A method to determine residue-specific unfolded-state pKa values from analysis of stability changes in single mutant cycles.
A method to determine residue-specific unfolded-state pKa values from analysis of stability changes in single mutant cycles.
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一种通过分析单突变体循环稳定性变化来确定残基特异性未折叠状态 pKa 值的方法。
DOI：
10.1021/ja101761m
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发表时间：
2010
影响因子：
15
通讯作者：
Shen,JanaK
中科院分区：
文献类型：
--
作者：
Shen,JanaK
现在广泛认识到，在折叠条件下与天然状态平衡的蛋白质的未折叠状态可能含有显著的残余结构。然而，由于技术上的困难，在未折叠状态下的残基特异性相互作用仍然难以捉摸。在这里，我们介绍了一种方法来自怀曼-坦福德理论，以确定残基特定的pKa的展开状态。该方法需要平衡稳定性测量的野生型和单点突变体，其中可滴定的残基被替换为电荷中性的两个pH条件下。所提出的方法的应用揭示了在NTL 9蛋白的未折叠状态下Asp 8的高度抑制的pKa。展开状态的pKa的知识，使蛋白质稳定性的展开状态的静电效应的定量估计。它也提供了有价值的基准力场的改进和验证微观信息的分子动力学模拟。
It is now widely recognized that the unfolded state of a protein in equilibrium with the native state under folding conditions may contain significant residual structures. However, due to technical difficulties residue-specific interactions in the unfolded state remain elusive. Here we introduce a method derived from the Wyman−Tanford theory to determine residue-specific pKa’s in the unfolded state. This method requires equilibrium stability measurements of the wild type and single-point mutants in which titrable residues are replaced with charge-neutral ones under two pH conditions. Application of the proposed approach reveals a highly depressed pKafor Asp8 in the unfolded state of the NTL9 protein. Knowledge of unfolded-state pKa’s enables quantitative estimation of the unfolded-state electrostatic effects on protein stability. It also provides valuable benchmarks for the improvement of force fields and validation of microscopic information from molecular dynamics simulations.
蛋白质变性状态下的 pKA 值受到干扰。
影响因子：
5.6
作者：
Y. Tan;M. Oliveberg;B. Davis;A. Fersht
通讯作者：
A. Fersht
影响因子：
5.6
作者：
Cho, JH;Raleigh, DP
通讯作者：
Raleigh, DP