GroES in the asymmetric GroEL14-GroES7 complex exchanges via an associative mechanism

The interaction of the chaperonin GroEL(14) with its cochaperonin GroES(7) is dynamic, involving stable. asymmetric 1:1 complexes (GroES(7). GroEL(7)-GroEL(7)) and transient, metastable symmetric 2:1 complexes [GroES(7). GroES(7)-GroEL(7). GroES(7)]. The transient formation of a 2:1 complex permits exchange of free GroES(7) for GroES(7) bound in the stable 1:1 complex, Electrophoresis in the presence of ADP was used to resolve free GroEL(14) from the GroES(7)-GroEL(14) complex. Titration of GroEL(14) with radiolabeled GroES(7) to molar ratios of 32:I demonstrated a 1:1 Limiting stoichiometry in a stable complex. No stable 2:1 complex was detected Preincubation of the asymmetric GroES(7). GroEL(7)-GroEL(7) complex with excess unlabeled GroES7 in the presence of ADP demonstrated GroES(7) exchange, The rates of GroES(7) exchange were proportional to the concentration of unlabeled free GroES(7). This concentration dependence points to an associative mechanism in which exchange of GroES(7) occurs by way of a transient 2:1 complex and excludes a dissociative mechanism in which exchange occurs by way Of free GroEL(14) Exchange of radiolabeled ADP from 1:1 complexes was much slower than the exchange of GroES(7), In agreement with recent structural studies, this indicates that conformational changes in GroEL(14) following the dissociation of GroES(7) must precede ADP release, These results explain how the GroEL(14) cavity can become reversibly accessible to proteins under in vivo conditions that favor 2:1 complexes.