STRUCTURAL STUDIES OF SITE SPECIFIC RECOMBINATION

描述：该提案旨在获得三维结构 关于λ催化的重组过程的信息 整合酶（Int）蛋白。Int催化噬菌体的整合 λ进入细菌宿主，以及切除基因组期间， 溶解该反应与I型催化剂催化的反应有一定的相似性 拓扑异构酶，其中在酪氨酸残基之间形成共价键， 蛋白质和DNA的磷酸骨架中。在以往工作的基础上 艾伦伯格博士建议用X射线晶体学来研究 λ整合酶的结构与DNA片段相结合， 反应循环的各个阶段。已经获得了以下二聚体的晶体： λ整合酶通过酪氨酸键与未切割的DNA结合。这 结构将解决有关的问题，是否酪氨酸残基的一个 整合酶蛋白催化其活性位点内的键断裂 分子，或者它是否在相邻分子中这样做。兴趣 在这个问题上的出现是因为一个根本的利益，了解如何 协同性用于驱动DNA切割和再连接。还力求 包括确定λ整合酶的结构 与四路霍利迪枢纽复杂，以及一个非常大的（在条款 由DNA弯曲蛋白稳定的DNA发夹结构 称为IHF。

DESCRIPTION: This proposal is aimed at obtaining three-dimensional structural information about the process of recombination catalyzed by the lambda integrase (Int) protein. Int catalyzes the integration of the bacteriophages lambda into the bacterial host, as well as the excision of the genome during lysis. The reaction has some similarity to that catalyzed by Type I topoisomerases, in that a covalent linkage is formed between a tyrosine residue in the protein and the phosphate backbone of DNA. Building on previous work from his laboratory, Dr. Ellenberger proposes to study by x-ray crystallography the structures of lambda integrase bound to pieces of DNA corresponding to various stages in the reaction cycle. Crystals have been obtained of a dimer of lambda integrase bound to uncleaved DNA through the tyrosine linkage. This structure will resolve issues relating to whether the tyrosine residue of one integrase protein catalyzes bond cleavage within the active site of the same molecule, or whether it does so in that of the neighboring molecule. Interest in this issue arises because of a fundamental interest in understanding how cooperativity is utilized in driving DNA cleavage and religation. Further aims of the proposal include the determination of the structure of lambda integrase complexed with a four way Holliday Junction, as well as a very large (in terms of crystallography) DNA hairpin structure stabilized by the DNA bending protein known as IHF.