Gene Center, Ludwig-Maximilians-Universität, 81377 Munich, Germany; Department of Biochemistry, Ludwig-Maximilians-Universität, 81377 Munich, Germany.
Gene Center, Ludwig-Maximilians-Universität, 81377 Munich, Germany; Department of Biochemistry, Ludwig-Maximilians-Universität, 81377 Munich, Germany.
Mol Cell. 2022 Sep 15;82(18):3513-3522.e6. doi: 10.1016/j.molcel.2022.07.019. Epub 2022 Aug 19.
DNA double-strand breaks (DSBs) threaten genome stability and are linked to tumorigenesis in humans. Repair of DSBs requires the removal of attached proteins and hairpins through a poorly understood but physiologically critical endonuclease activity by the Mre11-Rad50 complex. Here, we report cryoelectron microscopy (cryo-EM) structures of the bacterial Mre11-Rad50 homolog SbcCD bound to a protein-blocked DNA end and a DNA hairpin. The structures reveal that Mre11-Rad50 bends internal DNA for endonucleolytic cleavage and show how internal DNA, DNA ends, and hairpins are processed through a similar ATP-regulated conformational state. Furthermore, Mre11-Rad50 is loaded onto blocked DNA ends with Mre11 pointing away from the block, explaining the distinct biochemistries of 3' → 5' exonucleolytic and endonucleolytic incision through the way Mre11-Rad50 interacts with diverse DNA ends. In summary, our results unify Mre11-Rad50's enigmatic nuclease diversity within a single structural framework and reveal how blocked DNA ends and hairpins are processed.
DNA 双链断裂 (DSBs) 威胁着基因组的稳定性，并与人类的肿瘤发生有关。DSBs 的修复需要通过 Mre11-Rad50 复合物的一种尚未完全了解但在生理上至关重要的内切酶活性，去除附着的蛋白质和发夹。在这里，我们报告了与蛋白封闭的 DNA 末端和 DNA 发夹结合的细菌 Mre11-Rad50 同源物 SbcCD 的冷冻电子显微镜 (cryo-EM) 结构。这些结构揭示了 Mre11-Rad50 使内部 DNA 弯曲以进行内切酶切割，并展示了内部 DNA、DNA 末端和发夹如何通过类似的 ATP 调节构象状态进行处理。此外，Mre11-Rad50 与被封闭的 DNA 末端结合，Mre11 指向远离封闭物的方向，这解释了 3'→5' 外切核酸酶和内切核酸酶切割的不同生物化学特性，因为 Mre11-Rad50 与不同的 DNA 末端相互作用的方式。总之，我们的结果在单个结构框架内统一了 Mre11-Rad50 神秘的核酸酶多样性，并揭示了被封闭的 DNA 末端和发夹的处理方式。
