CAREER: Linking Structure, Stability and Protection in Protamine Packaged DNA

职业：连接鱼精蛋白包装 DNA 的结构、稳定性和保护

• 批准号: 1453168
• 负责人: Jason DeRouchey
• 金额: $ 69.16万
• 依托单位: University of Kentucky Research Foundation
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-03-15 至 2024-02-29
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1453168&HistoricalAwards=false
• 关键词: CAREER; Linking; Structure; Stability; Protection

Chromatin (the material of chromosomes) is much more compact and highly organized in the nucleus of sperm cells than in the nucleus of other cells. This high degree of compaction, brought about by the association of the DNA with a class of proteins called protamines, is thought to help protect the DNA from damage. Despite previous research, the mechanisms underlying the tight packaging of DNA by protamines remain poorly understood. The goals of this project are to use a multidisciplinary array of tools to establish fundamental knowledge of the mechanisms by which nature utilizes protamines to package, protect and store DNA in sperm cells. The project spans the disciplines of molecular biology, biochemistry, biophysics and analytical chemistry. The unique properties of protamines also have promise for new biotechnological applications in the creation of novel devices for DNA delivery and the development of biosensor technologies. The multidisciplinary nature of the research will create unique training opportunities for undergraduate and graduate student researchers. The project also includes education in chemistry and biophysics, including the development of massive open online courses (MOOCs). Summer workshops will be created for high school teachers to learn about and actively participate in refining and assessing the proposed MOOC courses to best meet the needs of teachers and their students nationwide.Spermiogenesis is a unique multi-step process resulting ultimately in the replacement of histones by protamines in sperm nuclei to a final volume roughly 1/20th that of a somatic nucleus. The near crystalline organization of DNA in mature sperm is crucial for both DNA delivery and the protection of genetic information due to the absence of DNA repair. The long-term goal of the project is to understand the link between physical compaction, interaction energetics and biochemical driving forces in protamine-DNA complexes. The project will use a variety of structural, biophysical and biochemical methods to evaluate protamine-DNA structure and stability to free radicals in vitro and in vivo. These experiments will provide a realistic model of how protamine chemistry controls sperm chromatin organization and quantitative knowledge of the link between DNA packaging and susceptibility of DNA to reactive oxidative species. Protamines exhibit a combination of phosphorylation sites, disulfide bonds, and zinc ions that make them a uniquely rich system to investigate the physical chemistry and structure of protein/DNA interactions and the structure of DNA in the compacted state. These novel aspects of protamine have potential for expoitation in synthetic systems, and the models developed and tested in this project will make contributions to our understanding of protamine-DNA compaction in sperm chromatin as well as providing insights for biotechnological exploitation of DNA devices.

染色质(染色体的物质)在精子细胞的核中比在其他细胞的核中更紧密和高度组织化。这种高度的紧凑性是由DNA与一种名为鱼精蛋白的蛋白质结合而产生的，被认为有助于保护DNA免受损害。尽管之前有过研究，但鱼精蛋白紧密包装DNA的机制仍然知之甚少。这个项目的目标是使用一系列多学科的工具来建立关于自然界利用鱼精蛋白包装、保护和储存精子细胞中DNA的机制的基本知识。该项目涉及分子生物学、生物化学、生物物理学和分析化学等学科。鱼精蛋白的独特性质也有望在创造新的DNA输送装置和开发生物传感器技术方面获得新的生物技术应用。这项研究的多学科性质将为本科生和研究生研究人员创造独特的培训机会。该项目还包括化学和生物物理学方面的教育，包括开发大规模开放在线课程(MOOC)。将为高中教师创建暑期研讨会，以了解并积极参与改进和评估拟议的MOOC课程，以最大限度地满足全国教师和学生的需求。精子发生是一个独特的多步骤过程，最终导致精子核中的组蛋白被精蛋白取代，最终体积约为体细胞核的1/20。由于缺乏DNA修复，成熟精子中DNA的近晶体结构对于DNA传递和遗传信息保护都是至关重要的。该项目的长期目标是了解鱼精蛋白-DNA复合体中物理压实、相互作用能量学和生化驱动力之间的联系。该项目将使用各种结构、生物物理和生化方法来评估鱼精蛋白-DNA的结构和在体外和体内对自由基的稳定性。这些实验将提供一个现实的模型，说明鱼精蛋白化学如何控制精子染色质组织，并定量了解DNA包装与DNA对活性氧化物种的敏感性之间的联系。鱼精蛋白表现出磷酸化位点、二硫键和锌离子的组合，使它们成为研究蛋白质/DNA相互作用的物理化学和结构以及紧凑状态下DNA结构的独特丰富的体系。鱼精蛋白的这些新方面具有在合成系统中进行探索的潜力，本项目中开发和测试的模型将有助于我们理解精子染色质中鱼精蛋白与DNA的紧密结合，并为DNA装置的生物技术开发提供见解。