Department of Chemistry, Virginia Tech, Blacksburg, Virginia 24060, United States.
Macromolecules Innovation Institute (MII),Virginia Tech, Blacksburg, Virginia 24060, United States.
J Phys Chem B. 2023 Nov 23;127(46):9936-9942. doi: 10.1021/acs.jpcb.3c05749. Epub 2023 Nov 14.
The role cysteine residues play in proteins is mediated by their protonation state, whereby the thiolate form of the side chain is highly reactive while the thiol form is more inert. However, the p of cysteine residues is hard to predict as it can differ widely from its reference value in solution, an effect that is accentuated by local effects in the heterogeneous protein environment. Here, we present a new approach to the prediction of cysteine reactivity based on electric field calculations at the thiol/thiolate group. We validated our approach by predicting the protonation state of cysteine residues in different protein environments (in the active site, at the protein surface, and buried within the protein interior), including Cys-25 in papaya protease omega, which was proven problematic for the more traditional constant pH molecular dynamics (MD) technique. We predict p shifts consistent with experimental observations, and the decomposition of the electric fields into contributions from molecular fragments provides a direct handle to rationalize local pH and p effects in proteins without introducing parameters other than those of the force field used for MD simulations.
半胱氨酸残基在蛋白质中的作用是通过其质子化状态介导的，其中侧链的硫醇盐形式具有高反应性，而硫醇形式则更惰性。然而，半胱氨酸残基的 p 值很难预测，因为它在溶液中的参考值可能有很大差异，这种效应在异质蛋白质环境中的局部效应中更加突出。在这里，我们提出了一种基于巯基/硫醇基团电场计算的半胱氨酸反应性预测新方法。我们通过预测不同蛋白质环境中（活性部位、蛋白质表面和埋藏在蛋白质内部）半胱氨酸残基的质子化状态来验证我们的方法，包括木瓜蛋白酶ω中的 Cys-25，这对半胱氨酸残基的质子化状态预测对于更传统的恒定 pH 分子动力学（MD）技术来说是一个难题。我们预测的 p 值位移与实验观察结果一致，并且将电场分解为分子片段的贡献为在不引入除用于 MD 模拟的力场参数之外的参数的情况下，提供了一种直接的方法来合理化蛋白质中的局部 pH 和 p 值效应。
