Single-molecule spectroscopy of protein folding in a chaperonin cage.
Molecular chaperones are known to be essential for avoiding protein aggregation in vivo, but it is still unclear how they affect protein folding mechanisms. We use single-molecule Förster resonance energy transfer to follow the folding of a protein inside the GroEL/GroES chaperonin cavity over a time range from milliseconds to hours. Our results show that confinement in the chaperonin decelerates the folding of the C-terminal domain in the substrate protein rhodanese, but leaves the folding rate of the N-terminal domain unaffected. Microfluidic mixing experiments indicate that strong interactions of the substrate with the cavity walls impede the folding process, but the folding hierarchy is preserved. Our results imply that no universal chaperonin mechanism exists. Rather, a competition between intra- and intermolecular interactions determines the folding rates and mechanisms of a substrate inside the GroEL/GroES cage.
Hofmann
Hagen
H
Biochemisches Institut, Universität Zürich, Winterthurerstrasse 190, 8057 Zürich, Switzerland.
Hillger
Frank
F
Pfeil
Shawn H
SH
Hoffmann
Armin
A
Streich
Daniel
D
Haenni
Dominik
D
Nettels
Daniel
D
Lipman
Everett A
EA
Schuler
Benjamin
B
D055592:Biophysical Phenomena; D018835:Chaperonin 10 / Q000737:chemistry; D018834:Chaperonin 60 / Q000737:chemistry; D018833:Chaperonins / Q000737:chemistry*; D029968:Escherichia coli Proteins / Q000737:chemistry; D031541:Fluorescence Resonance Energy Transfer; D005456:Fluorescent Dyes; D007700:Kinetics; D044085:Microfluidics; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013816:Thermodynamics; D013884:Thiosulfate Sulfurtransferase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
D018835:Chaperonin 10; D018834:Chaperonin 60; D029968:Escherichia coli Proteins; D005456:Fluorescent Dyes; D011994:Recombinant Proteins; D013884:Thiosulfate Sulfurtransferase; D018833:Chaperonins