Table 3. Thermodynamic properties of WT and DCase-M3 by urea denaturation (25 °C).

Unfolding of WT and DCase-M3, induced by the addition of urea, was monitored by the intrinsic fluorescence of tryptophan. The data containing the fraction of protein in the folded and unfolded state were calculated as the two-state unfolding model (ΔG=−RT·lnK_u). ΔG_H2O is the Gibbs free energy in the absence of denaturant, ΔG=ΔG_H2O−m[urea]. C_m represents denaturant concentration at the midpoint of the denaturant transition. The m value represents the dependence of the free energy charge of unfolding on denaturant concentration, that is, the slope of plots shown in Figure 4(B).

Protein	ΔGH2O (kcal/mol)	Cm (M)	m (kcal/mol per M)
WT	5.72±0.41	3.56±0.03	1.61±0.10
DCase-M3	6.13±0.60	3.21±0.10	1.91±0.13