Urea-induced unfolding of the alpha subunit of tryptophan synthase: evidence for a multistate process.
Urea-induced unfolding of the alpha subunit of tryptophan synthase: evidence for a multistate process.
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DOI：
10.1021/bi00507a021
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发表时间：
1981-02
期刊：
影响因子：
2.9
通讯作者：
Matthews Cr;Crisanti Mm
中科院分区：
文献类型：
--
作者：
Matthews Cr;Crisanti Mm
利用光谱学和尿素梯度凝胶电泳技术对大肠杆菌色氨酸合成酶α亚基的展开进行了监测。三个独立的证据线支持该过程中存在一种或多种稳定中间体的结论：(i)从286 nm的差谱和222 nm的圆二色光谱得到的平衡展开跃迁的满意拟合需要一个除了天然和未展开形式之外还包括稳定中间体的模型。（ii）消光系数在286 nm处变化的动力学研究表明，虽然展开可以用单指数变化很好地描述，但再折叠动力学是复杂的。再折叠动力学对尿素初始浓度依赖性的性质支持了至少存在一种稳定中间体的结论。尿素梯度凝胶电泳对α亚基的实验结果表明，至少有一种或可能有两种稳定的中间体参与其中；中间体具有明显不同程度的致密性。可以建立一个与所有这些结果一致的α亚基折叠的动力学模型。
The urea-induced unfolding of the alpha subunit of tryptophan synthase from E. coli was monitored by optical spectroscopy and by urea-gradient gel electrophoresis. Three independent lines of evidence support the conclusion that one or more stable intermediates are present in this process: (i) Satisfactory fits of the equilibrium unfolding transitions obtained from difference spectroscopy at 286 nm and circular dichroism spectroscopy at 222 nm require a model which involves a stable intermediate in addition to the native and unfolded forms. (ii) Kinetic studies of the change in the extinction coefficient at 286 nm show that while the unfolding is well described by a single exponential change the refolding kinetics are complex. The nature of the dependence of the refolding kinetics on the initial concentration of urea supports the conclusion that at least one stable intermediate exists. (iii) The patterns obtained from urea-gradient gel electrophoresis experiments on the alpha subunit show that at least one and possibly two stable intermediates are involved; the intermediates have markedly different degrees of compactness. A kinetic model for the folding of the alpha subunit, consistent with all of these results, can be formulated.