Modulated control of DNA supercoiling balance by the DNA-wrapping domain of bacterial gyrase
Modulated control of DNA supercoiling balance by the DNA-wrapping domain of bacterial gyrase
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DOI：
10.1093/nar/gkz1230
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发表时间：
2020-02-28
影响因子：
14.9
通讯作者：
Berger, James M.
中科院分区：
文献类型：
--
作者：
Hobson, Matthew J.;Bryant, Zev;Berger, James M.
DNA 旋转酶的负超螺旋对于维持细菌中的染色体压缩、转录编程和遗传完整性至关重要。关于不同物种的回旋酶如何在动力学、效率和负超螺旋程度方面产生巨大差异，仍然存在疑问。为了探讨这个问题，我们分析了大肠杆菌促旋酶（“CTD”）的 GyrA 亚基的 C 端 DNA 包裹结构域中的同源定向突变。添加或去除选定的保守碱性残基会显着影响酶的核苷酸依赖性 DNA 包裹和超螺旋。 CTD-DNA 相互作用的减弱会减慢超螺旋，损害 DNA 依赖性 ATP 水解，并限制 DNA 超螺旋的程度，同时增强去连接和超螺旋松弛。相反，加强 DNA 包裹不会导致更广泛的超螺旋 DNA 产物，但会部分解开 ATP 周转与链通道的耦合，表现为无效循环。我们的研究结果表明，大肠杆菌旋转酶的催化循环以高热力学效率运行，并且 CTD 包裹 DNA 的稳定性为 DNA 超螺旋引入提供了一个限制，超出该限制，链通道将与 ATP 依赖性超螺旋松弛竞争。这些结果强调了旋转酶可以以物种特异性方式进化出不同的稳态超螺旋设定点的方法。
Negative supercoiling by DNA gyrase is essential for maintaining chromosomal compaction, transcriptional programming, and genetic integrity in bacteria. Questions remain as to how gyrases from different species have evolved profound differences in their kinetics, efficiency, and extent of negative supercoiling. To explore this issue, we analyzed homologydirected mutations in the C-terminal, DNA-wrapping domain of the GyrA subunit of Escherichia coli gyrase (the `CTD'). The addition or removal of select, conserved basic residues markedly impacts both nucleotide-dependent DNA wrapping and supercoiling by the enzyme. Weakening CTD-DNA interactions slows supercoiling, impairs DNA-dependent ATP hydrolysis, and limits the extent of DNA supercoiling, while simultaneously enhancing decatenation and supercoil relaxation. Conversely, strengthening DNA wrapping does not result in a more extensively supercoiled DNA product, but partially uncouples ATP turnover from strand passage, manifesting in futile cycling. Our findings indicate that the catalytic cycle of E. coli gyrase operates at high thermodynamic efficiency, and that the stability of DNA wrapping by the CTD provides one limit to DNA supercoil introduction, beyond which strand passage competes with ATP-dependent supercoil relaxation. These results highlight a means by which gyrase can evolve distinct homeostatic supercoiling setpoints in a species-specific manner.