Carra J H, Privalov P L
Department of Biology and Biocalorimetry Center, Johns Hopkins University, Baltimore, Maryland 21218, USA.
FASEB J. 1996 Jan;10(1):67-74. doi: 10.1096/fasebj.10.1.8566550.
A valuable approach to understanding the forces that maintain protein structure is to analyze the thermodynamic effects of mutations on protein folding. The folding process is most often described using an energetic model that assumes a two-state transition between the native and denatured states. However, some results obtained using this approach for mutants of the protein staphylococcal nuclease have contradicted expectations from our current understanding of protein energetics. The application of differential scanning calorimetry to a set of mutant nuclease proteins allowed us to measure directly the effects of mutations on the enthalpy and heat capacity changes of unfolding, as well as on the cooperativity. We found that most of these effects can be understood with a three-state model of folding including a distinct intermediate, but not with the two-state model. Use of a three-state instead of a two-state model leads to large differences in conclusions about the stability effects of some mutations, suggesting that reevaluation of the effects of mutations on this and other proteins may be necessary to achieve an accurate description of folding energetics. The two-state assumption commonly used in protein stability studies may be an oversimplification in many cases.
一种理解维持蛋白质结构作用力的有效方法是分析突变对蛋白质折叠的热力学效应。折叠过程通常使用一种能量模型来描述，该模型假定天然态和变性态之间存在两态转变。然而，用这种方法对葡萄球菌核酸酶突变体进行研究所得的一些结果与我们目前对蛋白质能量学的理解相悖。将差示扫描量热法应用于一组突变核酸酶蛋白，使我们能够直接测量突变对解折叠的焓变和热容变化以及协同性的影响。我们发现，这些效应中的大多数可以用包含一个独特中间体的三态折叠模型来解释，但不能用两态模型来解释。使用三态模型而非两态模型会导致关于某些突变稳定性效应的结论有很大差异，这表明可能有必要重新评估突变对该蛋白及其他蛋白的影响，以准确描述折叠能量学。蛋白质稳定性研究中常用的两态假设在许多情况下可能过于简单化了。
