STRUCTURAL STUDY OF THE DNA DECAMER DUPLEXES CONTAINING DOUBLE T-G MISMATCHES

这个子项目是许多研究子项目中的一个 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 了解DNA光损伤和修复过程的机制将是找到减少光损伤和增强修复的解决方案的第一步。顺式环丁烷嘧啶二聚体（cis-syn cyclobutane pyrimidine dimer，CPD）是紫外光诱导的一类重要的细胞毒性、致突变性和致癌性DNA光产物。尽管由于UV照射而形成CPD的情况非常频繁，但是CPD损伤很难被修复酶XPC-hHR 23 B识别，所述修复酶XPC-hHR 23 B是核苷酸切除修复（NER）途径的主要损伤识别蛋白。有趣的是，由于存在双T-G错配，CPD/GG双链体代表异常严重的螺旋扭曲，并且与通常的CPD损伤（CPD/TT）相比，XPC-hHR 23 B蛋白对CPD/GG双链体具有相当高的结合亲和力。因此，重要的是要了解这种蛋白质的靶底物是以何种方式形成的，以及T-G错配如何影响这一过程。为了实现这一目标，我们进行了广角X射线散射（WAXS）测量，以研究在CPD形成、双T-G错配以及与溶液中XPC-hHR 23 B蛋白相互作用下DNA双链体的结构变化。由于X射线溶液散射提供了关于结构的丰富信息，我们期望看到受损和未受损DNA之间以及正常和错配DNA之间的本质区别。我们成功地从DNA系统中获得了高质量的WAXS数据，实验数据与理论预测非常吻合。进一步的分析，以解决破坏和修复过程的性质仍在进行中。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Understanding the mechanism responsible for the DNA photodamage and the repair process would be a first step toward finding a solution to reduce the photodamage and enhance the repair. The cis-syn cyclobutane pyrimidine dimer (CPD) is one of the major classes of cytotoxic, mutagenic and carcinogenic DNA photoproducts induced by UV light. Though CPD formation due to the UV irradiation occurs quite frequently, CPD lesion is poorly recognized by repair enzyme XPC-hHR23B which is the primary damage recognition protein of the nucleotide excision repair (NER) pathway. Intriguingly, CPD/GG duplex represents unusually severe helical distortion due to the presence of double T-G mismatches, and XPC-hHR23B protein has a considerably higher binding affinity for CPD/GG duplex compared with the usual CPD lesion (CPD/TT). Therefore it is of importance to understand in which manner the target substrate for this protein is formed and how the T-G mismatches affect this process. Toward this goal we conducted wide-angle X-ray scattering (WAXS) measurement to investigate structural change of DNA duplex under CPD formation, double T-G mismatches and interaction with XPC-hHR23B protein in solution. Since X-ray solution scattering provides rich information about the structure, we expected to see the essential difference between damaged and undamaged DNA and between normal and mismatched DNA. We successfully obtained high-quality WAXS data from the DNA systems, with the experimental data in very good agreement with theoretical prediction. Further analysis to solve the nature of damaging and repairing process is still ongoing.