Characterization of the ferrous iron transporter Feo in Vibrio cholerae

描述（由申请人提供）：铁参与多种细胞反应，是几乎所有生物体的基本元素。对于致病菌来说，铁的获取是一个问题，因为致病菌必须与宿主竞争有限数量的可用铁，而且细菌通常表达对其铁或亚铁配体具有特异性的高亲和力运输系统。Feo是原核生物中主要的亚铁转运系统。它广泛分布于真细菌和古细菌中，表明它是一类非常古老的铁转运体。此外，FeoB蛋白具有与真核G蛋白相似的GTPase和GDI结构域，Feo可能代表现代G蛋白的原始祖先。尽管其普遍存在的性质和潜在的作用在细菌的发病机制，知之甚少的结构，转运机制和生物学作用。因此，表征该蛋白及其在亚铁运输中的作用是至关重要的。这些研究将在病原体霍乱弧菌中进行，霍乱弧菌是人类发病和死亡的主要原因。霍乱弧菌是这些研究的理想模型，因为我们有试剂和遗传工具来回答关于Feo的基本问题。此外，我们可以将这些研究扩展到包括Feo在宿主定植和疾病中的作用的问题，这将为开发阻断霍乱弧菌感染的新方法奠定基础。我们的第一个具体目标是利用遗传和生化方法来表征Feo的结构及其运输机制。这些研究将有助于确定Feo转运体的基本蛋白质成分以及每种蛋白质中的关键区域和氨基酸。基于这些结果，我们将能够测试Feo结构和功能的模型。我们的第二个具体目标是表征辅助亚铁转运蛋白VciB。这可能为了解Feo如何在周质中获得亚铁配体提供见解。第三，我们将明确feo操纵子的表达调控机制，确定细菌在宿主体内时其表达模式。这些数据将为致病菌如何协调铁转运基因的表达提供新的信息，以允许最佳利用可用的铁源，也将有助于确定细菌在宿主中遇到的环境信号。

DESCRIPTION (provided by applicant): Iron participates in a wide variety of cellular reactions and is an essential element for nearly all organisms. Iron acquisition is problematic for pathogenic bacteria, which must compete with the host for limited amounts of available iron, and bacteria typically express high affinity transport systems that are specific for their ferric or ferous iron ligands. Feo is the major ferrous iron transport system in prokaryotic organisms. It is widely distributed in eubacteria and in the Archaea, suggesting that it is a very ancient class of iron transporter. Further, the FeoB protein has GTPase and GDI domains similar to eukaryotic G proteins, and Feo may represent a primitive ancestor of modern G proteins. Despite its ubiquitous nature and potential role in bacterial pathogenesis, little is known about its structure, mechanism of transport, and biological role. Thus, it is critical to characterize this protein and its role in ferrous iron transport. These studies will be done in the pathogen Vibrio cholerae, a major cause of human morbidity and mortality. V. cholerae is an ideal model for these studies as we have the reagents and genetic tools to answer basic questions about Feo. Further, we can extend these studies to include questions about the role of Feo in host colonization and disease, which will lay the groundwork for developing new methods of blocking infection by V. cholerae. Our first specific aim is to use genetic and biochemical approaches to characterize Feo structure and its mechanism of transport. These studies will help define the essential protein components of the Feo transporter and the critical regions and amino acids within each protein. Based on these results, we will be able to test models for Feo structure and function. Our second specific aim is to characterize the accessory ferrous iron transport protein, VciB. This may provide insight into how Feo obtains its ferrous iron ligand in the periplasm. Third, we will define the mechanism of regulation of expression of the feo operon and determine its pattern of expression when the bacteria are within the host. These data will provide new information about how pathogenic bacteria coordinate the expression of their iron transport genes to allow optimal utilization of available iron sources and will also help define the environmental signals the bacteria encounter in the host. PUBLIC HEALTH RELEVANCE: Obtaining essential iron from the host is a critical step in production of disease by bacterial pathogens. Characterizing the ferrous iron uptake system Feo in Vibrio cholerae will provide important information about an iron transporter found in many bacterial pathogens, and it will further our knowledge of the organism that causes cholera. Because no system comparable to Feo is found in the human, this iron transport system may represent a new class of targets for antimicrobial therapy.