FREE-ENERGY CALCULATIONS ON PROTEIN STABILITY - THR-157-] VAL-157 MUTATION OF T4 LYSOZYME
FREE-ENERGY CALCULATIONS ON PROTEIN STABILITY - THR-157-] VAL-157 MUTATION OF T4 LYSOZYME
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DOI：
10.1021/ja00204a027
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发表时间：
1989-10-25
影响因子：
15
通讯作者：
KOLLMAN, PA
中科院分区：
文献类型：
--
作者：
DANG, LX;MERZ, KM;KOLLMAN, PA
我们对天然T4溶菌酶、T4溶菌酶的Thr-157-Val-157突变体以及代表天然和突变酶变性状态的四肽模型进行了自由能微扰计算。这是在可以与实验进行精确比较的情况下，第一次对蛋白质稳定性进行自由能微扰计算。天然模型和变性模型对天然模型和Thr-Val突变体的相对计算自由能差为1.9±1.1千卡/摩尔。这一值与1.6kcal/mol的实验值符合得很好，支持我们的方法。与预期相反，范德瓦尔斯而不是计算自由能的静电分量是这种微分稳定性的主要决定因素。突变后模拟出来的结构与X射线结构有重要的不同。这就强调了用不完整的蛋白质再现X射线结构的困难，以及有限的模拟时间(这里是40ps)。尽管如此，模拟结构中的氢键模式是合理的。T4溶菌酶已经成为研究蛋白质稳定性与蛋白质序列和三维结构关系的范例。1蛋白质和许多突变体的X射线结构和变性热力学都是可用的。2由此提出的问题是：理论分子动力学/自由能微扰方法能否模拟蛋白质稳定性的相对自由能以及天然和突变酶的X射线结构的差异？这些自由能方法在研究溶剂化自由能、3·4对蛋白质-配基结合药物结构的影响、5以及定点突变对酶配基结合和催化的影响方面是非常有用的。6·7
We present free energy perturbation calculations on the X-ray structure of native T4 lysozyme, the Thr-157—Val-157 mutant of T4 lysozyme, and on a tetrapeptide model that represents the denaturedstate of the native and mutant enzymes. This is the first free energy perturbation calculation on protein stability in a case where a precise comparison with experiment can be made. The relative calculated free energy difference between native and denatured models for nativeand the Thr— Val mutant was 1.9±1.1 kcal/mol. Thisvalue is in good agreement with the experimental value of 1.6 kcal/mol, supporting our approach. Contrary to expectation, the van der Waals rather than the electrostatic component of the computed free energy is the dominant determinant of this differential stability. Thestructures that emerge from the simulations after mutation differ in important ways from the X-ray structures. This emphasizes the difficulties in reproducing X-ray structure with incomplete protein representation as well as limited simulation times (40 ps here). Nonetheless, the hydrogen-bonding patterns in the simulated structures are physically reasonable.T4 lysozyme has become the paradigm for the study of the dependence of protein stability on protein sequence and threedimensional structure. 1 Both the X-ray structures and the thermodynamics of denaturation of the protein and many mutants are available. 2 Thus the question can be posed: Can theoretical molecular dynamics/free energy perturbation methods simulate the relative free energies of protein stability and the differences in X-ray structures of native and mutant enzymes? These free energy methods have been shown to be very useful in studies of solvation free energies, 3· 4 effect on drug structure of protein-ligand binding, 5 and effect of site-specific mutations on enzyme ligand binding andcatalysis. 6· 7