Institute of Biostructure and Bioimaging, National Research Council, Via P. Castellino, 80131 Naples, Italy.
Department of Chemical Sciences, University of Naples Federico II, Via Cintia, 80126 Naples, Italy.
Int J Mol Sci. 2024 Aug 29;25(17):9360. doi: 10.3390/ijms25179360.
By performing differential scanning calorimetry(DSC) measurements on RNase A, we studied the stabilization provided by the addition of potassium aspartate(KAsp) or potassium glutamate (KGlu) and found that it leads to a significant increase in the denaturation temperature of the protein. The stabilization proves to be mainly entropic in origin. A counteraction of the stabilization provided by KAsp or KGlu is obtained by adding common denaturants such as urea, guanidinium chloride, or guanidinium thiocyanate. A rationalization of the experimental data is devised on the basis of a theoretical approach developed by one of the authors. The main contribution to the conformational stability of globular proteins comes from the gain in translational entropy of water and co-solute ions and/or molecules for the decrease in solvent-excluded volume associated with polypeptide folding (i.e., there is a large decrease in solvent-accessible surface area). The magnitude of this entropic contribution increases with the number density and volume packing density of the solution. The two destabilizing contributions come from the conformational entropy of the chain, which should not depend significantly on the presence of co-solutes, and from the direct energetic interactions between co-solutes and the protein surface in both the native and denatured states. It is the magnitude of the latter that discriminates between stabilizing and destabilizing agents.
通过对 RNase A 进行差示扫描量热法 (DSC) 测量，我们研究了添加天门冬氨酸钾 (KAsp) 或谷氨酸钾 (KGlu) 提供的稳定性，并发现这导致蛋白质变性温度显著升高。这种稳定性主要来源于熵的增加。通过添加常见的变性剂，如尿素、盐酸胍或硫氰酸胍，可以抵消 KAsp 或 KGlu 提供的稳定性。基于其中一位作者提出的理论方法，对实验数据进行了合理化解释。球状蛋白质构象稳定性的主要贡献来自于水和共溶剂离子和/或分子的平移熵的增加，以及与多肽折叠相关的溶剂排除体积的减少（即溶剂可及表面积有很大减少）。这种熵贡献的大小随着溶液的数密度和体积堆积密度的增加而增加。两个失稳贡献来自于链的构象熵，它不应显著依赖于共溶剂的存在，以及来自于共溶剂和蛋白质表面在天然和变性状态下的直接能量相互作用。正是后者的大小区分了稳定和不稳定的试剂。
