Institute of Advanced Energy, Kyoto University, Uji, Kyoto 611-0011, Japan.
J Chem Phys. 2013 Jun 28;138(24):245101. doi: 10.1063/1.4811287.
It is experimentally known that the heat-denaturation temperature of a protein is raised (i.e., its thermal stability is enhanced) by sugar addition. In earlier works, we proposed a physical picture of thermal denaturation of proteins in which the measure of the thermal stability is defined as the solvent-entropy gain upon protein folding at 298 K normalized by the number of residues. A multipolar-model water was adopted as the solvent. The polyatomic structures of the folded and unfolded states of a protein were taken into account in the atomic detail. A larger value of the measure implies higher thermal stability. First, we show that the measure remains effective even when the model water is replaced by the hard-sphere solvent whose number density and molecular diameter are set at those of real water. The physical picture is then adapted to the elucidation of the effects of sugar addition on the thermal stability of a protein. The water-sugar solution is modeled as a binary mixture of hard spheres. The thermal stability is determined by a complex interplay of the diameter of sugar molecules dC and the total packing fraction of the solution η: dC is estimated from the volume per molecule in the sugar crystal and η is calculated using the experimental data of the solution density. We find that the protein is more stabilized as the sucrose or glucose concentration becomes higher and the stabilization effect is stronger for sucrose than for glucose. These results are in accord with the experimental observations. Using a radial-symmetric integral equation theory and the morphometric approach, we decompose the change in the measure upon sugar addition into two components originating from the protein-solvent pair and protein-solvent many-body correlations, respectively. Each component is further decomposed into the excluded-volume and solvent-accessible-surface terms. These decompositions give physical insights into the microscopic origin of the thermal-stability enhancement by sugar addition. As an example, the higher stability of the native state relative to that of the unfolded state is found to be attributable primarily to an increase in the solvent crowding caused by sugar addition. Due to the hydrophilicity of sugar molecules, the addition of sugar by a larger amount or that with a larger molecular size leads to an increase in η which is large enough to make the solvent crowding more serious.
实验表明，添加糖会提高蛋白质的热变性温度（即增加其热稳定性）。在早期的工作中，我们提出了蛋白质热变性的物理图像，其中热稳定性的度量定义为在 298 K 下蛋白质折叠时溶剂熵的增加，除以残基数。采用多极模型水作为溶剂。在原子细节上考虑了蛋白质折叠和展开状态的多原子结构。该度量的较大值意味着较高的热稳定性。首先，我们表明，即使模型水被其密度和分子直径设定为真实水的硬球溶剂取代，该度量仍然有效。然后，我们将该物理图像应用于阐明糖添加对蛋白质热稳定性的影响。将糖水溶液建模为硬球的二元混合物。热稳定性由糖分子直径 dC 和溶液总堆积分数 η 的复杂相互作用决定：dC 由糖晶体中每个分子的体积估算，η 使用溶液密度的实验数据计算。我们发现，随着蔗糖或葡萄糖浓度的增加，蛋白质的稳定性越高，且蔗糖的稳定效果强于葡萄糖。这些结果与实验观察结果一致。使用径向对称积分方程理论和形态计量学方法，我们将糖添加引起的度量变化分解为分别源自蛋白质-溶剂对和蛋白质-溶剂多体相关的两个分量。每个分量进一步分解为排斥体积和溶剂可及表面积项。这些分解为糖添加增强热稳定性的微观起源提供了物理见解。例如，与展开状态相比，天然状态的更高稳定性主要归因于糖添加引起的溶剂拥挤增加。由于糖分子的亲水性，添加更多量或更大分子尺寸的糖会导致 η 的增加足够大，从而使溶剂拥挤更加严重。
