通过黄素腺嘌呤二核苷酸（FAD）诱导的顺磁弛豫增强对DNA光解酶修复机制的核磁共振研究。
NMR study of repair mechanism of DNA photolyase by FAD-induced paramagnetic relaxation enhancement.
作者信息
Ueda Takumi, Kato Akira, Ogawa Yuuta, Torizawa Takuya, Kuramitsu Seiki, Iwai Shigenori, Terasawa Hiroaki, Shimada Ichio
机构信息
Graduate School of Pharmaceutical Sciences, the University of Tokyo, Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
出版信息
J Biol Chem. 2004 Dec 10;279(50):52574-9. doi: 10.1074/jbc.M409942200. Epub 2004 Oct 1.
Cyclobutane pyrimidine dimer (CPD) photolyases, which contain FAD as a cofactor, use light to repair CPDs. We performed structural analyses of the catalytic site of the Thermus thermophilus CPD photolyase-DNA complex, using FAD-induced paramagnetic relaxation enhancement (PRE). The distances between the tryptophan residues and the FAD calculated from the PRE agree well with those observed in the x-ray structure (with an error of <3 A). Subsequently, a single-stranded DNA containing 13C-labeled CPD was prepared, and the FAD-induced PRE of the NMR resonances from the CPD lesion in complex with the CPD photolyase was investigated. The distance between the FAD and the CPD calculated from the PRE is 16 +/- 3 A. The FAD-induced PRE was also observed in the CPD photolyase-double-stranded DNA complex. Based on these results, a model of the CPD photolyase-DNA complex was constructed, and the roles of Arg-201, Lys-240, Trp-247, and Trp-353 in the CPD-repair reaction are discussed.
含有黄素腺嘌呤二核苷酸（FAD）作为辅因子的环丁烷嘧啶二聚体（CPD）光解酶利用光来修复CPD。我们使用FAD诱导的顺磁弛豫增强（PRE）对嗜热栖热菌CPD光解酶-DNA复合物的催化位点进行了结构分析。根据PRE计算出的色氨酸残基与FAD之间的距离与X射线结构中观察到的距离非常吻合（误差<3 Å）。随后，制备了含有13C标记CPD的单链DNA，并研究了与CPD光解酶形成复合物的CPD损伤的NMR共振的FAD诱导的PRE。根据PRE计算出的FAD与CPD之间的距离为16±3 Å。在CPD光解酶-双链DNA复合物中也观察到了FAD诱导的PRE。基于这些结果，构建了CPD光解酶-DNA复合物的模型，并讨论了精氨酸-201、赖氨酸-240、色氨酸-247和色氨酸-353在CPD修复反应中的作用。
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