Simulation and experiment at high temperatures: Ultrafast folding of a thermophilic protein by nucleation-condensation
Simulation and experiment at high temperatures: Ultrafast folding of a thermophilic protein by nucleation-condensation
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DOI：
10.1016/j.jmb.2004.12.061
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发表时间：
2005-04-08
影响因子：
5.6
通讯作者：
Fersht, AR
中科院分区：
文献类型：
--
作者：
Ferguson, N;Day, R;Fersht, AR
我们用Phi值分析法确定了一个嗜热蛋白质在325 K相对高温下折叠的过渡态。在不存在化学变性剂的情况下，通过温度跳跃实验测定嗜热脂肪芽孢杆菌（E3 BD）的三螺旋束、外周亚基结合结构域的折叠的Phi（F）值。E3 BD折叠在微秒通过一个高度扩散的过渡状态。在373 K下，实验结果与八个（独立的）分子动力学模拟的解折叠结果之间观察到极好的一致性。我们使用了杂波NMR实验和分子动力学模拟相结合的变性合奏，并发现它包含很少的持久性，残留结构。然而，通过模拟发现，在天然状态下采用螺旋结构的那些区域在变性状态下准备形成螺旋。这些区域在折叠的过渡态中也具有显著的结构。整个折叠途径似乎是成核-缩合。(c)2005爱思唯尔有限公司保留所有权利。
We used Phi-value analysis to characterise the transition state for folding of a thermophilic protein at the relatively high temperature of 325 K. Phi(F) values for the folding of the three-helix bundle, peripheral subunit binding domain from Bacillus stearothermophilus (E3BD) were determined by temperature-jump experiments in the absence of chemical denaturants. E3BD folded in microseconds through a highly diffuse transition state. Excellent agreement was observed between experiment and the results from eight (independent) molecular dynamics simulations of unfolding at 373 K. We used a combination of heteronuclear NMR experiments and molecular dynamics simulations to characterise the denatured ensemble, and found that it contained very little persistent, residual structure. However, those regions that adopt helical structure in the native state were found by simulation to be poised for helix formation in the denatured state. These regions also had significant structure in the transition state for folding. The overall folding pathway appears to be nucleation-condensation. (c) 2005 Elsevier Ltd. All rights reserved.