title: Four-colour FRET reveals directionality in the Hsp90 multicomponent machinery.
abstract: In living organisms, most proteins work in complexes to form multicomponent protein machines. The function of such multicomponent machines is usually addressed by dividing them into a collection of two state systems at equilibrium. Many molecular machines, like Hsp90, work far from equilibrium by utilizing the energy of ATP hydrolysis. In these cases, important information is gained from the observation of the succession of more than two states in a row. We developed a four-colour single-molecule FRET system to observe the succession of states in the heat shock protein 90 (Hsp90) system, consisting of an Hsp90 dimer, the cochaperone p23 and nucleotides. We show that this multicomponent system is a directional ATP-dependent machinery. This reveals a previously undescribed mechanism on how cochaperones can modify Hsp90, namely by strengthening of the coupling between ATP hydrolysis and a kinetic step involved in the Hsp90 system resulting in a stronger directionality.
authors.affiliation: 1] Physik-Department E22 and IMETUM, Technische Universität München, München 85748, Germany [2].
authors.forename: C
authors.initials: C
authors.lastname: Ratzke
authors.affiliation: Physik-Department E22 and IMETUM, Technische Universität München, München 85748, Germany.
authors.forename: B
authors.initials: B
authors.lastname: Hellenkamp
authors.affiliation: Physik-Department E22 and IMETUM, Technische Universität München, München 85748, Germany.
authors.forename: T
authors.initials: T
authors.lastname: Hugel
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D003001:Cloning, Molecular; D019281:Dimerization; D004926:Escherichia coli; D005454:Fluorescence Polarization; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods*; D005456:Fluorescent Dyes; D018841:HSP90 Heat-Shock Proteins / Q000378:metabolism*; D018832:Molecular Chaperones / Q000235:genetics / Q000378:metabolism*; D046912:Multiprotein Complexes / Q000378:metabolism*; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism*; D029702:Schizosaccharomyces pombe Proteins / Q000235:genetics / Q000378:metabolism*; D015003:Yeasts
chemical_list: D005456:Fluorescent Dyes; D018841:HSP90 Heat-Shock Proteins; D018832:Molecular Chaperones; D046912:Multiprotein Complexes; D010750:Phosphoproteins; D029702:Schizosaccharomyces pombe Proteins; C490848:wos2 protein, S pombe; D000255:Adenosine Triphosphate