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The unfolding and attempted refolding of citrate synthase from pig heart
S M West1, S M Kelly, N C Price
1School of Molecular and Biological Sciences, University of Stirling, U.K.
Abstract:
The unfolding of the dimeric enzyme citrate synthase from pig heart in solutions of guanidinium chloride (GdnHCl) was studied. Data from fluorescence, circular dichroism (CD) and thiol group reactivity studies indicated that the enzyme was almost completely unfolded at GdnHCl concentrations greater than or equal to 4 M. On dilution of GdnHCl, essentially no reactivation of the enzyme occurred. The implications of this finding for the process of folding and assembly in vivo of this and other mitochondrial enzymes are discussed. Exposure of the enzyme to high pH (9-10) led to only a small loss of secondary structure and partial reactivation could be observed on readjustment of the pH to 8.0.