Equilibrium Stability of Double-CI2
A. GdnHCl denatured
fluorescence curve.
Left: double-CI2
Right: WT-CI2
(Ex 280nm, Em 356nm)

B.By stopped-flow fluorescence:

(Right hand side)
Top: Double-CI2
Meddle: "Slower phase" double-CI2
Bottom: WT-CI2

The free energy of Double-CI2 didn't change a lot! Although the Cm is smaller
than WT-CI2, but the slope "m-value" is big!

The mutants "destabilized" the double-CI2!

Double-CI2 showed a remarkable curvture at the low [GdnHCl] , suggesting that
a stable intermediate accumulated in the refolding process of double-CI2.

ie. Double-CI2 is not simple two-state refolding!