Understanding Homology in Genetic Recombination

了解基因重组中的同源性

基本信息

批准号：
7624427
负责人：
SCOTT F SINGLETON
金额：
$ 32.4万
依托单位：
UNIV OF NORTH CAROLINA CHAPEL HILL
依托单位国家：
美国
项目类别：
财政年份：
1998
资助国家：
美国
起止时间：
1998-10-01 至 2009-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7624427
关键词：
ATP Hydrolysis Achievement Address Adjuvant Affect Affinity Amino Acid Sequence Anti-Bacterial Agents Antibiotic Resistance Antibiotics Attenuated Bacillus subtilis Bacteria Bacterial Drug Resistance Binding Sites Biochemical Biological Assay Biological Process Cells Cellular Assay Communicable Diseases Competence Complex Coupled DNA DNA Binding Data Depth Development Escherichia coli F Factor Filament Funding Gene Mutation Gene Transfer Genes Genetic Recombination In Vitro Individual Interdisciplinary Study Kinetics Laboratories Legal patent Life Ligand Binding Light Literature Mediating Methods Modeling Molecular Mutagenesis Mutation Neisseria gonorrhoeae Organism Pathogenicity Peptide Sequence Determination Phenotype Physiological Play Polymerase Gene Process Property Protocols documentation Pseudomonas aeruginosa Public Health Range Rate Reaction Rec A Recombinases Recombinants Research Resolution Role SOS Response Son of Sevenless Proteins Stress Structure Structure-Activity Relationship Testing Therapeutic Index Thermodynamics Translating Uncertainty Variant Zidovudine analog bactericide biodefense biothreat cell killing chemotherapy homologous recombination in vivo inhibitor/antagonist insight intein interest novel pathogen pathogenic bacteria programs rapid technique research study response small molecule tool transmission process

项目摘要

Antibiotic resistance is an escalating problem for modern chemotherapy of bacterial infectious diseases, and, in combination with the deteriorating pipeline of new antibacterials, is creating a clear and urgent danger to public health and national biodefense. Although the mechanisms that facilitate the de novo development, clonal spread, and horizontal transfer of resistance factors are not fully understood, the rapid rate at which antibiotic-resistant bacteria arise is likely due to a combination of mutations introduced during SOS mutagenesis and gene transfer between organisms. Recently, the Escherichia coli RecA protein¿s activities in SOS induction and homologous recombination have revealed RecA as a crucial player in these phenomena. A combination of primary literature, patent data, and unpublished results demonstrate that RecA mediates a range of phenomena related to bacterial pathogenecity, particularly the development and transmission of antibiotic resistance genes. Although the high conservation of RecA among bacterial species compellingly suggests the possibility that RecA may play similar roles in species other than E. coli, many questions remain as to how the properties of individual variants are related to their specific biological functions. To delimit possible models for the RecA-mediated activities that occur in pathogenic bacteria, we propose three Specific Aims to exploit our recently developed methods for rapid, parallel purification and rigorous characterization of RecA proteins to elucidate the relationships between RecA structure, in vitro activities, and physiologic functions. Briefly, we will (1) systematically define and evaluate structure-function relationships among RecA proteins from 31 pathogenic bacteria using biochemical and cellular activity assays; (2) provide insight into the species-specific molecular mechanisms of RecADNA filament activation using directed mutagenesis and substrate analogs; and (3) demonstrate that RecA effectors can be delivered into living bacteria to produce physiological consequences. The successful realization of the Aims will provide (1) substantial and novel insights into the molecular mechanisms by which different RecA proteins from select bacterial pathogens carry out their biological functions; (2) a novel microbiological toolbox that will be central to teasing apart the various roles of RecA in pathogenicity; and (3) novel methods for the delivery of small-molecule RecA effectors into bacterial pathogens.

抗生素耐药性是现代细菌化疗面临的一个日益严重的问题传染病，并且结合新的不断恶化的管道抗菌药物正在对公共卫生和国家造成明显而紧迫的危险虽然促进从头发展的机制，克隆。传播和横向转移的阻力因素尚未完全了解，迅速抗生素耐药性细菌出现的速度可能是由于突变的组合最近在 SOS 诱变和生物体之间的基因转移过程中引入。大肠杆菌 RecA 蛋白¿ SOS 诱导和同源活性重组表明 RecA 在这些现象中发挥着关键作用。原始文献、专利数据和未发表结果的结合表明 RecA 介导一系列与细菌致病性相关的现象，特别是抗生素抗性基因的发展和传播虽然较高。 RecA 在细菌物种中的保守性令人信服地表明以下可能性： RecA 可能在大肠杆菌以外的物种中发挥类似的作用，但仍有许多问题各个变体的特性如何与其特定的生物学功能相关。界定致病性中发生的 RecA 介导的活动的可能模型细菌，我们提出了三个具体目标来利用我们最近开发的方法 RecA 蛋白的快速、平行纯化和严格表征，以阐明 RecA结构、体外活性和生理功能之间的关系。简而言之，我们将（1）系统地定义和评估结构-功能关系利用生化和细胞活性对来自 31 种病原菌的 RecA 蛋白进行分析 (2) 深入了解 RecADNA 的物种特异性分子机制使用定向诱变和底物类似物激活丝；和（3）证明 RecA 效应器可以被传递到活细菌中以产生目标的成功实现将提供（1）对不同 RecA 的分子机制有实质性和新颖的见解来自选定细菌病原体的蛋白质发挥其生物学功能；（2）一种新的微生物学工具箱将成为梳理 RecA 的各种作用的核心致病性；(3) 递送小分子 RecA 效应物的新方法转化为细菌性病原体。