IRB-BSC Joint Research Program in Computational Biology, Barcelona Supercomputing Center, Torre Girona, C/Jordi Girona 31, Barcelona 08034, Spain.
J Am Chem Soc. 2011 Aug 10;133(31):12154-61. doi: 10.1021/ja204053n. Epub 2011 Jul 15.
The continually emerging functional significance of intrinsic disorder and conformational flexibility in proteins has challenged the long-standing dogma of a well-defined structure contributing to a specific function. Molten-globular states, a class of proteins with significant secondary-structure but a fluid hydrophobic core, is one such example. They have however been difficult to characterize due to the complexity of experimental data and lack of computational avenues. Here, we dissect the folding mechanism of the α-helical molten-globular protein NCBD from three fundamentally different approaches: statistical-mechanical variable barrier model, C(α)-based Gō-model and explicit water all-atom molecular dynamics (MD) simulations. We find that NCBD displays the characteristics of a one-state globally downhill folder but is significantly destabilized. Using simulation techniques, we generate a highly constrained but a heterogeneous native ensemble of the molten-globule for the first time that is consistent with experimental data including small angle X-ray scattering (SAXS), circular dichroism (CD), and nuclear magnetic resonance (NMR). The resulting native ensemble populates conformations reported in other bound-forms providing direct evidence to the mechanism of conformational selection for binding multiple partners in this domain. Importantly, our simulations reveal a connection between downhill folding and large conformational flexibility in this domain that has been evolutionarily selected and functionally exploited resulting in large binding promiscuity. Finally, the multimodel approach we employ here serves as a powerful methodology to study mechanisms and suggests that the thermodynamic features of molten-globules fall within the array of folding mechanisms available to small single-domain proteins.
蛋白质中固有无序和构象灵活性的不断涌现的功能意义，挑战了明确的结构有助于特定功能的长期存在的教条。具有显著二级结构但具有流体疏水性核心的熔融球蛋白状态就是这样的一个例子。然而，由于实验数据的复杂性和缺乏计算途径，它们一直难以表征。在这里，我们从三个根本不同的方法来剖析α-螺旋熔融球蛋白 NCBD 的折叠机制：统计力学变势垒模型、基于 C(α)的 Gō 模型和显式水全原子分子动力学（MD）模拟。我们发现 NCBD 显示出单态全局下坡折叠的特征，但稳定性显著降低。使用模拟技术，我们首次生成了一个高度约束但具有异质天然集合的熔融球蛋白，与实验数据包括小角度 X 射线散射（SAXS）、圆二色性（CD）和核磁共振（NMR）一致。所得的天然集合体填充了在其他结合形式中报告的构象，为该结构域中与多个配体结合的构象选择机制提供了直接证据。重要的是，我们的模拟揭示了该结构域中下坡折叠和大构象灵活性之间的联系，这种联系是经过进化选择和功能利用的，导致了大的结合多态性。最后，我们在这里采用的多模型方法为研究机制提供了一种强大的方法，并表明熔融球蛋白的热力学特征属于小单域蛋白可用的折叠机制范围。
