Opening of a double stranded DNA replication fork by a hexameric helicase

通过六聚体解旋酶打开双链 DNA 复制叉

• 批准号: BB/K019252/1
• 负责人: C Sanders
• 金额: $ 36.88万
• 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: 2014
• 资助国家: 英国
• 起止时间: 2014 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FK019252%2F1
• 关键词: Opening; double; stranded; DNA; replication

The inner workings of mammalian cells involve a number of interrelated, small, "nanomachines". Individual machines are usually composed of a number of proteins that work together to perform one of the cell's functions in life, and determine its ability to survive and replicate. Understanding how individual machines function is crucial because when these machines breakdown the consequences can be catastrophic. For example, some cancers can be attributed directly to the failure of machines that process the genetic information (DNA) of a cell. In addition, invading microorganisms such as viruses bring their own machines that subvert or destroy a cell as it is hijacked for the pathogen's own advantage. The identification and understanding of how these machines function can therefore be a starting point for combating such disease states. The biosciences can inspire nanotechnology, which aims to develop small mechanical devices based on protein machines that can be harnessed to drive or modulate molecular processes from which we benefit. Significant examples are found in biotechnology, such as emerging nanopore DNA sequencing devices that can operate at the single molecule level. The precise understanding of cellular nanomachines at the atomic level is absolutely dependent on the availability of precise structural information. For large molecular assemblies this information comes from X-ray structural analysis and, for very large assemblies, it is vastly assisted by powerful electron microscopes that can image single particles. Alone, electron microscopy can guide our understanding, but for the most part lacks the resolving power to provide critical detail. E1, the protein that we will study, is from a large group of pathogens, the papillomaviruses that cause warts and cancer in animals and man. Papillomavirus-derived diseases are of significant health and economic importance. E1 is often referred to as a "motor protein" that forms a six membered (hexameric) ring-like assembly, a machine that unwinds the two strands of the DNA molecule during replication (reproduction) of viral DNA. It can be viewed as a small radial engine that can move along DNA and separate its two strands and as such is a prototypic nanomachine and important target for drugs that could inhibit viral replication and disease.Here we aim to understand how the E1 machine unwinds DNA. The objectives are to carry out detailed X-ray structural studies on E1 to determine its overall structure and how it interacts with DNA. We intend to verify these results and test models of function with complementary biochemical and biophysical studies. As a leading tractable model, it has great promise for being the first hexameric DNA-unwinding machine that we fully understand. The potential applications of our work include formulating strategies to target E1 and related helicases through structure based drug design. They will help understand replication processes in our own cells that are more complex and challenging to define. There are potential applications in nanobiotechnology and in medicine. Consequently, we envision that our studies will have a wide impact, with health benefits and improved living and economic standards nationally and internationally.

哺乳动物细胞的内部运作涉及许多相互关联的小型“纳米机器”。单个机器通常由许多蛋白质组成，这些蛋白质共同工作以执行细胞在生命中的功能之一，并决定其生存和复制的能力。了解单个机器的功能至关重要，因为当这些机器发生故障时，后果可能是灾难性的。例如，一些癌症可以直接归因于处理细胞遗传信息（DNA）的机器故障。此外，入侵的微生物，如病毒，带来了自己的机器，破坏或摧毁细胞，因为它是为了病原体自己的利益而劫持的。因此，识别和理解这些机器如何运作可以成为对抗这些疾病状态的起点。生物科学可以激发纳米技术，其目的是开发基于蛋白质机器的小型机械装置，可以利用这些装置来驱动或调节我们从中受益的分子过程。在生物技术中发现了重要的例子，例如可以在单分子水平上操作的新兴纳米孔DNA测序设备。在原子水平上对细胞纳米机器的精确理解完全取决于精确结构信息的可用性。对于大分子组装，这些信息来自X射线结构分析，对于非常大的组装，它极大地帮助了强大的电子显微镜，可以成像单个粒子。电子显微镜可以单独指导我们的理解，但在大多数情况下缺乏提供关键细节的分辨率。我们将要研究的蛋白质E1来自一大群病原体，即在动物和人类中引起疣和癌症的乳头状瘤病毒。乳头状瘤病毒衍生的疾病具有重要的健康和经济意义。E1通常被称为“马达蛋白”，它形成六元（六聚体）环状组装体，这是一种在病毒DNA复制（繁殖）过程中解开DNA分子两条链的机器。它可以被看作是一个小的径向引擎，可以沿着沿着DNA移动并分离它的两条链，因此是一个原型纳米机器和药物的重要目标，可以抑制病毒复制和疾病。目的是对E1进行详细的X射线结构研究，以确定其整体结构以及它如何与DNA相互作用。我们打算验证这些结果和测试功能的补充生化和生物物理研究模型。作为一个领先的易处理的模型，它有很大的希望成为第一个六聚体DNA解旋机器，我们完全理解。我们工作的潜在应用包括通过基于结构的药物设计来制定针对E1和相关解旋酶的策略。它们将有助于理解我们自己细胞中的复制过程，这些过程更复杂，更难以定义。在纳米生物技术和医学方面有潜在的应用。因此，我们设想，我们的研究将产生广泛的影响，与健康的好处和提高生活和经济水平的国家和国际。

项目成果

DNA substrate recognition and processing by the full-length human UPF1 helicase.

• DOI: 10.1093/nar/gkx478
• 发表时间: 2017-07-07
• 期刊: Nucleic acids research
• 影响因子: 14.9
• 作者: Dehghani-Tafti S;Sanders CM
• 通讯作者: Sanders CM

Structural basis for DNA strand separation by a hexameric replicative helicase.

• DOI: 10.1093/nar/gkv778
• 发表时间: 2015-09-30
• 期刊: Nucleic acids research
• 影响因子: 14.9
• 作者: Chaban Y;Stead JA;Ryzhenkova K;Whelan F;Lamber EP;Antson A;Sanders CM;Orlova EV
• 通讯作者: Orlova EV