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

项目摘要 铁硫原子团簇是由多个铁和硫原子组成的古老的辅因子。他们是 对生命各个领域的许多生物过程至关重要。由于丰富的、可调的氧化还原反应性， 和选择性，Fe-S蛋白在生理和生理条件下的氧化还原控制中发挥着多方面的作用， 应力条件Fe-S蛋白在氧化还原调控中的作用对于维持正常的细胞功能是至关重要的。 功能和细胞存活，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、结构性和机械性的 了解许多铁硫蛋白在细胞控制氧化还原稳态，特别是在三个核心 本MIRA提案中涉及的方面：i）Fe-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;和iii）Fe-S蛋白与低分子- 氧化还原止血中的重量（LMW）硫醇。几种Fe-S蛋白介导的氧化还原调控机制 分枝杆菌将作为例子来阐述我们的研究目标和方法在这个建议， 包括i）通过独特的Fe-S簇结合转录家族的氧化还原感测和转录调节 ii）通过SUF系统组装、转移和修复Fe-S簇;和iii） 真菌硫醇在Fe-S簇内稳态中的作用。拟议的研究计划是建立在独特的组合， 技能和丰富的研究经验，这对表征氧敏感性至关重要。 金属结合蛋白我们最近确定了第一个和长期期望的Fe-S团簇结合结构， 结核分枝杆菌单体转录因子WhilB 1，建立了一种新的 由该蛋白介导的细菌转录调节。通过结合结构，光谱和 生物化学方法在体外与分子生物学在体内，我们准备确定：i）结构基础 的氧化还原反应性和配体选择性的Fe-S簇; ii）的机制，氧化还原状态和完整性 的Fe-S簇允许这些蛋白质感测氧化还原状态和调节转录; iii）结构的 Fe-S簇生物合成和调节的生物化学;和iv）非蛋白质硫醇在 调节Fe-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 在氧化还原稳态的一个未充分研究的方面进行了突破性的研究。制定的战略 将有助于研究新发现的Fe-S团簇系统， 与那些在DNA修复和RNA代谢中对氧化应激的反应一样。因为他们的关键角色 在氧化还原控制中，对这些Fe-S蛋白的新机制的研究不仅将阐明 Fe-S蛋白介导的氧化还原控制的基本分子机制，但也有显着的 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.