Silow M, Oliveberg M
Biochemistry, Chemical Centre, Lund, Sweden.
Biochemistry. 1997 Jun 24;36(25):7633-7. doi: 10.1021/bi970210x.
Recent controversy about the role of populated intermediates in protein folding emphasizes the need to better characterize other events on the folding pathway. A complication is that these involve high-energy states which are difficult to target experimentally since they do not accumulate kinetically. Here, we explore the energetics of high-energy states and map out the shape of the free-energy profile for folding of the two-state protein U1A. The analysis is based on nonlinearities in the GdnHCl dependence of the activation energy for unfolding, which we interpret in terms of structural changes of the protein-folding transition state. The result suggests that U1A folds by high-energy channeling where most of the conformational search takes place isoenergetically at transition-state level. This is manifested in a very broad and flat activation barrier, the top of which covers more than 60% of the reaction coordinate. The interpretation favors a folding mechanism where the pathway leading to the native protein is determined by the sequence's ability to stabilize productive transition states.
近期关于折叠中间体在蛋白质折叠过程中作用的争议凸显了更好地表征折叠途径上其他事件的必要性。一个复杂之处在于，这些事件涉及高能态，由于它们在动力学上不会积累，因此很难通过实验进行靶向研究。在此，我们探究了高能态的能量学，并绘制了两态蛋白U1A折叠的自由能分布图。该分析基于去折叠活化能对盐酸胍（GdnHCl）依赖性的非线性关系，我们根据蛋白质折叠过渡态的结构变化对其进行解释。结果表明，U1A通过高能通道进行折叠，其中大部分构象搜索在过渡态水平上等能进行。这表现为一个非常宽且平坦的活化能垒，其顶部覆盖了超过60%的反应坐标。这种解释支持一种折叠机制，即通往天然蛋白质的途径由序列稳定有效过渡态的能力决定。
