Xu Yan, Tashiro Ryu, Sugiyama Hiroshi
Department of Chemistry, Graduate School of Science, Kyoto University, Sakyo, Kyoto 606-8502, Japan.
Nat Protoc. 2007;2(1):78-87. doi: 10.1038/nprot.2006.467.
The various conformations of DNA are thought to have important biological roles. Investigation of the local DNA conformational changes associated with biological events is therefore essential to an understanding of the functions of DNA. We have reported the photoreactivities of 5-halouracil in the five characteristic local DNA structures: the A, B and Z forms, protein-induced DNA kinks and the G-quadruplex form. These studies demonstrate the detailed relationships between the local DNA structures and the photochemical products of photoinduced hydrogen abstraction by the resulting uracil-5-yl radicals, and show that this photochemical method can be used to detect DNA structures. Here, we describe in detail procedures that have been developed in our laboratory for probing DNA conformations by product analysis of photoirradiated 5-halouracil-containing DNA. The protocol includes the preparation of 5-halouracil-containing DNA and the characterization of the photoproducts, and it can be completed in 2 weeks.
人们认为DNA的各种构象具有重要的生物学作用。因此，研究与生物事件相关的局部DNA构象变化对于理解DNA的功能至关重要。我们已经报道了5-卤代尿嘧啶在五种特征性局部DNA结构中的光反应性：A、B和Z型、蛋白质诱导的DNA弯折以及G-四链体结构。这些研究揭示了局部DNA结构与由生成的尿嘧啶-5-基自由基光诱导氢提取的光化学产物之间的详细关系，并表明这种光化学方法可用于检测DNA结构。在此，我们详细描述了我们实验室开发的通过对光照射含5-卤代尿嘧啶的DNA进行产物分析来探测DNA构象的程序。该方案包括含5-卤代尿嘧啶的DNA的制备和光产物的表征，并且可以在2周内完成。
