The structure of ClpB: A molecular chaperone that rescues proteins from an aggregated state
ABSTRACT Molecular chaperones assist protein folding by facilitating their "forward" folding and preventing aggregation. However, once aggregates have formed, these chaperones cannot facilitate protein disaggregation. Bacterial ClpB and its eukaryotic homolog Hsp104 are essential proteins of the heat-shock response, which have the remarkable capacity to rescue stress-damaged proteins from an aggregated state. We have determined the structure of Thermus thermophilus ClpB (TClpB) using a combination of X-ray crystallography and cryo-electron microscopy (cryo-EM). Our single-particle reconstruction shows that TClpB forms a two-tiered hexameric ring. The ClpB/Hsp104-linker consists of an 85 A long and mobile coiled coil that is located on the outside of the hexamer. Our mutagenesis and biochemical data show that both the relative position and motion of this coiled coil are critical for chaperone function. Taken together, we propose a mechanism by which an ATP-driven conformational change is coupled to a large coiled-coil motion, which is indispensable for protein disaggregation.
- SourceAvailable from: Clarisse Brígido
- "The bacterial ClpB monomer contains an N-terminal domain and two AAA+ nucleotide-binding domains (NBD) separated by a coiled-coil middle-domain (M-domain) (Barnett et al. 2000; DeSantis and Shorter 2012; Doyle and Wickner 2009). Several studies indicate that the M-domain is essential for the chaperone activity (Barnett et al. 2005; Kedzierska et al. 2003; Lee et al. 2005; Mogk et al. 2003; Schirmer et al. 2004). Protein disaggregation by ClpB in vitro requires the collaboration of a second ATP-dependent molecular chaperone, DnaK, to promote the solubilization and reactivation of proteins that misfold and aggregate following heat shock (Goloubinoff et al. 1999; Motohashi et al. 1999; Zolkiewski 1999). "
Dataset: Brigido 2012b
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- "We used the homology modeling approach, implemented in the MODELLER [Sali and Blundell, 1993] 9v10 program, to build models of torsinAwt, delE302, delE303, F205I and R288Q based on the 3D structure of the ClpB second AAA+ domain of Thermus thermophilus (T. thermophilus) [Lee et al., 2003]. The protocol used to perform the molecular modeling experiments was: generation of 10 models, from which one model for each torsinA sequence was selected. "
ABSTRACT: Early-onset dystonia is associated with the deletion of one of a pair of glutamic acid residues (c.904_906delGAG/c.907_909delGAG; p.Glu302del/Glu303del; ΔE 302/303) near the carboxyl-terminus of torsinA, a member of the AAA+ protein family that localizes to the endoplasmic reticulum (ER) lumen and nuclear envelope (NE). This deletion commonly underlies early-onset DYT1 dystonia. While the role of the disease-causing mutation, torsinAΔE, has been established through genetic association studies, it is much less clear whether other rare human variants of torsinA are pathogenic. Two missense variations have been described in single patients; R288Q (c.863G>A; p.Arg288Gln; R288Q) identified in a patient with onset of severe generalized dystonia and myoclonus since infancy, and F205I (c.613T>A, p.Phe205Ile; F205I) in a psychiatric patient with late-onset focal dystonia. In this study, we have undertaken a series of analyses comparing the biochemical and cellular effects of these rare variants to torsinAΔE and wild-type (wt) torsinA in order to reveal whether there are common dysfunctional features. The results revealed that the variants, R288Q and F205I, are more similar in their properties to torsinAΔE protein than to torsinAwt. These findings provide functional evidence for the potential pathogenic nature of these rare sequence variants in the TOR1A gene, thus implicating these pathologies in the development of dystonia. This article is protected by copyright. All rights reserved.Human Mutation 09/2014; DOI:10.1002/humu.22602 · 5.05 Impact Factor
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- "As described above, the dynein hexamer exists as a singlelayered asymmetric ring. In contrast, the two covalently linked ATPase domains of p97, NSF, and ClpB oligomerize into a two-tiered structure of stacked hexameric rings     . In such twolayered barrels, one of the homohexamers is responsible for the ATP hydrolysis while the second homohexamer is only capable of nucleotide binding   . "
ABSTRACT: Among protein secretion systems there are specialized ATPases that serve different functions such as substrate recognition, substrate unfolding, and assembly of the secretory machinery. ESX protein secretion systems require FtsK/SpoIIIE family ATPases but the specific function of these ATPases is poorly understood. The ATPases of ESX secretion systems have a unique domain architecture among proteins of the FtsK/SpoIIIE family. All well-studied FtsK family ATPases to date have one ATPase domain and oligomerize to form a functional molecular machine, most commonly a hexameric ring. In contrast, the ESX ATPases have three ATPase domains, either encoded by a single gene or by two operonic genes. It is currently unknown which of the ATPase domains is catalytically functional and whether each domain plays the same or a different function. Here we focus on the ATPases of two ESX systems, the ESX-1 system of Mycobacterium tuberculosis and the yuk system of Bacillus subtilis. We show that ATP hydrolysis by the ESX ATPase is required for secretion, suggesting that this enzyme at least partly fuels protein translocation. We further show that individual ATPase domains play distinct roles in substrate translocation and complex formation. Comparing the single chain and split ESX ATPases we reveal differences in the requirements of these unique secretory ATPases.Journal of Molecular Biology 06/2014; 427(5). DOI:10.1016/j.jmb.2014.06.013 · 4.33 Impact Factor