Slow refolding kinetics in yeast iso-2 cytochrome c.
Slow refolding kinetics in yeast iso-2 cytochrome c.
复制标题
酵母 iso-2 细胞色素 c 中的缓慢重折叠动力学。
DOI：
10.1021/bi00348a024
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发表时间：
1985
期刊：
影响因子：
2.9
通讯作者：
Nall,BT
中科院分区：
文献类型：
--
作者：
OsterhoutJr,JJ;Nall,BT
摘要：在酿酒酵母（Saccharomyces cerevisiae）的iso-2细胞色素c的再折叠过程中，存在rla和rlb两种缓慢折叠反应。慢反应中较慢的rla=100-200 s，仅通过吸光度变化观察到，而rlb（10-20倍快）通过荧光变化检测到。测量了这些反应速率的温度依赖性：在折叠-展开过渡区（pH 7.2, 0.3 M盐酸胍，5-30℃）以下结束的动力学实验中，rla的活化焓为AH*= 27 keal/mol， rlb的活化焓为21 keal/mol。双跳（展开，然后再折叠）实验表明，在展开条件下（3 M盐酸胍，pH 7.2, 20℃），两组负责缓慢折叠反应的物质以不同的速率缓慢生成。最后，作为对不同展开条件下慢再折叠物种种群变化的测试，慢再折叠的振幅作为初始展开条件的函数进行了测量，最终再折叠条件保持不变。在没有其他反应干扰的可测量范围内，荧光检测（alb）和吸光度检测（ala）缓慢折叠的振幅与初始条件下盐酸胍的浓度和pH无关。虽然完整的描述需要一个更复杂的解释，但许多慢折叠物种的性质是脯氨酸酰亚胺键异构化所期望的。
Revised Manuscript Received August 26, 1985 abstract: In refolding of iso-2 cytochrome c from Saccharomyces cerevisiae, there are two slow folding reactions, rla and rlb. The slower of the slow reactions, rla=100-200 s, is observed only by absorbance changes, while rlb (10-20-fold faster) is detected by fluorescence changes. The temperature dependence of the rates of these reactions has been measured: for kinetic experiments endingbelow the folding-unfolding transition zone (pH 7.2, 0.3 M guanidine hydrochloride, 5-30 C), the activation enthalpies are AH*= 27 keal/mol for rla and 21 keal/mol for rlb. Double-jump (unfolding, then refolding) experiments demonstrate that thetwo sets of species responsible for the slow folding reactions are generated slowly but at different rates under unfolding conditions (3 M guanidine hydrochloride, pH 7.2, 20 C). Finally, as a test for changes in the population of the slow refolding species under different unfolding conditions, the amplitudes for slow refolding have been measured as a function of the initial unfoldingconditions with the final refoldingconditions held constant. Over the range accessible to measurement in the absence of interference from other reactions, the amplitudes for fluorescence-detected (alb) and absorbance-detected (ala) slow folding are independent of guanidine hydrochloride concentration and pH in the initial conditions. Although a full description requires a more complex explanation, many of the properties of the slow folding species are those expected for proline imide bond isomerization.