Iron-sulfur cluster biosynthesis : Molucular mechanism of the assembly of intermediate cluster and its transfer to apo proteins.

铁硫簇生物合成：中间簇组装及其转移到apo蛋白的分子机制。

• 批准号: 17570112
• 负责人: TAKAHASHI Yasuhiro
• 金额: $ 2.24万
• 依托单位: Osaka University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2005
• 资助国家: 日本
• 起止时间: 2005 至 2006
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-17570112/
• 关键词: iron-sulfur cluster; iron-sulfur protein; biosynthesis; gene; protein; molecular chaperone; X-ray crystal structure; 鉄硫黄蛋白質; 蛋白質

Iron-sulfur (Fe-S) proteins are present in almost all living organisms and exhibit diverse functions, which include electron transport, redox and non-redox catalysis, and sensing for regulatory processes. They contain clusters of iron and sulfur atoms with variable complexity, such as [2Fe-2S], [3Fe-4S], and [4Fe-4S] clusters. We have recently found two independent systems, called ISC and SUF, essential for the biosynthesis of Fe-S clusters, although still very little is known about the mechanistic details. In this study we have focused on IcsU, a central key player of the ISC machinery working as a scaffold for a nascent Fe-S cluster, and undertaken genetic, biochemical and structural approaches to elucidate the complex mechanism by which the Fe-S clusters are assembled and transferred to target proteins.1. We have isolated and analyzed 14 psuedorevertants generated from a mutant strain lacking HscA and HscB of the ISC machinery, and found that the responsible suppressor mutations are all located in IscU. The mutated IscUs were conformationally altered from WT IscU at the secondary structure level, suggesting that, in wild-type cells, HscA and HscB induce the structural changes of IscU thereby facilitating the cluster transfer from IscU to target proteins.2. In contrast to the notoriously unstable Fe-S cluster intermediate in the scaffold proteins, we found that the IscU homolog from the hyperthermophilic bacterium Aquifex aeolicus (AaIscU) carries a stable Fe-S cluster. AaIscU was purified, characterized and crystallized for the first time under anaerobic conditions. The crystal structure has revealed that the holo-AaIscU is an asymmetric trimer carrying only one [2Fe-2S] cluster at the interface between two structurally altered protomers. The structure of the asymmetric trimer provides valuable insight into the molecular event: the assembly, maintenance and transfer of the nascent Fe-S cluster is tightly coupled to the conversion of the oligomeric state of IscU.

铁硫蛋白（Fe-S）几乎存在于所有生物体内，并具有多种功能，包括电子传递、氧化还原和非氧化还原催化以及调节过程的传感。它们含有不同复杂程度的铁和硫原子簇，如[2Fe-2S]、[3Fe-4S]和[4Fe-4S]簇。我们最近发现了两个独立的系统，称为ISC和SUF，对Fe-S簇的生物合成至关重要，尽管对其机制细节知之甚少。在这项研究中，我们将重点放在IcsU上，IcsU是ISC机制的核心关键角色，作为新生Fe-S簇的支架，并采用遗传、生化和结构方法来阐明Fe-S簇组装和转移到靶蛋白的复杂机制。我们分离并分析了从缺乏HscA和ISC机制的HscB的突变株中产生的14个伪复归物，发现负责的抑制突变都位于IscU。突变的IscU在二级结构水平上与WT型IscU构象发生了改变，这表明在野生型细胞中，HscA和HscB诱导了IscU的结构变化，从而促进了IscU向靶蛋白的簇转移。与支架蛋白中众所周知的不稳定的Fe-S簇相比，我们发现来自超嗜热细菌Aquifex aeolicus （AaIscU）的IscU同源物携带稳定的Fe-S簇。首次在厌氧条件下对aiscu进行了纯化、表征和结晶。晶体结构表明，全息aaiscu是一种不对称三聚体，在两个结构改变的原聚体之间的界面上只携带一个[2Fe-2S]簇。不对称三聚体的结构为分子事件提供了有价值的见解：新生Fe-S簇的组装，维持和转移与IscU低聚态的转化紧密相关。

项目成果

Crystallization and preliminary x-ray crystallographic studies of the oxidative-stress sensor SoxR and its complex with DNA

氧化应激传感器 SoxR 及其与 DNA 复合物的结晶和初步 X 射线晶体学研究

• 发表时间: 2006
• 期刊: Acta Cryst. F62
• 作者: Otsuka;M.;Otsuka M.;Otsuka M;Takaki Nimura et al.;Yoshinori Hagiwara;Shintaro Kitaoka;Akifumi Kamata et al.;Yoshinori Hagiwara;Satoshi Watanabe
• 通讯作者: Satoshi Watanabe

Crystal structure of Escherichia coli SufC, an ABC-type ATPase component of the SUF iron-sulfur cluster assembly machinery

• DOI: 10.1016/j.febslet.2005.11.058
• 发表时间: 2006-01-09
• 期刊: FEBS LETTERS
• 影响因子: 3.5
• 作者: Kitaoka, S;Wada, K;Takahashi, Y
• 通讯作者: Takahashi, Y

Structural study of ferredoxin-dependent bilin reductase.

铁氧还蛋白依赖性胆碱还原酶的结构研究。

• 发表时间: 2006
• 期刊: J.Cryst.Soc.Jpn 48-4
• 作者: K.Matsuoka;H.Tabunoki;T.Kawai;S.Ishikawa;M.Yamamoto;R.Sato;T.Ando;Satoshi Watanabe;Hiroshi Tokumitsu et al.;安藤 哲;Masakazu Sugishima
• 通讯作者: Masakazu Sugishima

フェレドキシン依存性ビリン還元酵素の構造研究

铁氧还蛋白依赖性胆碱还原酶的结构研究

• 发表时间: 2006
• 期刊: 日本結晶学会誌 48
• 作者: T.Fujii;M.G Suzuki;T.Kawai;K.Tsuneizumi;A.Ohnishi;M.Kurihara;S.Matsumoto;T.Ando;杉島正一
• 通讯作者: 杉島正一

Crystal structure of Escherichia coli YfhJ protein, a member of the ISC machinery involved in assembly of iron-sulfur clusters

• DOI: 10.1002/prot.20481
• 发表时间: 2005-08-15
• 期刊: PROTEINS-STRUCTURE FUNCTION AND BIOINFORMATICS
• 影响因子: 2.9
• 作者: Shimomura, Y;Takahashi, Y;Fukuyama, K
• 通讯作者: Fukuyama, K