Brzezinski Peter, Johansson Ann-Louise
Department of Biochemistry and Biophysics, The Arrhenius Laboratories for Natural Sciences, Stockholm University, SE-106 91 Stockholm, Sweden.
Biochim Biophys Acta. 2010 Jun-Jul;1797(6-7):710-23. doi: 10.1016/j.bbabio.2010.02.020. Epub 2010 Feb 23.
Cytochrome c oxidase is a multisubunit membrane-bound enzyme, which catalyzes oxidation of four molecules of cytochrome c2+ and reduction of molecular oxygen to water. The electrons are taken from one side of the membrane while the protons are taken from the other side. This topographical arrangement results in a charge separation that is equivalent to moving one positive charge across the membrane for each electron transferred to O2. In this reaction part of the free energy available from O2 reduction is conserved in the form of an electrochemical proton gradient. In addition, part of the free energy is used to pump on average one proton across the membrane per electron transferred to O2. Our understanding of the molecular design of the machinery that couples O2 reduction to proton pumping in oxidases has greatly benefited from studies of so called "uncoupled" structural variants of the oxidases. In these uncoupled oxidases the catalytic O2-reduction reaction may display the same rates as in the wild-type CytcO, yet the electron/proton transfer to O2 is not linked to proton pumping. One striking feature of all uncoupled variants studied to date is that the (apparent) pKa of a Glu residue, located deeply within a proton pathway, is either increased or decreased (from 9.4 in the wild-type oxidase). The altered pKa presumably reflects changes in the local structural environment of the residue and because the Glu residue is found near the catalytic site as well as near a putative exit pathway for pumped protons these changes are presumably important for controlling the rates and trajectories of the proton transfer. In this paper we summarize data obtained from studies of uncoupled structural oxidase variants and present a hypothesis that in quantitative terms offers a link between structural changes, modulation of the apparent pKa and uncoupling of proton pumping from O2 reduction.
细胞色素c氧化酶是一种多亚基膜结合酶，它催化四分子细胞色素c2+的氧化以及分子氧还原为水的反应。电子从膜的一侧获取，而质子从另一侧获取。这种拓扑结构导致电荷分离，相当于每将一个电子转移到O2时，就有一个正电荷穿过膜。在这个反应中，O2还原所释放的部分自由能以电化学质子梯度的形式被保存下来。此外，部分自由能用于每将一个电子转移到O2时平均将一个质子泵过膜。我们对氧化酶中使O2还原与质子泵浦相偶联的机制的分子设计的理解，很大程度上受益于对氧化酶所谓“解偶联”结构变体的研究。在这些解偶联的氧化酶中，催化性的O2还原反应可能与野生型细胞色素c氧化酶具有相同的速率，但电子/质子向O2的转移与质子泵浦不相关联。迄今为止研究的所有解偶联变体的一个显著特征是，位于质子传导途径深处的一个Glu残基的（表观）pKa要么升高要么降低（相对于野生型氧化酶中的9.4）。pKa的改变大概反映了该残基局部结构环境的变化，并且由于Glu残基既位于催化位点附近，又位于推测的泵出质子的出口途径附近，所以这些变化大概对于控制质子转移的速率和轨迹很重要。在本文中，我们总结了从解偶联结构氧化酶变体研究中获得的数据，并提出了一个假设，该假设从定量角度提供了结构变化、表观pKa的调节以及质子泵浦与O2还原解偶联之间的联系。
