Two-dimensional 1H NMR studies of cytochrome c: hydrogen exchange in the N-terminal helix.
Two-dimensional 1H NMR studies of cytochrome c: hydrogen exchange in the N-terminal helix.
复制标题
细胞色素 c 的二维 1H NMR 研究：N 末端螺旋中的氢交换。
DOI：
10.1021/bi00353a025
复制
发表时间：
1986
期刊：
影响因子：
2.9
通讯作者：
Englander,SW
中科院分区：
文献类型：
--
作者：
Wand,AJ;Roder,H;Englander,SW
摘要：利用二维核磁共振方法研究了马心脏细胞色素c n端螺旋段在还原和氧化两种形式下的氢交换行为。前6个残基的酰胺质子不成氢键，与溶剂质子交换迅速。大多数n端氢键基团- Lys-7到ph -10的酰胺NH表现出动态磨损行为的急剧交换率梯度，符合统计力学原理。这在氧化的和氧化的细胞色素c中都是一样的。在氧化的形式中，形成最后一个螺旋转的残基11-14都以相似的速率交换，比自由暴露的肽NH的速率慢大约一百万倍，即使一些在螺旋的水表面上，而另一些则完全被掩埋。这些和其他几种蛋白质的类似观察似乎记录了局部协同展开反应作为蛋白质H交换反应的决定因素。细胞色素c的n端片段对血红素氧化还原状态不敏感，如在晶体学模型中，除了最靠近血红素的残基（Cys-14和Ala-15），它们在还原形式下的交换速度要慢约15倍。细胞色素碳氢交换的结果可以进一步考虑天然形态和瞬时展开形态的构象以及它们在还原态和氧化态下的自由能关系。
Department of Biochemistry and Biophysics, University of Pennsylvania, Philadelphia, Pennsylvania 19104 Received June 11, 1985 abstract: The hydrogen exchange behavior of the N-terminal helical segment in horse heart cytochrome c was studied in both the reduced and the oxidized forms by use of two-dimensional nuclear magnetic resonance methods. The amide protons of the first six residues are not H bonded and exchange rapidly with solvent protons. The most N-terminal H-bonded groups—the amide NH of Lys-7 to Phe-10—exhibit a sharp gradient in exchange rate indicative of dynamic fraying behavior, consistent with statistical-mechanical principles. This occurs identically in bothreduced and oxidized cytochrome c. In the oxidized form, residues 11-14, which form the last helical turn, all exchange with a similar rate, about one million times slower than the rate characteristic of freely exposed peptide NH, even though some are on the aqueous face of the helix and others are fully buried. These and similar observations in several other proteins appear to document local cooperative unfolding reactions as determinants of protein H exchange reactions. The N-terminal segment of cytochrome c is insensitive to the heme redox state, as in the crystallographic model, except for residues closest to the heme (Cys-14 and Ala-15), which exchange about 15-fold more slowly in the reduced form. The cytochrome c H exchange results can be further considered in terms of the conformation of the native and the transiently unfolded formsand their free energy relationships in both the reduced and the oxidized states.