Department of Chemical and Biomolecular Engineering, The Johns Hopkins University, Baltimore, Maryland.
Department of Chemical and Biomolecular Engineering, The Johns Hopkins University, Baltimore, Maryland; Program in Molecular Biophysics, The Johns Hopkins University, Baltimore, Maryland.
Biophys J. 2012 Aug 8;103(3):587-595. doi: 10.1016/j.bpj.2012.06.044.
We developed a Rosetta-based Monte Carlo method to calculate the pK(a) values of protein residues that commonly exhibit variable protonation states (Asp, Glu, Lys, His, and Tyr). We tested the technique by calculating pK(a) values for 264 residues from 34 proteins. The standard Rosetta score function, which is independent of any environmental conditions, failed to capture pK(a) shifts. After incorporating a Coulomb electrostatic potential and optimizing the solvation reference energies for pK(a) calculations, we employed a method that allowed side-chain flexibility and achieved a root mean-square deviation (RMSD) of 0.83 from experimental values (0.68 after discounting 11 predictions with an error over 2 pH units). Additional degrees of side-chain conformational freedom for the proximal residues facilitated the capture of charge-charge interactions in a few cases, resulting in an overall RMSD of 0.85 pH units. The addition of backbone flexibility increased the overall RMSD to 0.93 pH units but improved relative pK(a) predictions for proximal catalytic residues. The method also captures large pK(a) shifts of lysine and some glutamate point mutations in staphylococcal nuclease. Thus, a simple and fast method based on the Rosetta score function and limited conformational sampling produces pK(a) values that will be useful when rapid estimation is essential, such as in docking, design, and folding.
我们开发了一种基于 Rosetta 的蒙特卡罗方法，用于计算通常表现出可变质子化状态的蛋白质残基的 pK(a) 值（Asp、Glu、Lys、His 和 Tyr）。我们通过计算 34 种蛋白质中的 264 个残基的 pK(a) 值来测试该技术。标准的 Rosetta 评分函数，它不依赖于任何环境条件，无法捕捉到 pK(a) 偏移。在结合库仑静电势并优化 pK(a) 计算的溶剂参考能后，我们采用了一种允许侧链灵活性的方法，并实现了与实验值的均方根偏差 (RMSD) 为 0.83（扣除 11 个误差超过 2 pH 单位的预测值后为 0.68）。近端残基的侧链构象自由度的额外增加促进了电荷-电荷相互作用的捕捉，在少数情况下导致整体 RMSD 为 0.85 pH 单位。增加骨架灵活性会将整体 RMSD 提高到 0.93 pH 单位，但会改善近端催化残基的相对 pK(a) 预测。该方法还可以捕捉到枯草溶菌素核酶中赖氨酸和一些谷氨酸点突变的大 pK(a) 偏移。因此，基于 Rosetta 评分函数和有限构象采样的简单快速方法产生的 pK(a) 值在快速估计至关重要的情况下将非常有用，例如在对接、设计和折叠中。
