Compressing the free energy range of substructure stabilities in iso-1-cytochrome c
Compressing the free energy range of substructure stabilities in iso-1-cytochrome c
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DOI：
10.1002/pro.120
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发表时间：
2009-06-01
期刊：
影响因子：
8
通讯作者：
Bowler, Bruce E.
中科院分区：
文献类型：
--
作者：
Duncan, Michael G.;Williams, Michael D.;Bowler, Bruce E.
通过有限的蛋白水解、碱性构象转变和盐酸胍的整体解折叠，研究了酵母 iso-1-细胞色素 c 和充分表征的马细胞色素 c 之间亚结构的进化保守性。对 iso-1-细胞色素 c 的有限蛋白水解裂解产物的质谱分析表明，其最不稳定的亚结构与马细胞色素 c 相同。有限的蛋白水解数据产生 3.8 +/- 6 0.4 kcal mol(-1) 的自由能来展开最不稳定的子结构，而 iso-1-细胞色素 c 的整体展开需要 5.05 +/- 0.30 kcal mol(-1) 的自由能。因此，iso-1-细胞色素c 的亚结构稳定性跨度仅与1.2 kcal mol(-1) 相似，而马细胞色素c 的亚结构稳定性与8 kcal mol(-1) 相似。与马蛋白预期的较少合作折叠一致，马蛋白与酵母细胞色素 c 的胍-HCl m 值与 3 kcal mol(-1) M-1 相似，而与 4.5 kcal mol(-1) M-1 相似。酵母细胞色素 c 亚结构的紧密自由能间距表明其折叠比马细胞色素 c 具有更多的分支点。对具有 H26N 突变的 iso-1-细胞色素 c 变体的研究表明，与马细胞色素 c 一样，最不稳定和最稳定的亚结构依次展开，并且两个最不稳定的亚结构独立展开。因此，马细胞色素 c 亚结构的重要方面，尽管被大力压缩，但在酵母 iso-1-细胞色素 c 中仍然在进化上得以保留。
Evolutionary conservation of substructure architecture between yeast iso-1-cytochrome c and the well-characterized horse cytochrome c is studied with limited proteolysis, the alkaline conformational transition and global unfolding with guanidine-HCl. Mass spectral analysis of limited proteolysis cleavage products for iso-1-cytochrome c show that its least stable substructure is the same as horse cytochrome c. The limited proteolysis data yield a free energy of 3.8 +/- 6 0.4 kcal mol(-1) to unfold the least stable substructure compared with 5.05 +/- 0.30 kcal mol(-1) for global unfolding of iso-1-cytochrome c. Thus, substructure stabilities of iso-1-cytochrome c span only similar to 1.2 kcal mol(-1) compared with similar to 8 kcal mol(-1) for horse cytochrome c. Consistent with the less cooperative folding thus expected for the horse protein, the guanidine-HCl m-values are similar to 3 kcal mol(-1) M-1 versus similar to 4.5 kcal mol(-1) M-1 for horse versus yeast cytochrome c. The tight free energy spacing of the yeast cytochrome c substructures suggests that its folding has more branch points than for horse cytochrome c. Studies on a variant of iso-1-cytochrome c with an H26N mutation indicate that the least and most stable substructures unfold sequentially and the two least stable substructures unfold independently as for horse cytochrome c. Thus, important aspects of the substructure architecture of horse cytochrome c, albeit compressed energetically, are preserved evolutionally in yeast iso-1-cytochrome c.