Understanding DNA transport by topo IV and gyrase to counter antimicrobial resistance

了解拓扑 IV 和旋转酶的 DNA 转运以对抗抗菌药物耐药性

• 批准号: MR/T000848/1
• 负责人: Larry Mark Fisher
• 金额: $ 107.91万
• 依托单位: St George's, University of London
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2019
• 资助国家: 英国
• 起止时间: 2019 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=MR%2FT000848%2F1
• 关键词: Understanding; DNA; transport; topo; IV

Antibiotic resistance is a worldwide problem and a major challenge for the treatment of infection. Bacteria are becoming resistant to our most important antibiotics jeopardising even straightforward medical procedures. One serious threat is posed by Streptococcus pneumoniae, a global pathogen that causes life-threatening pneumonia and meningitis in childern and the elderly. Increasingly, S. pneumoniae isolates are found to be resistant to penicillins and to fluoroquinolones, which are key members of our antibiotic arsenal. It is known that fluoroquinolones interfere with DNA breakage by gyrase and topoisomerase (topo) IV, two enzymes that untangle DNA and are required for bacterial DNA replication and growth. Remarkably, these enzymes make a transient break in DNA and cross another DNA through the break. By this means, gyrase can remove twists when DNA is copied in the cell, and topo IV can unlink tangled chromosomes ahead of cell division. By using X-ray crystallography to solve the structure of these enzymes bound to DNA and fluoroquinolones, we now understand in detail how quinolones bind and trap the DNA break and how resistance arises. However, almost nothing is known about the nature of the transported DNA, how it is recognised, captured and then crossed through the DNA break. To fill this gap in knowledge, and guided by our recent structure of a topo IV-transport DNA complex, we shall use a combination of techniques including X-ray crystallography, cryo-electron microscopy, fluorescence and protein biochemistry to establish the mechanism of DNA transport by topo IV and gyrase from S. pneumoniae and from Klebsiella pneumoniae, another pathogen highly resistant to quinolones. Completion of the work will be a major contribution in understanding these fascinating molecular machines and will provide new opportunites for design of new drugs that overcome resistance by targeting DNA transport. New drugs will be essential in addressing the global emergency of antimicrobial resistance.

抗生素耐药性是一个世界性的问题，也是治疗感染的主要挑战。细菌正在对我们最重要的抗生素产生抗药性，甚至危及直接的医疗程序。一种严重威胁来自肺炎链球菌，这是一种全球性病原体，可在儿童和老年人中引起危及生命的肺炎和脑膜炎。越来越多的肺炎链球菌分离株被发现对青霉素和氟喹诺酮类药物耐药，这是我们抗生素库的主要成员。众所周知，氟喹诺酮类药物干扰螺旋酶和拓扑异构酶（topo） IV的DNA断裂，这两种酶能解开DNA，是细菌DNA复制和生长所必需的。值得注意的是，这些酶在DNA中制造一个短暂的断裂，并通过这个断裂使另一个DNA交叉。通过这种方法，当DNA在细胞中复制时，gyrase可以去除扭曲，topo IV可以在细胞分裂前解开缠结的染色体。通过使用x射线晶体学来解决这些与DNA和氟喹诺酮类药物结合的酶的结构，我们现在详细了解了喹诺酮类药物是如何结合和捕获DNA断裂的，以及耐药性是如何产生的。然而，对于运输DNA的性质，以及它是如何被识别、捕获并通过DNA断裂交叉的，几乎一无所知。为了填补这一知识空白，并在我们最近构建的topo IV-transport DNA复合物结构的指导下，我们将结合x射线晶体学、冷冻电子显微镜、荧光和蛋白质生物化学等技术，建立来自肺炎链球菌和肺炎克雷伯菌（另一种对喹诺酮类药物高度耐药的病原体）的topo IV和gyrase的DNA转运机制。这项工作的完成将对理解这些令人着迷的分子机器做出重大贡献，并将为设计通过靶向DNA转运来克服耐药性的新药提供新的机会。新药对于解决抗菌素耐药性的全球紧急情况至关重要。