Unfolding and refolding of the NAD(+)-dependent isocitrate dehydrogenase from yeast.

The unfolding of the NAD(+)-dependent isocitrate dehydrogenase from yeast in guanidinium chloride (GdnHCl) has been monitored by changes in c.d. and fluorescence. Major structural changes occur over the range of GdnHCl concentrations from 0.5 to 1.5 M, although loss of catalytic activity is complete at 0.3 M. After incubation in GdnHCl, activity can be regained on dilution; however, the extent of this regain is dependent on the initial concentration of GdnHCl and is very small at a concentration of 2 M or above. Under these conditions there is only limited regain of the secondary and tertiary structure of the enzyme. Considerably more structure and activity can be regained if the concentration of GdnHCl is lowered by dialysis. The implications of these results for the folding and assembly of the enzyme are discussed.