Transcription-coupled repair (TCR) is a sub-pathway of nucleotide excision repair (NER) that preferentially removes lesions from the template-strand (t-strand) that stall RNA polymerase (RNAP) elongation complexes (EC). Mfd mediates TCR in bacteria by removing the stalled RNAP concealing the lesion and recruiting Uvr(A)BC. We used cryo-electron microscopy to visualize Mfd engaging with a stalled EC and attempting to dislodge the RNAP. We visualized seven distinct Mfd-EC complexes in both ATP and ADP-bound states. The structures explain how Mfd is remodeled from its repressed conformation, how the UvrA-interacting surface of Mfd is hidden during most of the remodeling process to prevent premature engagement with the NER pathway, how Mfd alters the RNAP conformation to facilitate disassembly, and how Mfd forms a processive translocation complex after dislodging the RNAP. Our results reveal an elaborate mechanism for how Mfd kinetically discriminates paused from stalled ECs and disassembles stalled ECs to initiate TCR.

中文翻译：

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Mfd转位酶转录复合物破坏的结构基础

转录偶联修复 (TCR) 是核苷酸切除修复 (NER) 的一个子途径，它优先从模板链 (t 链) 中去除使 RNA 聚合酶 (RNAP) 延伸复合物 (EC) 停滞的损伤。Mfd 通过去除隐藏病变的停滞 RNAP 并招募 Uvr(A)BC 来介导细菌中的 TCR。我们使用低温电子显微镜来观察 Mfd 与停滞的 EC 接合并试图移除 RNAP。我们在 ATP 和 ADP 结合状态下可视化了七种不同的 Mfd-EC 复合物。这些结构解释了 Mfd 如何从其受抑制的构象中重塑，在大多数重塑过程中如何隐藏 Mfd 的 UvrA 相互作用表面以防止与 NER 途径过早接合，Mfd 如何改变 RNAP 构象以促进拆卸，以及 Mfd 在去除 RNAP 后如何形成进行性易位复合物。我们的研究结果揭示了 Mfd 如何在动力学上区分暂停和停滞的 EC 并拆卸停滞的 EC 以启动 TCR 的详细机制。