A new electron transport mechanism in mitochondrial steroid hydroxylase systems based on structural changes upon the reduction of adrenodoxin
A new electron transport mechanism in mitochondrial steroid hydroxylase systems based on structural changes upon the reduction of adrenodoxin
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DOI：
10.1021/bi0160361
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发表时间：
2002-06-25
期刊：
影响因子：
2.9
通讯作者：
Rüterjans, H
中科院分区：
文献类型：
--
作者：
Beilke, D;Weiss, R;Rüterjans, H
肾上腺铁氧还蛋白是一种酸性的14.4 kDa[2Fe-2S]铁氧还蛋白，属于脊椎动物铁氧还蛋白家族。它参与黄素酶NADPH-肾上腺素还原酶向细胞色素P-450(SCC)和P-450(11beta)的电子传递。氧化还原伙伴之间在电子传递过程中的相互作用尚未完全确定。确定电子转移蛋白的三级结构可能有助于理解电子转移的机制。在目前的工作中，我们报告了用高分辨核磁共振波谱研究牛肾上腺素(BAdx)在溶液中的氧化和还原形式的结构。在大肠杆菌中表达该蛋白，分别用N-15或C-13/N-15进行单标和双标。约70%的N-15、C-13和H-1共振可分别归属于还原的和氧化的bAdx。利用NOE距离信息确定了还原和氧化态的二级结构和三级结构。H-1(N)-T-1，某些残基的驰豫时间被用来获得对[2Fe-2S]团簇的额外距离约束。结果表明，氧化态ADX的溶液结构与X-射线结构十分相似。然而，如核磁共振测量所示，当[2Fe-2S]团簇减少时，结构发生变化。可以证明，这些构象变化，特别是在C-末端区域，导致ADX二聚体在还原时解离。提出了一种新的电子传输机制，该机制通过一种改进的穿梭机制进行，其中单体和二聚体同时作为电子载体。
The adrenal ferredoxin (adrenodoxin, Adx) is an acidic 14.4-kDa [2Fe-2S] ferredoxin that belongs to the vertebrate ferredoxin family. It is involved in the electron transfer from the flavoenzyme NADPH-adrenodoxin-reductase to cytochromes P-450(scc) and P-450(11beta). The interaction between the redox partners during electron transport has not yet been fully established. Determining the tertiary structure of an electron-transfer protein may be very helpful in understanding the transport mechanism. In the present work, we report a structural study on the oxidized and reduced forms of bovine adrenodoxin (bAdx) in solution using high-resolution NMR spectroscopy. The protein was produced in Escherichia coli and singly or doubly labeled with N-15 or C-13/N-15, respectively. Approximately 70 and 75% of the N-15, C-13, and H-1 resonances could be assigned for the reduced and the oxidized bAdx, respectively. The secondary and tertiary structures of the reduced and oxidized states were determined using NOE distance information. H-1(N)-T-1, relaxation times of certain residues were used to obtain additional distance constraints to the [2Fe-2S] cluster. The results suggest that the solution structure of oxidized Adx is quite similar to the X-ray structure. However, structural changes occur upon reduction of the [2Fe-2S] cluster, as indicated by NMR measurements. It could be shown that these conformational changes, especially in the C-terminal region, cause the dissociation of the Adx dimer upon reduction. A new electron transport mechanism proceeding via a modified shuttle mechanism, with both monomers and dimers acting as electron carriers, is proposed.