在酶活性部位的X射线和NMR结晶学:在托合成酶中的印林化物中间体
Jinfeng Lai1, Dimitri Niks, Yachong Wang
1Department of Chemistry, University of California, Riverside, California 92521, USA.
Journal of the American Chemical Society
|December 15, 2010
概括
固态NMR和X射线结晶学结合起来,揭示了托合成酶中一个关键中间体的详细结构. 这种方法通过在催化过程中定义质子化状态来澄清酶机制.
科学领域:
- 生物化学 生物化学
- 结构生物学 结构生物学
- 酶催化酶的催化作用
背景情况:
- 了解酶机制需要详细的化学信息,如质子化和杂交状态.
- 仅靠X射线晶体学,即使在高分辨率下,也往往难以辨别这些细微的化学细节.
- 核磁共振 (NMR) 光谱的化学转移对当地的化学环境非常敏感.
研究的目的:
- 在活性催化过程中,确定氧-5'-酸盐依赖的酸合成酶中的印林化物中间体的化学丰富的晶体结构.
- 展示结合固态NMR光谱与X射线晶体学进行详细的结构分析的力量.
主要方法:
- 采用了结合X射线晶体学和固态NMR光谱学的协同方法.
- 使用ab initio计算化学的反应基质模拟的优化模型.
- 在优化过程中,在结晶学上确定的坐标处固定侧链残留.
- 使用 (13) C 和 (15) N 标签来测量基板上特定位置的化学转移.
主要成果:
- 成功确定了印林诺化物中间体的化学细节结构.
- 对基质和催化残留物进行电荷和质子化状态的各种模型进行区分.
- 确定了计算化学转移对特定标记位置的重大影响.
结论:
- 结合的NMR和晶体学方法提供了化学详细的三维结构.
- 这项研究强调了基质的双相体形式之间的平衡.
- 较大的复合体形式的质子化状态对于指导随后的催化步骤至关重要.
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