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首页> 外文期刊>Proceedings of the National Academy of Sciences of the United States of America >Architecture of recombination intermediates visualized by in-gel FRET of λ integrase-Holliday junction-arm DNA complexes
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Architecture of recombination intermediates visualized by in-gel FRET of λ integrase-Holliday junction-arm DNA complexes

机译：λ整合酶-霍利迪连接臂DNA复合物的凝胶内FRET可视化重组中间体的体系结构

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摘要

λ Integrase (Int) mediates recombination between attachment sites on phage and Escherichia coli DNA. Int is assisted by accessory protein-induced DNA loops in bridging pairs of distinct "arm-type" and "core-type" DNA sites to form synapsed recombination complexes that subsequently recombine by means of a Holliday junction (HJ) intermediate. An in-gel FRET assay was developed and used to measure 15 distances between six points in two Int-HJ complexes containing arm-DNA oligonucleotides, and 3D maps of these complexes were derived by distance-geometry calculations. The maps reveal unexpected positions for the arm-type DNAs relative to core sites on the HJ and a new Int conformation in the HJ tetramer. The results show how the position of arm DNAs determines the bias of catalytic activities responsible for directional resolution, provide insights into the organization of Int higher-order complexes, and lead to models of the structure of the full HJ recombination intermediates.
机译：λ整合酶（Int）介导噬菌体和大肠杆菌DNA附着位点之间的重组。在辅助的蛋白质诱导的DNA环的协助下，将不同的“臂型”和“核心型” DNA位点桥接起来，形成突触重组复合体，随后通过霍利迪结（HJ）中间体进行重组，从而实现了Int的合成。开展了凝胶内FRET分析，并用于测量两个包含arm-DNA寡核苷酸的Int-HJ复合物中六个点之间的15个距离，并通过距离几何计算得出了这些复合物的3D图。这些图揭示了臂型DNA相对于HJ核心位点的意外位置以及HJ四聚体中的新Int构象。结果表明，臂DNA的位置如何确定负责方向拆分的催化活性的偏倚，提供有关Int高阶复合物组织的见解，并导致建立完整的HJ重组中间体的结构模型。

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