PH-DEPENDENCE OF THE UREA AND GUANIDINE-HYDROCHLORIDE DENATURATION OF RIBONUCLEASE-A AND RIBONUCLEASE-T1
PH-DEPENDENCE OF THE UREA AND GUANIDINE-HYDROCHLORIDE DENATURATION OF RIBONUCLEASE-A AND RIBONUCLEASE-T1
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DOI：
10.1021/bi00462a019
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发表时间：
1990-03-13
期刊：
影响因子：
2.9
通讯作者：
THOMSON, JA
中科院分区：
文献类型：
--
作者：
PACE, CN;LAURENTS, DV;THOMSON, JA
为了研究核糖核酸酶A和T1的构象稳定性与pH的依赖关系，测定了在pH 2-10范围内尿素和盐酸胍的变性曲线。核糖核酸酶T1的最大构象稳定性在pH 4.5附近，核糖核酸酶A的最大构象稳定性在pH 7~9之间，表明带电基团之间的静电相互作用对蛋白质构象稳定性的贡献相对较小。核糖核酸酶A对尿素浓度的依赖性从高pH时的约1200卡·摩尔~(-1)·M~(-1)增加到低pH时的约2400卡·摩尔~(-1)·M~(-1)。这表明，随着分子上净电荷的增加，核糖核酸酶A的未折叠构象更容易与尿素结合。对于核糖核酸酶T1，在pH 6附近，ΔG对尿素浓度的依赖性最小，在较高和较低pH下都会增加。用Tanford发展的模型分析几种蛋白质的这一类型的信息化学。SoC。86,2050-2059]表明蛋白质在尿素和GdnHCl溶液中的未折叠状态可能在它们与变性剂相互作用的程度上有很大的不同。因此，未折叠蛋白质的构象可能至少在一定程度上取决于蛋白质的氨基酸序列。
To investigate the pH dependence of the conformational stability of ribonucleases A and T1, urea and guanidine hydrochloride denaturation curves have been determined over the pH range 2-10. The maximum conformational stability of both proteins is about 9 kcal/mol and occurs near pH 4.5 for ribonuclease T1 and between pH 7 and 9 for ribonuclease A. The pH dependence suggests that electrostatic interactions among the charged groups make a relatively small contribution to the conformational stability of these proteins. The dependence of .DELTA.G on urea concentration increases from about 1200 cal mol-1 M-1 at high pH to about 2400 cal mol-1 M-1 at low pH for ribonuclease A. This suggests that the unfolded conformations of RNase A become more accessible to urea as the net charge on the molecule increases. For RNase T1, the dependence of .DELTA.G on urea concentration is minimal near pH 6 and increases at both higher and lower pH. An analysis of information of this type for several proteins in terms of a model developed by Tanford [Tanford, C. (1964) J. Am. Chem. Soc. 86, 2050-2059] suggests that the unfolded states of proteins in urea and GdnHC1 solutions may differ significantly in the extent of their interaction with denaturants. Thus, the conformations assumed by unfolded proteins may depend to at least some extent on the amino acid sequence of the protein.