Department of Molecular Biosciences, Northwestern University, Evanston, United States.
Division of Physical Biochemistry, MRC National Institute for Medical Research, London, United Kingdom.
Elife. 2018 Nov 20;7:e41215. doi: 10.7554/eLife.41215.
Gyrase is a unique type IIA topoisomerase that uses ATP hydrolysis to maintain the negatively supercoiled state of bacterial DNA. In order to perform its function, gyrase undergoes a sequence of conformational changes that consist of concerted gate openings, DNA cleavage, and DNA strand passage events. Structures where the transported DNA molecule (T-segment) is trapped by the A subunit have not been observed. Here we present the cryoEM structures of two oligomeric complexes of open gyrase A dimers and DNA. The protein subunits in these complexes were solved to 4 Å and 5.2 Å resolution. One of the complexes traps a linear DNA molecule, a putative T-segment, which interacts with the open gyrase A dimers in two states, representing steps either prior to or after passage through the DNA-gate. The structures locate the T-segment in important intermediate conformations of the catalytic cycle and provide insights into gyrase-DNA interactions and mechanism.
回旋酶是一种独特的 IIA 拓扑异构酶，利用 ATP 水解来维持细菌 DNA 的负超螺旋状态。为了发挥其功能，回旋酶经历了一系列构象变化，包括协同门的打开、DNA 切割和 DNA 链通过事件。尚未观察到被 A 亚基捕获的转运 DNA 分子（T 片段）的结构。在这里，我们展示了两个开放回旋酶 A 二聚体和 DNA 的寡聚复合物的 cryoEM 结构。这些复合物中的蛋白质亚基分别解析到 4 Å 和 5.2 Å 的分辨率。其中一个复合物捕获了一个线性 DNA 分子，即一个假定的 T 片段，该片段与开放的回旋酶 A 二聚体以两种状态相互作用，分别代表穿过 DNA 门之前或之后的步骤。这些结构将 T 片段定位在催化循环的重要中间构象中，并提供了对回旋酶-DNA 相互作用和机制的深入了解。
