Structural studies of a folding intermediate of bovine pancreatic ribonuclease A by continuous recycled flow.
Structural studies of a folding intermediate of bovine pancreatic ribonuclease A by continuous recycled flow.
复制标题
通过连续循环流对牛胰腺核糖核酸酶 A 的折叠中间体进行结构研究。
DOI：
10.1021/bi00407a033
复制
发表时间：
1988
期刊：
影响因子：
2.9
通讯作者：
Scheraga,HA
中科院分区：
文献类型：
--
作者：
Adler,M;Scheraga,HA
摘要：连续循环流（CRF）光谱技术被用于观察半衰期大于等于10 s的热诱导可逆反应的中间体。该结构可以用任何不干扰系统的光谱方法来探测。核糖核酸酶A延长8小时的信号采集是可能的。CRF用于研究化学完整核糖核酸酶A （RNase A）在酸性条件下热展开/折叠过程中缓慢折叠中间体的结构。利用质子核磁共振和远紫外圆二色光谱对一种折叠中间体进行了分析，得出以下结论：(a)所检测到的折叠中间体的构象与热变性蛋白的构象相似。只有有限的新结构形成。(B) n端螺旋在这些条件下是部分稳定的，并且与变性构象处于快速平衡（< 10 ms）。(C) n端螺旋的疏水残基与蛋白质的其余部分之间存在远程相互作用。这些相互作用远高于熔点。(D)一个脂肪族甲基报告在n端区域外形成了一个新结构。(E)在化学修饰的、非折叠形式的rna酶A中检测到的结构也存在于折叠中间体中。然而，对于化学完整的RNase A来说，还有一些额外的相互作用是独一无二的。
Revised Manuscript Received November 5, 1987 abstract: A new technique, continuous recycled flow (CRF) spectroscopy, has been developed for observing intermediates of any thermally induced, reversible reaction with a half-life of 10 s or longer. The structure can be probedby any spectroscopic method which does not perturb the system. Prolonged signal acquisitions of 8 h for ribonuclease A are possible. CRF was used toinvestigate the structure of the slow-folding intermediates of chemically intact ribonuclease A (RNase A) during thermal unfolding/folding under acidic conditions. The following conclusions were reached on the basis of the proton nuclear magnetic resonance and far-ultraviolet circular dichroism spectra of a folding intermediate (s):(A) The conformation of the detected folding intermediate (s) is similar to that of the heat-denatured protein. There is only limited formation of new structures.(B) The N-terminal-helix is partially stable under these conditions and is in rapid (< 10 ms) equilibrium with the denatured conformation.(C) There are long-range interactions between the hydrophobic residues of the N-terminal-helix and the rest of the protein. These interactions persist well above the melting point.(D) An aliphatic methyl group reports on the formation of a new structure (s) that lie (s) outside of the N-terminal region.(E) Thestructures detected in chemically modified, nonfolding forms of the RNase A are also present in the folding intermediate (s). There are, however, additional interactions that are unique to chemically intact RNase A.