Several observations have implicated oxidative stress and aggregation of the presynaptic protein α-synuclein in the pathogenesis of Parkinson disease. α-Synuclein has been shown to have affinity for unsaturated fatty acids and membranes enriched in polyunsaturated fatty acids, which are especially sensitive to oxidation under conditions of oxidative stress. One of the most important products of lipid oxidation is 4-hydroxy-2-nonenal (HNE), which has been implicated in the pathogenesis of Parkinson disease. Consequently, we investigated the effects of the interaction of HNE with α-synuclein. Incubation of HNE with α-synuclein at pH 7.4 and 37 °C resulted in covalent modification of the protein, with up to six HNE molecules incorporated as Michael addition products. Fourier transform infrared and CD spectra indicated that HNE modification of α-synuclein resulted in a major conformational change involving increased β-sheet. HNE modification of α-synuclein led to inhibition of fibrillation in an HNE concentration-dependent manner. This inhibition of fibrillation was shown to be due to the formation of soluble oligomers based on size exclusion high pressure liquid chromatography and atomic force microscope data. Small angle x-ray scattering analysis indicated that the HNE-induced oligomers were compact and tightly packed. Treatment with guanidinium chloride demonstrated that the HNE-induced oligomers were very stable with an extremely slow rate of dissociation. Addition of 5 μm HNE-modified oligomers to primary mesencephalic cultures caused marked neurotoxicity because the integrity of dopaminergic and GABAergic neurons was reduced by 95 and 85%, respectively. Our observations indicate that HNE modification of α-synuclein prevents fibrillation but may result in toxic oligomers, which could therefore contribute to the demise of neurons subjected to oxidative damage.
This research was supported by grant NS39985 from the National Institutes of Health. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
