Been M D, Champoux J J
J Mol Biol. 1984 Dec 15;180(3):515-31. doi: 10.1016/0022-2836(84)90025-1.
Eukaryotic type 1 DNA topoisomerases break single-stranded DNA at specific sites. A preferred site for rat liver topoisomerase breakage in single-stranded phi X174 DNA was located within a region of the DNA with the potential for duplex formation. To investigate the relationship between sites of breakage in duplex and single-stranded DNA, a restriction fragment containing sequences from the transcriptional regulatory and enhancer region of the simian virus 40 genome was used as a substrate for topoisomerase. While different patterns of breakage in the native and denatured forms of the DNA were observed, some sites of breakage were common to both forms. The break sites in the denatured DNA were a subset of the break sites in the duplex DNA and were located in regions which had the potential for intrastrand base-pairing due to distal complementary sequences. A series of single-stranded fragments were generated with the distal complementary sequences deleted and these fragments were used as substrates for topoisomerase breakage. The lack of detectable breakage at a site when the complementary sequence was deleted, suggests that topoisomerase acts at duplex regions in the single-stranded DNA and that it is not active on regions of single-stranded DNA that are not base-paired.
真核生物1型DNA拓扑异构酶在特定位点切断单链DNA。大鼠肝脏拓扑异构酶在单链φX174 DNA中切断的首选位点位于DNA中具有形成双链可能性的区域内。为了研究双链DNA和单链DNA中切断位点之间的关系，将一个包含猿猴病毒40基因组转录调控和增强子区域序列的限制性片段用作拓扑异构酶的底物。虽然观察到DNA天然形式和变性形式中不同的切断模式，但两种形式都有一些共同的切断位点。变性DNA中的切断位点是双链DNA中切断位点的一个子集，并且位于由于远端互补序列而具有链内碱基配对可能性的区域。生成了一系列删除了远端互补序列的单链片段，这些片段用作拓扑异构酶切断的底物。当互补序列被删除时，在一个位点未检测到切断，这表明拓扑异构酶作用于单链DNA中的双链区域，并且它对未碱基配对的单链DNA区域没有活性。
