A mutant chaperonin with rearranged inter-ring electrostatic contacts and temperature-sensitive dissociation.
The chaperonin GroEL assists protein folding through ATP-dependent, cooperative movements that alternately create folding chambers in its two rings. The substitution E461K at the interface between these two rings causes temperature-sensitive, defective protein folding in Escherichia coli. To understand the molecular defect, we have examined the mutant chaperonin by cryo-EM. The normal out-of-register alignment of contacts between subunits of opposing wild-type rings is changed in E461K to an in-register one. This is associated with loss of cooperativity in ATP binding and hydrolysis. Consistent with the loss of negative cooperativity between rings, the cochaperonin GroES binds simultaneously to both E461K rings. These GroES-bound structures were unstable at higher temperature, dissociating into complexes of single E461K rings associated with GroES. Lacking the allosteric signal from the opposite ring, these complexes cannot release their GroES and become trapped, dead-end states.
Electron Microscope Unit and Department of Chemistry, University of Cape Town, Rondebosch, South Africa.
B Trevor
BT
Sewell
Robert B
RB
Best
Shaoxia
S
Chen
Alan M
AM
Roseman
George W
GW
Farr
Arthur L
AL
Horwich
Helen R
HR
Saibil
D000251:Adenosine Triphosphatases / Q000737:chemistry; D000255:Adenosine Triphosphate / Q000737:chemistry; D018835:Chaperonin 10 / Q000737:chemistry* / Q000378:metabolism; D018834:Chaperonin 60 / Q000737:chemistry* / Q000378:metabolism*; D018833:Chaperonins / Q000235:genetics*; D020285:Cryoelectron Microscopy; D004305:Dose-Response Relationship, Drug; D004926:Escherichia coli / Q000378:metabolism; D006868:Hydrolysis; D008958:Models, Molecular; D009154:Mutation; D011485:Protein Binding; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D015398:Signal Transduction; D055672:Static Electricity; D013696:Temperature
D018835:Chaperonin 10; D018834:Chaperonin 60; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases; D018833:Chaperonins