Labhardt A M
Proc Natl Acad Sci U S A. 1984 Dec;81(24):7674-8. doi: 10.1073/pnas.81.24.7674.
It is shown that circular dichroism (CD) can distinguish between the S-peptide and the S-protein fragments of RNase S at 225 nm and 235 nm. The conformational source for the strong CD at 225 nm is the S-peptide alpha-helix. The structural assignment of the CD at 235 nm is not clear but it is shown to be largely due to the S-protein moiety. This situation is utilized to monitor the kinetics of pH-induced unfolding and refolding of the two moieties. It is observed that major changes occur both in the fast and slow phases of unfolding as well as refolding. Specifically, the S-peptide alpha-helix unzippering is a fast reaction, followed by slow kinetics only at 235 nm. These latter kinetics parallel the appearance of the slow-folding species commonly attributed to the accumulation of non-native proline isomers. In refolding, a large fraction of the CD of S-protein at 235 nm recovers rapidly. The S-peptide alpha-helix zippers up last. These results are unexpected and their implications for the folding mechanism of ribonuclease are discussed.
结果表明，圆二色性（CD）在225nm和235nm处能够区分核糖核酸酶S的S肽和S蛋白片段。225nm处强CD信号的构象来源是S肽的α-螺旋。235nm处CD信号的结构归属尚不清楚，但已表明其主要归因于S蛋白部分。利用这种情况来监测pH诱导的两个部分展开和重折叠的动力学。观察到在展开和重折叠的快速和慢速阶段均发生了主要变化。具体而言，S肽α-螺旋的解链是一个快速反应，仅在235nm处随后出现慢速动力学。这些后者的动力学与通常归因于非天然脯氨酸异构体积累的慢折叠物种的出现平行。在重折叠过程中，235nm处S蛋白的大部分CD信号迅速恢复。S肽α-螺旋最后形成拉链结构。这些结果出乎意料，并讨论了它们对核糖核酸酶折叠机制的影响。
