Reversible Covalent Crosslinking of DNA to Befuddle Its Repair

DNA 可逆共价交联扰乱其修复

• 批准号: 1405123
• 负责人: Steven Rokita
• 金额: $ 45.95万
• 依托单位: Johns Hopkins University
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-08-15 至 2018-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1405123&HistoricalAwards=false
• 关键词: Reversible; Covalent; Crosslinking; DNA; Befuddle

The Chemistry of Life Processes Program in the Division of Chemistry is funding Steven Rokita of Johns Hopkins University for research aimed at increasing our understanding of the way in which our cells react to toxins in the environment. In particular, the investigators are looking at how cells repair damage those toxins might inflict. One way the damage occurs is through a process known as DNA alkylation, in which small chemical groups are added to the molecules that make up genes. Food preservatives, natural flavorings and even drugs can induce DNA alkylation. The current project is designed to assess the biochemical consequences of toxins that interact reversibly with DNA. The reversibility of the deleterious interactions between these molecules and DNA creates the possibility that the effects of the toxins are amplified and may overwhelm the repair capacity of cells. To study the process in detail and identify the most dramatic effects of the reversibility, a series of model compounds is being prepared and tested for each step of the DNA repair mechanism. This work is having a broader impact on scientist's ability to evaluate the toxicology of substances prior to their market release. It is having a further impact on the training of the next generation of scientists, both undergraduate and graduate students, by encouraging them to recognize and pursue problems that transcend the traditional barriers between organic chemistry and biochemistry and are important for the human health.In this project, the structural determinants required for efficient diffusion of covalent cross-linking within duplex DNA are being identified. This is being accomplished through the synthesis and evaluation of quinone methide intermediates that express a range of reaction kinetics and DNA affinities. The principles gleaned from this first activity will be used for challenging enzymes responsible for DNA repair with reversible and dynamic cross-links. The entities formed during each step of cross-link repair will be mapped by diagnostic fragmentation and separation using electrophoresis and chromatography. Initial attention will be directed to isolated DNA and then progress to nucleosomes and finally embrace the complexity of crude cell extracts.

化学系的生命过程化学计划资助约翰霍普金斯大学的史蒂文·罗基塔进行研究，旨在增加我们对细胞对环境中毒素的反应方式的了解。特别是，调查人员正在研究这些毒素可能造成的细胞修复损害。损伤发生的一种方式是通过一种称为DNA烷基化的过程，在这个过程中，组成基因的分子中添加了一些小的化学基团。食品防腐剂、天然调味料甚至药物都可以诱导DNA烷基化。目前的项目旨在评估与DNA可逆相互作用的毒素的生化后果。这些分子和DNA之间有害相互作用的可逆性造成了毒素影响被放大的可能性，并可能压倒细胞的修复能力。为了详细研究这一过程并确定可逆性最显著的影响，正在为DNA修复机制的每一步准备和测试一系列模型化合物。这项工作正在对科学家在物质上市前评估其毒理学的能力产生更广泛的影响。它对下一代科学家的培训产生了进一步的影响，包括本科生和研究生，鼓励他们认识和追求超越有机化学和生物化学之间的传统障碍并对人类健康重要的问题。在这个项目中，正在确定在双链DNA中有效扩散共价交联所需的结构决定因素。这是通过合成和评价表达一系列反应动力学和DNA亲和力的甲基苯醌中间体来实现的。从第一个活动中收集到的原理将用于挑战负责DNA修复的酶，这些酶具有可逆和动态的交联性。在交联链修复的每个步骤中形成的实体将通过诊断碎片和使用电泳层析法进行分离来绘制图谱。最初的注意力将被引导到分离的DNA上，然后进展到核小体，最后接受粗细胞提取物的复杂性。