Targeting Twist: Single-molecule insights into supercoiled DNA-topoisomerase interactions for drug discovery

拓扑异构酶是存在于从人类到细菌的所有生物体中的酶，由于它们能够解开打结和扭曲的DNA，因此对于生命至关重要。我们的DNA的打结和扭曲发生在2米长的DNA折叠成细胞核时;比人类头发的宽度要窄得多。这是加剧了我们的细胞中的分子机制，它沿着沿着我们的DNA，把它分开，并操纵它，在一个紧凑的环境。如果没有拓扑异构酶，我们的DNA会不可逆转地打结和扭曲，细胞就会死亡。因此，关键的治疗药物，如抗癌药物和抗生素药物，分别针对人类和细菌的拓扑异构酶，杀死癌细胞或细菌细胞。制药管道正在不断努力，以改善我们现有的治疗方法，并创造新的治疗方法。为了改善现有的治疗方法，我们需要更好地了解它们是如何工作的。这一领域仍有很大的发展空间。靶向拓扑异构酶的抗癌治疗剂目前缺乏选择性。除此之外，还有许多未开发的抗菌靶标，它们具有显著的潜力，但尚未被靶向。我建议使用单分子技术，如原子力显微镜，当针在表面上扫描时，它上下移动，逐行构建表面外观的图片（很像唱片播放器通过感觉其表面的轮廓来读取唱片）。这些将由更快的单分子技术来补充，例如磁镊子，其中一段DNA一端被拴在磁性球上，另一端被拴在玻璃表面上。通过旋转磁球，DNA被扭曲成更复杂的状态。DNA的长度是通过测量球的高度来确定的。这给出了拓扑异构酶如何能够解旋DNA的非常快的读数，以及这个过程如何被抗癌治疗剂和抗生素等药物破坏。该项目的目的是提高我们对拓扑异构酶如何解开DNA的理解，以及拓扑异构酶抑制剂如何阻止这种情况，以帮助开发新的或改进的抗癌药物和抗生素。

Topoisomerases are enzymes, present in all organisms from humans to bacteria, which are essential for life due to their ability to untangle knotted and twisted DNA. The knotting and twisting of our DNA occurs as 2 metres of our DNA is folded into the cell nucleus; much narrower than the width of a human hair. This is exacerbated by the molecular machinery in our cells which travel along our DNA, pulling it apart and manipulating it, in a compact environment. Without topoisomerases, our DNA becomes irreversibly knotted and twisted, and the cell will die. For this reason, key therapeutics such as anticancer and antibiotic drugs target human and bacterial topoisomerases respectively either killing cancerous or bacterial cells.The pharmaceutical pipeline is continually working both to improve the therapeutics that we have, and to create novel therapeutics. To improve existing therapeutics, we require better understanding of how they work. There remains much room for development in this field. Anti-cancer therapeutics that target topoisomerases currently suffer from a lack of selectivity. Further to this, there remain a number of unexplored antibacterial targets, which have significant potential, but are as yet untargeted. I propose to use single molecule techniques such as Atomic Force Microscopy, where a needle moves up and down as it is scanned over a surface, building up a picture of what the surface looks like, line by line, (much like a record player reads out a record by feeling the contours of its surface). These will be complemented by faster single molecule techniques, such as magnetic tweezers where a piece of DNA is tethered to a magnetic ball at one end, and a glass surface at the other. The DNA is twisted into a more complex state, by rotating the magnetic ball. The length of DNA is determined by measuring the height of the ball. This gives a very fast readout of how topoisomerases are able to untwist DNA, and how this process can be disrupted by drugs such as anti-cancer therapeutics and antibiotics. The aim of the project is to improve our understanding of how topoisomerases untwist DNA, and how this is prevented by topoisomerase inhibitors, to aid in the development of new or improved anti-cancer drugs and antibiotics.