在2-基乙酸二氧化原酶和甲基酸合成酶的催化过程中常见的晚期中间体
Spencer C Peck1, Jonathan R Chekan, Emily C Ulrich
1Howard Hughes Medical Institute and Department of Chemistry, University of Illinois at Urbana-Champaign , 600 South Mathews Avenue, Urbana, Illinois 61801, United States.
一种双功能突变的2-基乙酸氧化酶 (HEPD) 模仿甲基酸合成酶 (MPnS) 的活动. 这表明,涉及一个共同的中间体的同位素敏感分支机制决定了这些非血红素铁氧酶的产品形成.
科学领域:
- 生物化学 生物化学
- 酶学 是一种酶学.
- 结构生物学 结构生物学
背景情况:
- 非血铁氧酶催化了关键的C-C键裂解反应.
- 2-基酸二氧化原酶 (HEPD) 和甲基酸合成酶 (MPnS) 是不同的酶,具有重叠的基质特异性.
- 之前的研究提出了一个共同的中间产品决定了产品的身份.
研究的目的:
- 设计一种双功能酶,结合HEPD和MPnS活动.
- 调查差异产品形成的机制基础.
- 阐明活性部位残留在酶功能的作用.
主要方法:
- 使用局部导向突变发生来产生双功能HEPD突变体 (E176H).
- 用同位素标记的基质进行了酶活性测试.
- 采用X射线晶体学来确定突变酶的结构.
主要成果:
- 该E176H突变体表现出HEPD和MPnS活动.
- 产品分销对基质同位素效应敏感,支持分支机制.
- 结构分析表明,引入的histidine不协调活性部位的金属.
结论:
- 在HEPD和MPnS催化中都涉及一种共同的中间体.
- 产品的结果是由对同位素敏感的分支阶段决定的.
- 活性部位残留物替代可以改变酶的功能和机制.
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