Structure, Function and Evolution of DNA Repair Enzymes

描述(由申请人提供)：DNA修复酶的结构、功能和进化计划的总体目标是了解支撑DNA糖基酶和RecA重组酶的第N超家族和FPG家族的基本机制，这些酶处理电离辐射诱导的DNA损伤，已知的损伤启动致癌过程。支持该计划项目的中心假设是，所讨论的DNA修复酶家族中的每一个都有一个结构框架，该结构框架支持特异性或调节特性的各种显着变化，只有少量的序列改变。为了验证这一假设，将使用一种新的系统发育/结构分析来开发算法，不仅可以识别同源但具有不同活性的天然蛋白质变体，而且还可以确定特定蛋白质中的哪些氨基酸应该改变，以潜在地改变底物特异性或其他蛋白质功能。由于启动碱基切除修复的DNA糖基酶和参与双链断裂修复的RecA重组酶在所有三个王国中都高度保守，它们特别适合这种方法。拟议的方案项目由三个项目和三个核心组成。核心A，生物信息学核心，将使用许多方法来选择要在项目中检查的蛋白质，以检验我们的假设。项目1旨在描述和改变核心A所针对的氧化DNA糖基酶的底物特异性。项目2将侧重于特定的DNA糖基酶并确定其晶体结构。项目3将使用类似的计算、生化和结构方法来了解RecA重组酶的生化特性的变化。这些项目直接依靠核心A来提供对天然蛋白质序列的分析，并为实验设计提供支持，并将检验所得出的假设，以便在项目之间进行持续迭代。所有三个项目都将得到表达、表征和结晶核心(核心B)和管理核心(核心C)的支持。

DESCRIPTION (provided by applicant): The overall goal of the Program Project Structure, Function and Evolution of DNA Repair Enzymes is to understand the fundamental mechanisms underpinning the Nth Superfamily and the Fpg Family of DNA glycosylases and the RecA recombinases, enzymes that process ionizing radiation-induced DNA damages, damages known to initiate the carcinogenic process. The central hypothesis underpinning this Program Project is that the families of the DNA repair enzymes in question each have a structural framework that supports a variety of significant changes in specificity or regulatory properties with only a small number of sequence alterations. To test this hypothesis a novel phylogenetic/structural analysis will be used to develop algorithms not only to identify natural protein variants that are orthologous, but have different activities, but also to determine which amino acids in a particular protein should be varied to potentially alter substrate specificity or other protein functions. Since the DNA glycosylases that initiate Base Excision Repair and the RecA recombinases involved in Double Strand Break Repair are highly conserved across all three kingdoms, they are particularly suited to this approach. The proposed Program Project consists of three Projects and three Cores. Core A, the Bioinformatics Core, will use a number of methodologies to select the proteins to be examined in the projects to test our hypothesis. Project 1 is designed to delineate and alter the substrate specificities of the oxidative DNA glycosylases targeted by Core A. Project 2 will focus on particular DNA glycosylases and determine their crystal structures. Project 3 will use similar computational, biochemical and structural approaches to understand variations in the biochemical properties of the RecA recombinases. The projects directly depend on Core A to provide the analysis of the natural protein sequences and to provide support for experimental design and will test the hypotheses derived so that there is continued iteration among the projects. All three projects will be supported by the Expression, Characterization, and Crystallization Core (Core B) and the Administrative Core (Core C).