Department of Chemistry, State University of New York at Potsdam, Potsdam, NY 13676, United States.
Mutat Res. 2012 Jul 1;735(1-2):1-11. doi: 10.1016/j.mrfmmm.2012.05.005. Epub 2012 Jun 6.
The catechol-mediated DNA damage in the presence of Cu(II) ions involves oxidation of guanine to 8-oxoguanine (8-oxoG) and DNA strand scission. It proceeds through the reactive oxygen species (ROS) generation. The mutagenicity of 8-oxoG lesions is due to its miscoding propensity reflected in GC→TA transversion taking place during the DNA repair process. To gain new insights into the nature of catechol-mediated DNA damage and its prevention, we have investigated the changes in DNA melting characteristics and 8-oxoG formation as the indicators of DNA damage in a model calf-thymus DNA system. A novel fluorescence method for DNA melting temperature determination, based on DAPI fluorescent-probe staining, has been proposed. The DNA melting-onset temperature has been found to be more sensitive to DNA damage than the standard melting temperature due to the increased width of the melting transition observed in oxidatively damaged DNA. We have found that the efficiency of Fenton cascade in generating DNA-damaging ROS is higher for catechol than for GSH, two strong antioxidants, mainly due to the much longer distance between ROS-generating radical group in GS to nucleobases than that of semiquinone radical group to nucleobases (2.1nm vs. 0.27nm), making the ROS transport from GSH an order of magnitude less likely to damage DNA because of short lifetime of HO radicals. The antioxidant and DNA-protecting behaviors of GSH have been elucidated. We have found that the redox potential of GSH/GSSG couple is lower than that of catechol/semiquinone couple. Hence, GSH keeps catechol in the reduced state, thereby shutting down the initial step of the catechol-mediated Fenton cascade. The catechol-induced DNA damage in the presence of Cu(II) ions has also been confirmed in studies of ON-OFF hairpin-oligonucleotide beacons.
在 Cu(II)离子存在下，儿茶酚介导的 DNA 损伤涉及鸟嘌呤氧化为 8-氧代鸟嘌呤（8-oxoG）和 DNA 链断裂。它通过活性氧物种（ROS）的生成进行。8-oxoG 损伤的致突变性是由于其在 DNA 修复过程中发生的 GC→TA 颠换反映了其错配倾向。为了深入了解儿茶酚介导的 DNA 损伤的性质及其预防，我们研究了小牛胸腺 DNA 模型系统中 DNA 熔融特性和 8-oxoG 形成的变化，作为 DNA 损伤的指标。提出了一种基于 DAPI 荧光探针染色的新的 DNA 熔融温度测定荧光方法。由于在氧化损伤的 DNA 中观察到熔融转变的宽度增加，因此 DNA 熔融起始温度比标准熔融温度对 DNA 损伤更为敏感。我们发现，Fenton 级联在生成 DNA 损伤 ROS 方面，儿茶酚比两种强抗氧化剂 GSH 更有效，主要是因为 GS 中产生 ROS 的自由基基团与碱基之间的距离比半醌自由基基团与碱基之间的距离长得多（2.1nm 比 0.27nm），使得 ROS 从 GSH 运输到 DNA 损伤的可能性小一个数量级，因为 HO 自由基的寿命短。阐明了 GSH 的抗氧化和 DNA 保护作用。我们发现 GSH/GSSG 对的氧化还原电位低于儿茶酚/半醌对的氧化还原电位。因此，GSH 将儿茶酚保持在还原状态，从而阻止儿茶酚介导的 Fenton 级联的初始步骤。在 Cu(II)离子存在下，儿茶酚诱导的 DNA 损伤也在 ON-OFF 发夹寡核苷酸信标研究中得到了证实。
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