在阿波和基质模仿状态下,mus-ms酶运动的保存
Heather Beach1, Roger Cole, Michelle L Gill
1Department of Chemistry, Yale University, New Haven, Connecticut 06520, USA.
Journal of the American Chemical Society
|June 23, 2005
概括
核磁共振 (NMR) 旋转放松实验显示,无论是以apo形式还是与基质模仿物结合,Ribonuclease A (RNase A) 动态保持一致. 带结合稳定了,但没有诱导酶的结构变化.
科学领域:
- 生物化学 生物化学
- 结构生物学 结构生物学
- 生物物理学的生物物理.
背景情况:
- 核糖酶A (RNase A) 是生物系统中一个关键的酶.
- 了解酶动态是阐明催化机制的关键.
- 基质结合可以显著改变酶的结构和灵活性.
研究的目的:
- 为了比较RNase A在其apo形式中的分子动力学 (mus-ms) 与与基质模拟物 (pTppAp) 复合时的分子动力学 (mus-ms).
- 调查连接体结合在调节酶灵活性和构成状态中的作用.
- 确定控制RNase A动态的能量和运动参数.
主要方法:
- 溶液核磁共振 (NMR) 旋转放松实验,包括放松补偿的卡尔-普尔塞尔-梅布姆-吉尔 (CPMG) 实验.
- 对NMR自旋放松分散曲线的定量分析.
- 取决于温度的测量以确定激活障碍.
- 确定RNase A/pTppAp复合物的晶体结构.
主要成果:
- 结晶结构证实pTppAp与RNase A活性部位结合.
- 在CPMG的实验中,在apo和结合的RNase A.中都发现了柔性残留物.
- 核磁共振回旋-放松分散分析显示,在pTppAp结合时,运动时间表没有变化.
- 运动的激活障碍与apo (5.2 ± 1.0 kcal/mol) 和绑定 (4.5 ± 1.2 kcal/mol) RNase A. 的激活障碍相似.
- 化学转移数据表明,在两个状态中,运动的大小相似.
结论:
- RNase A存在于开放式和闭合式构造之间的动态平衡中.
- 联体结合稳定了结合的适配体,但没有诱导它.
- 酶样本的apo-like形状,即使当连接体被绑定,表明先前存在的灵活性.
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