Protein interactions regulate iron storage and utilization in bacteria

该项目的成果将填补我们对病原体铜绿假单胞菌中细菌铁代谢的基本理解的主要空白。这种病原体的几种菌株已经产生了多药耐药性，对烧伤患者、癌症患者和患有慢性肺部感染的囊性纤维化患者的医院感染的治疗造成了严峻的挑战。目前的建议，虽然在本质上是基本的，也可能暴露细菌铁代谢作为一个脆弱性，可以利用在未来的发展中的治疗，目标细菌铁稳态。该项目将在多学科环境中培养研究生和本科生，该环境来自化学，生物化学，光谱学，结构生物学和微生物学的不同领域。此外，堪萨斯大学（KU）的“收获希望领导学院”（HHLA）为来自移民农场工作背景的高中生举办了一个暑期住宿学院。项目总监，在与HHLA项目总监合作，将组织和核心研究实验室，在那里学生将有机会会见和互动的核心实验室主任的图尔斯旅游带领学生。成为KU本科生的HHLA学生在第一年由KU的大学援助移民计划（CAMP）组织指导。每年夏天，CAMP项目将招募两名优秀学生参加与该项目相关的研究活动。该项目将测试细菌铁稳态的新模型，铜绿假单胞菌中两种蛋白质之间的相互作用（BfrB和Bfd）使细菌细胞能够正确地感知细胞内游离铁水平的变化，并有效地将铁结合到利用铁的蛋白质中。BfrB是铁储存蛋白，Bfd是铁从BfrB动员所需的铁氧还蛋白。公认的细菌铁稳态模型假设，铁储存蛋白的功能只是作为铁载体，积累的铁被动员到细胞质只有当细菌细胞面临铁限制条件。 新模型假定，BfrB：Bfd相互作用通过促进铁流出BfrB，使细胞溶质中的游离铁与BfrB中储存的铁之间达到动态平衡，这对于细胞感知游离铁水平的变化并做出反应至关重要。修正后的模型还表明，从BfrB的铁动员所需的铁有效地掺入到铁利用蛋白质，因此，抑制BfrB：Bfd相互作用将导致无效的铁掺入到铁利用酶，这将不利地影响重要的生理过程，依赖于含铁的蛋白质和酶。这条途径与公认的模型不同，在公认的模型中，利用铁的蛋白质的铁来源被假定为细胞质中的游离铁。 研究人员还发现了一种抑制BfrB：Bfd相互作用的小分子探针，并获得了这种分子增强某些现有抗生素的杀伤活性的证据。因此，该项目还旨在从根本上了解BfrB：Bfd相互作用的抑制剂如何增强抗生素活性。 这些研究将由以下想法指导：由BfrB：Bfd相互作用的小分子抑制剂引起的BfrB中铁的不可逆积累严重限制了支持抵御干旱诱导的应激所需的关键代谢过程所需的细胞内铁。

The project outcomes will fill a major gap in our fundamental understanding of bacterial iron metabolism in the pathogen Pseudomonas aeruginosa. Several strains of this pathogen have developed multidrug resistance, causing severe challenges for the treatment of hospital infections that affect burn victims, cancer patients and cystic fibrosis patients with chronic lung infections. The present proposal, while fundamental in nature, is also likely to expose bacterial iron metabolism as a vulnerability that can be exploited in future development of therapies that target bacterial iron homeostasis. The project will train graduate and undergraduate students in a multidisciplinary environment that draws from the diverse fields of chemistry, biochemistry, spectroscopy, structural biology and microbiology. In addition, the Harvest of Hope Leadership Academy (HHLA) at the University of Kansas (KU) hosts a summer residential academy for high school students from migrant farm-work background. The Project Director, in collaboration with the HHLA Program Director, will organize and lead students on tours of Core Research laboratories, where students will have the opportunity to meet and interact with the Core Laboratory Directors. HHLA students who become KU undergraduates are mentored during their first year by the College Assistance Migrant Program (CAMP) organization at KU. Two deserving students from the CAMP program will be recruited each summer to participate in research activities related to the project.The project will test a new model for bacterial iron homeostasis, whereby the interactions between two proteins in P. aeruginosa (BfrB and Bfd) enable the bacterial cells to correctly sense changes in intracellular free iron levels and to efficiently incorporate iron into iron-utilizing proteins. BfrB is an iron storage protein and Bfd is a ferredoxin required for iron mobilization from BfrB. The accepted model of bacterial iron homeostasis assumes that iron storage proteins function simply as iron accumulators and that accumulated iron is mobilized to the cytosol only when bacterial cells face iron-limiting conditions. The new model posits that the BfrB:Bfd interaction, by facilitating iron flux out of BfrB, enables a dynamic equilibrium between free iron in the cytosol and iron stored in BfrB, which is crucial for the cells to sense and react to changes in free iron levels. The revised model also suggests that iron mobilization from BfrB is required for the efficient incorporation of iron into iron-utilizing proteins, so that inhibition of the BfrB:Bfd interaction will result in inefficient iron incorporation into iron-utilizing enzymes, which will adversely affect important physiological process that depend on iron-containing proteins and enzymes. This proposed path is different from that in the accepted model, where the source of iron for iron-utilizing proteins is assumed to be the free iron in the cytosol. The investigators have also discovered a small molecule probe that inhibits the BfrB:Bfd interaction, and obtained evidence that this molecule enhances the killing activity of some existing antibiotics. Hence, the project also aims to obtain a fundamental understanding how the antibiotic activity is enhanced by inhibitors of the BfrB:Bfd interaction. These studies will be guided by the idea that irreversible accumulation of iron in BfrB, which is caused by the small molecule inhibitor of the BfrB:Bfd interaction, severely restricts the intracellular iron needed to support crucial metabolic processes required to fend antibiotic-induced stress.