Sensing Superfund Chemicals with Recombinant Systems

使用重组系统感测 Superfund 化学品

• 批准号: 6932249
• 负责人: Sylvia Daunert
• 金额: $ 25.96万
• 依托单位: UNIVERSITY OF KENTUCKY
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-04-01 至 2008-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6932249
• 关键词: DVD /CD ROM; biohazard detection; biosensor device; biotechnology; environmental toxicology; halobiphenyl /halotriphenyl compound; high throughput technology; microfluidics; molecular /cellular imaging; operon; oxygenases; reporter genes; technology /technique development; toxin metabolism

Detection of Superfund chemicals is a first step towards eliminating them from the environment or studying their effect on biological processes. To that end, we propose to develop molecular tools for the highly sensitive and direct detection of Superfund chemicals such as PCBs. These molecular tools will include whole cell- and protein-based optical sensing systems. For that, we propose to utilize naturally available operons induced in the presence PCBs and the catabolic enzymes involved in their degradation. We plan to modify these operons by incorporating the gene for the reporter, which will allow us to monitor their induction in the presence of PCBs. Catabolic enzymes involved in the degradation of PCBs, in particular the enzyme biphenyl dioxygenase (BphAt), will be isolated and labeled with signal transduction moieties for the analysis of these compounds. In addition, to improve the selectivity of these catabolic enzymes towards PCB congeners, mutagenesis studies aimed at modifying the active site of the enzyme are proposed. In order to make these biosensing systems amenable to field studies and high throughput sample analysis, we propose to incorporate them into a microfluidics platform. This miniaturized analytical platform should find applications in the on-site monitoring of Superfund chemicals. Specifically, we plan to adapt our biosensing systems for PCBs to a compact disc centrifugal microfluidics platform. The advantages of our proposed biosensing systems include sensitivity, portability, and the feasibility of analysis of PCBs in different matrices, such as soil, water, and biological samples.

检测超级基金化学品是将其从环境中消除或研究其对生物过程影响的第一步。为此，我们建议开发分子工具，用于高灵敏度和直接检测超级基金化学品，如多氯联苯。这些分子工具将包括基于全细胞和蛋白质的光学传感系统。为此，我们建议利用自然可用的操纵子诱导存在的多氯联苯和参与其降解的分解代谢酶。我们计划通过整合报告基因来修改这些操纵子，这将使我们能够在多氯联苯的存在下监测它们的诱导。参与降解多氯联苯的分解代谢酶，特别是酶联苯双加氧酶（BphAt），将被分离和标记的信号转导部分，这些化合物的分析。此外，为了提高这些分解代谢酶的选择性， 针对多氯联苯同系物，提出了旨在改变酶活性位点的诱变研究。为了使这些生物传感系统适合现场研究和高通量样品分析，我们建议将它们纳入微流体平台。这种小型化的分析平台应能应用于超级基金化学品的现场监测。具体来说，我们计划将我们的PCB生物传感系统适应于光盘离心微流体平台。我们提出的生物传感系统的优点包括灵敏度，便携性，和不同的矩阵，如土壤，水和生物样品中的多氯联苯分析的可行性。