Interrupted unfolding of CMPK wt.

The mixing-scheme of the interrupted unfolding reaction is displayed in (a). After unfolding in 6.0 M urea with incubation time t1, the protein is diluted into 1.2 M urea. The subsequent refolding is recorded as function of refolding time t2 (b). For short incubation times (F2(IU) can be observed. For long incubation times, the fast and slow refolding phases λF1(RS) and λF3(RS) described in the single jump experiments sufficiently describe the observed transitions λF1(IU) and λF3(IU). A secondary plot of the amplitudes AF1(IU) (•), AF2(IU) (□) and AF3(IU) (×) corresponding to the rate constants λF1(IU), λF2(IU) and λF3(IU) is shown in (c). A global fit performed on these amplitudes yields rate constants of ΛU2(IU) = 0.14 s−1 and ΛU3(IU) = 0.012 s−1. The fits are depicted as solid (AF1(IU)), dashed (AF2(IU)) and dotted (AF3(IU)) lines. To account for the different scales in amplitude changes, the y-axis is splitted at 0.1 a.u.中断型展开反应（interrupted unfolding reaction）的混合流程如图(a)所示。将蛋白质置于6.0 M尿素中完成展开反应并孵育t₁时长后，将其稀释至1.2 M尿素体系中。后续的复性过程以复性时长t₂为变量进行记录，结果如图(b)所示。当孵育时长较短时，可观测到F2(IU)复性相。当孵育时长较长时，单跳实验中所表征的快、慢复性相λF1(RS)与λF3(RS)，可充分拟合实验中观测到的λF1(IU)与λF3(IU)相变过程。图(c)展示了对应于速率常数λF1(IU)、λF2(IU)与λF3(IU)的振幅AF1(IU)（•）、AF2(IU)（□）及AF3(IU)（×）的副图。对上述振幅进行整体拟合后，得到速率常数ΛU2(IU) = 0.14 s⁻¹与ΛU3(IU) = 0.012 s⁻¹。拟合结果分别以实线（对应AF1(IU)）、虚线（对应AF2(IU)）与点线（对应AF3(IU)）绘制。为适配振幅变化的不同尺度，y轴在0.1任意单位（a.u.，arbitrary units）处进行了拆分。