在Escherichia coli pyruvate dehydrogenase复合体上确定稳定状态前的速率常数显示,循环运动控制了速率限制步骤
Anand Balakrishnan1, Natalia S Nemeria, Sumit Chakraborty
1Department of Chemistry, Rutgers the State University, Newark, New Jersey 07102, United States.
Journal of the American Chemical Society
|October 24, 2012
概括
酸盐脱酶复合体 (PDHc) E1p组件中的动态区域对于催化非常重要. 破坏活点环节显著减缓了中间形成,显示这些动态控制了共价催化.
科学领域:
- 酶学和酶动力学
- 酶复合体的结构生物学
- 代谢生物化学 代谢生物化学
背景情况:
- 酸盐脱酶多酶复合体 (PDHc) 是碳代谢的核心.
- 假设E1p组件的动态区域调节PDHc的催化循环.
- 了解中间体形成是阐明PDHc功能的关键.
研究的目的:
- 在PDHc及其变体中的共价中间体的动态特征.
- 研究E1p组件在PDHc催化中的动态区域的作用.
- 为了确定PDHc组件是否影响E1p循环动态和催化.
主要方法:
- 在E1p上结合胺二酸盐 (ThDP) 的中间体的动态表征.
- 在PDHc和变体内对E2p的胺胺结合中间体的分析.
- 在E1p组件中活动部位循环的部位定向突变发生.
主要成果:
- 鉴定出C2α-lactylthiamin二酸盐 (LThDP) 中间体的形成是限制速率的.
- 活跃地点环中的突变使LThDP形成率降低了多达900倍.
- 将E401K变种组装成PDHc部分恢复了LThDP形成速度.
结论:
- E1p循环动力学对于控制涉及ThDP的共价催化是必不可少的.
- PDHc组件调节这些动态,可能通过选择特定的循环形状.
- 这种界面动态区域控制机制可以在2-oxoacid脱酶复合体中保持.
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