Molecular mechanisms used by chaperones to reduce the toxicity of aberrant protein oligomers

Abstract

Chaperones are the primary regulators of the proteostasis network and are known to facilitate protein folding, inhibit protein aggregation, and promote disaggregation and clearance of misfolded aggregates inside cells. We have tested the effects of five chaperones on the toxicity of misfolded oligomers preformed from three different proteins added extracellularly to cultured cells. All the chaperones were found to decrease oligomer toxicity significantly, even at very low chaperone/protein molar ratios, provided that they were added extracellularly rather than being overexpressed in the cytosol. Infrared spectroscopy and site-directed labeling experiments using pyrene ruled out structural reorganizations within the discrete oligomers. Rather, confocal microscopy, SDS-PAGE, and intrinsic fluorescence measurements indicated tight binding between oligomers and chaperones. Moreover, atomic force microscopy imaging indicated that larger assemblies of oligomers are formed in the presence of the chaperones. This suggests that the chaperones bind to the oligomers and promote their assembly into larger species, with consequent shielding of the reactive surfaces and a decrease in their diffusional mobility. Overall, the data indicate a generic ability of chaperones to neutralize extracellular misfolded oligomers efficiently and reveal that further assembly of protein oligomers into larger species can be an effective strategy to neutralize such extracellular species.protein homeostasis | protein misfolding | protein aggregates | amyloid | extracellular chaperones

Rational design of potent domain antibody inhibitors of amyloid fibril assembly

“…We next sought to determine how substoichiometric concentrations of inhibitory gammabodies (1:10 gammabody:Aβ molar ratio) render excess Aβ in a state that is incompetent for amyloid formation. Interestingly, some chaperones, aromatic small molecules, and peptides with antiaggregation activity have also been shown to completely prevent amyloid formation at low substoichiometric concentrations (≤1:10 inhibitor: monomer molar ratios) by converting monomers into unstructured, nonamyloid complexes (11)(12)(13)(14)(15)(16)(17). Thus, we posited that gammabodies convert Aβ fibrillar intermediates and monomers into similar complexes that are incompetent for amyloid formation.…”

Antiapoptotic Properties of α-Crystallin–Derived Peptide Chaperones and Characterization of Their Uptake Transporters in Human RPE Cells

“…Neutralization of the extracellular misfolded oligomers and suppression of oligomer toxicity by sHSPs, including aBcrystallin, were reported in a recent study, thus providing a valuable strategy for therapeutic interventions against diseases stemming from extracellular protein aggregation. 48 We have shown recently that the a-crystallin chaperone counteracts the cataractogenic effect of a-crystallin mutants. 49 Future work is likely to offer valuable insight into the beneficial, therapeutic use of a-crystallin minichaperones in the prevention of retinal diseases involving oxidative stress such as AMD.…”

Extracellular Chaperones and Proteostasis

“…In the context of amyloid formation, ECs interact most strongly with oligomers formed early in the aggregation pathway (50,51,86), probably via the exposed hydrophobic residues thought to responsible for cellular toxicity. Indeed, it is likely that this is a common mechanism by which a range of ECs, such as clusterin, α 2 M, haptoglobin and ApoE protect cells from misfolded or aggregated proteins (51,173,174). Recent insights into the mechanism of these interactions have come from studies exploiting advanced microscopy techniques.…”