Although bacterial gyrase and topoisomerase IV have critical interactions with positively supercoiled DNA, little is known about the actions of these enzymes on overwound substrates. Therefore, the abilities of Bacillus anthracis and Escherichia coli gyrase and topoisomerase IV to relax and cleave positively supercoiled DNA were analyzed. Gyrase removed positive supercoils ∼10-fold more rapidly and more processively than it introduced negative supercoils into relaxed DNA. In time-resolved single-molecule measurements, gyrase relaxed overwound DNA with burst rates of ∼100 supercoils per second (average burst size was 6.2 supercoils). Efficient positive supercoil removal required the GyrA-box, which is necessary for DNA wrapping. Topoisomerase IV also was able to distinguish DNA geometry during strand passage and relaxed positively supercoiled substrates ∼3-fold faster than negatively supercoiled molecules. Gyrase maintained lower levels of cleavage complexes with positively supercoiled (compared with negatively supercoiled) DNA, whereas topoisomerase IV generated similar levels with both substrates. Results indicate that gyrase is better suited than topoisomerase IV to safely remove positive supercoils that accumulate ahead of replication forks. They also suggest that the wrapping mechanism of gyrase may have evolved to promote rapid removal of positive supercoils, rather than induction of negative supercoils.

中文翻译：

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旋回酶和拓扑异构酶IV对超螺旋DNA的活性

尽管细菌促旋酶和拓扑异构酶IV与超螺旋阳性DNA具有关键的相互作用，但对于这些酶在过度酶切的底物上的作用知之甚少。因此，炭疽杆菌和大肠杆菌的能力分析了旋松酶和拓扑异构酶IV来放松和切割阳性超螺旋DNA。与将负超螺旋引入松弛的DNA相比，旋涡酶能更快，更有效率地去除正超螺旋约10倍。在时间分辨的单分子测量中，回旋酶松弛了DNA，猝灭速率约为每秒100个超螺旋（平均猝发大小为6.2个超螺旋）。有效地清除超螺旋需要GyrA-box，这对于DNA包装是必需的。拓扑异构酶IV还能够区分链通过过程中的DNA几何形状，并且使正超螺旋底物的松弛速度比负超螺旋分子快约3倍。与正超螺旋（与负超螺旋）相比，回旋酶可保持较低水平的裂解复合物，拓扑异构酶IV在两种底物上产生相似的水平。结果表明，回旋酶比拓扑异构酶IV更适合安全地去除积聚在复制叉之前的阳性超螺旋。他们还暗示，回旋酶的包裹机制可能已经进化为促进快速清除阳性超螺旋，而不是诱导阴性超螺旋。