RECOMBINATION INTERMEDIATES--MECHANISMS IN RESOLUTION

本项目源于我们对以下机制的长期兴趣： 哪些遗传物质被重新组合和重新排列。 我们建议研究 霍利迪复合的形成、性质和解决方法 中间体的 我们正在使用的系统， 噬菌体λ的途径，已经在两个领域进行了广泛的研究， 基因和生化水平。 因为霍利迪的中间人物 在这么多的重组途径中， 建议的研究应该具有普遍意义-除了它们的 位点特异性重组的重要性，以及（不断增长的）Int家族 重组酶。 我们提出的实验依赖于我们最近的发现， 通过一对顺序的高度有序的互补链 交流 他们还利用我们的证明， “合成的”重组中间体提供了一个非常有用的工具， 研究霍利迪路口 本建议涉及六个具体问题：1）什么是 DNA的作用：DNA同源性在启动链转移？ 2)有哪些 分支迁移在分辨率中的作用和性质？ 3)有哪些 结合模式和Int蛋白之间的相互作用结合在 霍利迪路口？ 4)臂型远端部位在 霍利迪路口的解决方案？ 5)结构特点是什么 霍利迪交叉点与重组的关系 6)Int-mediated 链转移具有与非法转移相关机械特征 重组？

This project is derived from our long term interest in the mechanisms by which genetic material is reassorted and rearranged. We propose to study the formation, properties and resolution of Holliday recombination intermediates. The system we are using, the Int-dependent site-specific pathway of bacteriophage lambda, has been extensively studied at both the genetic and biochemical levels. Because the Holliday intermediate figures prominently in so many recombination pathways, the results obtained in the proposed studies should be of general significance -- in addition to their importance for site-specific recombination, and the (growing) Int family of recombinases. The proposed experiments depend upon our recent finding that this pathway proceeds via a sequential highly ordered pair of reciprocal strand exchanges. They also draw on our demonstration that the appropriate "synthetic" recombination intermediates afford a very useful tool for studying Holliday junctions. Six specific questions are addressed in this proposal: 1) What is the role of DNA:DNA homology in initiating strand transfer? 2) What are the role and properties of branch migration in resolution? 3) What are the binding patterns and interactions among the Int proteins bound at the Holliday junction? 4) What is the role of arm-type distal sites in the resolution of Holliday junctions? 5) what are the structural features of the Holliday junction in relation to recombination? 6) Does Int-mediated strand transfer have mechanistic features related to illegitimate recombination?