Chaperonin-Ring's Twist is Critical for Folding Activity of Group II Chaperonin

Abstract

Group II chaperonin is an indispensable protein in archaea or in the eukaryotic cytosol that captures an unfolded protein and refolds it to the correct conformation in an ATP dependent manner. Chaperonin-mediated protein folding is achieved on the closure and opening of a built-in lid under ATP cycle. Recent study with cryo-electron microscopy and single particle analysis suggested that the ring structure of chaperonin twisted to seal off the central cavity. However such dynamics could not be traced experimentally since the motion was expected to be too small to detect by visible light technology.Here we show that ATP dependent dynamic motion of group II chaperonin from Thermococcus strain KS-1 at single molecular level with high accurate rotational view by diffracted X-ray tracking (DXT). DXT has been considered as a powerful technique in biological science for detecting subtle dynamic motion of the target protein at single molecular level. The dynamics of a single protein can be monitored through trajectory of the Laue spot from the nanocrystal which was labeled on the objective protein immobilized on the substrate surface. UV-light triggered DXT using caged ATP revealed that the chaperonin’s lid closed partially within one second after ATP binding, the closed ring twisted counter clock-wisely from top to bottom view of chaperonin with 130 m rad/sec in angular speed within 2 to 6 seconds, and the twisted ring turned back to the original open state with 80 m rad/sec clockwise twisting motion. Our analyses with precise rotational and macroscopic views for chaperonin’s dynamics show that there are distinct two modes in lid-closure process, in-cooperative closure and cooperative counter-clockwise twisting motion. Moreover, we found that the ring’s twisting motion correlated to the folding activity of group II chaperonin.