Gast K, Zirwer D, Müller-Frohne M, Damaschun G
Max-Delbrück-Center for Molecular Medicine, Berlin, Germany.
Protein Sci. 1999 Mar;8(3):625-34. doi: 10.1110/ps.8.3.625.
The trifluoroethanol (TFE)-induced structural changes of two proteins widely used in folding experiments, bovine alpha-lactalbumin, and bovine pancreatic ribonuclease A, have been investigated. The experiments were performed using circular dichroism spectroscopy in the far- and near-UV region to monitor changes in the secondary and tertiary structures, respectively, and dynamic light scattering to measure the hydrodynamic dimensions and the intermolecular interactions of the proteins in different conformational states. Both proteins behave rather differently under the influence of TFE: alpha-lactalbumin exhibits a molten globule state at low TFE concentrations before it reaches the so-called TFE state, whereas ribonuclease A is directly transformed into the TFE state at TFE concentrations above 40% (v/v). The properties of the TFE-induced states are compared with those of equilibrium and kinetic intermediate states known from previous work to rationalize the use of TFE in yielding information about the folding of proteins. Additionally, we report on the properties of TFE/water and TFE/buffer mixtures derived from dynamic light scattering investigations under conditions used in our experiments.
研究了三氟乙醇（TFE）诱导的两种在折叠实验中广泛使用的蛋白质——牛α-乳白蛋白和牛胰核糖核酸酶A的结构变化。实验通过在远紫外和近紫外区域使用圆二色光谱分别监测二级和三级结构的变化，并通过动态光散射测量不同构象状态下蛋白质的流体动力学尺寸和分子间相互作用。在TFE的影响下，这两种蛋白质的行为有很大不同：α-乳白蛋白在达到所谓的TFE状态之前，在低TFE浓度下呈现熔融球状状态，而核糖核酸酶A在TFE浓度高于40%（v/v）时直接转变为TFE状态。将TFE诱导状态的性质与先前工作中已知的平衡态和动力学中间态的性质进行比较，以合理化TFE在获取蛋白质折叠信息方面的应用。此外，我们报告了在我们实验所用条件下，通过动态光散射研究得出的TFE/水和TFE/缓冲液混合物的性质。
