Department of Biophysics, Zhejiang University School of Medicine, Hangzhou 310058, China.
Department of Pathology, Sir Run Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou 310058, China.
Nucleic Acids Res. 2020 Nov 18;48(20):11762-11772. doi: 10.1093/nar/gkaa904.
Mfd-dependent transcription termination plays an important role in transcription-coupled DNA repair, transcription-replication conflict resolution, and antimicrobial resistance development. Despite extensive studies, the molecular mechanism of Mfd-dependent transcription termination in bacteria remains unclear, with several long-standing puzzles. How Mfd is activated by stalled RNA polymerase (RNAP) and how activated Mfd translocates along the DNA are unknown. Here, we report the single-particle cryo-electron microscopy structures of T. thermophilus Mfd-RNAP complex with and without ATPγS. The structures reveal that Mfd undergoes profound conformational changes upon activation, contacts the RNAP β1 domain and its clamp, and pries open the RNAP clamp. These structures provide a foundation for future studies aimed at dissecting the precise mechanism of Mfd-dependent transcription termination and pave the way for rational drug design targeting Mfd for the purpose of tackling the antimicrobial resistance crisis.
Mfd 依赖的转录终止在转录偶联的 DNA 修复、转录-复制冲突解决以及抗菌药物耐药性发展中发挥着重要作用。尽管已经进行了广泛的研究，但细菌中 Mfd 依赖的转录终止的分子机制仍不清楚，存在一些长期存在的难题。Mfd 如何被停滞的 RNA 聚合酶（RNAP）激活，以及激活的 Mfd 如何沿着 DNA 迁移，这些都还不清楚。在这里，我们报道了含有和不含有 ATPγS 的 T. thermophilus Mfd-RNAP 复合物的单颗粒冷冻电镜结构。这些结构表明，Mfd 在被激活时会发生深刻的构象变化，与 RNAP 的β1 结构域及其夹子接触，并撬动 RNAP 夹子。这些结构为进一步研究 Mfd 依赖的转录终止的精确机制提供了基础，并为针对 Mfd 的合理药物设计铺平了道路，以应对抗菌药物耐药性危机。
