变性剂结合模型的新证据。
New evidence for the denaturant binding model.
作者信息
Wu J W, Wang Z X
机构信息
National Laboratory of Biomacromolecules, Institute of Biophysics, Academia Sinica, Beijing, People's Republic of China.
出版信息
Protein Sci. 1999 Oct;8(10):2090-7. doi: 10.1110/ps.8.10.2090.
Denaturation with guanidine hydrochloride (GdnHCl) or urea is one of the primary ways of measuring the conformational stability of proteins and comparing the stability of mutant proteins. Despite the widespread use of these two denaturants to provide quantitative data for the free energies of unfolding, the mode of action of these agents is not well understood. We are not even certain whether the action of these agents on proteins is direct and can be regarded as ligand binding, or indirect and involves a change in the properties of solvent (water) in the presence of GdnHCl and urea. In this paper, an extensive kinetic study of the inhibition of ribonuclease A and papain by urea has been performed. The results suggest that the effect of urea on activities of these enzymes can be well described by the denaturant binding model. The binding constants of urea determined by the present method are nearly identical to that determined from a variety of different studies on model compounds and proteins.
用盐酸胍（GdnHCl）或尿素进行变性是测量蛋白质构象稳定性以及比较突变蛋白稳定性的主要方法之一。尽管广泛使用这两种变性剂来提供蛋白质解折叠自由能的定量数据，但这些试剂的作用方式尚未得到很好的理解。我们甚至不确定这些试剂对蛋白质的作用是直接的，可被视为配体结合，还是间接的，涉及在GdnHCl和尿素存在下溶剂（水）性质的变化。本文对尿素抑制核糖核酸酶A和木瓜蛋白酶进行了广泛的动力学研究。结果表明，变性剂结合模型可以很好地描述尿素对这些酶活性的影响。通过本方法测定的尿素结合常数与从对模型化合物和蛋白质的各种不同研究中测定的结果几乎相同。
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