STABILITY OF THE ASYMMETRIC ESCHERICHIA-COLI CHAPERONIN COMPLEX - GUANIDINE CHLORIDE CAUSES RAPID DISSOCIATION

The chaperonin proteins, GroEL(14), and GroES(7), inhibit protein aggr egation and assist in protein folding in a potassium/ATP-dependent man ner. In vitro, assays for chaperonin activity typically involve adding a denatured substrate protein to the chaperonins and measuring the ap pearance of correctly folded substrate protein. The influence of denat urant is generally ignored. Low concentrations of guanidinium chloride (<100 mM) had a profound effect on the activity/structure of the chap eronins. Guanidinium decreased the ATPase activity of GroEL and attenu ated the inhibition of GroEL ATP hydrolysis by GroES. The stable, asym metric chaperonin complex which forms in the presence of GroES and ADP (GroES(7) . ADP(7) . GroEL(7)-GroEL()) rapidly dissociated upon addit ion of 80 mM guanidinium chloride. Dissociation was enhanced at high i onic strength, but rapid dissociation was guanidinium-specific. Accele rated release of the GroES from the complex was also demonstrated. Unf olded proteins alone had no effect on complex stability. Residual guan idinium depressed the rate of Rhodospirillum rubrum ribulose-1,5-bisph osphate carboxylase (Rubisco) folding; an increased aggregation rate a lso decreased the yield of folded Rubisco. Chaperonin-assisted folding is therefore best studied using proteins denatured by means other tha n guanidinium chloride.