摘要/Abstract

摘要： 目的· 研究SUMO 化修饰对胸腺嘧啶DNA 糖基化酶（thymine DNA glycosylase，TDG）蛋白的结构、稳定性及活性的影响。
方法· 建立体外表达纯化体系，获取可用于晶体筛选及活性检测的SUMO-1-TDG 蛋白。通过晶体筛选、衍射数据收集及结构解析，
分析SUMO-1-TDG 的分子结构。利用蛋白热稳定性实验检测在SUMO 化修饰前后TDG 稳定性的变化。建立TDG 活性测试体系，探
讨SUMO 化修饰对TDG 活性产生的影响。结果· 通过结构解析得到SUMO-1-TDG 分子结构。稳定性、活性实验检测发现TDG 蛋白
熔解温度（Tm）值提高约16℃，催化活性提升约9.70 %。结论· SUMO-1 分子结合TDG 对其进行修饰使得结合位点附近氨基酸形成
分子间相互作用，并进一步参与调控TDG 蛋白的稳定性及催化活性。
关键词: 胸腺嘧啶DNA 糖基化酶, SUMO 化, 晶体结构, 稳定性实验, 活性检测
Abstract:
Objective · To study the effect of sumoylation on the structure, stability and activity of human thymine DNA glycosylase (TDG).
Methods · Expression and purification systems were established for obtaining SUMO-1-TDG protein with high purity which can be used for crystal
screening and activity detection. Structure of SUMO-1-TDG was solved after crystal screening, diffraction data collection and structure analysis. The
change of TDG stability led by sumoylation was detected through a protein thermal shift assay. In addition, an activity assay was applied to investigate the
effect of sumoylation on the activity of TDG. Results · A high-resolution structure of SUMO-1-TDG which could clearly describe the interaction between
TDG and SUMO-1 was solved. The melting temperature (Tm) value of SUMO-1-TDG increased by about 16 ℃ and the catalytic activity increased by 9.70%,
comparing with TDG protein. Conclusion · SUMO-1 binds to TDG to modify the intermolecular interaction of amino acids near the binding site, and
further participates in the regulation of the stability and catalytic activity of TDG protein.
Key words: thymine DNA glycosylase, sumoylation, crystal structure, thermal shift assay, activity assay


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