Stability of the asymmetric Escherichia coli chaperonin complex. Guanidine chloride causes rapid dissociation

Abstract

The chaperonin proteins, GroEL14 and GroES7, inhibit protein aggregation and assist in protein folding in a potassium/ATP-dependent manner. In vitro, assays for chaperonin activity typically involve adding a denatured substrate protein to the chaperonins and measuring the appearance of correctly folded substrate protein. The influence of denaturant is generally ignored. Low concentrations of guanidinium chloride (< 100 mM) had a profound effect on the activity/structure of the chaperonins. Guanidinium decreased the ATPase activity of GroEL and attenuated the inhibition of GroEL ATP hydrolysis by GroES. The stable, asymmetric chaperonin complex which forms in the presence of GroES and ADP (GroES7.ADP7.GroEL7-GroEL7) rapidly dissociated upon addition of 80 mM guandinium chloride. Dissociation was enhanced at high ionic strength, but rapid dissociation was guanidinium-specific. Accelerated release of the GroES from the complex was also demonstrated. Unfolded proteins alone had no effect on complex stability. Residual guanidinium depressed the rate of Rhodospirillum rubrum ribulose-1,5-bisphosphate carboxylase (Rubisco) folding; an increased aggregation rate also decreased the yield of folded Rubisco. Chaperonin-assisted folding is therefore best studied using proteins denatured by means other than guanidinium chloride.