Morgan Ian L, Jian Jeffrey Y, Osheroff Neil, Neuman Keir C
National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892, USA.
Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, TN 37232, USA.
bioRxiv. 2025 Jul 28:2025.07.28.667256. doi: 10.1101/2025.07.28.667256.
The type II topoisomerases, gyrase and topoisomerase IV, are essential enzymes in nearly all bacteria and are the targets of fluoroquinolones, which are some of the most widely prescribed broad-spectrum antibacterials in clinical use. As part of their catalytic cycle, gyrase and topoisomerase IV transiently cleave DNA in a sequence-dependent manner. However, it is unclear whether this sequence-dependence is species-specific. Therefore, using our recently developed SHAN-seq method, we mapped and compared cleavage sites for type II topoisomerases from three different pathogenic bacterial species, , , and in the presence of the fluoroquinolone, ciprofloxacin. We found that the enzymes have substantially different DNA cleavage specificities that vary between gyrase and topoisomerase IV, across species, with supercoil chirality, and in response to ciprofloxacin. Our results demonstrate that bacterial species fine-tune the DNA cleavage specificity of their type II topoisomerases. This finding suggests that cleavage specificity may play important physiological roles and, in turn, may affect the susceptibility of bacteria to fluoroquinolone antibacterials.
II型拓扑异构酶，即促旋酶和拓扑异构酶IV，是几乎所有细菌中的必需酶，也是氟喹诺酮类药物的作用靶点，氟喹诺酮类药物是临床使用中处方最广泛的一些广谱抗菌药物。作为其催化循环的一部分，促旋酶和拓扑异构酶IV以序列依赖的方式瞬时切割DNA。然而，尚不清楚这种序列依赖性是否具有物种特异性。因此，我们使用最近开发的SHAN-seq方法，在氟喹诺酮类药物环丙沙星存在的情况下，对三种不同致病细菌物种（分别为……、……和……，此处原文未给出具体物种名称）的II型拓扑异构酶的切割位点进行了定位和比较。我们发现，这些酶具有显著不同的DNA切割特异性，这些特异性在促旋酶和拓扑异构酶IV之间、不同物种之间、超螺旋手性之间以及对环丙沙星的反应中均有所不同。我们的结果表明，细菌物种会微调其II型拓扑异构酶的DNA切割特异性。这一发现表明，切割特异性可能发挥重要的生理作用，进而可能影响细菌对氟喹诺酮类抗菌药物的敏感性。
