Structures and Mechanisms of Iron-Sulfur Proteins in Redox Control and Stress Response

项目总结 铁-硫(Fe-S)团簇是由多个铁和硫原子组成的古老的协因子。他们是 是生命各个领域中众多生物过程的基础。由于丰富的、可调节的氧化还原反应性 和选择性，Fe-S蛋白在生理和非生理条件下都在氧化还原控制中发挥多方面的作用。 压力条件。铁-S蛋白在氧化还原控制中的作用对维持正常细胞至关重要 功能和细胞生存，因此它们与健康和疾病，如癌症和糖尿病密切相关 人类和细菌感染。这与缺乏功能性、结构性和机械性形成鲜明对比。 许多铁-S蛋白在细胞内控制氧化还原动态平衡中的作用，特别是在三核 米拉建议涉及的方面：i)铁-S蛋白的氧化还原传感和转录调控； II)Fe-S簇的组装、转移和修复；以及III)Fe-S蛋白质与低分子蛋白质之间的串扰. 氧化还原止血中的重量(LMW)硫醇。铁-S蛋白介导的几种氧化还原控制机制 本提案将以分枝杆菌为例，阐述我们的研究目标和方法， 包括i)一个独特的Fe-S簇结合转录家族对氧化还原的感知和转录调控 WhiB样家族中的因素；ii)通过超滤系统组装、转移和修复Fe-S簇；以及iii) 真菌硫醇在铁-S集群内的动态平衡。拟议的研究计划建立在以下独特组合的基础上 在我的研究团队中的技能和丰富的研究经验，对于表征氧气敏感的 金属结合蛋白。我们最近确定了第一个也是人们期待已久的Fe-S团簇束缚结构 从结核分枝杆菌中分离到单体转录因子WhilB1，并建立了一种新的作用机制 由该蛋白介导的细菌转录调控。通过将结构、光谱和 体外生化方法结合体内分子生物学，我们准备确定：i)结构基础 Fe-S团簇的氧化还原反应性和配体选择性的研究；II)氧化还原状态和完整性 Fe-S簇的作用使这些蛋白质能够感知氧化还原状态并调节转录；iii)结构 铁-S簇合物生物合成和调控的生物化学；以及IV)非蛋白硫醇在 调节铁-S簇介导的氧化还原控制。总之，拟议的研究计划将建立一个新的 在氧化还原动态平衡方面研究不足的开创性研究。制定的战略 对新发现的Fe-S团簇系统的研究将是有益的。 如在DNA修复和RNA新陈代谢中对氧化应激的反应。因为他们的关键角色 在氧化还原控制方面，对这些Fe-S蛋白新机制的研究不仅将有助于阐明 铁-S蛋白介导的氧化还原调控的基本分子机制，也具有显著意义 对改善健康和防治传染病的影响。

PROJECT SUMMARY Iron-sulfur (Fe-S) clusters are ancient cofactors composed of multiple iron and sulfur atoms. They are fundamental to numerous biological processes in all domains of life. Owing to the rich, tunable redox reactivity and selectivity of the cluster, Fe-S proteins play multifaced roles in redox control under both physiological and stress conditions. The roles of Fe-S proteins in redox control are vital for the maintenance of normal cellular functions and cell survival, and thus they are tightly linked to health and disease such as cancer and diabetes in human and bacterial infection. This contrasts vividly with the lack of functional, structural and mechanistic understanding of many Fe-S proteins in the cellular control of redox homeostasis, particularly in the three core aspects that are addressed in this MIRA proposal: i) redox sensing and transcriptional regulation by Fe-S proteins; ii) assembly, transfer and repair of Fe-S clusters; and iii) crosstalk between Fe-S proteins and the low-molecular- weight (LMW) thiols in redox hemostasis. Several Fe-S proteins-mediated mechanisms for redox control in mycobacteria will be used as examples to elaborate our research goals and approaches in this proposal, including i) redox sensing and transcriptional regulation by a unique family of Fe-S cluster-bound transcription factors in the WhiB-like family; ii) assembly, transfer and repair of Fe-S clusters by the SUF system; and iii) mycothiol in Fe-S cluster homeostasis. The proposed research program is built on the unique combination of skills and rich research experience in my research team that are crucial for characterizing the oxygen-sensitive metal-binding proteins. We recently determined the first and long-desired Fe-S cluster-bound structure of the monomeric transcription factor WhilB1 from Mycobacterium tuberculosis and established a new mechanism of bacterial transcriptional regulation mediated by this protein. By combining structural, spectroscopic and biochemical approaches in vitro with molecular biology in vivo, we are poised to determine: i) the structural basis of redox reactivity and ligand selectivity in Fe-S clusters; ii) the mechanism by which the redox state and integrity of the Fe-S cluster allows these proteins to sense redox state and regulate transcription; iii) the structural biochemistry of Fe-S cluster biosynthesis and regulation; and iv) the role of non-proteinaceous thiols in modulating Fe-S cluster-mediated redox control. Altogether, the proposed research program will establish a new line of ground-breaking research in an understudied aspect of redox homeostasis. The strategies developed from the proposed program will be instrumental for studies on the newly discovered Fe-S cluster system, such as those in the DNA repair and RNA metabolism in response to oxidative stress. Because of their critical roles in redox control, the study of the novel mechanisms of these Fe-S proteins will not only shed light on the fundamental molecular mechanism governing Fe-S protein-mediated redox control, but also have a significant impact on improving health and combating infectious diseases.