Oxidative folding of small Tims is mediated by site-specific docking onto Mia40 in the mitochondrial intermembrane space
Oxidative folding of small Tims is mediated by site-specific docking onto Mia40 in the mitochondrial intermembrane space
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DOI：
10.1111/j.1365-2958.2007.05880.x
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发表时间：
2007-09-01
影响因子：
3.6
通讯作者：
Tokatlidis, Kostas
中科院分区：
文献类型：
--
作者：
Sideris, Dionisia P.;Tokatlidis, Kostas
线粒体膜间隙（IMS）中的氧化折叠对于某些富含半胱氨酸的IMS蛋白的输入至关重要。必需蛋白质Mia 40和Erv 1是该机制的关键组分，其作为二硫蛋白级联发挥作用，通过将电子穿梭到CytC上而在功能上与呼吸链连接。分子伴侣复合物Tim 9-Tim 10的亚基需要Mia 40用于其生物合成。以前，它表明，四个半胱氨酸的Tim 10是至关重要的折叠和组装，它们是连接成一个内部和外部的二硫桥分子内，内部的二硫键在这些过程中有一个更突出的作用。在这里，我们表明与Mia 40的相互作用是一个位点特异性事件：（i）前体的N-末端第一个半胱氨酸是通过混合二硫键对接到Mia 40上的关键;（ii）释放是由C-末端半胱氨酸到N-末端半胱氨酸的二硫键配对触发的;以及（iii）在第二和第三个半胱氨酸之间的内部二硫化物的形成显然先于释放反应，并且对于与Tim 9的组装是关键的。Tim 10-Mia 40的相互作用是独立的二价阳离子，任何其他线粒体蛋白或膜，并被证明有效地发生在细胞器和体外。
Oxidative folding in the mitochondrial intermembrane space (IMS) is crucial for the import of certain cysteine-rich IMS proteins. The essential proteins Mia40 and Erv1 are key components for this mechanism functioning as a disulphide protein cascade that is functionally linked to the respiratory chain by shuttling electrons onto CytC. The subunits of the chaperone complex Tim9-Tim10 require Mia40 for their biogenesis. Previously, it was shown that the four cysteines of Tim10 are crucial for folding and assembly, that they are connected intramolecularly into an inner and an outer disulphide bridge, and that the inner disulphide has a more prominent role in these processes. Here we show that interaction with Mia40 is a site-specific event: (i) the N-terminal first cysteine of the precursor is crucial for docking onto Mia40 via a mixed disulphide; (ii) release is triggered by disulphide pairing of the C-terminal cysteine onto the N-terminal one; and (iii) formation of the inner disulphide between the second and third cysteines apparently precedes the release reaction and is critical for assembly with Tim9. The Tim10-Mia40 interaction is independent of divalent cations, any other mitochondrial proteins or membranes, and is shown to occur efficiently in organello and in vitro.