催化空间工程作为一种在哺乳动物基因组中激活C-H氧化的策略
Sushma Sappa1, Debasis Dey1, Babu Sudhamalla1
1Department of Chemistry, University of Pittsburgh, Pittsburgh, Pennsylvania 15260, United States.
Journal of the American Chemical Society
|July 29, 2021
概括
蛋白质工程现在可以通过改变活性位点大小来控制酶催化. 这项研究表明,扩展活性位点增强了TET酶在5-甲基细胞素 (5mC) 修饰的活性.
科学领域:
- 酶工程
- 生物化学
- 分子生物学
背景情况:
- 通过蛋白质工程控制酶催化是一项挑战.
- 十一转位 (TET) 酶家族在DNA脱甲基化中起着至关重要的作用.
- 调节TET酶活性是理解和操纵基因表达的关键.
研究的目的:
- 开发一种新型的活性位点工程策略,以指导TET酶催化.
- 研究活性位体体积对TET酶的催化效率的影响.
- 确定TET酶家族中负责催化方向的关键残留物.
主要方法:
- 有修改的活性部位腔 (扩大或拥挤) 的TET酶的工程变体.
- 对5-甲基细胞素 (5mC) 多步C-H氧化的催化活性进行评估.
- 使用结构分析和突变生成来识别进化保存的催化指导残留物.
主要成果:
- 具有扩大活性位点的变体显著增强了5mC的多步C-H氧化.
- 一个拥挤的活跃部位导致了一步的催化机制.
- 发现了一种具有显著催化指导能力的保存残留物.
- 在染色体DNA中成功氧化了5mC,增强了基因表达.
结论:
- 活跃站点工程是控制TET酶催化结果的可行策略.
- 调节活性位点架构允许微调5mC氧化路径.
- 可以使用TET酶的向工程来调节基因表达,在癌症治疗中具有潜在的应用.
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