CAREER: Iron-sulfur cluster assembly in Bacillus subtilis

所有形式的生命都需要它们的许多蛋白质中加入被称为辅因子的非蛋白质部分，以便它们能够正常发挥作用。其中许多辅因子都是由饮食中重要的维生素和矿物质构成的。一个这样的辅因子是铁-硫(铁-S)簇，它允许蛋白质执行许多重要类型的化学。因为游离铁和S对细胞有毒性，所以使用复杂的蛋白质系统来构建这些辅因子。一旦它们被构建，Fe-S簇必须被贩运到需要它们的蛋白质中，并插入到需要它们的蛋白质中。不能正确构建铁-S簇或成熟铁-S蛋白的细胞会出现慢性并发症，这往往会导致细胞死亡。然而，这种细胞机制的性质还没有被很好地理解，关于生物体如何建立铁-S簇并将它们运送到需要它们发挥功能的蛋白质上，仍然存在许多问题。新的铁-S蛋白成熟蛋白继续被发现，这突显了这样一个事实，即全球对这一过程缺乏了解。该项目将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十名本科生提供独特的教育机会。这些受训人员将接受技术和智力培训，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。学员将重点提高沟通技能、技术技能、教学技能和人员管理技能。革兰氏阳性菌枯草杆菌利用SufCDSUB Fe-S集群生物合成系统和至少两个Fe-S集群载体。其他一些在铁-S蛋白成熟过程中起作用的因子已经被发现，包括Suft和Ylan；然而，这些分子的功能尚不清楚。此外，用于SuF定向Fe-S团簇合成的Fe和电子给体以及额外的载体分子尚不清楚。为了正确理解铁-S蛋白是如何成熟的，必须确定相关的成分并确定它们的功能。该项目将经典遗传学和生物化学与最新、新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和交替Fe-S簇组装因子对需要Fe-S酶的特定代谢途径功能的必要性，3)赋予Suft和Ylan生化功能，4)模拟和预测有缺陷的Fe-S蛋白成熟的影响，并在体内验证这些影响。这一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智力优势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.