Equilibrium and kinetic analysis on the folding of hen egg lysozyme in the aqueous-glycerol solution

In the present study, the equilibrium and kinetic analysis on the folding of hen egg lysozyme in the aqueous-glycerol solutions have been reported by means of fluorescence and circular dichroism spectra. Addition of glycerol increases the stability of lysozyme on the guanidine-induced denaturation, and the unfolding transition of lysozyme is from a two-state into a three-state mechanism. On the kinetic experiments, the folding pathway of lysozyme has been changed in the presence of glycerol. An overshot phenomenon in fluorescence spectra appears and the preformed ellipticity within the dead time is weakened. These results reveal that a new intermediate state with a non-native hydrophobic core but lack of the stable secondary structure is populated in the folding pathway of lysozyme, and the conversion of it to the native state is decelerated due to the excess of non-native hydrophobic interactions.