Single-molecule visualization and mechanisms of DNA recombination and intersecting DNA transactions
单分子可视化以及 DNA 重组和交叉 DNA 事务的机制
基本信息
- 批准号:10374054
- 负责人:
- 金额:$ 80.25万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2019
- 资助国家:美国
- 起止时间:2019-04-01 至 2024-03-31
- 项目状态:已结题
- 来源:
- 关键词:AddressBacteriaBehaviorBiochemicalBiological ProcessBuffersChromosome StructuresChromosomesComplexDNADNA RepairDNA SequenceDNA biosynthesisDNA-Protein InteractionDetectionEnzymesEscherichia coli ProteinsFilamentGenetic RecombinationImageIndividualMaintenanceMediator of activation proteinNucleoproteinsProcessProteinsReactionRegulationSaccharomyces cerevisiaeSequence HomologyTimeTransactVisualizationbiophysical techniqueschromatin remodelinghelicasehomologous recombinationreconstitutionsingle molecule
项目摘要
Project Description
The broad objective of this proposal is to understand the mechanism of homologous recombination and its
relationship to chromosome maintenance in both Bacteria and Eukarya. Our approach is to reconstitute
increasingly more complex reactions that recapitulate the steps of the homologous recombination process,
using purified proteins from Escherichia coli and from Saccharomyces cerevisiae, and to use sophisticated
biochemical and biophysical methods of analysis. In particular, we will use single-molecule visualization to
image key steps and intermediates of these complex biological processes. We have developed new
experimental strategies for studying protein-DNA interactions at the single-molecule level. We devised several
approaches wherein we could easily manipulate an individual DNA molecule and expose it to a variety of
different proteins, enzymes, and buffers. Using fluorescent detection, we can image individual proteins on DNA
or detect their action on individual DNA molecules, typically in real-time. We propose to continue to use these
approaches to study unresolved aspects of recombinational DNA repair and its interface with the important
biological processes of chromatin remodeling and DNA replication. We seek to continue to address unresolved
questions regarding the dynamic behavior of RecA/Rad51 nucleoprotein filaments and their regulation by
protein mediators and modulators;; the mechanism by which RecA and Rad51 find DNA sequence homology;;
the reconstitution increasingly more complex recombination processes using purified proteins;; the mechanism
and function of DNA helicases and their partner proteins;; and understanding the mechanism of DNA
transactions that intersect with recombination, namely, chromatin remodeling and DNA replication.
项目描述
这项建议的主要目的是了解同源重组的机制及其对基因表达的影响。
与细菌和真核生物染色体维持的关系。我们的方法是重建
越来越复杂的反应重演了同源重组过程的步骤,
使用来自大肠杆菌和酿酒酵母的纯化蛋白质,并使用复杂的
生物化学和生物物理学的分析方法。特别是,我们将使用单分子可视化,
对这些复杂生物过程的关键步骤和中间体进行成像。我们开发了新的
在单分子水平上研究蛋白质-DNA相互作用的实验策略。我们设计了几个
我们可以很容易地操纵一个单独的DNA分子,并将其暴露在各种各样的环境中,
不同的蛋白质,酶和缓冲液。使用荧光检测,我们可以在DNA上成像单个蛋白质
或检测它们对单个DNA分子的作用,通常是在真实的实时。我们建议继续使用这些
研究重组DNA修复的未解决问题及其与重要的
染色质重塑和DNA复制的生物过程。我们寻求继续解决未解决的
关于RecA/Rad 51核蛋白丝的动态行为及其通过
蛋白质介质和调节剂; RecA和Rad 51发现DNA序列同源性的机制;
重组越来越复杂的重组过程中使用纯化的蛋白质;
DNA解旋酶及其伴侣蛋白的功能;了解和理解DNA解旋酶的作用机制
与重组交叉的交易,即染色质重塑和DNA复制。
项目成果
期刊论文数量(0)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
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Stephen Charles Kowalczykowski其他文献
Stephen Charles Kowalczykowski的其他文献
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{{ truncateString('Stephen Charles Kowalczykowski', 18)}}的其他基金
Functions of BRCA1 and RAD51 Paralogs in Genome Stability and Tumor Suppression via Homologous Recombination
BRCA1 和 RAD51 旁系同源物在基因组稳定性和同源重组肿瘤抑制中的功能
- 批准号:
10565421 - 财政年份:2023
- 资助金额:
$ 80.25万 - 项目类别:
Single-molecule visualization and mechanisms of DNA recombination and intersecting DNA transactions
单分子可视化以及 DNA 重组和交叉 DNA 事务的机制
- 批准号:
10595678 - 财政年份:2019
- 资助金额:
$ 80.25万 - 项目类别:
Single-molecule visualization and mechanisms of DNA recombination and intersecting DNA transactions
单分子可视化以及 DNA 重组和交叉 DNA 事务的机制
- 批准号:
9900833 - 财政年份:2019
- 资助金额:
$ 80.25万 - 项目类别:
Molecular Functions of Recombination in Genome Stability and Tumor Suppression
重组在基因组稳定性和肿瘤抑制中的分子功能
- 批准号:
9311800 - 财政年份:2017
- 资助金额:
$ 80.25万 - 项目类别:
Molecular Functions of Recombination in Genome Stability and Tumor Suppression
重组在基因组稳定性和肿瘤抑制中的分子功能
- 批准号:
10092116 - 财政年份:2017
- 资助金额:
$ 80.25万 - 项目类别:
Single-Molecule Assembly of Protein-DNA Complexes
蛋白质-DNA 复合物的单分子组装
- 批准号:
7937183 - 财政年份:2009
- 资助金额:
$ 80.25万 - 项目类别:
Single-Molecule Assembly of Protein-DNA Complexes
蛋白质-DNA 复合物的单分子组装
- 批准号:
8338808 - 财政年份:2002
- 资助金额:
$ 80.25万 - 项目类别:
Single-Molecule Assembly of Protein-DNA Complexes
蛋白质-DNA 复合物的单分子组装
- 批准号:
9276464 - 财政年份:2002
- 资助金额:
$ 80.25万 - 项目类别:
Single-Molecule Assembly of Protein-DNA Complexes
蛋白质-DNA 复合物的单分子组装
- 批准号:
8690091 - 财政年份:2002
- 资助金额:
$ 80.25万 - 项目类别:
Single-Molecule Assembly of Protein-DNA Complexes
蛋白质-DNA 复合物的单分子组装
- 批准号:
6422004 - 财政年份:2002
- 资助金额:
$ 80.25万 - 项目类别:
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