C ENDOR脂氧酶基质复合物的光谱揭示了道化C-H激活的结构基础
Masaki Horitani1, Adam R Offenbacher, Cody A Marcus Carr
1Department of Chemistry, Northwestern University , Evanston, Illinois 60208, United States.
Journal of the American Chemical Society
|January 26, 2017
概括
酶C-H激活需要接近道. 这项研究揭示了大豆氧酶通过构造性采样实现这一点,而不是使用Mn2+和ENDOR光谱的不寻常的活性位点结构.
科学领域:
- 生物化学
- 量子生物学
- 酵素学
背景情况:
- 酶性C-H激活依赖于道化,需要一个短的捐赠者-接受器距离 (DAD) 进行有效的催化.
- 豆氧化酶 (SLO) 是研究非adiabatic道的一个关键系统,但其地面状态的活性位点结构和实现所需的DAD的机制尚不清楚.
- 之前的研究表明,要么采用不寻常的活性部位安排,要么采用构造性采样,以达到约2.7 Å的最佳DAD.
研究的目的:
- 阐明大豆脂氧酶 (SLO) 的基态活性位结构,并确定它如何实现非adiabatic道化所需的 donor-acceptor距离.
- 调查是否来自固定的活跃点几何或动态形状采样.
- 验证基质定位和活性位动力学在SLO的催化机制中的作用的动力学和理论预测.
主要方法:
- 使用Mn2+作为SLO中原生Fe离子的旋探替代物,通过X射线衍射证实了结构保真性.
- 采用13C和1H电子核双共振 (ENDOR) 光谱来精确地定位与金属中心相对的基质原子 (亚麻酸).
- 集成的ENDOR衍生控制装置与完全溶解的SLO模型的分子动力学模拟,以生成3D活动部位表示.
主要成果:
- 基态SLO活性位点存在于适配体的混合物中,包括具有~3.1 Å的DAD的反应状态 (接近范德瓦尔斯接触) 和具有较长DAD的不活性状态.
- 这表明随机形态采样,而不是静态的异常活位结构,对于实现高效道化所需的短DAD至关重要.
- 对道工程损害的SLO变体的分析显示了增加的DAD和改变的基板定位/刚度,证实了动力学和理论发现.
结论:
- 通过其活性位点的动态构造采样,大豆氧酶实现了非adiabatic道的最佳气供体-接受器距离.
- 该研究验证了SLO中质子合电子转移的非adiabatic量子处理的预测能力.
- 在测试,检测和证实动力学预测趋势时,ENDOR光谱非常灵敏,揭示了SLO活跃站点架构和反应性的关键方面.
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