Activities of nucleoprotein complexes visualized in single-molecule experiments

项目摘要 对生物的机械理解需要我们理解蛋白质和DNA如何相互作用 共同产生功能性染色体。这种理解对于保护人类健康至关重要， 处理遗传疾病和对抗病原体。拟议项目的重点是 单分子分析允许直接观察生物分子相互作用，并可用于 详细分析蛋白质-DNA相互作用。该提案的目的包括仔细研究一个便利的 从DNA中去除蛋白质的解离“交换”机制，这表明对 蛋白质周转的传统描述，并将影响基因的广泛研究 调节蛋白和染色体结构蛋白。第二和第三个目标是研究 大的“染色体结构维持”（SMC）蛋白复合物介导的机制， 真核细胞中染色体的折叠。第二个目标是了解 Smc 5/6复合物的作用是对DNA的选择和压实，在分离中起着关键作用。 在细胞分裂过程中复制染色体，而第三个目的涉及直接显微镜 观察Smc 5/6压缩DNA的机制。第二和第三个目标还包括 Smc 5/6和更好理解的缩合复合物的并排比较。高度 提出的DNA加工机制的机制分析将使我们更好地理解 细胞如何解释，折叠和改变它们的基因组，从而更好地理解病理学， 这些功能受损，更好地了解如何在致病性疾病中靶向这些功能， 有机体

Project Summary Mechanistic understanding of living things requires our understanding of how proteins and DNA interact together to generate functional chromosomes. This understanding is central to preserving human health, dealing with genetic disorders, and fighting pathogenic organisms. The proposed projects are focused on single-molecule analyses which permit direct visualization of biomolecule interactions, and can be used to analyze protein-DNA interactions in detail. The aims of the proposal include careful study of a facilitated dissociation "exchange" mechanism for removal of proteins from DNA that suggests a major revision of conventional descriptions of protein turnover on DNA, and which will affect a wide range of studies of gene regulatory and chromosome structural proteins. The second and third aims are focused on study of mechanisms of large "Structural Maintenance of Chromosomes" (SMC) protein complexes which mediate the folding of chromosomes in eukaryote cells. The second aim concerns understanding the mechanics of compaction and DNA substrate selection by the Smc5/6 complex, which plays a key role in separating replicated chromosomes from one another during cell division, while the third aim concerns direct microscopy observation of the mechanism by which Smc5/6 compacts DNA. The second and third aim will also include side-by-side comparisons of Smc5/6 and the more well-understood condensin complex. The highly mechanistic analyses of DNA processing machinery that are proposed will give us a stronger understanding of how cells interpret, fold and change their genomes, leading to a better understanding of pathologies where those functions are impaired, and better understanding of how to target those functions in pathogenic organisms.