Folding trajectories of human dihydrofolate reductase inside the GroEL GroES chaperonin cavity and free in solution.
The chaperonin GroEL binds non-native polypeptides in an open ring via hydrophobic contacts and then, after ATP and GroES binding to the same ring as polypeptide, mediates productive folding in the now hydrophilic, encapsulated cis chamber. The nature of the folding reaction in the cis cavity remains poorly understood. In particular, it is unclear whether polypeptides take the same route to the native state in this cavity as they do when folding spontaneously free in solution. Here, we have addressed this question by using NMR measurements of the time course of acquisition of amide proton exchange protection of human dihydrofolate reductase (DHFR) during folding in the presence of methotrexate and ATP either free in solution or inside the stable cavity formed between a single ring variant of GroEL, SR1, and GroES. Recovery of DHFR refolded by the SR1/GroES-mediated reaction is 2-fold higher than in the spontaneous reaction. Nevertheless, DHFR folding was found to proceed by the same trajectories inside the cis folding chamber and free in solution. These observations are consistent with the description of the chaperonin chamber as an "Anfinsen cage" where polypeptide folding is determined solely by the amino acid sequence, as it is in solution. However, if misfolding occurs in the confinement of the chaperonin cavity, the polypeptide chain cannot undergo aggregation but rather finds its way back to a productive pathway in a manner that cannot be accomplished in solution, resulting in the observed high overall recovery.
Horst
Reto
R
Department of Molecular Biology and The Skaggs Institute for Chemical Biology, The Scripps Research Institute, La Jolla, CA 92037, USA.
Fenton
Wayne A
WA
Englander
S Walter
SW
Wüthrich
Kurt
K
Horwich
Arthur L
AL
D000255:Adenosine Triphosphate / Q000737:chemistry; D018835:Chaperonin 10 / Q000737:chemistry*; D018834:Chaperonin 60 / Q000737:chemistry*; D018833:Chaperonins / Q000737:chemistry; D006801:Humans; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D011485:Protein Binding; D011489:Protein Denaturation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D012996:Solutions; D012997:Solvents / Q000737:chemistry; D013762:Tetrahydrofolate Dehydrogenase / Q000737:chemistry*
D018835:Chaperonin 10; D018834:Chaperonin 60; D012996:Solutions; D012997:Solvents; D000255:Adenosine Triphosphate; D013762:Tetrahydrofolate Dehydrogenase; D018833:Chaperonins