Calculated proton uptake on anaerobic reduction of cytochrome C oxidase: is the reaction electroneutral?
Calculated proton uptake on anaerobic reduction of cytochrome C oxidase: is the reaction electroneutral?
复制标题
计算细胞色素 C 氧化酶厌氧还原的质子摄取：该反应是电中性的吗？
DOI：
10.1021/bi052183d
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发表时间：
2006
期刊：
影响因子：
2.9
通讯作者：
Gunner,MR
中科院分区：
文献类型：
--
作者：
Song,Yifan;Michonova-Alexova,Ekaterina;Gunner,MR
细胞色素氧化酶是一种跨膜质子泵，利用 O2 还原产生的化学能建立电化学梯度。使用多构象连续静电 (MCCE) 在从完全氧化到完全还原的七种厌氧氧化酶氧化还原状态下计算所有残留物的电离态。一个长期存在的问题是质子吸收如何与活性位点双核中心（BNC）的减少相结合。 BNC 有两个辅助因子： hemea3 和 CuB。如果蛋白质需要保持电中性，那么当 BNC 在反应周期的还原半程中被 2 个电子还原时，将结合 2 个质子。在球形红细菌细胞色素氧化酶中评估 BNC 周围可电离残基的有效 pKa。在 pH 7 时，只有与 CuB 配位的氢氧化物才会将其 pKa 从低于 7 变为高于 7，因此当 hemea3 和 CuB 还原时会拾取质子。 Glu I-286、Tyr I-288、His I-334 和 hemea3 上的第二种氢氧化物在所有氧化还原态下的 pKa 均高于 7，尽管它们的去质子化能量成本仅为 1.6−3.5 ΔpKunits。因此，在平衡时，它们被质子化并且不能充当质子受体。 BNC 附近的丙酸被去质子化，pKa 远低于 7。它们在阴离子状态下非常稳定，并且在 BNC 还原时不会结合质子。这表明 BNC 中的电中性在厌氧还原过程中并未保持。 CuA、hemea、hemea3 和 CuB 还原时的质子吸收显示每 4 个电子约 2.5 个质子，这与之前的实验一致。一个质子与 BNC 中的羟基结合，其余质子与远离 BNC 的基团结合。血红蛋白的电化学中点电位 (Em) 是在完全氧化的蛋白质和 BNC 中具有 1 或 2 个电子的情况下计算的。当 BNC 减少时，血红素的 Em 会向下移动，这与之前的实验一致。如果 BNC 还原是电中性的，则 hemeaEmis 与 BNC 氧化还原态无关。
Cytochromecoxidase is a transmembrane proton pump that builds an electrochemical gradient using chemical energy from the reduction of O2. Ionization states of all residues were calculated with Multi-Conformation Continuum Electrostatics (MCCE) in seven anaerobic oxidase redox states ranging from fully oxidized to fully reduced. One long-standing problem is how proton uptake is coupled to the reduction of the active site binuclear center (BNC). The BNC has two cofactors: hemea3and CuB. If the protein needs to maintain electroneutrality, then 2 protons will be bound when the BNC is reduced by 2 electrons in the reductive half of the reaction cycle. The effective pKas of ionizable residues around the BNC are evaluated inRhodobacter sphaeroidescytochromecoxidase. At pH 7, only a hydroxide coordinated to CuBshifts its pKafrom below 7 to above 7 and so picks up a proton when hemea3and CuBare reduced. Glu I-286, Tyr I-288, His I-334, and a second hydroxide on hemea3all have pKas above 7 in all redox states, although they have only 1.6−3.5 ΔpKunits energy cost for deprotonation. Thus, at equilibrium, they are protonated and cannot serve as proton acceptors. The propionic acids near the BNC are deprotonated with pKas well below 7. They are well stabilized in their anionic state and do not bind a proton upon BNC reduction. This suggests that electroneutrality in the BNC is not maintained during the anaerobic reduction. Proton uptake on reduction of CuA, hemea, hemea3, and CuBshows ≈2.5 protons bound per 4 electrons, in agreement with prior experiments. One proton is bound by a hydroxyl group in the BNC and the rest to groups far from the BNC. The electrochemical midpoint potential (Em) of hemeais calculated in the fully oxidized protein and with 1 or 2 electrons in the BNC. TheEmof hemeashifts down when the BNC is reduced, which agrees with prior experiments. If the BNC reduction is electroneutral, then the hemeaEmis independent of the BNC redox state.