ABSTRACT
The molecular chaperone Hsp90 is an important regulator of proteostasis. It has remained unclear why S. cerevisiae possesses two Hsp90 isoforms, the constitutively expressed Hsc82 and the stress-inducible Hsp82. Here, we report distinct differences despite a sequence identity of 97%. Consistent with its function under stress conditions, Hsp82 is more stable and refolds more efficiently than Hsc82. The two isoforms also differ in their ATPases and conformational cycles. Hsc82 is more processive and populates closed states to a greater extent. Variations in the N-terminal ATP-binding domain modulate its dynamics and conformational cycle. Despite these differences, the client interactomes are largely identical, but isoform-specific interactors exist both under physiological and heat shock conditions. Taken together, changes mainly in the N-domain create a stress-specific, more resilient protein with a shifted activity profile. Thus, the precise tuning of the Hsp90 isoforms preserves the basic mechanism but adapts it to specific needs.
Subject(s)
HSP90 Heat-Shock Proteins/chemistry , Molecular Chaperones/chemistry , Protein Isoforms/chemistry , Saccharomyces cerevisiae Proteins/chemistry , Saccharomyces cerevisiae/metabolism , Adenosine Triphosphatases/metabolism , Amino Acid Sequence , HSP90 Heat-Shock Proteins/genetics , HSP90 Heat-Shock Proteins/metabolism , HSP90 Heat-Shock Proteins/physiology , Heat-Shock Response/physiology , Ligands , Models, Molecular , Molecular Chaperones/metabolism , Protein Binding , Protein Conformation , Protein Folding , Protein Isoforms/metabolism , Protein Stability , Saccharomyces cerevisiae Proteins/genetics , Saccharomyces cerevisiae Proteins/metabolism , Sequence Alignment , Stress, PhysiologicalABSTRACT
The heat-shock protein 90 (Hsp90) molecular chaperones are highly conserved across species. However, their dynamic properties can vary significantly from organism to organism. Here we used high-precision optical tweezers to analyze the mechanical properties and folding of different Hsp90 orthologs, namely bacterial Hsp90 (HtpG) and Hsp90 from the endoplasmic reticulum (ER) (Grp94), as well as from the cytosol of the eukaryotic cell (Hsp82). We find that the folding rates of Hsp82 and HtpG are similar, while the folding of Grp94 is slowed down by misfolding of the N-terminal domain. Furthermore, the domain interactions mediated by the charged linker, involved in the conformational cycles of all three orthologs, are much stronger for Grp94 than for Hsp82, keeping the N-terminal domain and the middle domain in close proximity. Thus, the ER resident Hsp90 ortholog differs from the cytosolic counterparts in basic functionally relevant structural properties.
Subject(s)
Cytosol/chemistry , Endoplasmic Reticulum/chemistry , Escherichia coli Proteins/chemistry , HSP70 Heat-Shock Proteins/chemistry , HSP90 Heat-Shock Proteins/chemistry , Membrane Proteins/chemistry , Saccharomyces cerevisiae Proteins/chemistry , Amino Acid Sequence , Animals , Cloning, Molecular , Crystallography, X-Ray , Cytosol/metabolism , Dogs , Endoplasmic Reticulum/metabolism , Escherichia coli/genetics , Escherichia coli/metabolism , Escherichia coli Proteins/genetics , Escherichia coli Proteins/metabolism , Gene Expression , Genetic Vectors/chemistry , Genetic Vectors/metabolism , HSP70 Heat-Shock Proteins/genetics , HSP70 Heat-Shock Proteins/metabolism , HSP90 Heat-Shock Proteins/genetics , HSP90 Heat-Shock Proteins/metabolism , Kinetics , Membrane Proteins/genetics , Membrane Proteins/metabolism , Optical Tweezers , Protein Conformation, alpha-Helical , Protein Conformation, beta-Strand , Protein Folding , Protein Interaction Domains and Motifs , Protein Multimerization , Recombinant Proteins/chemistry , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , Saccharomyces cerevisiae/genetics , Saccharomyces cerevisiae/metabolism , Saccharomyces cerevisiae Proteins/genetics , Saccharomyces cerevisiae Proteins/metabolism , Spectrum Analysis/methods , ThermodynamicsABSTRACT
In this issue, Voth et al. (2014) reveal that Get3, the GET pathway targeting factor shuttling TA-proteins from the ribosome to the ER membrane, moonlights as a chaperone under oxidizing conditions in a manner reminiscent of bacterial Hsp33.
Subject(s)
Adenosine Triphosphatases/physiology , Guanine Nucleotide Exchange Factors/physiology , Oxidative Stress , Saccharomyces cerevisiae Proteins/physiology , Saccharomyces cerevisiae/metabolismABSTRACT
Huntington disease (HD), a dominantly inherited neurodegenerative disorder caused by the expansion of a CAG-encoded polyglutamine (polyQ) repeat in huntingtin (Htt), displays a highly heterogeneous etiopathology and disease onset. Here, we show that the translation of expanded CAG repeats in mutant Htt exon 1 leads to a depletion of charged glutaminyl-transfer RNA (tRNA)(Gln-CUG) that pairs exclusively to the CAG codon. This results in translational frameshifting and the generation of various transframe-encoded species that differently modulate the conformational switch to nucleate fibrillization of the parental polyQ protein. Intriguingly, the frameshifting frequency varies strongly among different cell lines and is higher in cells with intrinsically lower concentrations of tRNA(Gln-CUG). The concentration of tRNA(Gln-CUG) also differs among different brain areas in the mouse. We propose that translational frameshifting may act as a significant disease modifier that contributes to the cell-selective neurotoxicity and disease course heterogeneity of HD on both cellular and individual levels.
