DNA supercoiling must be precisely regulated by topoisomerases to prevent DNA entanglement. The interaction of type IIA DNA topoisomerases with two DNA molecules, enabling the transport of one duplex through the transient double-stranded break of the other, remains elusive owing to structures derived solely from single linear duplex DNAs lacking topological constraints. Using cryo–electron microscopy, we solved the structure of Escherichia coli DNA gyrase bound to a negatively supercoiled minicircle DNA. We show how DNA gyrase captures a DNA crossover, revealing both conserved molecular grooves that accommodate the DNA helices. Together with molecular tweezer experiments, the structure shows that the DNA crossover is of positive chirality, reconciling the binding step of gyrase-mediated DNA relaxation and supercoiling in a single structure.

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大肠杆菌 DNA 旋转酶 DNA 交叉捕获的结构基础

DNA 超螺旋必须由拓扑异构酶精确调节，以防止 DNA 缠结。 IIA 型 DNA 拓扑异构酶与两个 DNA 分子的相互作用，使一个双链体能够通过另一个双链体的瞬时双链断裂进行运输，但由于仅源自缺乏拓扑约束的单个线性双链体 DNA 的结构，仍然难以捉摸。使用冷冻电子显微镜，我们解析了大肠杆菌DNA 旋转酶与负超螺旋小环 DNA 结合。我们展示了 DNA 旋转酶如何捕获 DNA 交叉，揭示容纳 DNA 螺旋的两个保守分子凹槽。结合分子镊子实验，该结构表明DNA交叉具有正手性，在单一结构中协调了旋转酶介导的DNA松弛和超螺旋的结合步骤。