为进行双链DNA移行，II型拓扑异构酶产生一个共价酶剪切DNA复合物（即切割复合物）。尽管该复合物是必需的酶中间体，但它对基因组稳定性本质上是危险的。因此，切割复合物是数种临床相关的抗癌和抗菌药物的靶点。人类拓扑异构酶 IIα 和IIβ以及细菌陀螺酶在负超螺旋DNA底物上维持更高水平的切割复合物，而相反，细菌拓扑异构酶 IV 在区分DNA超螺旋手性方面的能力较弱。尽管超螺旋几何对 II 类拓扑异构酶的活性非常重要，但在DNA切割期间的超螺旋手性识别基础尚未被表征。根据台面实验和快速减速流动动力学实验的结果，切割前速率是影响 II型拓扑异构酶α/IIβ，陀螺酶和拓扑异构酶IV区分超螺旋手性的决定因素，在存在抗癌/抗菌药物的情况下，该能力可以通过与负超螺旋DNA形成更稳定的切割复合物来增强。最后，酶介导的DNA连接速率并不对DNA超螺旋几何识别在切割过程中起作用。我们的研究结果更深入地揭示了 II型拓扑异构酶如何识别其DNA底物。本文由牛津大学出版社代表核酸研究于2023年发表。

To perform double-stranded DNA passage, type II topoisomerases generate a covalent enzyme-cleaved DNA complex (i.e. cleavage complex). Although this complex is a requisite enzyme intermediate, it is also intrinsically dangerous to genomic stability. Consequently, cleavage complexes are the targets for several clinically relevant anticancer and antibacterial drugs. Human topoisomerase IIα and IIβ and bacterial gyrase maintain higher levels of cleavage complexes with negatively supercoiled over positively supercoiled DNA substrates. Conversely, bacterial topoisomerase IV is less able to distinguish DNA supercoil handedness. Despite the importance of supercoil geometry to the activities of type II topoisomerases, the basis for supercoil handedness recognition during DNA cleavage has not been characterized. Based on the results of benchtop and rapid-quench flow kinetics experiments, the forward rate of cleavage is the determining factor of how topoisomerase IIα/IIβ, gyrase and topoisomerase IV distinguish supercoil handedness in the absence or presence of anticancer/antibacterial drugs. In the presence of drugs, this ability can be enhanced by the formation of more stable cleavage complexes with negatively supercoiled DNA. Finally, rates of enzyme-mediated DNA ligation do not contribute to the recognition of DNA supercoil geometry during cleavage. Our results provide greater insight into how type II topoisomerases recognize their DNA substrates.Published by Oxford University Press on behalf of Nucleic Acids Research 2023.