Alsner J, Sorensen H V, Schmidt V K, Sorensen B S, Westergaard O
Department of Molecular and Structural Biology, University of Aarhus, Denmark.
J Mol Biol. 1996 Jun 14;259(3):317-24. doi: 10.1006/jmbi.1996.0321.
To determine the specific interaction sites of topoisomerase II within the DNA region defined by the footprint of the enzyme, we have investigated the cleavage reaction on double-stranded DNA substrates containing nicks and deletions. Topoisomerase II-mediated cleavage of the DNA substrates is suicidal as the enzyme is unable to religate the cleaved DNA due to diffusion of the small nucleotides 5' to the cleavage position. Thus, suicidal cleavage is obtained with substrates having one, two or three nucleotides 5' to the cleavage position. The enzyme requires interaction with three distinct regions of double-stranded DNA for cleavage to occur, one region spanning the eight nucleotides located around the cleavage position and two distal regions each spanning approximately six nucleotides. A model is proposed, where these data are taken to imply that two distinct regions of interactions exist between each topoisomerase II subunit and its DNA substrate. The model is discussed in relation to the recently solved three-dimensional structure of yeast topoisomerase II.
为了确定拓扑异构酶II在该酶足迹所定义的DNA区域内的特定相互作用位点，我们研究了含切口和缺失的双链DNA底物上的切割反应。拓扑异构酶II介导的DNA底物切割是自杀性的，因为由于小核苷酸向切割位点5'端扩散，该酶无法重新连接切割后的DNA。因此，在切割位点5'端有一个、两个或三个核苷酸的底物上可获得自杀性切割。该酶需要与双链DNA的三个不同区域相互作用才能发生切割，一个区域跨越切割位点周围的八个核苷酸，另外两个远端区域各跨越约六个核苷酸。我们提出了一个模型，这些数据表明每个拓扑异构酶II亚基与其DNA底物之间存在两个不同的相互作用区域。结合最近解析的酵母拓扑异构酶II的三维结构对该模型进行了讨论。
