Collaborative Proposal: Elucidating Chemical Mechanisms of DNA Repair Using Transition State Analogs

由于细胞代谢和环境暴露而造成的DNA碱基损伤，可能会侵蚀DNA蓝图中保存的生命信息。DNA碱基的修改导致DNA序列的变化，称为突变。这些突变会对细胞过程产生负面影响，但它们可以通过DNA修复酶的作用得到纠正。通过这一奖项，化学部门的生命过程化学项目资助了加州大学戴维斯分校的Sheila David博士和犹他大学的Martin Horvath博士，以揭示受损DNA碱基是如何被检测和修复的分子细节。这项研究利用合成DNA化学和x射线晶体学来捕捉和可视化DNA修复过程中的步骤，否则这些步骤太短暂而无法研究，并生成DNA修复过程的详细分子图。研究生和本科生学习如何结合不同的化学和生物学方法来揭示和理解基本生命过程的关键特征。这个项目也被整合到一个独特的“基于课程的研究体验”（CURE）实验课程中，该课程由加州大学戴维斯分校和犹他大学共同教授。通过本课程的学习，本科生将提高批判性思维能力，并对生物体的关键信息如何在DNA中编码并由DNA修复酶维持有更好的概念性知识。本研究项目描述了碱基切除修复（BER）糖基酶的化学机制，以阐明这些酶如何有效地发现罕见的修饰DNA碱基。模拟过渡态的修饰核苷酸是通过合成化学制备的。对含有这些过渡态模拟物的DNA结合的几种BER糖基酶的结构研究提供了对碱基切除催化的深入了解。事实上，通过三种BER糖基酶（即MutY、AlkD和MBD4）发现的令人惊讶的特征，对这些糖基酶和相关糖基酶提出了新的问题，并促使David和Horvath研究小组进行新的创新结构和功能研究。参与该项目的学生将接受跨学科的培训，包括合成核酸化学、核酸酶学和dna -蛋白质晶体学。培育有利于女性和少数族裔学生成长的环境是该奖项支持的工作的核心目标。该项目具有积极而广泛的教育影响，因为它与两门本科实验课程相关联，向新生和高年级本科生介绍化学生物学中使用的现代研究技术和方法，重点是DNA损伤和修复。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

The information held within the DNA blueprint for life can be eroded by damage to DNA bases due to cellular metabolism and environmental exposures. Modified DNA bases lead to changes in the DNA sequence called mutations. These mutations can negatively impact cellular processes but they can be corrected by the action of DNA repair enzymes. With this award, the Chemistry of Life Processes Program in the Chemistry Division is funding Dr. Sheila David at the University of California Davis and Dr. Martin Horvath at the University of Utah to uncover molecular details of how damaged DNA bases are detected and repaired. The research makes uses of synthetic DNA chemistry and X-ray crystallography to trap and visualize steps in the DNA repair process that are otherwise too fleeting to study and to generate a detailed molecular picture of the DNA repair process. Graduate and undergraduate students learn how to combine different chemistry and biology approaches to reveal and understand key features of fundamental life processes. This project is also integrated into a unique "Course-Based Research Experience" (CURE) laboratory course taught at both the University of California Davis and the University of Utah. Undergraduates emerge from the course with improved critical thinking skills and a better conceptual knowledge of how information crucial for living organisms is encoded in their DNA and maintained by DNA repair enzymes. This research project delineates chemical mechanisms of base excision repair (BER) glycosylases to illuminate how these enzymes efficiently find rare modified DNA bases. Modified nucleotides that mimic transition states are prepared through synthetic chemistry. Structural studies of several BER glycosylases bound to DNA containing these transition state mimics provide insight into the catalysis of base excision. Indeed, surprising features uncovered with three BER glycosylases, namely MutY, AlkD and MBD4, raise new questions on these and related glycosylases and prompt new innovative structural and functional studies that will be pursued by the David and Horvath research groups. Students working on this project receive interdisciplinary training spanning synthetic nucleic acid chemistry, nucleic acid enzymology, and DNA-protein crystallography. Fostering environments in which women and minority students thrive is a core goal of the work supported by this award. This project has a positive broader educational impact because it is tied to two undergraduate laboratory courses that introduce incoming and advanced undergraduate students to modern research techniques and approaches used in chemical biology with a focus on DNA damage and repair.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.