Powers L, Chance B
J Inorg Biochem. 1985 Mar-Apr;23(3-4):207-17. doi: 10.1016/0162-0134(85)85027-3.
X-ray absorption studies have been used to investigate the structure of the four redox centers (2Fe, 2Cu) of the terminal enzyme in the respiratory chain, cytochrome c oxidase in the resting oxidized form as well as in the functional intermediates that are freeze-trapped. Methods of x-ray fluorescence detection for these low-concentration samples together with low-temperature cryostats and simultaneous optical monitoring were developed to ensure good signal-to-noise data and sample integrity. The resting oxidized form contains a sulfur bridge between the copper and iron of the active site which are separated by approximately 3.8 A. This separation of the active site metal atoms was uniquely identified by comparison of both the iron and copper EXAFS data and iron EXAFS of the copper-depleted enzyme. In the reduced state, the CO or O2 is bound to the active site iron having a structure identical to CO or oxy hemoglobin while the sulfur remains with the active site copper. Little change in structure is observed for the other iron and copper. It is the sulfur bridged active site form that is isolated by the Yonetani and Caughy methods with greater than or equal to 85% homogeneity but not the Hartzell-Beinert or similar methods. Another form observed in the redox cycle is also fully oxidized but lacks the sulfur bridged active site with the iron of the active site having a structure identical to that of the peroxidases. This form exhibits peroxidase as well as oxidase activity, and a stable intermediate is formed with hydrogen and ethylhydrogen peroxide in which the iron of the active site is structurally similar to that of the peroxidase intermediate. The active site copper, however, does not participate in the peroxidatic role and the structures of the other iron and copper are identical to those of the sulfur bridged resting oxidized form. Thus this unique enzyme has peroxidase activity which may serve to safeguard its main oxidase function.
X射线吸收研究已被用于研究呼吸链末端酶细胞色素c氧化酶的四个氧化还原中心（2Fe、2Cu）的结构，包括其静止氧化形式以及冷冻捕获的功能中间体。针对这些低浓度样品开发了X射线荧光检测方法，结合低温恒温器和同步光学监测，以确保获得良好的信噪比数据和样品完整性。静止氧化形式在活性位点的铜和铁之间含有一个硫桥，它们之间的距离约为3.8埃。通过比较铁和铜的扩展X射线吸收精细结构（EXAFS）数据以及铜耗尽酶的铁EXAFS，独特地确定了活性位点金属原子的这种分离。在还原状态下，CO或O2与活性位点的铁结合，其结构与CO或氧合血红蛋白相同，而硫则与活性位点的铜结合。其他铁和铜的结构变化不大。通过米谷和考希方法分离出的硫桥连活性位点形式的同质性大于或等于85%，而哈茨尔-贝纳特或类似方法则不行。在氧化还原循环中观察到的另一种形式也是完全氧化的，但缺乏硫桥连活性位点，活性位点的铁具有与过氧化物酶相同的结构。这种形式表现出过氧化物酶和氧化酶活性，并且与氢和乙基过氧化氢形成稳定的中间体，其中活性位点的铁在结构上与过氧化物酶中间体相似。然而，活性位点的铜不参与过氧化物作用，其他铁和铜的结构与硫桥连静止氧化形式的结构相同。因此，这种独特的酶具有过氧化物酶活性，这可能有助于保护其主要的氧化酶功能。
