Evidence for dynamic in vivo interconversion of the conformational states of IscU during iron?sulfur cluster biosynthesis
Evidence for dynamic in vivo interconversion of the conformational states of IscU during iron?sulfur cluster biosynthesis
复制标题
铁硫簇生物合成过程中 IscU 构象状态动态体内相互转化的证据
DOI：
10.1111/mmi.14646
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发表时间：
2020
影响因子：
3.6
通讯作者：
Takahashi Yasuhiro
中科院分区：
文献类型：
--
作者：
Sato Sakiko;Matsushima Yumeka;Kanazawa Miaki;Tanaka Naoyuki;Fujishiro Takashi;Kunichika Kouhei;Nakamura Ryosuke;Tomioka Hiroaki;Wada Kei;Takahashi Yasuhiro
IscU是ISC机制的核心组成部分，并作为Fe-S簇从头组装的支架。由Hsp 70伴侣HscA和J蛋白辅助伴侣HscB组成的专用伴侣系统与IscU协同相互作用，并促进从IscU到受体载脂蛋白的簇转移。在这里，我们报告说，否则HscA和HscB的重要作用可以绕过在体内由一些单一的氨基酸取代IscU。能够进行这种旁路活性的变体IscU蛋白的CD光谱研究揭示了两种构象之间的动态相互转换：分别在不存在和存在Zn 2+的情况下的变性（D）和结构化（S）状态，这比在野生型IscU中观察到的相互转换要突出得多。此外，我们发现无论是IscU的S移位（更结构化）变体还是永久变性变体都不能发挥其体内作用，无论伴侣系统是否存在。因此，本研究首次提供了IscU体内D-状态存在的证据，并暗示支架蛋白的S-和D-状态之间的构象相互转换是Fe-S簇组装和转移的基本要求。
IscU is a central component of the ISC machinery and serves as a scaffold for de novo assembly of Fe–S clusters. The dedicated chaperone system composed of the Hsp70‐chaperone HscA and the J‐protein cochaperone HscB synergistically interacts with IscU and facilitates cluster transfer from IscU to recipient apo‐proteins. Here, we report that the otherwise essential roles of HscA and HscB can be bypassed in vivo by a number of single amino acid substitutions in IscU. CD spectroscopic studies of the variant IscU proteins capable of this bypass activity revealed dynamic interconversion between two conformations: the denatured (D) and the structured (S) state in the absence and presence of Zn2+, respectively, which was far more prominent than interconversion observed in wild‐type IscU. Furthermore, we found that neither the S‐shifted (more structured) variants of IscU nor the perpetually denatured variants could perform their in vivo role regardless of whether the chaperone system was present or not. The present study thus provides for the first time evidence that an in vivo D‐state of IscU exists and implies that conformational interconversion between the S‐ and D‐states of the scaffolding protein is a fundamental requirement for the assembly and transfer of the Fe–S cluster.