Mechanism of GroEL action: Productive release of polypeptide from a sequestered position under groes

Abstract

The chaperonin GroEL is a large, double-ring structure that, together with ATP and the cochaperonin GroES, assists protein folding in vivo. GroES forms an asymmetric complex with GroEL in which a single GroES ring binds one end of the GroEL cylinder. Cross-linking studies reveal that polypeptide binding occurs exclusively to the GroEL ring not occupied by GroES (trans). During the folding reaction, however, released GroES can rebind to the GroEL ring containing polypeptide (cis). The polypeptide is held tightly in a proteolytically protected environment in cis complexes, in the presence of ADP. Single turnover experiments with ornithine transcarbamylase reveal that polypeptide is productively released from the cis but not the trans complex. These observations suggest a two-step mechanism for GroEL-mediated folding. First, GroES displaces the polypeptide from its initial binding sites, sequestering it in the GroEL central cavity. Second, ATP hydrolysis induces release of GroES and productive release of polypeptide.

GroEL‐Mediated protein folding

“…1B) are also critical for binding GroES . Such apparent competition by GroES for the peptide binding sites leads, in the presence of ATP, to complete release of polypeptide into the central channel, accompanied by initiation of productive folding (Weissman et al, 1995; see below).…”

“…Our current understanding of the pathway of a chaperonin reaction is summarized in Figure 4. (1) Non-native polypeptide (Iuc) is bound by an asymmetric GroEL-GroES complex, a dynamic species formed in the presence of physiological levels of ATP/ADP, in which the GroEL central channel at one end of the cylinder is capped by GroES, whereas the channel at the other end remains accessible to polypeptide (Jackson et al, 1993;Martin et al, 1993;Burston et al, 1995;Weissman et al, 1995; and see below). One subunit is shown in purple, and the single resolved mobile loop is shown in red.…”

“…forming a productive cis ternary complex Weissman et al, 1995;Mayhew et al, 1996). Such a cis ternary complex could be formed either by release of GroES from the frons ring, followed by binding in cis, or by binding of a second GroES in cis.…”

“…Interestingly, the association rate to form the complex was lower than those reported for some polypeptide-GroEL interactions (above), varying from 1 X IO5 M" s-] (Jackson et al, 1993) to 4 X IO5 M" s -' , both in the presence of ADP. Finally, dissociation constants for ternary GroELGroES-polypeptide complexes that take into account cis and trans topologies (see below) have not been reported, but the ability to isolate such complexes by gel filtration (Weissman et al, 1995) suggests that their dissociation constants must be submicromolar as well.…”

Conformational dynamics of the molecular chaperone Hsp90

“…Hsp60 (GroEL) and Hsp70 (DnaK) represent two well studied members of this class of proteins. Both Hsp60 and Hsp70 interact with nascent polypeptide chains and promote their folding by interacting with hydrophobic surfaces on substrate proteins (Gomez-Puertas et al, 2004;Kurt et al, 2006;Rudiger et al, 1997;Weissman et al, 1995). Once substrates are bound, the chaperone undergoes subsequent rounds of ATP hydrolysis causing conformational changes within the chaperone (Young et al, 2004).…”