Abstract
We have identified and reconstituted a multicomponent redox-chaperone network that appears to be designed to protect proteins against stress-induced unfolding and to refold proteins when conditions return to normal. The central player is Hsp33, a redox-regulated molecular chaperone. Hsp33, which is activated by disulfide bond formation and subsequent dimerization, works as an efficient chaperone holdase that binds to unfolding protein intermediates and maintains them in a folding competent conformation. Reduction of Hsp33 is catalyzed by the glutaredoxin and thioredoxin systems in vivo, and leads to the formation of highly active, reduced Hsp33 dimers. Reduction of Hsp33 is necessary but not sufficient for substrate protein release. Substrate dissociation from Hsp33 is linked to the presence of the DnaK/DnaJ/GrpE foldase system, which alone, or in concert with the GroEL/GroES system, then supports the refolding of the substrate proteins. Upon substrate release, reduced Hsp33 dimers dissociate into inactive monomers. This regulated substrate transfer ultimately links substrate release and Hsp33 inactivation to the presence of available DnaK/DnaJ/GrpE, and, therefore, to the return of cells to non-stress conditions.
MeSH Terms
Animals
Cattle
Citrate (si)-Synthase/metabolism
Cysteine/chemistry
Dimerization
Disulfides/chemistry
Dithiothreitol/pharmacology
Fluorescence Polarization
Heat-Shock Proteins/chemistry,genetics,metabolism,physiology
Luciferases/metabolism
Models, Biological
Molecular Chaperones/chemistry,genetics,metabolism,physiology
Oxidation-Reduction
Protein Conformation
Protein Denaturation
Protein Folding
Protein Processing, Post-Translational
Reducing Agents/pharmacology
Serum Albumin, Bovine/metabolism
Substrate Specificity
Swine
Temperature
Time Factors
Chemicals
Disulfides
Heat-Shock Proteins
Molecular Chaperones
Reducing Agents
Serum Albumin, Bovine
Luciferases
Citrate (si)-Synthase
Cysteine
Dithiothreitol
Authors & Affiliations
5 authors, click to expand affiliations / ORCID
Hoffmann Jörg H
Department of Molecular, Cellular and Developmental Biology, University of Michigan, Ann Arbor, MI, USA.
Linke Katrin
Graf Paul C F
Lilie Hauke
Jakob Ursula
References (20)
20 references, click to expand
-
Identification of thermolabile Escherichia coli proteins: prevention and reversion of aggregation by DnaK and ClpB.
EMBO J. 1999 Dec 15;18(24):6934-49
PMID: 10601016
-
Activation of the OxyR transcription factor by reversible disulfide bond formation.
Science. 1998 Mar 13;279(5357):1718-21
PMID: 9497290
-
Redox switch of hsp33 has a novel zinc-binding motif.
J Biol Chem. 2000 Dec 8;275(49):38302-10
PMID: 10976105
-
Protein disulfide isomerase acts as a redox-dependent chaperone to unfold cholera toxin.
Cell. 2001 Mar 23;104(6):937-48
PMID: 11290330
-
Activation of the redox-regulated molecular chaperone Hsp33--a two-step mechanism.
Structure. 2001 May 9;9(5):377-87
PMID: 11377198
-
DnaK dependence of mutant ethanol oxidoreductases evolved for aerobic function and protective role of the chaperone against protein oxidative damage in Escherichia coli.
Proc Natl Acad Sci U S A. 2002 Apr 2;99(7):4626-31
PMID: 11917132
-
Trigger Factor and DnaK possess overlapping substrate pools and binding specificities.
Mol Microbiol. 2003 Mar;47(5):1317-28
PMID: 12603737
-
Refolding of substrates bound to small Hsps relies on a disaggregation reaction mediated most efficiently by ClpB/DnaK.
J Biol Chem. 2003 Aug 15;278(33):31033-42
PMID: 12788951
-
How chaperones fold proteins.
Biol Chem. 1998 Mar;379(3):245-59
PMID: 9563819
-
The small heat-shock protein IbpB from Escherichia coli stabilizes stress-denatured proteins for subsequent refolding by a multichaperone network.
J Biol Chem. 1998 May 1;273(18):11032-7
PMID: 9556585
-
Chaperone activity with a redox switch.
Cell. 1999 Feb 5;96(3):341-52
PMID: 10025400
-
Interaction of DnaK with native proteins and membrane proteins correlates with their accessible hydrophobicity.
Gene. 1999 Apr 16;230(2):163-70
PMID: 10216254
-
Regulation of the OxyR transcription factor by hydrogen peroxide and the cellular thiol-disulfide status.
Proc Natl Acad Sci U S A. 1999 May 25;96(11):6161-5
PMID: 10339558
-
The molecular chaperone concept.
Biochem Soc Symp. 1989;55:145-53
PMID: 2695089
-
DnaK, DnaJ, and GrpE heat shock proteins negatively regulate heat shock gene expression by controlling the synthesis and stability of sigma 32.
Genes Dev. 1990 Dec;4(12A):2202-9
PMID: 2269429
-
GroE facilitates refolding of citrate synthase by suppressing aggregation.
Biochemistry. 1991 Feb 12;30(6):1586-91
PMID: 1671555
-
A conserved loop in the ATPase domain of the DnaK chaperone is essential for stable binding of GrpE.
Nat Struct Biol. 1994 Feb;1(2):95-101
PMID: 7656024
-
Equilibrium intermediates in the reversible unfolding of firefly (Photinus pyralis) luciferase.
J Biol Chem. 1997 Mar 14;272(11):7099-105
PMID: 9054403
-
The role of the thioredoxin and glutaredoxin pathways in reducing protein disulfide bonds in the Escherichia coli cytoplasm.
J Biol Chem. 1997 Jun 20;272(25):15661-7
PMID: 9188456
-
Mass spectrometry unravels disulfide bond formation as the mechanism that activates a molecular chaperone.
J Biol Chem. 2000 Jun 23;275(25):18759-66
PMID: 10764757