CAREER: Iron-sulfur cluster assembly in Bacillus subtilis

所有形式的生命都需要在它们的许多蛋白质中加入非蛋白质部分，即所谓的辅助因子，这样它们才能正常工作。许多这些辅助因子是由饮食中重要的维生素和矿物质组成的。其中一个辅助因子是铁硫（Fe-S）簇，它允许蛋白质进行许多重要的化学反应。由于游离铁和硫对细胞是有毒的，精细的蛋白质系统被用来构建这些辅助因子。一旦它们被构建，Fe-S簇就必须被运送到需要它们的蛋白质上，并被插入蛋白质中。不能正常构建Fe-S簇或不能成熟Fe-S蛋白的细胞会产生慢性并发症，通常会导致细胞死亡。然而，这种细胞机制的性质尚不清楚，关于生物体如何构建Fe-S簇并将其传递给需要它们发挥功能的蛋白质，仍存在许多问题。新的Fe-S蛋白成熟蛋白不断被发现，这凸显了对这一过程缺乏全局理解的事实。在这个项目中进行的研究将影响对从细菌到人类的铁依赖过程的理解。该项目还将为两名博士后学者和最多10名本科生提供独特的教育机会。这些受训者将接受技术和智力方面的训练，并将得到必要的指导和资源，以帮助他们过渡到职业生涯的下一个阶段。学员将重点提高他们的沟通技巧、技术技能、教学技能和人事管理技能。革兰氏阳性细菌枯草芽孢杆菌利用SufCDSUB Fe-S簇生物合成系统和至少两个Fe-S簇载体。已经确定了在Fe-S蛋白成熟中起作用的其他因素，包括SufT和YlaN；然而，这些分子的功能是未知的。此外，Fe和su定向Fe- s簇合成的电子给体以及附加的载流子分子都是未知的。为了正确理解Fe-S蛋白是如何成熟的，必须确定所涉及的成分并定义其功能。该项目将经典遗传学和生物化学与最先进的下一代技术和计算生物学相结合，以1)确定SufT， YlaN和Fe-S簇合成，组装和靶蛋白之间的物理相互作用；2)量化SufT和其他Fe-S簇组装因子在特定代谢途径中需要Fe-S酶的功能的必要性；3)分配生化功能给SufT和YlaN。4)建立并预测Fe-S蛋白成熟缺陷的影响，并在体内验证其影响。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

All forms of life require that many of their proteins have non-protein parts known as cofactors added to them so that they can function properly. Many of these cofactors are built from vitamins and minerals that are important in the diet. One such cofactor is the iron-sulfur (Fe-S) cluster, which allows proteins to perform many important types of chemistry. Because free Fe and S are toxic to cells, elaborate protein systems are used to build these cofactors. Once they are built, the Fe-S clusters must be trafficked to, and inserted into, proteins that require them. Cells that cannot properly build Fe-S clusters or maturate Fe-S proteins have chronic complications, which often results in cell death. The nature of this cellular machinery, however, is not well understood, and many questions remain about how organisms build Fe-S clusters and deliver them to proteins that require them for function. Novel Fe-S protein maturation proteins continue to be discovered, highlighting the fact that a global understanding of this process is lacking. The studies to be conducted in this project will impact the understanding of iron-dependent processes from bacteria to humans. The project will also provide unique educational opportunities for two postdoctoral scholars and up to ten undergraduate students. These trainees will be technically and intellectually trained and will receive the necessary guidance and resources to aid their transition into the next phases of their careers. The trainees will focus on enhancing their communication skills, technical skills, teaching skills, and personnel management skills.The Gram-positive bacterium Bacillus subtilis utilizes the SufCDSUB Fe-S cluster biosynthetic system and at least two Fe-S cluster carriers. Additional factors have been identified that have roles in Fe-S protein maturation, including SufT and YlaN; however, the functions of these molecules are unknown. Moreover, the Fe and electron donors for Suf-directed Fe-S cluster synthesis, as well as additional carrier molecules, are unknown. To properly understand how Fe-S proteins are maturated, the components involved must be identified and their functions defined. This project combines classic genetics and biochemistry with state-of-the-art, next-generation technologies and computational biology to 1) determine the physical interactions between SufT, YlaN, and Fe-S cluster synthesis, assembly, and target proteins, 2) quantify the necessity of SufT and alternate Fe-S cluster assembly factors for the function of specific metabolic pathways that require Fe-S enzymes, 3) assign biochemical functions to SufT and YlaN, and 4) model and predict the effect of faulty Fe-S protein maturation and validate the effects in vivo.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.