Urea denaturation of barnase: pH dependence and characterization of the unfolded state.
Urea denaturation of barnase: pH dependence and characterization of the unfolded state.
复制标题
芽孢杆菌RNA酶的尿素变性：pH依赖性和展开状态的表征。
DOI：
10.1021/bi00125a013
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发表时间：
1992
期刊：
影响因子：
2.9
通讯作者：
Erickson,RE
中科院分区：
文献类型：
--
作者：
Pace,CN;Laurents,DV;Erickson,RE
摘要：为了研究藤壶酶构象稳定性与pH的依赖关系，测定了pH为2-10范围内的尿素变性曲线。蔗糖酶的最大构象稳定性为9kcal mol~(-1)，在pH为5~6之间。AG对尿素浓度的依赖性从高pH时的1850卡·mol~(-1)·M~(-1)增加到pH为3附近的约3000卡·mol~(-1)·M~(-1)。这表明，随着分子上净电荷的增加，其未折叠的构象更容易与尿素结合。以前的研究表明，在8M的尿素中，即使二硫键断裂，尿素的棒状酶也比核糖核酸酶TL更完整地展开[Pace，C.N.，Laurents，DV，&Thomson，J.A.(1990)BioChemical 29,2564-2572]。溶剂扰动差示光谱表明，在8M尿素中，棒状酶中的Trp和Tyr残基比核糖核酸酶TL中的Trp和Tyr残基更容易被二甲基亚砜扰动，二硫键断裂。
Revised Manuscript Received December 23, 1991 abstract: To investigate the pH dependence of the conformationalstability of barnase, urea denaturation curves were determined over the pH range 2-10. The maximum conformational stability of barnase is 9 kcal mol'1 and occurs between pH 5 and 6. The dependence of AG on urea concentration increases from 1850 cal mol'1 M'1 at high pH to about 3000 cal mol'1 M'1 near pH 3. This suggests that the unfolded conformations of barnase become more accessible to urea as the net charge on the molecule increases. Previous studies suggested that in 8 M urea barnase unfolds more completely than ribonuclease Tl, even with the disulfide bonds broken [Pace, C. N., Laurents, DV, & Thomson, J. A.(1990) Biochemistry 29, 2564-2572]. In support of this, solvent perturbation difference spectroscopy showed that in 8 M urea the Trp and Tyr residues in barnase are more accessible to perturbation by dimethyl sulfoxide than in ribonuclease Tl with the disulfide bonds broken.