Figure 10.
Kinetic models for refolding and unfolding. A, refolding kinetics model for MSG under strong native conditions. Rates for unobservable kinetic steps are hypothetical values and are shown in red. The values in black represent rate-determining steps of refolding and are adjusted according to hypothetical rate values. IN behaves as an off-pathway intermediate whose partial unfolding to IP is a prerequisite for correct folding. The values in parentheses indicate the dependence of natural log of rates on urea concentration. The microscopic rates for reverse reactions are neglected due to the assumption of refolding under strong native conditions. B, unfolding kinetic model consists of initial rapid partition toward two unfolding pathways. The hypothetical rates (red) for initial partition are adjusted to obtain amplitudes corresponding to phase-1 (fast phase) and -2 (slow phase) for unfolding at 7 m urea within <100 ms dead time of mixing. The rest of the values (black) are proposed unfolding rates in the absence of urea whose variation with the denaturant is tuned by values shown in parentheses. A common kinetic model satisfying both refolding and unfolding data requires the inclusion of additional species in the system, and will be unreliable due to a large number of hypothetical rates. C, fitting of urea dependence of theoretical rates on the chevron plot. Where R1, R2, R3, R4, and R5 are the refolding model rates corresponding to U → IM, IM → ID, ID → IP, IP → IN, IN → IP steps, respectively. U1, U2, U3, and U4 are the unfolding model rates corresponding to steps, N → IY, IY → IP, IP → U, and IX → U, respectively. D, population variation of individual species; U (red), IM (orange), IN (blue), and N (purple) with time during refolding at 0.1 m urea (dashed lines). ID and IP do not populate to appreciable amounts during refolding (at 0.1 m urea). The net resultant signal (thick gray line) is hypothesized based on optical signals for individual species, where the fluorescence for U, IM, and N have been fixed in accordance with observed kinetic trace.