Solvation of the folding-transition state in Pseudomonas aeruginosa azurin is modulated by metal: Solvation of azurin's folding nucleus.
Solvation of the folding-transition state in Pseudomonas aeruginosa azurin is modulated by metal: Solvation of azurin's folding nucleus.
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铜绿假单胞菌天青蛋白折叠过渡态的溶剂化受金属调节：天青蛋白折叠核的溶剂化。
DOI：
10.1110/ps.051838206
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发表时间：
2006
期刊：
影响因子：
--
通讯作者：
Wittung-Stafshede,Pernilla
中科院分区：
文献类型：
--
作者：
Wilson,CoreyJ;Apiyo,David;Wittung-Stafshede,Pernilla
水在蛋白质折叠中的作用，特别是它在过渡态结构中的存在与否，是一个悬而未决的问题。有两类常见的折叠过渡态：扩散过渡态，其中几乎所有的侧链都具有类似的较低的Phi(φ)值；以及极化过渡态，其显示出不同的子结构，具有非常高的φ值。脱氧型和锌型铜绿假单胞菌Apo型和锌型铜绿假单胞菌都在两态平衡和动力学反应中折叠；脱氧型铜绿假单胞菌呈现极化过渡态，而锌型铜绿假单胞菌需要扩散的、移动的过渡态。为了研究这两种折叠过渡态中水的存在，我们探讨了在天青的六个位置用等位苏氨酸取代核心缬氨酸的平衡和动力学结果。与常规的疏水到丙氨酸的φ值分析相反，缬氨酸到苏氨酸的突变不会破坏核心填充，而是稳定了未折叠状态，并可用于评估与常规φ值结合时折叠过渡态的溶剂化程度。我们发现，脱氧天青素的折叠过渡态看起来完全干燥，而锌天青素的过渡态包括与水分子接触的部分形成的相互作用。根据自由能垒的形状，可以合理地解释远地折叠核和全息折叠核之间的这种明显差异。
The role of water in protein folding, specifically its presence or not in the transition‐state structure, is an unsolved question. There are two common classes of folding‐transition states: diffuse transition states, in which almost all side chains have similar, rather low phi (φ) values, and polarized transition states, which instead display distinct substructures with very high φ‐values. Apo‐and zinc‐forms ofPseudomonas aeruginosaazurin both fold in two‐state equilibrium and kinetic reactions; while the apo‐form exhibits a polarized transition state, the zinc form entails a diffuse, moving transition state. To examine the presence of water in these two types of folding‐transition states, we probed the equilibrium and kinetic consequences of replacing core valines with isosteric threonines at six positions in azurin. In contrast to regular hydrophobic‐to‐alanine φ‐value analysis, valine‐to‐threonine mutations do not disrupt the core packing but stabilize the unfolded state and can be used to assess the degree of solvation in the folding‐transition state upon combination with regular φ‐values. We find that the transition state for folding of apo‐azurin appears completely dry, while that for zinc‐azurin involves partially formed interactions that engage water molecules. This distinct difference between the apo‐and holo‐folding nuclei can be rationalized in terms of the shape of the free‐energy barrier.