Yeeles Joseph T P, Dillingham Mark S
DNA, Protein Interactions Unit, Department of Biochemistry, School of Medical Sciences, University of Bristol, University Walk, Bristol, UK.
J Mol Biol. 2007 Aug 3;371(1):66-78. doi: 10.1016/j.jmb.2007.05.053. Epub 2007 May 25.
Nature has devised many strategies for repairing DNA breaks. In homology-dependent pathways, the break is first processed to a 3'-ssDNA overhang that serves as a substrate for DNA strand exchange. Here, we demonstrate a distinct biochemical mechanism for DNA break processing employed by the AddAB class of helicase-nuclease. We show that this enzyme complex contains two active nuclease domains, each of which is dedicated to cleavage of one specific DNA strand. The nuclease activity responsible for cleavage in the 3'-->5' direction is attenuated when the enzyme encounters a recombination hotspot sequence, whereas cleavage in the 5'-->3' direction is unaffected, resulting in the production of recombinogenic 3'-terminated ssDNA tails. Finally, we show that the molecular events that underlie the recognition and response to recombination hotspots can be uncoupled: mutant proteins that are unable to cleave at recombination hotspots retain the ability to form stable complexes with the hotspot sequence.
大自然已经设计出许多修复DNA断裂的策略。在同源依赖性途径中，断裂首先被加工成3'-单链DNA突出端，作为DNA链交换的底物。在这里，我们展示了AddAB类解旋酶-核酸酶用于DNA断裂加工的独特生化机制。我们表明，这种酶复合物包含两个活性核酸酶结构域，每个结构域专门负责切割一条特定的DNA链。当该酶遇到重组热点序列时，负责3'→5'方向切割的核酸酶活性会减弱，而5'→3'方向的切割不受影响，从而产生具有重组活性的3'末端单链DNA尾巴。最后，我们表明，识别和响应重组热点的分子事件可以被分离：在重组热点处无法切割的突变蛋白仍保留与热点序列形成稳定复合物的能力。
