Chemical Chaperones Modulate the Formation of Metabolite Assemblies

Abstract

The formation of amyloid-like structures by metabolites is associated with several inborn errors of metabolism (IEMs). These structures display most of the biological, chemical and physical properties of protein amyloids. However, the molecular interactions underlying the assembly remain elusive, and so far, no modulating therapeutic agents are available for clinical use. Chemical chaperones are known to inhibit protein and peptide amyloid formation and stabilize misfolded enzymes. Here, we provide an in-depth characterization of the inhibitory effect of osmolytes and hydrophobic chemical chaperones on metabolite assemblies, thus extending their functional repertoire. We applied a combined in vivo-in vitro-in silico approach and show their ability to inhibit metabolite amyloid-induced toxicity and reduce cellular amyloid content in yeast. We further used various biophysical techniques demonstrating direct inhibition of adenine self-assembly and alteration of fibril morphology by chemical chaperones. Using a scaffold-based approach, we analyzed the physiochemical properties of various dimethyl sulfoxide derivatives and their role in inhibiting metabolite self-assembly. Lastly, we employed whole-atom molecular dynamics simulations to elucidate the role of hydrogen bonds in osmolyte inhibition. Our results imply a dual mode of action of chemical chaperones as IEMs therapeutics, that could be implemented in the rational design of novel lead-like molecules.

The Metabolostasis Network and the Cellular Depository of Aggregation‐Prone Metabolites

“…Yet, even under such extreme conditions, modulation of the metabolite assemblies could be achieved by the administration of compounds that compete with the hydrophobic interactions that lead to the formation of supramolecular assemblies. [15,78] While these inhibitors reduced the aggregate content and improved cell viability, the overall intracellular adenine content remained unaffected thus reinforcing the notion that the toxicity results from metabolite aggregation, rather than the mere accumulation of monomeric metabolite molecules. [77] However, further research is required to elucidate the cellular pathways that are affected by metabolostasis collapse and the cellular consequences of metabolite aggregation, which could serve as novel drug targets for IEMs.…”

“…Such conditions represent extreme cases in which the concentrations of metabolites can be orders of magnitude higher than those found under physiological conditions. Yet, even under such extreme conditions, modulation of the metabolite assemblies could be achieved by the administration of compounds that compete with the hydrophobic interactions that lead to the formation of supramolecular assemblies [15, 78] . While these inhibitors reduced the aggregate content and improved cell viability, the overall intracellular adenine content remained unaffected thus reinforcing the notion that the toxicity results from metabolite aggregation, rather than the mere accumulation of monomeric metabolite molecules [77] .…”

“…[9][10][11] The cytotoxic effect of the assemblies formed by the physiological enantiomer of phenylalanine (L-Phe) could be modulated by various small molecules including the D-enantiomer of phenylalanine (D-Phe), [12] doxycycline, [13] various polyphenols such as epigallocatechin gallate (EGCG) and tannic acid, [14] and chemical chaperons. [15] Later studies demonstrated that these phenomena are not unique to phenylalanine but are shared among metabolites in general. Proteinogenic and non-proteinogenic amino acids, nucleobases, various metabolic intermediates, and even sphingolipids were shown to form ordered amyloid-like assemblies with structural and biological properties similar to those observed for phenylalanine structures.…”

“…Such arrangements may provide a mechanistic explanation for the high toxicity of the assemblies, possibly mediated by membrane destabilization and demyelination [9–11] . The cytotoxic effect of the assemblies formed by the physiological enantiomer of phenylalanine (L‐Phe) could be modulated by various small molecules including the D‐enantiomer of phenylalanine (D‐Phe), [12] doxycycline, [13] various polyphenols such as epigallocatechin gallate (EGCG) and tannic acid, [14] and chemical chaperons [15] …”

Engineered Riboswitch Nanocarriers as a Possible Disease-Modifying Treatment for Metabolic Disorders

“…To provide further support for this hypothesis, we used the ProteoStat dye that was previously shown to detect metabolite amyloid-like structures in vitro and in vivo. 46,47 The staining was followed by flow cytometry analysis, which allowed the detection of a decrease in the aggregation in the presence of the riboswitch-VLPs (Figure 5e,f) as well as by confocal microscopy for the identification of an intracellular localization of the aggregates (Figure 5g). The stained dots were not detected in the presence of the riboswitch-VLPs.…”