The Mechanism of Cyclobutylpyrimidine Dimer Repair by Photoinduced Electron Transfer in DNA Photolyase: Protein and Model Studies

StanleyMCB 9982532许多生物体已经开发出DNA修复机制，其基础是光诱导的超快电子从黄素蛋白（称为DNA光裂合酶）转移到紧密结合的环丁基嘧啶二聚体（CPD）损伤。 本研究的目的是阐明详细的动力学和动力学的电子转移和二聚体裂解反应的光解酶-CPD复合物，使用近紫外/紫外飞秒吸收光谱。对这些实验的分析将表明CPD是否是通过协调或顺序机制修复的，以及涉及哪些中间体。将合成黄素-DNA模型系统，以分离和突出酶促反应的各种动力学细节，包括蛋白质在修复中的作用。荧光标记技术将用于获得CPD裂解反应伴随的结构变化的实时信息。紫外线辐射能够交联DNA中相邻的核酸碱基，导致突变和细胞死亡。最常见和致命的DNA光损伤是环丁基嘧啶二聚体（CPD）。为了应对这些损伤，许多生物体使用黄素蛋白来修复受损的DNA。这些蛋白质被称为光解酶，它们的特殊之处在于它们需要可见光来消除诱变紫外线辐射造成的损伤。在吸收可见光后，修复过程在2纳秒内完成。这项研究旨在了解光驱动修复反应背后的分子机制。

StanleyMCB 9982532Many organisms have developed DNA repair mechanisms based on photo-induced ultrafast electron transfer from a flavoprotein, called DNA photolyase, to the tightly bound cyclobutylpyrimidine dimer (CPD) lesion. The objective of this research is to elucidate the detailed kinetics and dynamics of the electron transfer and dimer splitting reactions of the photolyase-CPD complex, using near-UV/UV femtosecond absorption spectroscopy. An analysis of these experiments will show whether the CPD is repaired by a concerted or sequential mechanism and what intermediates are involved. Flavin-DNA model systems will be synthesized to isolate and highlight various kinetic details of the enzymatic reaction, including the role of the protein in the repair. Fluorescent labeling technology will be used to gain real-time information on the structural changes that accompany the CPD splitting reaction.Ultraviolet radiation is capable of crosslinking adjacent nucleic acid bases in DNA, leading to mutation and cell death. The most common and lethal DNA photo-lesion is the cyclobutylpyrimidine dimer (CPD). In response to these lesions many organisms use flavoproteins to repair the damaged DNA. These proteins are called photolyases and are unusual in that they require visible light to undo the damage wrought by the mutagenic UV radiation. Upon absorption of visible light the repair process is complete within 2 nanoseconds. This research is aimed at understanding the molecular mechanism behind the light-driven repair reaction.