High-throughput fluorescence screening for inhibitors of TonB-dependent iron transport

高通量荧光筛选 TonB 依赖性铁转运抑制剂

基本信息

批准号：
8969952
负责人：
PHILLIP E KLEBBA
金额：
$ 23.75万
依托单位：
KANSAS STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-07-15 至 2017-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8969952
关键词：
2,4-Dinitrophenol Animals Antibiotics Assimilations Bacteria Bacterial Infections Bacteriophages Binding Binding Sites Biochemical Biological Assay Biological Factors Cell surface Cells Chemicals Collection Complex Development Dinitrophenols Enterobacter aerogenes bacterium Enterobactin Escherichia coli FDA approved Ferrichrome Fluorescein Fluorescence Fluorescence Spectroscopy Fluorescent Probes Genetic Engineering Gram-Negative Bacteria Growth Haemophilus influenzae Human Hydrazones Individual Iron Kansas Klebsiella pneumonia bacterium Knowledge Label Laboratories Lead Libraries Life Location Maleimides Measurement Membrane Membrane Proteins Membrane Transport Proteins Metabolism Metals Methods Microbe Molecular Monitor Motion Nature Neisseria meningitidis Pathogenesis Pathogenicity Pathway interactions Peptidoglycan Performance Physiology Preparation Procedures Process Proton-Motive Force Pseudomonas aeruginosa Reaction Research Salmonella typhi Salmonella typhimurium Siderophores Site Staging Structure Surface Temperature Testing Text Time Transport Reaction Universities Validation Vibrio cholerae Vitamin B 12 aerobactin bacteriocin base biophysical analysis cell envelope cell preparation fluorophore high throughput screening in vivo inhibitor/antagonist insight killings mesoxalonitrile microbial novel pathogen permease public health relevance receptor research study scaffold screening uptake vibriobactin

项目摘要

DESCRIPTION (provided by applicant): The ability of microbes to acquire iron is a key aspect of microbial invasiveness and pathogenicity. TonB-dependent transporters in the Gram-negative bacterial outer membrane catalyze the uptake of iron complexes called ferric siderophores, and the assimilation of these metal chelates is a determinant of the pathogenesis of many species, including Escherichia coli, Salmonella typhi, Neisseria meningitidis, Vibrio cholerae and others. Unfortunately, the full molecular understanding of these transport reactions is lacking, which impairs our ability to conceive of potential inhibitors. Nevertheless, it's known that the E. coli outer membrane protein FepA accomplishes active iron transport by a process that involves extensive conformational motion. The proposed research capitalizes on this knowledge with a fluorescence spectroscopic assay that monitors uptake of the siderophore ferric enterobactin. The experiments will encompass high-throughput screening of validation and primary libraries of compounds at the University of Kansas High-Throughput Screening facility. The three libraries include standards and test chemicals that total ~17,500 unique molecules. These tests, performed in 384-well microtiter plates, will identify compounds that inhibit TonB-dependent iron transport by Gram- negative bacteria, creating a panel of candidates for further testing as antibiotics against bacterial pathogenesis.

描述（由适用提供）：微生物获得铁的能力是微生物侵入性和致病性的关键方面。革兰氏阴性细菌中的TONB依赖性转运蛋白外膜外膜催化了铁配合物的吸收，称为铁载体，这些金属樱桃的同化是许多物种发病机理的决定因素，包括大肠杆菌，salmonella typhi，salmonella typhi，neisseria meningitisis，vibrio chibio cholerae，其他。不幸的是，缺乏对这些转运反应的全部分子理解，这会损害我们想象潜在抑制剂的能力。然而，众所周知，大肠杆菌外膜蛋白FEPA通过涉及广泛的会议运动的过程来实现主动铁运输。拟议的研究通过荧光光谱评估来利用这一知识，该评估可以监测辅助铁的摄取。实验将包括堪萨斯大学高通量筛查设施的化合物验证和主要库的高通量筛选。这三个文库包括标准和测试化学物质，总计约17,500个独特的分子。这些测试在384孔微量滴定板上进行，将鉴定出革兰氏阴性细菌抑制TONB依赖的铁转运的化合物，从而创建了一组候选者，以进一步测试作为针对细菌发病机理的抗生素。