Modulation of the active site conformation by site-directed mutagenesis in cytochrome c oxidase from Paracoccus denitrificans.
Modulation of the active site conformation by site-directed mutagenesis in cytochrome c oxidase from Paracoccus denitrificans.
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DOI：
10.1016/j.jinorgbio.2009.11.011
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发表时间：
2010-03
影响因子：
3.9
通讯作者：
Rousseau, Denis L.
中科院分区：
文献类型：
--
作者：
Ji, Hong;Das, Tapan K.;Puustinen, Anne;Wikstrom, Marten;Yeh, Syun-Ru;Rousseau, Denis L.
利用共振拉曼光谱研究了副球菌细胞色素c氧化酶活性位点突变体（PdCcO）的结构和功能特性。基于Fe-CO伸缩振动模式和低频血红素振动模式，确定了酶中处于平衡状态的两种构象（α-和β-形式）。α-构象异构体是野生型酶中的优势物种，与β-构象异构体相比，α-构象异构体具有更短的血红素a3铁-CuB距离和更扭曲的血红素，β-构象异构体具有更松弛和开放的远端口袋。一般来说，突变导致α-构象异构体的数量减少，这伴随着催化活性的降低，表明α-构象异构体是酶的活性形式。这些数据表明，天然结构的酶是在一个微妙的平衡的分子内相互作用。我们提出了一个模型，其中突变使α-构象不稳定，相对于β-构象，并提高了两种构象之间相互转换的活化势垒。在CcO的构象空间中的两种构象的可及性在酶的催化循环过程中的氧化还原反应与质子易位的耦合中起着至关重要的作用。
The structural and functional properties of active site mutants of cytochrome c oxidase from Paracoccus denitrificans (PdCcO) were investigated with resonance Raman spectroscopy. Based on the Fe-CO stretching modes and low frequency heme modes, two conformers (α- and β-forms) were identified that are in equilibrium in the enzyme. The α-conformer, which is the dominant species in the wild type enzyme, has a shorter heme a3 iron-CuB distance and a more distorted heme, as compared to the β-conformer, which has a more relaxed and open distal pocket. In general, the mutations caused a decrease in the population of the α-conformer, which is concomitant with a decreased in the catalytic activity, indicating that the α-conformer is the active form of the enzyme. The data suggest that the native structure of the enzyme is in a delicate balance of intramolecular interactions. We present a model in which the mutations destabilize the α-conformer, with respect to the β-conformer, and raise the activation barrier for the inter-conversion between the two conformers. The accessibility of the two conformers in the conformational space of CcO plausibly plays a critical role in coupling the redox reaction to proton translocation during the catalytic cycle of the enzyme.
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影响因子：
3
作者：
Pfitzner, U;Odenwald, A;Richter, OMH
通讯作者：
Richter, OMH
影响因子：
2.9
作者：
Qin, Ling;Liu, Jian;Ferguson-Miller, Shelagh
通讯作者：
Ferguson-Miller, Shelagh
影响因子：
3.4
作者：
Das, TK;Tomson, FL;Rousseau, DL
通讯作者：
Rousseau, DL
影响因子：
64.8
作者：
IWATA, S;OSTERMEIER, C;MICHEL, H
通讯作者：
MICHEL, H
DOI：
10.1073/pnas.78.1.234
发表时间：
1981-01-01
期刊：
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA-BIOLOGICAL SCIENCES
影响因子：
--
作者：
ALBEN, JO;MOH, PP;ALTSCHULD, RA
通讯作者：
ALTSCHULD, RA