title: Folding and assembly of the large molecular machine Hsp90 studied in single-molecule experiments.
abstract: Folding of small proteins often occurs in a two-state manner and is well understood both experimentally and theoretically. However, many proteins are much larger and often populate misfolded states, complicating their folding process significantly. Here we study the complete folding and assembly process of the 1,418 amino acid, dimeric chaperone Hsp90 using single-molecule optical tweezers. Although the isolated C-terminal domain shows two-state folding, we find that the isolated N-terminal as well as the middle domain populate ensembles of fast-forming, misfolded states. These intradomain misfolds slow down folding by an order of magnitude. Modeling folding as a competition between productive and misfolding pathways allows us to fully describe the folding kinetics. Beyond intradomain misfolding, folding of the full-length protein is further slowed by the formation of interdomain misfolds, suggesting that with growing chain lengths, such misfolds will dominate folding kinetics. Interestingly, we find that small stretching forces applied to the chain can accelerate folding by preventing the formation of cross-domain misfolding intermediates by leading the protein along productive pathways to the native state. The same effect is achieved by cotranslational folding at the ribosome in vivo.
authors.affiliation: Physik-Department, Technische Universität München, 85748 Garching, Germany;
authors.forename: Markus
authors.initials: M
authors.lastname: Jahn
authors.affiliation: Department Chemie, Technische Universität München, 85748 Garching, Germany; Munich Center for Integrated Protein Science, 81377 München, Germany;
authors.forename: Johannes
authors.initials: J
authors.lastname: Buchner
authors.affiliation: Institute of Physical Chemistry, University of Freiburg, 79104 Freiburg, Germany.
authors.forename: Thorsten
authors.initials: T
authors.lastname: Hugel
authors.affiliation: Physik-Department, Technische Universität München, 85748 Garching, Germany; Munich Center for Integrated Protein Science, 81377 München, Germany; mrief@ph.tum.de.
authors.forename: Matthias
authors.initials: M
authors.lastname: Rief
mesh_terms: D019281:Dimerization; D018841:HSP90 Heat-Shock Proteins / Q000737:chemistry*; D007700:Kinetics; D017510:Protein Folding
chemical_list: D018841:HSP90 Heat-Shock Proteins
keywords: misfolding; off-pathway; optical tweezers; rough energy landscape