Department of Chemistry - BMC , Uppsala University , BMC Box 576, 751 23 Uppsala , Sweden.
Institute of Complex Systems: Structural Biochemistry (ICS-6) , Forschungszentrum Jülich , 52425 Jülich , Germany.
J Am Chem Soc. 2018 Nov 21;140(46):15889-15903. doi: 10.1021/jacs.8b09378. Epub 2018 Nov 8.
Conformational changes are crucial for the catalytic action of many enzymes. A prototypical and well-studied example is loop opening and closure in triosephosphate isomerase (TIM), which is thought to determine the rate of catalytic turnover in many circumstances. Specifically, TIM loop 6 "grips" the phosphodianion of the substrate and, together with a change in loop 7, sets up the TIM active site for efficient catalysis. Crystal structures of TIM typically show an open or a closed conformation of loop 6, with the tip of the loop moving ∼7 Å between conformations. Many studies have interpreted this motion as a two-state, rigid-body transition. Here, we use extensive molecular dynamics simulations, with both conventional and enhanced sampling techniques, to analyze loop motion in apo and substrate-bound TIM in detail, using five crystal structures of the dimeric TIM from Saccharomyces cerevisiae. We find that loop 6 is highly flexible and samples multiple conformational states. Empirical valence bond simulations of the first reaction step show that slight displacements away from the fully closed-loop conformation can be sufficient to abolish most of the catalytic activity; full closure is required for efficient reaction. The conformational change of the loops in TIM is thus not a simple "open and shut" case and is crucial for its catalytic action. Our detailed analysis of loop motion in a highly efficient enzyme highlights the complexity of loop conformational changes and their role in biological catalysis.
构象变化对于许多酶的催化作用至关重要。三磷酸甘油醛异构酶（TIM）中的环的打开和关闭就是一个典型的、研究充分的例子，它被认为在许多情况下决定了催化周转率。具体来说，TIM 环 6“抓住”底物的磷酸二阴离子，并与环 7 的变化一起，为高效催化作用建立 TIM 活性位点。TIM 的晶体结构通常显示环 6 的开放或闭合构象，环的尖端在构象之间移动约 7 Å。许多研究将这种运动解释为两态、刚体跃迁。在这里，我们使用广泛的分子动力学模拟，包括传统和增强采样技术，使用来自酿酒酵母的二聚 TIM 的五个晶体结构，详细分析无配体和底物结合的 TIM 中的环运动。我们发现环 6 具有高度的灵活性，并能采样多种构象状态。第一步反应的经验价键模拟表明，稍微偏离完全闭环构象就足以消除大部分催化活性；完全闭环对于高效反应是必需的。因此，TIM 中环的构象变化不是一个简单的“开和关”的情况，而是其催化作用的关键。我们对高效酶中环运动的详细分析突出了环构象变化的复杂性及其在生物催化中的作用。
