The thiol of human serum albumin: Acidity, microenvironment and mechanistic insights on its oxidation to sulfenic acid
The thiol of human serum albumin: Acidity, microenvironment and mechanistic insights on its oxidation to sulfenic acid
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DOI：
10.1016/j.freeradbiomed.2017.04.021
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发表时间：
2017-07-01
影响因子：
7.4
通讯作者：
Laura Coitino, E.
中科院分区：
文献类型：
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作者：
Bonanata, Jenner;Turell, Lucia;Laura Coitino, E.
人血清白蛋白（HSA）具有单一的半胱氨酸还原残基Cys34，其酸度一直存在争议。采用三种实验方法（对过氧化氢反应性的ph依赖性、紫外滴定法和红外光谱法）确定降解HSA在37℃和0.1 M离子强度下的K-P(a)值为8.1 +/- 0.2。亚微秒时间尺度的HSA分子动力学模拟表明，硫在溶剂中的暴露是有限的，硫醇形式的硫暴露是波动的，但硫酸盐中的硫暴露是增加的，硫酸盐中含有Tyr84和桥接水到Asp38和Gln33主链的持久氢键（HB）网络稳定了硫的暴露。通过包括量子水分子在内的ONIOM（QM:MM）模型，深入了解H2O2氧化Cys34的机理。反应通过稍微不同步的S(N)2过渡态（TS）进行，计算出298 K下三角*G和三角*H势垒分别为59和54 kJ mol(-1)（后者在实验值的化学精度范围内）。ts后质子转移导致HSA-SO-和水作为产物。结构反应位点束缚H2O2，为硫酸盐提供强HB。在到达TS之前失去HB会调节Cys34的亲核性，并导致H2O2的不稳定。TS的差异稳定所需的反应现场特征（正电荷，H2O2 HB强化）的缺乏解释了在其他蛋白质（如过氧化物还毒素）中相同反应的动力学效率的显著差异。围绕HSA-SH的结构化HB网络与隔离的水对势垒高度产生熵罚。这些研究有助于加深对人类血浆中最丰富的硫醇HSA-SH的反应性的理解，并从更广泛的角度提供了调节硫醇对H2O2反应性的关键方面的线索。
Human serum albumin (HSA) has a single reduced cysteine residue, Cys34, whose acidity has been controversial. Three experimental approaches (pH-dependence of reactivity towards hydrogen peroxide, ultraviolet titration and infrared spectroscopy) are used to determine that the K-P(a) value in delipidated HSA is 8.1 +/- 0.2 at 37 degrees C and 0.1 M ionic strength. Molecular dynamics simulations of HSA in the sub-microsecond timescale show that while sulfur exposure to solvent is limited and fluctuating in the thiol form, it increases in the thiolate, stabilized by a persistent hydrogen-bond (HB) network involving Tyr84 and bridging waters to Asp38 and Gln33 backbone. Insight into the mechanism of Cys34 oxidation by H2O2 is provided by ONIOM(QM:MM) modeling including quantum water molecules. The reaction proceeds through a slightly asynchronous S(N)2 transition state (TS) with calculated triangle*G and triangle*H barriers at 298 K of respectively 59 and 54 kJ mol(-1) (the latter within chemical accuracy from the experimental value). A post-TS proton transfer leads to HSA-SO- and water as products. The structured reaction site cages H2O2, which donates a strong HB to the thiolate. Loss of this HB before reaching the TS modulates Cys34 nucleophilicity and contributes to destabilize H2O2. The lack of reaction-site features required for differential stabilization of the TS (positive charges, H2O2 HB strengthening) explains the striking difference in kinetic efficiency for the same reaction in other proteins (e.g. peroxiredoxins). The structured HB network surrounding HSA-SH with sequestered waters carries an entropic penalty on the barrier height. These studies contribute to deepen the understanding of the reactivity of HSA-SH, the most abundant thiol in human plasma, and in a wider perspective, provide clues on the key aspects that modulate thiol reactivity against H2O2.