Correlation between the stability of the GroEL-protein ligand complex and the release mechanism.
The protein-protein interactions during GroE-mediated protein refolding are of crucial importance for understanding how the assisted refolding of non-native proteins is achieved. Since GroEL seems to be a rather promiscuous polypeptide-binding protein it is not surprising that conditions for efficient dissociation from GroEL are promiscuous as well. To understand assisted protein refolding it is necessary to elucidate the underlying principles of the different partial steps of the functional cycle. Here we show a correlation between the overall stability of the complex between GroEL and ligand protein and the conditions for functional release from the chaperonin. As a model system, differently denatured species of an antibody Fab fragment were used. While weakly bound Fab fragments are functionally released in the absence of GroES, stably associated non-native forms of the same protein are dependent on the presence of the co-chaperonin for optimal GroE-mediated reactivation, suggesting that complex stability determines the release requirement. However, the observed overall stability of the complex between GroEL and substrate protein may be regarded as the net product of constant binding and rebinding of the ligand protein, once associated with GroEL, as shown by competition experiments.
Institut für Biophysik und Physikalische Biochemie, Universität Regensburg, Germany.
M
M
Schmidt
U
U
Bücheler
B
B
Kaluza
J
J
Buchner
D018835:Chaperonin 10 / Q000737:chemistry / Q000378:metabolism; D018834:Chaperonin 60 / Q000737:chemistry* / Q000378:metabolism; D004355:Drug Stability; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000378:metabolism; D019791:Guanidine; D006146:Guanidines; D007140:Immunoglobulin Fab Fragments / Q000096:biosynthesis / Q000737:chemistry* / Q000302:isolation & purification; D007700:Kinetics; D008024:Ligands; D011485:Protein Binding; D017510:Protein Folding; D017433:Protein Structure, Secondary; D017384:Sequence Deletion; D013050:Spectrometry, Fluorescence; D013997:Time Factors
D018835:Chaperonin 10; D018834:Chaperonin 60; D006146:Guanidines; D007140:Immunoglobulin Fab Fragments; D008024:Ligands; D019791:Guanidine