Sugita Y, Kitao A
Department of Chemistry, Graduate School of Science, Kyoto University, Kyoto 606-8502, Japan.
Biophys J. 1998 Nov;75(5):2178-87. doi: 10.1016/S0006-3495(98)77661-1.
Free energy calculations were carried out to understand the effect of the I56V mutation of human lysozyme on its thermal stability. In the simulation of the denatured state, a short peptide including the mutation site in the middle is employed. To study the dependence of the stability on the denatured-state structure, five different initial conformations, native-like, extended, and three random-coil-like conformations, were examined. We found that the calculated free energy difference, DeltaDeltaGcal, depends significantly on the structure of the denatured state. When native-like structure is employed, DeltaDeltaGcal is in good agreement with the experimental free energy difference, DeltaDeltaGexp, whereas in the other four models, DeltaDeltaGcal differs sharply from DeltaDeltaGexp. It is therefore strongly suggested that the structure around the mutation site takes a native-like conformation rather than an extended or random-coil conformation. From the free energy component analysis, it has been shown that free energy components originating from Lennard-Jones and covalent interactions dominantly determine DeltaDeltaGcal. The contribution of protein-protein interactions to the nonbonded component of DeltaDeltaGcal is about the same as that from protein-water interactions. The residues that are located in a hydrophobic core (F3, L8, Y38, N39, T40, and I89) contribute significantly to the nonbonded free energy component of DeltaDeltaGcal. We also propose a general computational strategy for the study of protein stability that is equally conscious of the denatured and native states.
进行了自由能计算，以了解人溶菌酶的I56V突变对其热稳定性的影响。在变性状态的模拟中，使用了一个在中间包含突变位点的短肽。为了研究稳定性对变性态结构的依赖性，研究了五种不同的初始构象，即天然样、伸展型和三种随机卷曲样构象。我们发现，计算得到的自由能差ΔΔGcal，显著取决于变性态的结构。当采用天然样结构时，ΔΔGcal与实验自由能差ΔΔGexp吻合良好，而在其他四种模型中，ΔΔGcal与ΔΔGexp差异很大。因此，强烈表明突变位点周围的结构采取天然样构象而非伸展或随机卷曲构象。通过自由能成分分析表明，源自 Lennard-Jones 和共价相互作用的自由能成分主要决定了ΔΔGcal。蛋白质-蛋白质相互作用对ΔΔGcal非键合成分的贡献与蛋白质-水相互作用的贡献大致相同。位于疏水核心中的残基（F3、L8、Y38、N39、T40和I89）对ΔΔGcal的非键合自由能成分有显著贡献。我们还提出了一种研究蛋白质稳定性的通用计算策略，该策略对变性态和天然态同样重视。
