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The Route from the Folded to the Amyloid State: Exploring the Potential Energy Surface of a Drug-Like Miniprotein.

ABSTRACT: The amyloid formation of the folded segment of a variant of Exenatide (a marketed drug for type-2 diabetes mellitus) was studied by electronic circular dichroism (ECD) and NMR spectroscopy. We found that the optimum temperature for E5 protein amyloidosis coincides with body temperature and requires well below physiological salt concentration. Decomposition of the ECD spectra and its barycentric representation on the folded-unfolded-amyloid potential energy surface allowed us to monitor the full range of molecular transformation of amyloidogenesis. We identified points of no return (e.g.; T=37?°C, pH?4.1, cE5 =250??m, cNaCl =50?mm, t>4-6?h) that will inevitably gravitate into the amyloid state. The strong B-type far ultraviolet (FUV)-ECD spectra and an unexpectedly strong near ultraviolet (NUV)-ECD signal (??275-285 ? nm ) indicate that the amyloid phase of E5 is built from monomers of quasi-elongated backbone structure (??-145°, ??+145°) with strong interstrand Tyr?Trp interaction. Misfolded intermediates and the buildup of "toxic" early-stage oligomers leading to self-association were identified and monitored as a function of time. Results indicate that the amyloid transition is triggered by subtle misfolding of the ?-helix, exposing aromatic and hydrophobic side chains that may provide the first centers for an intermolecular reorganization. These initial clusters provide the spatial closeness and sufficient time for a transition to the ?-structured amyloid nucleus, thus the process follows a nucleated growth mechanism.

SUBMITTER: Taricska N

PROVIDER: S-EPMC7028080 | biostudies-literature | 2020 Feb

REPOSITORIES: biostudies-literature

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The Route from the Folded to the Amyloid State: Exploring the Potential Energy Surface of a Drug-Like Miniprotein.

Taricska Nóra N Horváth Dániel D Menyhárd Dóra K DK Ákontz-Kiss Hanna H Noji Masahiro M So Masatomo M Goto Yuji Y Fujiwara Toshimichi T Perczel András A

Chemistry (Weinheim an der Bergstrasse, Germany) 20191227 9

The amyloid formation of the folded segment of a variant of Exenatide (a marketed drug for type-2 diabetes mellitus) was studied by electronic circular dichroism (ECD) and NMR spectroscopy. We found that the optimum temperature for E5 protein amyloidosis coincides with body temperature and requires well below physiological salt concentration. Decomposition of the ECD spectra and its barycentric representation on the folded-unfolded-amyloid potential energy surface allowed us to monitor the full ...[more]

PMID: 31647140

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Project description:Recent in vitro and in vivo studies suggest that destabilized proteins with defective folding induce aggregation and toxicity in protein-misfolding diseases. One such unstable protein state is called amyloid oligomer, a precursor of fully aggregated forms of amyloid. Detection of various amyloid oligomers with A11, an anti-amyloid oligomer conformation-specific antibody, revealed that the amyloid oligomer represents a generic conformation and suggested that toxic beta-aggregation processes possess a common mechanism. By using A11 antibody as a probe in combination with mass spectrometric analysis, we identified GroEL in bacterial lysates as a protein that may potentially have an amyloid oligomer conformation. Surprisingly, A11 reacted not only with purified GroEL but also with several purified heat shock proteins, including human Hsp27, 40, 70, 90; yeast Hsp104; and bovine Hsc70. The native folds of A11-reactive proteins in purified samples were characterized by their anti-beta-aggregation activity in terms of both functionality and in contrast to the beta-aggregation promoting activity of misfolded pathogenic amyloid oligomers. The conformation-dependent binding of A11 with natively folded Hsp27 was supported by the concurrent loss of A11 reactivity and anti-beta-aggregation activity of heat-treated Hsp27 samples. Moreover, we observed consistent anti-beta-aggregation activity not only by chaperones containing an amyloid oligomer conformation but also by several A11-immunoreactive non-chaperone proteins. From these results, we suggest that the amyloid oligomer conformation is present in a group of natively folded proteins. The inhibitory effects of A11 antibody on both GroEL/ES-assisted luciferase refolding and Hsp70-mediated decelerated nucleation of Abeta aggregation suggested that the A11-binding sites on these chaperones might be functionally important. Finally, we employed a computational approach to uncover possible A11-binding sites on these targets. Since the beta-sheet edge was a common structural motif having the most similar physicochemical properties in the A11-reactive proteins we analyzed, we propose that the beta-sheet edge in some natively folded amyloid oligomers is designed positively to prevent beta aggregation.

Dataset Information

The Route from the Folded to the Amyloid State: Exploring the Potential Energy Surface of a Drug-Like Miniprotein.

Publications

The Route from the Folded to the Amyloid State: Exploring the Potential Energy Surface of a Drug-Like Miniprotein.

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