Head-to-tail interactions of the coiled-coil domains regulate ClpB activity and cooperation with Hsp70 in protein disaggregation.
The hexameric AAA+ chaperone ClpB reactivates aggregated proteins in cooperation with the Hsp70 system. Essential for disaggregation, the ClpB middle domain (MD) is a coiled-coil propeller that binds Hsp70. Although the ClpB subunit structure is known, positioning of the MD in the hexamer and its mechanism of action are unclear. We obtained electron microscopy (EM) structures of the BAP variant of ClpB that binds the protease ClpP, clearly revealing MD density on the surface of the ClpB ring. Mutant analysis and asymmetric reconstructions show that MDs adopt diverse positions in a single ClpB hexamer. Adjacent, horizontally oriented MDs form head-to-tail contacts and repress ClpB activity by preventing Hsp70 interaction. Tilting of the MD breaks this contact, allowing Hsp70 binding, and releasing the contact in adjacent subunits. Our data suggest a wavelike activation of ClpB subunits around the ring.DOI: http://dx.doi.org/10.7554/eLife.02481.001.
Department of Crystallography, Birkbeck College, University of London, London, United Kingdom.
Marta
M
Carroni
Zentrum für Molekulare Biologie, Universität Heidelberg, Heidelberg, Germany.
Eva
E
Kummer
Zentrum für Molekulare Biologie, Universität Heidelberg, Heidelberg, Germany.
Yuki
Y
Oguchi
Gene Center, Ludwig-Maximilians-University Munich, Munich, Germany.
Petra
P
Wendler
Department of Crystallography, Birkbeck College, University of London, London, United Kingdom.
Daniel K
DK
Clare
Biochemie-Zentrum, Universität Heidelberg, Heidelberg, Germany.
Irmgard
I
Sinning
Biochemie-Zentrum, Universität Heidelberg, Heidelberg, Germany.
Jürgen
J
Kopp
Zentrum für Molekulare Biologie, Universität Heidelberg, Heidelberg, Germany.
Axel
A
Mogk
Zentrum für Molekulare Biologie, Universität Heidelberg, Heidelberg, Germany.
Bernd
B
Bukau
Department of Crystallography, Birkbeck College, University of London, London, United Kingdom h.saibil@mail.cryst.bbk.ac.uk.
Helen R
HR
Saibil
D020816:Amino Acid Motifs; D020285:Cryoelectron Microscopy; D018360:Crystallography, X-Ray; D049071:Endopeptidase Clp; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000378:metabolism*; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism*; D006360:Heat-Shock Proteins / Q000737:chemistry* / Q000378:metabolism*; D021621:Imaging, Three-Dimensional; D056004:Molecular Dynamics Simulation; D050505:Mutant Proteins / Q000737:chemistry; D016624:Negative Staining; D066329:Protein Aggregates; D011485:Protein Binding; D017434:Protein Structure, Tertiary
D029968:Escherichia coli Proteins; D018840:HSP70 Heat-Shock Proteins; D006360:Heat-Shock Proteins; D050505:Mutant Proteins; D066329:Protein Aggregates; D049071:Endopeptidase Clp; C069775:ClpB protein, E coli
ClpB/Hsp104; protein unfolding; single particle EM