Evidence of stable monomeric species in the unfolding of Cu,Zn superoxide dismutase from Photobacterium leiognathi

The equilibrium unfolding process of Photobacterium leiognathi Cu,Zn supero xide dismutase has been quantitatively monitored through circular dichroism (CD) and fluorescence spectroscopy, upon increasing the guanidinium hydroc hloride concentration. The study has been undertaken for both the holo- and the copper-free derivative to work out the role of copper in protein stabi lity. In both cases the unfolding: was reversible. The denaturation curve d erived from CD and fluorescence spectroscopy was not coincident, suggesting that the denaturation process occurs through a three-state model with form ation of an intermediate monomeric species. The occurrence of an intermedia te species has been unambiguously demonstrated following CD and steady-stat e fluorescence spectra of the enzyme at various concentrations in presence of a fixed amounts of guanidinium hydrochloride, (C) 1999 Academic Press.