Unfolding-refolding transition of a hinge bending enzyme: horse muscle phosphoglycerate kinase induced by guanidine hydrochloride

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Abstract
The unfolding-refolding transition of horse muscle phosphoglycerate kinase induced by guanidine hydrochloride was studied under equilibrium conditions using 4 different signals: fluorescence intensity at 336 nm, UV difference absorbance at 286 nm, ellipticity at 220 nm and enzyme activity. From the following arguments, it was found that the process deviates from a 2-state model and intermediates are significantly populated even at equilibrium: the noncoincidence of the transition curves and the asymmetry of the transition curve obtained from CD [circular dichroism] measurements. From these different data and the thermodynamic analysis, it was suggested that the 2 domains of the horse muscle phosphoglycerate kinase refold independently of one another with different equilibrium constants, the most favorable constant referring to the folding of the C-terminal domain which contains all tryptophans.