Characterization of Pairs of Toxic and Nontoxic Misfolded Protein Oligomers Elucidates the Structural Determinants of Oligomer Toxicity in Protein Misfolding Diseases

Abstract

The aberrant misfolding and aggregation of peptides and proteins into amyloid aggregates occurs in over 50 largely incurable protein misfolding diseases. These pathologies include Alzheimer's and Parkinson's diseases, which are global medical emergencies owing to their prevalence in increasingly aging populations worldwide. Although the presence of mature amyloid aggregates is a hallmark of such neurodegenerative diseases, misfolded protein oligomers are increasingly recognized as of central importance in the pathogenesis of many of these maladies. These oligomers are small, diffusible species that can form as intermediates in the amyloid fibril formation process or be released by mature fibrils after they are formed. They have been closely associated with the induction of neuronal dysfunction and cell death. It has proven rather challenging to study these oligomeric species because of their short lifetimes, low concentrations, extensive structural heterogeneity, and challenges associated with producing stable, homogeneous, and reproducible populations. Despite these difficulties, investigators have developed protocols to produce kinetically, chemically, or structurally stabilized homogeneous populations of protein misfolded oligomers from several amyloidogenic peptides and proteins at experimentally ameneable concentrations. Furthermore, procedures have been established to produce morphologically similar but structurally distinct oligomers from the same protein sequence that are either toxic or nontoxic to cells. These tools offer unique opportunities to identify and investigate the structural determinants of oligomer toxicity by a close comparative inspection of their structures and the mechanisms of action through which they cause cell dysfunction.This Account reviews multidisciplinary results, including from our own groups, obtained by combining chemistry, physics, biochemistry, cell biology, and animal models for pairs of toxic and nontoxic oligomers. We describe oligomers comprised of the amyloid-β peptide, which underlie Alzheimer's disease, and α-synuclein, which are associated with Parkinson's disease and other related neurodegenerative pathologies, collectively known as synucleinopathies. Furthermore, we also discuss oligomers formed by the 91-residue N-terminal domain of [NiFe]-hydrogenase maturation factor from E. coli, which we use as a model non-disease-related protein, and by an amyloid stretch of Sup35 prion protein from yeast. These oligomeric pairs have become highly useful experimental tools for studying the molecular determinants of toxicity characteristic of protein misfolding diseases. Key properties have been identified that differentiate toxic from nontoxic oligomers in their ability to induce cellular dysfunction. These characteristics include solvent-exposed hydrophobic regions, interactions with membranes, insertion into lipid bilayers, and disruption of plasma membrane integrity. By using these properties, it has been possible to rationalize in model systems the responses to pairs of toxic and nontoxic oligomers. Collectively, these studies provide guidance for the development of efficacious therapeutic strategies to target rationally the cytotoxicity of misfolded protein oligomers in neurodegenerative conditions.

Figure 1

Illustration of the four pairs of misfolded protein oligomers described in this Account. (A) Generic schematic for oligomer formation. The solution conditions listed for HypF-N (B), Aβ₄₂ (C), αSyn (D), and Sup35 NM (E) lead to the formation of toxic and nontoxic oligomers that facilitate the interrogation of the physico-chemical origins of oligomer cytotoxicity. Ca²⁺ influx images of cells were adapted with permission from ref (1). Copyright 2010 Springer Nature. Created with biorender.com. OC+ and A11+ notes indicate oligomers that react with these specific antibodies. OC– and A11– notes indicate oligomers that do not react with them. Notes with specific compounds (e.g., Amphotericin B) indicate oligomers formed in their presence.

Figure 2

Physico-chemical factors regulating oligomer toxicity discussed in this Account. The cell binding mechanism refers to toxic type B* and nontoxic type A* oligomers of αSyn, but is also representative of other oligomer pairs with shared features. Oligomer models (left) are adapted with permission from ref (35). Copyright 2018 Royal Society of Chemistry. Oligomer–membrane models (middle) are adapted with permission from ref (3). Copyright 2017 American Association for the Advancement of Science. Created with biorender.com.