Hidden in Plain Sight: Subtle Effects of the 8-Oxoguanine Lesion on the Structure, Dynamics, and Thermodynamics of a 15-Base Pair Oligodeoxynucleotide Duplex
Hidden in Plain Sight: Subtle Effects of the 8-Oxoguanine Lesion on the Structure, Dynamics, and Thermodynamics of a 15-Base Pair Oligodeoxynucleotide Duplex
复制标题
DOI：
10.1021/bi201007t
复制
发表时间：
2011-10-04
期刊：
影响因子：
2.9
通讯作者：
Nunez, Megan E.
中科院分区：
文献类型：
--
作者：
Crenshaw, Charisse M.;Wade, Jacqueline E.;Nunez, Megan E.
碱基损伤8-氧代鸟嘌呤容易通过DNA氧化形成，可能导致G -> T颠换突变。尽管8-氧代鸟嘌呤-胞嘧啶碱基对与正常鸟嘌呤-胞嘧啶碱基对明显相似，但细胞碱基切除修复系统有效地识别病变碱基。在这里，我们应用几种技术来检查一个单一的8-oxoguanine病变的非回文15聚体双链体寡核苷酸的中心，以努力确定什么，如果有的话，区分8-oxoguanine-cytosine（8 oxoG-C）从一个正常的碱基对的碱基对。使用圆二色光谱和紫外线熔化热力学，病变双链体与未修饰的亲本双链体在全球几乎无法区分。检测DNA错配的光裂解剂Rh（bpy）（2）chrysi（3+）仅微弱且非特异性地裂解，揭示了8 oxoG-C对在单个碱基对水平上是局部稳定的。核磁共振谱也与保守的B型双链体结构一致。在二维核Overhauser效应光谱中，基糖和亚氨基-亚氨基交叉峰在母体和病变双链体之间惊人地相似。由于8 oxoG病变的化学位移的变化被定位到其互补的胞嘧啶和2-3 bp立即侧翼病变的病变的立场。残留物进一步从病变被证明是不受干扰的存在。值得注意的是，亚氨基交换实验表明，8-氧代鸟嘌呤-胞嘧啶对是强的和稳定的，具有等于其他内部鸟嘌呤-胞嘧啶碱基对的开放的表观平衡常数，在10（-6）的数量级。这些实验表明，8-氧代鸟嘌呤-胞嘧啶碱基对非常稳定，与天然碱基对相似。
The base lesion 8-oxoguanine is formed readily by oxidation of DNA, potentially leading to G -> T transversion mutations. Despite the apparent similarity of 8-oxoguanine-cytosine base pairs to normal guanine-cytosine base pairs, cellular base excision repair systems effectively recognize the lesion base. Here we apply several techniques to examine a single 8-oxoguanine lesion at the center of a nonpalindromic 15-mer duplex oligonucleotide in an effort to determine what, if anything, distinguishes an 8-oxoguanine-cytosine (8oxoG-C) base pair from a normal base pair. The lesion duplex is globally almost indistinguishable from the unmodified parent duplex using circular dichroism spectroscopy and ultraviolet melting thermodynamics. The DNA mismatch-detecting photocleavage agent Rh(bpy)(2)chrysi(3+) cleaves only weakly and nonspecifically, revealing that the 8oxoG-C pair is locally stable at the level of the individual base pairs. Nuclear magnetic resonance spectra are also consistent with a well-conserved B-form duplex structure. In the two-dimensional nuclear Overhauser effect spectra, base-sugar and imino-imino cross-peaks are strikingly similar between parent and lesion duplexes. Changes in chemical shift due to the 8oxoG lesion are localized to its complementary cytosine and to the 2-3 bp immediately flanking the lesion on the lesion stand. Residues further removed from the lesion are shown to be unperturbed by its presence. Notably, imino exchange experiments indicate that the 8-oxoguanine-cytosine pair is strong and stable, with an apparent equilibrium constant for opening equal to that of other internal guanine-cytosine base pairs, on the order of 10(-6). This collection of experiments shows that the 8-oxoguanine-cytosine base pair is incredibly stable and similar to the native pair.