RECEPTOR MEDIATED UPTAKE OF IRON COMPLEXES IN BACTERIA

这项研究的长期目标是阐明分子 大肠埃希氏菌和相关生物的活跃过程 将铁与铁载体或铁结合蛋白的络合物运输到 满足他们的营养铁需求。细菌在体内获得铁 不溶性铁络合物产铁的好氧环境 或它们直接利用宿主铁结合蛋白，如 转铁蛋白或氯化血红素。特定的外膜受体识别铁- 结合分子以允许其能量依赖的吸收。所需能量 穿过外膜的传输是通过耦合质子得到的。 从细胞质膜到这些配体结合受体的动力 通过TonB及其辅助蛋白ExbB和ExbD的活性。 TonB依赖是细菌铁摄取系统中的一个共同主题， 包括铁载体和宿主铁结合蛋白在 公认的细菌病原体。高选择性受体的功能 与同源运输许可相配合，使微生物能够 吸收足够的铁以在有氧环境中生存和生长 在动物寄主身上。 本研究重点研究了大肠杆菌肠杆蛋白转运系统 研究TonB受体分子细节的原型 依赖铁摄取。遗传选择策略与生化 技术将用于定义的拓扑和功能特征 FepA受体，探测其与TonB的相互作用，并确定 FEP渗透酶的基本结构和功能特性。 这些研究将涉及以下具体目标：(1)表位 标签和缺失突变以完善FepA的拓扑图； (2)确定FepA功能区的突变选择；(3)遗传 以及FepA-TonB相互作用的生化交联分析； 用于功能表征的结合能力和突变体选择 FEP渗透酶复合体的膜蛋白；和(5)描绘 酶促肠杆菌素除铁的酶促条件 酯酶Fes。因为铁的可获得性的限制是一个关键 微生物种群增长和传播的控制因素， 它被认为是毒力能力的重要组成部分 细菌病原体。了解细菌的铁摄取过程 因此，分子水平对于设计有效的策略是重要的 为潜在的有毒生物保留这种至关重要的营养物质。

The long-term objective of this study is to elucidate the molecular processes by which Escherichia coli and related organisms actively transport iron complexed to siderophores or iron-binding proteins to satisfy their nutritional iron requirements. Bacteria obtain iron in an aerobic environment from insoluble ferric complexes by producing ferric chelators or they directly utilize host iron-binding proteins like transferrin or hemin. Specific outer membrane receptors recognize iron- binding molecules to allow their energy-dependent uptake. The energy for transport across the outer membrane is derived by coupling the proton motive force from the cytoplasmic membrane to these ligand-bound receptors through the activity of TonB and its accessory proteins ExbB and ExbD. TonB dependence is a common theme in bacterial iron uptake systems, including those for siderophores and host iron-binding proteins in recognized bacterial pathogens. The highly selective receptors function in concert with cognate transport permeases to enable microbes to assimilate sufficient iron to survive and grow in aerobic environments or in an animal host. This study focuses on the E. coli enterobactin transport system as a prototype for investigating the molecular details of TonB-receptor- dependent iron uptake. Genetic selection strategies and biochemical techniques will be used to define topological and functional features of the FepA receptor, to probe its interactions with TonB, and to define basis structural and functional characteristics of the Fep permease. These studies will involve the following specific aims: (1) epitope tagging and deletion mutagenesis to refine the topological map of FepA; (2) selection of mutations defining FepA functional domains; (3) genetic and biochemical cross-linking analysis of the FepA-TonB interaction; (4) binding capabilities and mutant selection for functional characterization of membrane proteins of the Fep permease complex; and (5) delineation of enzymatic requirements for iron removal from enterobactin catalyzed by the esterase Fes. Since the limitation of iron availability is a critical control factor in the growth and dissemination of microbial populations, it is recognized as a significant component of virulence capacity for bacterial pathogens. Understanding the bacterial iron uptake process at a molecular level is therefore important to designing effective strategies to withhold this vital nutrient for potentially virulent organisms.