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首页> 外文期刊>Proceedings of the National Academy of Sciences of the United States of America >Observing spontaneous branch migration of Holliday junctions one step at a time
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Observing spontaneous branch migration of Holliday junctions one step at a time

机译：一次观察霍利迪结的自发分支迁移

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摘要

Genetic recombination occurs between homologous DNA molecules via a four-way (Holliday) junction intermediate. This ancient and ubiquitous process is important for the repair of double-stranded breaks, the restart of stalled replication forks, and the creation of genetic diversity. Once formed, the four-way junction alone can undergo the stepwise exchange of base pairs known as spontaneous branch migration. Conventional ensemble assays, useful for finding average migration rates over long sequences, have been unable to examine the affect of sequence and structure on the migration process. Here, we present a single-molecule spontaneous branch migration assay with single-base pair resolution in a study of individual DNA junctions that can undergo one step of migration. Junctions exhibit markedly different dynamics of exchange between stacking conformers depending on the point of strand exchange, allowing the moment at which branch migration occurs to be detected. The free energy landscape of spontaneous branch migration is found to be highly nonuniform and governed by two types of sequence-dependent barriers, with unmediated local migration being up to 10 times more rapid than the previously deduced average rate.
机译：遗传重组发生在同源DNA分子之间，通过四向（Holliday）连接中间物发生。这个古老而无处不在的过程对于修复双链断裂，停滞的复制叉的重新启动以及遗传多样性的建立非常重要。一旦形成，单独的四向连接点就可以逐步进行碱基对的逐步交换，即自发分支迁移。传统的集合检测法可用于寻找长序列的平均迁移率，但无法检查序列和结构对迁移过程的影响。在这里，我们提出了具有单个碱基对分辨率的单分子自发分支迁移测定法，用于研究可能经历迁移步骤的单个DNA连接。取决于链交换的点，结点在堆叠构象异构体之间表现出明显不同的交换动力学，从而可以检测到发生分支迁移的时刻。发现自发分支迁移的自由能态非常不均匀，并受两种类型的序列依赖性屏障的控制，未介导的局部迁移的速度比先前推导的平均速率快10倍。

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