将质子出口通道连接到表面。改变水合程度可能控制原本在能量上有利的质子从P侧的回流。
