Institute for Physical Chemistry, University of Münster, D-48149 Münster, Germany.
J Biol Chem. 2012 Mar 30;287(14):10916-21. doi: 10.1074/jbc.M111.308247. Epub 2012 Feb 16.
DNA gyrase catalyzes ATP-dependent negative supercoiling of DNA by a strand passage mechanism that requires coordinated opening and closing of three protein interfaces, the N-, DNA-, and C-gates. ATP binding to the GyrB subunits of gyrase causes dimerization and N-gate closure. The closure of the N-gate is a key step in the gyrase catalytic cycle, as it captures the DNA segment to be transported and poises gyrase toward strand passage. We show here that K(+) ions are required for DNA supercoiling but are dispensable for ATP-independent DNA relaxation. Although DNA binding, distortion, wrapping, and DNA-induced narrowing of the N-gate occur in the absence of K(+), nucleotide-induced N-gate closure depends on their presence. Our results provide evidence that K(+) ions relay small conformational changes in the nucleotide-binding pocket to the formation of a tight dimer interface at the N-gate by connecting regions from both GyrB monomers and suggest an important role for K(+) in synchronization of N-gate closure and DNA-gate opening.
DNA 拓扑异构酶通过链迁移机制催化 ATP 依赖性的 DNA 负超螺旋化，该机制需要三个蛋白质界面（N 门、DNA 门和 C 门）的协调打开和关闭。ATP 与拓扑异构酶的 GyrB 亚基结合导致二聚体形成和 N 门关闭。N 门的关闭是拓扑异构酶催化循环中的关键步骤，因为它捕获要转运的 DNA 片段，并使拓扑异构酶向链迁移方向转变。我们在这里表明，K(+) 离子对于 DNA 超螺旋化是必需的，但对于 ATP 非依赖性的 DNA 松弛是可有可无的。尽管在没有 K(+) 的情况下，DNA 结合、扭曲、包裹和 DNA 诱导的 N 门变窄会发生，但核苷酸诱导的 N 门关闭取决于它们的存在。我们的结果提供了证据，表明 K(+) 离子通过连接来自两个 GyrB 单体的区域，将核苷酸结合口袋中的小构象变化传递到 N 门的紧密二聚体界面的形成，这表明 K(+) 在 N 门关闭和 DNA 门打开的同步化中起着重要作用。
