Structure and biochemical mechanism of DNA replication initiation machines

DNA复制起始机的结构和生化机制

• 批准号: BB/R001685/1
• 负责人: C Sanders
• 金额: $ 44.27万
• 依托单位: University of Sheffield
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2017
• 资助国家: 英国
• 起止时间: 2017 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FR001685%2F1
• 关键词: Structure; biochemical; mechanism; DNA; replication

Proteins are essential components of cells that do all the jobs that ensure the continuity of life. Some of them assemble to form "nanomachines" that do work. This work often involves rearranging or remodelling small molecules, other proteins or the DNA that encodes the information of life. Some of these machines are made up from groups of proteins that work together but others are proteins that act alone. Understanding how individual biological machines function is crucial because if they fail the consequences can be catastrophic for living organisms. For example, some cancers and neurodegenerative disorders are caused by the failure of nanomachines that process the DNA in our cells. In addition, pathogens that invade the body, such as viruses, bring their own nanomachines that destroy healthy cells or hijack them to make the virus stronger. Identifying these machines and understanding how they function can therefore be a starting point for combating diseases. We are studying the DNA replication proteins from a group of pathogenic viruses, the papillomaviruses (PV), that cause warts and serious diseases such as cancer. Accurate replication of our own DNA is essential for genome stability and healthy ageing. These systems have many components and to understand the process we require knowledge of the underlying mechanisms. The viral proteins we are studying, known simply as E1 and E2, bind to DNA and change its structure so that it can be replicated. In our own cells a complex assembly of many different proteins is required to do the same jobs that the PV E1 protein does. The aim of our research is to determine how the viral proteins E1 and E2 function on DNA during replication, to understand processes that are essential for life where potential therapeutic strategies could emerge to improve health and well-being.To do this we have designed ways to characterize these exceptionally small protein machines. Just as complex man-made mechanical machines could never be understood without understanding how they are made up, we need to know the structure of protein nanomachines to know how they work. We use biochemistry, powerful electron microscopes and computer technology to generate images of protein complexes and then use this information to re-construct their detailed three-dimensional structures. In parallel we apply biochemical techniques to understand the reactions performed by these proteins and relate this information to the structures we observe. Ultimately, we expect that our studies will have a significant impact in the healthcare sector and wider economy. The potential applications of our work include designing drugs to target viral proteins required for their replication. This research is important since it is estimated that up to 5% of all cancers are caused by human papillomavirus (HPV) and up to 600 million people are infected with HPV at any one time but there are currently no drug treatments of proven efficacy. Our studies will also inform us of how the more complex cellular counterparts of the PV replication proteins work, where there are further therapeutic applications to treat cellular and microbial (e.g. fungal) diseases. There are also potential applications in nanobiotechnology. This branch of science aims to develop small mechanical devices based on protein machines that can be harnessed to drive or alter molecular processes for our benefit.

蛋白质是细胞的重要组成部分，它完成了确保生命连续性的所有工作。他们中的一些人聚集在一起，形成了能够工作的“纳米机器”。这项工作通常涉及重新排列或改造小分子、其他蛋白质或编码生命信息的DNA。这些机器中的一些是由一起工作的蛋白质组组成的，但另一些是单独作用的蛋白质。了解单个生物机器是如何运作的至关重要，因为如果它们失败了，对活着的有机体来说后果可能是灾难性的。例如，一些癌症和神经退行性疾病是由处理细胞中DNA的纳米机器失败引起的。此外，入侵人体的病原体，如病毒，会携带自己的纳米机器来摧毁健康细胞或劫持它们，以使病毒变得更强大。因此，识别这些机器并了解它们是如何工作的，可以成为抗击疾病的起点。我们正在研究一组致病病毒-乳头瘤病毒(PV)的DNA复制蛋白，这些病毒会导致尖锐湿疣和癌症等严重疾病。准确复制我们自己的DNA对于基因组的稳定和健康的衰老至关重要。这些系统有许多组件，要理解这一过程，我们需要了解基本机制。我们正在研究的病毒蛋白，简称为E1和E2，与DNA结合并改变其结构，使其能够复制。在我们自己的细胞中，需要许多不同蛋白质的复杂组装来完成与PV E1蛋白相同的工作。我们研究的目的是确定病毒蛋白E1和E2在复制过程中如何在DNA上发挥作用，了解对生命至关重要的过程，在这些过程中可能会出现潜在的治疗策略来改善健康和福祉。为此，我们设计了一些方法来表征这些异常小的蛋白质机器。就像复杂的人造机械在不了解它们是如何组成的情况下永远不可能被理解一样，我们需要知道蛋白质纳米机器的结构才能知道它们是如何工作的。我们使用生物化学、强大的电子显微镜和计算机技术来生成蛋白质复合体的图像，然后利用这些信息重建其详细的三维结构。同时，我们应用生化技术来了解这些蛋白质进行的反应，并将这些信息与我们观察到的结构联系起来。最终，我们预计我们的研究将对医疗保健行业和更广泛的经济产生重大影响。我们工作的潜在应用包括设计针对复制所需的病毒蛋白的药物。这项研究很重要，因为据估计，高达5%的癌症是由人类乳头瘤病毒(HPV)引起的，任何时候都有多达6亿人感染HPV，但目前还没有被证明有效的药物治疗。我们的研究还将告诉我们，光伏复制蛋白的更复杂的细胞对应物是如何工作的，在哪里有进一步的治疗应用来治疗细胞和微生物(例如真菌)疾病。在纳米生物技术方面也有潜在的应用。这一科学分支的目标是开发基于蛋白质机器的小型机械设备，这些设备可以被利用来驱动或改变分子过程，以造福于我们。