Structural Cell Biology of DNA Repair Machines

描述（由申请人提供）：我们正在研究DNA修复复合物的结构组织，该复合物切除DNA损伤以及通过突变和小分子抑制剂破坏这些蛋白质-蛋白质相互作用的功能后果。修复的替代途径可用于许多类型的DNA损伤，并且响应于DNA损伤产生的翻译后修饰控制修复复合物的差异组装，提供调节途径选择的机制。癌症相关的DNA维持活动的缺陷可以通过靶向剩余的修复活动与机制为基础的抑制剂在治疗上利用。我们的工作主要集中在碱基切除修复途径和核苷酸切除修复途径修复DNA单链断裂的机制。我们正在研究DNA损伤切除复合物在体外和培养细胞中的物理组装，以及DNA切割与末端加工和连接的耦合机制。纯化的DNA修复复合物的小角X-射线散射揭示了动态构象状态，我们建议是重要的DNA修复中间体，连续酶的通路中的baffs。高分辨率晶体结构和小分子筛选实验正被用于预测和鉴定修复蛋白相互作用的抑制剂，其是抗肿瘤疗法的候选者，并在DNA损伤反应期间充当细胞生理学的可逆化学探针。 这种综合方法利用了项目1，2和6中研究人员在检测和生物材料方面的广泛专业知识，以及SBDR计划的表达和分子生物学核心和结构细胞生物学核心的独特能力，以生产蛋白质并在结构上评估修复复合物。

DESCRIPTION (provided by applicant): We are studying the structural organization of DNA repair complexes that excise DNA damage and the functional consequences of disrupting these protein-protein interactions by mutation and small molecule inhibitors. Alternative pathways of repair are available for many types of DNA damage, and posttranslational modifications generated in response to DNA damage control the differential assembly of repair complexes, providing a mechanism for regulating pathway choice. Cancer-associated defects in DNA maintenance activities can be exploited therapeutically by targeting the remaining repair activities with mechanism-based inhibitors. Our work focuses on the mechanisms of repairing DNA single strand breaks generated by the base excision and nucleotide excision repair pathways. We are studying the physical assembly of DNA damage excision complexes in vitro and in cultured cells, and the mechanism of coupling DNA cleavage to end processing and ligation. Small angle x-ray scattering of purified DNA repair complexes reveals dynamic conformational states that we propose are important for handoffs of DNA repair intermediates to successive enzymes in a pathway. High resolution crystal structures and small molecule screening experiments are being used to predict and identify inhibitors of repair protein interactions, which are candidates for anti-tumor therapies and serve as reversible chemical probes of cellular physiology during DNA damage responses. This integrated approach takes advantage of the broad expertise of investigators in Projects 1, 2, and 6 for assays and biological materials, as well as the unique capabilities of the Expression and Molecular Biology Core and the Structural Cell Biology Core of the SBDR Program to produce proteins and structurally evaluate repair complexes.