Bacteria deploy weapons to kill their neighbours during competition for resources and to aid survival within microbiomes. Colicins were the first such antibacterial system identified, yet how these bacteriocins cross the outer membrane (OM) of Escherichia coli is unknown. Here, by solving the structures of translocation intermediates via cryo-EM and by imaging toxin import, we uncover the mechanism by which the Tol-dependent nuclease colicin E9 (ColE9) crosses the bacterial OM. We show that threading of ColE9’s disordered N-terminal domain through two pores of the trimeric porin OmpF causes the colicin to disengage from its primary receptor, BtuB, and reorganises the translocon either side of the membrane. Subsequent import of ColE9 through the lumen of a single OmpF subunit is driven by the proton-motive force, which is delivered by the TolQ-TolR-TolA-TolB assembly. Our study answers longstanding questions, such as why OmpF is a better translocator than OmpC, and reconciles the mechanisms by which both Tol- and Ton-dependent bacteriocins cross the bacterial outer membrane.

中文翻译：

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孔蛋白穿线驱动受体脱离并通过大肠杆菌OmpF 建立主动大肠菌素转运

细菌在争夺资源时部署武器杀死它们的邻居，并帮助微生物组内的生存。大肠菌素是第一个确定的此类抗菌系统，但这些细菌素如何穿过大肠杆菌的外膜 (OM)是未知的。在这里，通过冷冻电镜和毒素导入成像解决易位中间体的结构，我们揭示了 Tol 依赖性核酸酶大肠杆菌素 E9 (ColE9) 穿过细菌 OM 的机制。我们表明，将 ColE9 的无序 N 末端结构域穿过三聚体孔蛋白 OmpF 的两个孔会导致大肠菌素与其主要受体 BtuB 脱离，并重组膜两侧的易位子。随后通过单个 OmpF 亚基的腔导入 ColE9 是由 TolQ-TolR-TolA-TolB 组件提供的质子动力驱动的。我们的研究回答了长期存在的问题，例如为什么 OmpF 是比 OmpC 更好的转运蛋白，并调和了 Tol 和 Ton 依赖性细菌素穿过细菌外膜的机制。