earticle

영어

Group II chaperonin-mediated protein folding is critically dependent on the closure of a built-in lid. ATP drives the conformational change of the group II chaperonin from the open lid substrate-binding conformation to the closed lid conformation to encapsulate an unfolded protein in the central cavity. It is thought that the folding activity is strongly correlated with the ATP-dependent conformational change ability. In this study, we investigated that the group II chaperonin (PhCpn) from Pyrococcus horikoshii OT3 can ATP-dependently induce improvement of ATPase activity, thermal protection and inactivation of foreign proteins, such as alcohol dehydrogenase (ADH) from Saccharomyces cerevisiae and citrate synthase (CS) from porcine heart. To identify ATP dependency in ATPase and thermal activity, Phcpn, ATPbinding enhanced mutants (D64G and D393G) and ATP-binding impaired mutants (D64A, G65C, D393A, D64A/D393A) were constructed through positional change in ATP binding site by site-directed mutagenesis, respectively. The ATPase activities of enhanced mutants were the highest because of their high ATP-binding efficiency. Also, PhCpn prevented the thermal aggregation and inactivation of ADH and CS. Especially, the addition of ATP with PhCpn more extensively prevented thermal aggregation and inactivation. Therefore, these results reveal that ATPase activity of chaperonin is mediated through ATP dependent action.