Methionine oxidation accelerates the aggregation and enhances the neurotoxicity of the D178N variant of the human prion protein.

Boya Feng, Zonglin Wang, Ting Liu, Rui Jin, Shaobo Wang, Wei Wang, Gengfu Xiao, Zheng Zhou

Index: Biochim. Biophys. Acta 1842(12 Pt A) , 2345-56, (2014)

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Abstract

The D178N mutation of the prion protein (PrP) results in the hereditary prion disease fatal familial insomnia (FFI). Little is known regarding the effects of methionine oxidation on the pathogenesis of D178N-associated FFI. In the present study, we found that the D178N variant was more susceptible to oxidation than wild-type PrP, as indicated by reverse-phase high performance liquid chromatography (RP-HPLC) and mass spectrometry (MS) analysis. Circular dichroism (CD), differential scanning calorimetry (DSC), thioflavin T (ThT) binding assay studies demonstrated that methionine oxidation decreased the structural stability of the D178N variant, and the oxidized D178N variant exhibited a greater propensity to form β-sheet-rich oligomers and aggregates. Moreover, these aggregates of oxidized D178N PrP were more resistant to proteinase K (PK) digestion. Additionally, using fluorescence confocal microscopy, we detected a high degree of aggregation in D178N-transfected Neuro-2a (N2a) cells after treatment with hydrogen peroxide (H2O2). Furthermore, the oxidation and consequent aggregation of the D178N variant induced greater apoptosis of N2a cells, as monitored using flow cytometry. Collectively, these observations suggest that methionine oxidation accelerates the aggregation and enhances the neurotoxicity of the D178N variant, possibly providing direct evidence to link the pathogenesis of D178N-associated FFI with methionine oxidation. Copyright © 2014 Elsevier B.V. All rights reserved.