X-ray absorption studies on the mixed-valence and fully reduced forms of the soluble Cu-A domains of cytochrome c oxidase
X-ray absorption studies on the mixed-valence and fully reduced forms of the soluble Cu-A domains of cytochrome c oxidase
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DOI：
10.1021/ja970513e
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发表时间：
1997-07-02
影响因子：
15
通讯作者：
Fee, JA
中科院分区：
文献类型：
--
作者：
Blackburn, NJ;deVries, S;Fee, JA
细胞色素氧化酶是原核和真核细胞中的末端氧化酶，负责通过氧化磷酸化过程产生细胞能量：该酶包含两个 Fe 和三个 Cu 中心，它们共同提供将 O-2 还原为水的氧化还原机制。最近，X射线晶体学首次提供了金属中心配位球的三维描述：然而，结构以低分辨率显示金属位点，为了充分理解反应机理，需要以更高的精度确定度量细节（键长和角度）。 X 射线吸收光谱的独特之处在于其提供此类细节的能力，我们已应用该技术来确定 Cu-A 中心的结构，这是一种硫醇桥联的双核铜簇，其中铜由两个半胱氨酸配体桥接，并且具有类似于 2.4 埃的极短 Cu-Cu 距离。 X射线吸收光谱先前预测了较短的Cu-Cu距离，现已用于进一步细化嗜热栖热菌和枯草芽孢杆菌酶的混合价和完全还原形式中该位点的结构细节。结果将 Cu-A 核的结构定义为 Cu2S2 金刚石，Cu-S 键长为 2.3 埃，Cu-Cu = 2.44 埃，Cu-S-Cu 角为 65 度。一电子还原仅产生核心几何形状的微小变化，Cu-S和Cu-Cu键长分别增加到2.33和2.51埃，但Cu-S-Cu角保持在65度不变。异常高的 Cu-S Debye-Waller 项意味着 Cu2S2 金刚石核中存在因不等 Cu-S 键长而产生的显着不对称性。德拜-沃勒因子的度量参数和温度依赖性都表现出混合价和完全还原蛋白质之间的细微差异，这表明短距离可能部分是弱金属-金属键的结果。结果表明，不寻常的 Cu-A 簇的功能是提供一个在电子转移过程中发生结构扰动最小的位点。因此，它们为 Cu-A 中心观察到的极低重组能 lambda 提供了极好的合理化。
Cytochrome oxidase is the terminal oxidase in both prokaryotic and eukaryotic cells and is responsible for the generation of cellular energy via the process known as oxidative phosphorylation: The enzyme contains two Fe and three Cu centers which together provide the redox machinery for the reduction of O-2 to water. Recently, X-ray crystallography has provided the first three-dimensional description of the coordination spheres of the metal centers: However, the structures show the metal sites at low resolution, and in order to fully understand the mechanism of the reaction, it is desirable to determine the metrical details (bond lengths and angles) to much higher precision. X-ray absorption spectroscopy is unique in its ability to provide such detail, and we have applied the technique to determining the structure of the Cu-A center, a thiolate-bridged binuclear copper cluster in which the coppers are bridged by two cysteine ligands and have an extremely short Cu-Cu distance of similar to 2.4 Angstrom. X-ray absorption spectroscopy, which had previously predicted the short Cu-Cu distance, has been used to further refine the structural details of the site in both the mixed-valence and fully reduced forms of the enzymes from Thermus thermophilus and Bacillus subtilis. The results have defined the structure of the Cu-A core as a Cu2S2 diamond with Cu-S bond lengths of 2.3 Angstrom, Cu-Cu = 2.44 Angstrom, and very acute Cu-S-Cu angles of 65 degrees. One-electron reduction produces only minor changes in the core geometry, with the Cu-S and Cu-Cu bond lengths increasing to 2.33 and 2.51 Angstrom, respectively, but with the Cu-S-Cu angle remaining unchanged at 65 degrees. The unusually high Cu-S Debye-Waller terms imply that there is significant asymmetry in the Cu2S2 diamond core derived from inequivalent Cu-S bond lengths. Both the metrical parameters and the temperature dependence of the Debye-Waller factors exhibit subtle differences between the mixed-valence and fully reduced proteins which suggest that the short distance may be the result, in part, of a weak metal-metal bond. The results suggest that the function of the unusual Cu-A cluster is to provide a site with minimal structural perturbation occurring during electron transfer. Thus, they provide an excellent rationalization for the very low reorganizational energy, lambda, observed for the Cu-A center.