Vos Seychelle M, Lyubimov Artem Y, Hershey David M, Schoeffler Allyn J, Sengupta Sugopa, Nagaraja Valakunja, Berger James M
Department of Molecular and Cell Biology.
Deparment of Plant and Microbial Biology, University of California at Berkeley, Berkeley, California 94720, USA;
Genes Dev. 2014 Jul 1;28(13):1485-97. doi: 10.1101/gad.241984.114.
Precise control of supercoiling homeostasis is critical to DNA-dependent processes such as gene expression, replication, and damage response. Topoisomerases are central regulators of DNA supercoiling commonly thought to act independently in the recognition and modulation of chromosome superstructure; however, recent evidence has indicated that cells tightly regulate topoisomerase activity to support chromosome dynamics, transcriptional response, and replicative events. How topoisomerase control is executed and linked to the internal status of a cell is poorly understood. To investigate these connections, we determined the structure of Escherichia coli gyrase, a type IIA topoisomerase bound to YacG, a recently identified chromosomally encoded inhibitor protein. Phylogenetic analyses indicate that YacG is frequently associated with coenzyme A (CoA) production enzymes, linking the protein to metabolism and stress. The structure, along with supporting solution studies, shows that YacG represses gyrase by sterically occluding the principal DNA-binding site of the enzyme. Unexpectedly, YacG acts by both engaging two spatially segregated regions associated with small-molecule inhibitor interactions (fluoroquinolone antibiotics and the newly reported antagonist GSK299423) and remodeling the gyrase holoenzyme into an inactive, ATP-trapped configuration. This study establishes a new mechanism for the protein-based control of topoisomerases, an approach that may be used to alter supercoiling levels for responding to changes in cellular state.
精确控制超螺旋稳态对于诸如基因表达、复制和损伤应答等依赖DNA的过程至关重要。拓扑异构酶是DNA超螺旋的核心调节因子，通常认为它们在识别和调节染色体超结构时独立发挥作用；然而，最近的证据表明，细胞会严格调控拓扑异构酶的活性，以支持染色体动态变化、转录应答和复制事件。目前人们对拓扑异构酶的控制是如何执行以及与细胞内部状态如何关联了解甚少。为了研究这些联系，我们确定了大肠杆菌gyrase（一种与YacG结合的IIA型拓扑异构酶，YacG是最近鉴定出的一种染色体编码的抑制蛋白）的结构。系统发育分析表明，YacG经常与辅酶A（CoA）产生酶相关联，将该蛋白与代谢和应激联系起来。该结构以及支持性的溶液研究表明，YacG通过空间上阻断该酶的主要DNA结合位点来抑制gyrase。出乎意料的是，YacG通过同时结合与小分子抑制剂相互作用相关的两个空间分离区域（氟喹诺酮类抗生素和新报道的拮抗剂GSK299423）并将gyrase全酶重塑为无活性的、ATP捕获构型来发挥作用。这项研究建立了一种基于蛋白质的拓扑异构酶控制新机制，这种方法可用于改变超螺旋水平以应对细胞状态的变化。
