Starter Grant: Investigating the Role of a Tetraheme c-type Cytochrome in the Arsenate Respiration Pathway of Shewanella

已知铁（三价铁）和砷酸盐还原菌可促进砷从受污染的沉积物迁移到含水层。因此，饮用这些金属还原细菌活跃的水源的人群可能会因砷水平升高而增加患癌症和皮肤病变的风险。这项研究将发展的砷酸盐呼吸减少，这是一个主要的生物过程，认为促进砷动员的分子机制的理解。 该项目的目标是确定CymA如何与预测发生在砷酸盐还原希瓦氏菌物种的厌氧电子传递链中的其他氧化还原组分相互作用。具体目标是：（1）检验CymA表现出甲萘醌结合活性的假设和（2）检验CymA与砷酸呼吸还原酶ArrAB相互作用的假设。将使用可通过其光谱性质监测的甲基喹酚类似物测定CymA的甲基喹酚结合。将使用频率共振能量转移（FRET）方法研究CymA与ArrAB的体内蛋白质-蛋白质相互作用，并且CymA和ArrAB蛋白与已知表现出FRET信号的荧光蛋白融合。 这项研究的结果将直接应用于了解硝酸盐，富马酸盐和氧化铁代谢下游途径的类似相互作用。这项研究将为微生物介导的砷动员的潜在机制的环境调查提供新的见解。 这项工作的更广泛影响将包括促进本科生和研究生的教育。学生将接受厌氧微生物学，培养基制备和呼吸最终产物分析方法的实践培训。

Iron(III) and arsenate-reducing bacteria are known to promote the mobilization of arsenic from contaminated sediments into aquifers. Consequently, populations consuming water sources where these metal-reducing bacteria are active may be at increased risk for developing cancer and skin lesions due to elevated levels of arsenic. This research will develop a molecular-mechanistic understanding of arsenate respiratory reduction, which is a major biological process thought to promote arsenic mobilization. The goal of the project is to determine how CymA interacts with other redox components predicted to occur in the anaerobic electron transport chain of arsenate-reducing Shewanella species. The specific aims are to: (1) test the hypothesis that CymA exhibits menaquinol binding activity and (2) test the hypothesis that CymA interacts with the arsenate respiratory reductase, ArrAB. Menaquinol binding of CymA will be determined using menaquinol analogs that can be monitor by their spectroscopic properties. In vivo protein-protein interactions of CymA with ArrAB will be investigated using frequency resonance energy transfer (FRET) methods and CymA and ArrAB proteins fused with fluorescent proteins known to exhibit FRET signals. Results from this study will have direct applications to understanding similar interactions with downstream pathways for nitrate, fumarate, and iron oxide metabolisms. This research will provide new insight into environmental investigations of the underlying mechanism of microbial-mediated mobilization of arsenic. The broader impacts of the work will include promoting the education of undergraduate and graduate students. Students will receive hands-on training in anaerobic microbiology, preparation of media, and methods for analyzing the end products of respiration.