Localization and Characterization of the Fe(II) Oxidase Complex from Rhodopseudomonas palustris TIE-1

基于亚铁[Fe(II)]的光合作用被认为是地球历史早期的一个重要过程，但在分子水平上对这种代谢知之甚少。如果我们要在岩石记录中追踪这种代谢的起源（如记录在被称为带状铁地层的古代沉积沉积物中），那么了解哪些生物分子催化了这一过程以及它们是如何工作的将是至关重要的。带着这个目标，PI的实验室分离出一种遗传上易于处理的古红假单胞菌（TIE-1），一种革兰氏阴性的无氧光养菌，作为研究这种代谢的模型系统。最近，三个基因被确定为光营养性铁氧化所必需的：pioA， pioB和pioC。本项目的目标是探索这些基因产物的生物化学，具体目标有三个：1)研究Pio蛋白的定位。据推测，PioA和PioC存在于质周空间，而PioB存在于外膜。2)鉴定Pio蛋白的功能。该项目将测试PioA直接结合和氧化Fe（II）的假设，而PioC将电子从PioA携带到胞质膜内的电子传递链。此外，该项目将测试PioB是否与PioA相互作用，以及/或PioB是否参与铁（II）的运输。3)解出被证明是直接结合并氧化Fe（II）的蛋白的晶体结构。据推测，这种蛋白质是PioA。更广泛的影响本项目将为博士后和本科生提供分子地理生物学的高级培训。除了直接培训学生外，该项目还将支持社区外展活动，包括向少数族裔观众和公众发表公开演讲。最后，为了补充这个项目，PI将申请美国国家科学基金会国际旅行奖和/或美国微生物学会的国际教授职位，以支持与委内瑞拉同事的现有合作。

Photosynthesis based on ferrous iron [Fe(II)] is thought to have been an important process early in Earth history, yet very little is understood about this metabolism at the molecular level. If we are ever to trace the origin of this metabolism in the rock record (as recorded in ancient sedimentary deposits known as Banded Iron Formations), it will be essential to understand which biomolecules catalyze this process and how they work. With this goal in mind, the PI's laboratory isolated a genetically-tractable strain of Rhodopseudomonas palustris (TIE-1), a Gram-negative, anoxygenic phototroph to serve as a model system to study this metabolism. Recently, three genes were identified as essential for phototrophic iron oxidation: pioA, pioB and pioC. The goal of this project is to explore the biochemistry of the products of these genes, with three specific objectives: 1) To study the localization of the Pio proteins. It is hypothesized that PioA and PioC reside in the periplasmic space, whereas PioB resides in the outermembrane. 2) To characterize the functions of the Pio proteins. The project will test the hypothesis that PioA directly binds and oxidizes Fe(II), whereas PioC carries electrons from PioA to the electron transport chain in the intra cytoplasmic membrane. In addition, the project will test if PioB interacts with PioA, and/or if PioB is involved in Fe(II) transport. 3) To solve the crystal structure of whichever protein proves to be the one that directly binds and oxidizes Fe(II). It is postulated that this protein is PioA. Broader ImpactsThis project will enable the advanced training in molecular geobiology at postdoctoral and undergraduate student level. Beyond the direct training of students, this project will support community outreach activities including public speaking to minority audiences and the general public. Lastly, to complement this project, the PI will apply for an NSF-International travel award and/or an International Professorship from the American Society for Microbiology to support the continuation of an existing collaboration with a colleague in Venezuela.