Department of Biology, New York University, 100 Washington Square East, New York, NY 10003, USA.
Nucleic Acids Res. 2012 Mar;40(6):2506-17. doi: 10.1093/nar/gkr1087. Epub 2011 Dec 1.
The one-electron oxidation of guanine in DNA by carbonate radical anions, a decomposition product of peroxynitrosocarbonate which is associated with the inflammatory response, can lead to the formation of intrastrand cross-links between guanine and thymine bases [Crean et al. (Oxidation of single-stranded oligonucleotides by carbonate radical anions: generating intrastrand cross-links between guanine and thymine bases separated by cytosines. Nucleic Acids Res. 2008; 36: 742-755.)]. These involve covalent bonds between the C8 positions of guanine (G*) and N3 of thymine (T*) in 5'-d(…GpT…) and 5'-d(…GpCpT…) sequence contexts. We have performed nucleotide excision repair (NER) experiments in human HeLa cell extracts which show that the GCT intrastrand cross-link is excised with approximately four times greater efficiency than the GT cross-link embedded in 135-mer DNA duplexes. In addition, thermal melting studies reveal that both lesions significantly destabilize duplex DNA, and that the destabilization induced by the GCT cross-link is considerably greater. Consistent with this difference in NER, our computations show that both lesions dynamically distort and destabilize duplex DNA. They disturb Watson-Crick base-pairing and base-stacking interactions, and cause untwisting and minor groove opening. These structural perturbations are much more pronounced in the GCT than in the GT cross-link. Our combined experimental and computational studies provide structural and thermodynamic understanding of the features of the damaged duplexes that produce the most robust NER response.
DNA 中鸟嘌呤被碳酸盐自由基阴离子（过氧亚硝酰碳酸盐的分解产物，与炎症反应有关）单电子氧化，可导致鸟嘌呤和胸腺嘧啶碱基之间形成链内交联[Crean 等人（Oxidation of single-stranded oligonucleotides by carbonate radical anions: generating intrastrand cross-links between guanine and thymine bases separated by cytosines. Nucleic Acids Res. 2008; 36: 742-755.）]。这些交联涉及到 5′-d(…GpT…)和 5′-d(…GpCpT…)序列中鸟嘌呤（G*）的 C8 位和胸腺嘧啶（T*）的 N3 位之间的共价键。我们在人 HeLa 细胞提取物中进行了核苷酸切除修复（NER）实验，结果表明，GCT链内交联的切除效率比嵌入 135 个碱基对 DNA 双链中的 GT交联高约四倍。此外，热融解研究表明，这两种损伤都显著破坏双链 DNA 的稳定性，而 GCT交联引起的不稳定程度要大得多。与 NER 中的这种差异一致，我们的计算表明，这两种损伤都会动态扭曲和破坏双链 DNA。它们破坏沃森-克里克碱基配对和碱基堆积相互作用，并导致扭曲和小沟打开。与 GT交联相比，这种结构扰动在 GCT交联中更为明显。我们的综合实验和计算研究为产生最强大 NER 反应的损伤双链的结构和热力学特征提供了理解。
