RPG: Towards the Crystal Structure of Replication Protein A

9708028博格斯塔尔：这是一项针对女科学家和女工程师的研究计划补助金。Borgstahl博士将通过X射线结晶学研究复制蛋白A(RPA)的结构。RPA对于DNA的维护和复制是必不可少的。重组蛋白将在大肠杆菌中生产，并将使用最新的技术和既定的方案获得非常纯净的蛋白质样本。将探索一系列广泛的结晶条件来生长大而有序的晶体。将使用三种策略来获得衍射级晶体：完整酵母和人RPA的结晶；RPA结构域和亚复合体的结晶；修剪后的RPA杂三聚体的结晶。一旦获得晶体，将收集X射线衍射数据，以获得蛋白质的高分辨率结构。复制蛋白A是新陈代谢的基本分子，是真核生物生命所必需的。RPA在许多DNA代谢过程中是必不可少的，包括复制、核苷酸切除修复和同源重组。阐明RPA的三维结构应该有助于深入了解这种蛋白质如何与DNA相互作用，以及它如何在DNA代谢中发挥其重要的多学科作用。***

9708028 Borgstahl This is a Research Planning Grant for Women Scientists and Engineers. Dr. Borgstahl will study the structure of replication protein A (RPA) by X-ray crystallography. RPA is essential to the maintenance and replication of DNA. The recombinant protein will be produced in E.coli and very pure protein samples will be obtained using the latest techniques and established protocols. A wide range of crystallization conditions will be explored to grow large well-ordered crystals. Three strategies will be used to obtain diffraction quality crystals: crystallization of intact yeast and human RPA; crystallization of RPA domains and subcomplexes; crystallization of a trimmed RPA heterotrimer. Once crystals are obtained, X-ray diffraction data will be collected in order to obtain a high resolution structure of the protein. Replication protein A is a fundamental molecule in metabolism and is essential for eukaryotic life. RPA is absolutely required for many DNA metabolic processes, including replication, nucleotide excision repair and homologous recombination. The elucidation of the three-dimensional structure of RPA should provide insight into how this protein interacts with DNA and how it functions in its essential multidisciplinary role in DNA metabolism. ***