Jourdan M, Searle M S
School of Chemistry, University of Nottingham, University Park, Nottingham NG7 2RD, U.K.
Biochemistry. 2001 Aug 28;40(34):10317-25. doi: 10.1021/bi010767j.
The thermodynamics of the native<-->A state and native<-->unfolded transitions for ubiquitin have been characterized in detail using the denaturants methanol and guanidinium chloride (Gdn.HCl) both separately and in combination. Gdn.HCl destabilizes the partially folded alcohol-induced A state such that the effects of alcoholic solvents on the native<-->unfolded transition can be investigated directly via a two-state model. The combined denaturing effects of methanol and Gdn.HCl appear to conform to a simple additive model. We show that ubiquitin folds and unfolds cooperatively in all cases, forming the same "native" state; however, the thermodynamics of the N<-->U transition change dramatically between alcoholic and Gdn.HCl solutions, with folding in aqueous methanol associated with large negative enthalpy and entropy terms at 298 K with a gradual falloff in DeltaC(p) at higher methanol concentrations, as previously reported for the N<-->A transition (Woolfson, D. N., Cooper, A., Harding, M. M., Williams, D. H., and Evans, P. A. (1993) J. Mol. Biol. 229, 502-511.). Both the N<-->U and the N<-->A transitions are enthalpy driven to a similar extent. We conclude that under these conditions van der Waals interactions in the packing of the nonpolar protein core, which is common to both the N<-->U and the N<-->A transitions, appear to drive folding in the absence of entropic effects associated with release of ordered solvent (hydrophobic effect). Solvent transfer studies of hydrocarbons into alcoholic solvents, with and without Gdn.HCl, are consistent with a large enthalpic driving force for burial of a nonpolar surface, with a linear dependence of protein stability (DeltaG(N)(<-->)(U)) on cosolvent concentration reflected in a similar linear dependence of hydrocarbon solubility. The data demonstrate that the hydrophobic effect is not a prerequisite for specific stabilization of the native state or the A state and that van der Waals packing of the nonpolar core appears to be the dominant factor in stabilization of the native state.
已分别使用变性剂甲醇和氯化胍（Gdn.HCl）以及二者的组合，详细表征了泛素天然态与A态以及天然态与去折叠态转变的热力学性质。Gdn.HCl会使部分折叠的酒精诱导A态不稳定，从而可以通过双态模型直接研究醇类溶剂对天然态与去折叠态转变的影响。甲醇和Gdn.HCl的联合变性作用似乎符合简单的加和模型。我们表明，在所有情况下泛素都能协同折叠和去折叠，形成相同的“天然”态；然而，在醇类和Gdn.HCl溶液之间，N→U转变的热力学性质发生了显著变化，在298 K时，在甲醇水溶液中折叠伴随着较大的负焓和熵项，且在较高甲醇浓度下ΔC(p)逐渐下降，正如之前报道的N→A转变情况一样（伍尔夫森，D. N.，库珀，A.，哈丁，M. M.，威廉姆斯，D. H.，以及埃文斯，P. A.（1993年）《分子生物学杂志》229卷，502 - 511页）。N→U和N→A转变在焓驱动方面程度相似。我们得出结论，在这些条件下，非极性蛋白质核心堆积中的范德华相互作用，这在N→U和N→A转变中都存在，在没有与有序溶剂释放相关的熵效应（疏水效应）的情况下似乎驱动了折叠。在有和没有Gdn.HCl存在的情况下，将碳氢化合物转移到醇类溶剂中的溶剂转移研究表明，非极性表面埋藏存在很大的焓驱动力，蛋白质稳定性（ΔG(N)(<-->)(U)）对共溶剂浓度的线性依赖性反映在碳氢化合物溶解度的类似线性依赖性上。数据表明，疏水效应不是天然态或A态特异性稳定的先决条件，非极性核心的范德华堆积似乎是天然态稳定的主导因素。
