Hansson T, Nordlund P, Aqvist J
Department of Molecular Biology, Uppsala University Biomedical Centre, Sweden.
J Mol Biol. 1997 Jan 17;265(2):118-27. doi: 10.1006/jmbi.1996.0716.
The nucleophilic attack by cysteine 12 in the low-molecular-weight protein tyrosine phosphatase is believed to be carried out by the thiolate anion form of this residue. We here study the energetics of proton transfer between the thiol group of cysteine 12 and a substrate phosphate oxygen atom, to examine the effects of the enzymic environment on the stability of the thiolate nucleophile. This is done by molecular dynamics and free energy perturbation simulations, utilizing the empirical valence bond method to describe the potential surface of the system. The calculations show that the protein environment significantly stabilizes the thiolate ion, thereby setting the stage for the nucleophilic attack. We compare these results with those from further simulations of a mutant enzyme, and demonstrate the importance of serine 19 in thiolate stabilization.
低分子量蛋白质酪氨酸磷酸酶中半胱氨酸12的亲核攻击被认为是由该残基的硫醇盐阴离子形式进行的。我们在此研究半胱氨酸12的巯基与底物磷酸氧原子之间质子转移的能量学，以检查酶环境对硫醇盐亲核试剂稳定性的影响。这是通过分子动力学和自由能微扰模拟来完成的，利用经验价键方法描述系统的势能面。计算表明，蛋白质环境显著稳定了硫醇盐离子，从而为亲核攻击奠定了基础。我们将这些结果与突变酶的进一步模拟结果进行比较，并证明丝氨酸19在硫醇盐稳定中的重要性。
