A mobile loop order-disorder transition modulates the speed of chaperonin cycling.
Molecular machines order and disorder polypeptides as they form and dissolve large intermolecular interfaces, but the biological significance of coupled ordering and binding has been established in few, if any, macromolecular systems. The ordering and binding of GroES co-chaperonin mobile loops accompany an ATP-dependent conformational change in the GroEL chaperonin that promotes client protein folding. Following ATP hydrolysis, disordering of the mobile loops accompanies co-chaperonin dissociation, reversal of the GroEL conformational change, and release of the client protein. "High-affinity" GroEL mutants were identified by their compatibility with "low-affinity" co-chaperonin mutants and incompatibility with high-affinity co-chaperonin mutants. Analysis of binding kinetics using the intrinsic fluorescence of tryptophan-containing co-chaperonin variants revealed that excessive affinity causes the chaperonin to stall in a conformation that forms in the presence of ATP. Destabilizing the beta-hairpins formed by the mobile loops restores the normal rate of dissociation. Thus, the free energy of mobile-loop ordering and disordering acts like the inertia of an engine's flywheel by modulating the speed of chaperonin conformational changes.
Shewmaker
Frank
F
Department of Biochemistry, Tulane University Health Sciences Center, New Orleans, Louisiana 70112, USA.
Kerner
Michael J
MJ
Hayer-Hartl
Manajit
M
Klein
Gracjana
G
Georgopoulos
Costa
C
Landry
Samuel J
SJ
D000255:Adenosine Triphosphate / Q000737:chemistry; D019943:Amino Acid Substitution / Q000235:genetics*; D018835:Chaperonin 10 / Q000737:chemistry* / Q000235:genetics; D018834:Chaperonin 60 / Q000737:chemistry* / Q000235:genetics; D004926:Escherichia coli / Q000201:enzymology*; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics; D019906:Nuclear Magnetic Resonance, Biomolecular; D017354:Point Mutation / Q000235:genetics*; D017433:Protein Structure, Secondary; D015219:Substrate Cycling / Q000235:genetics; D020349:Surface Plasmon Resonance
D018835:Chaperonin 10; D018834:Chaperonin 60; D029968:Escherichia coli Proteins; D000255:Adenosine Triphosphate