4.8 Article

Changing protein-DNA interactions promote ORC binding- site exchange during replication origin licensing

出版社

NATL ACAD SCIENCES
DOI: 10.1073/pnas.2305556120

关键词

Mcm2-7 helicase; origin recognition complex (ORC); origin licensing; DNA replication initiation; single-molecule FRET

向作者/读者索取更多资源

This study reveals the importance of dynamic protein-DNA interactions in ensuring bidirectional DNA replication during origin licensing. The results show that ORC can ensure head-to-head helicase alignment by sequentially loading Mcm2-7, and that controlled DNA sliding allows ORC to access secondary DNA-binding sites at different locations relative to the initial binding site.
During origin licensing, the eukaryotic replicative helicase Mcm2-7 forms head-to-head double hexamers to prime origins for bidirectional replication. Recent single-molecule and structural studies revealed that one molecule of the helicase loader ORC (origin recognition complex) can sequentially load two Mcm2-7 hexamers to ensure proper head-to- head helicase alignment. To perform this task, ORC must release from its initial high-affinity DNA-binding site and flip to bind a weaker, inverted DNA site. However, the mechanism of this binding-site switch remains unclear. In this study, we used single-molecule Forster resonance energy transfer to study the changing interactions between DNA and ORC or Mcm2-7. We found that the loss of DNA bending that occurs during DNA deposition into the Mcm2-7 central channel increases the rate of ORC dissociation from DNA. Further studies revealed temporally controlled DNA sliding of helicase-loading intermediates and that the first sliding complex includes ORC, Mcm2-7, and Cdt1. We demonstrate that sequential events of DNA unbending, Cdc6 release, and sliding lead to a stepwise decrease in ORC stability on DNA, facilitating ORC dissociation from its strong binding site during site switching. In addition, the controlled sliding we observed provides insight into how ORC accesses secondary DNA-binding sites at different locations relative to the initial binding site. Our study highlights the importance of dynamic protein-DNA interactions in the loading of two oppositely oriented Mcm2-7 helicases to ensure bidirectional DNA replication.

作者

我是这篇论文的作者
点击您的名字以认领此论文并将其添加到您的个人资料中。

评论

主要评分

4.8
评分不足

次要评分

新颖性
-
重要性
-
科学严谨性
-
评价这篇论文

推荐

暂无数据
暂无数据