Abstract
α-Synuclein (aS) is a protein abundant in presynaptic nerve terminals in Parkinson disease (PD) and is a major component of intracellular Lewy bodies, the pathological hallmark of neurodegenerative disorders such as PD. Accordingly, the relationships between aS structure, its interaction with lipids, and its involvement in neurodegeneration have attracted great interest. Previously, we reported on the interaction of aS with brain polyunsaturated fatty acids, in particular docosahexaenoic acid (DHA). aS acquires an α-helical secondary structure in the presence of DHA and, in turn, affects DHA structural and aggregative properties. Moreover, aS forms a covalent adduct with DHA. Here, we provide evidence that His-50 is the main site of this covalent modification. To better understand the role of His-50, we analyzed the effect of DHA on aS-derived species: a naturally occurring variant, H50Q; an oxidized aS in which all methionines are sulfoxides (aS4ox); a fully lysine-alkylated aS (acetyl-aS); and aS fibrils, testing their ability to be chemically modified by DHA. We show, by mass spectrometry and spectroscopic techniques, that H50Q and aS4ox are modified by DHA, whereas acetyl-aS is not. We correlated this modification with aS structural features, and we suggest a possible functional role of aS in sequestering the early peroxidation products of fatty acids, thereby reducing the level of highly reactive lipid species. Finally, we show that fibrillar aS loses almost 80% of its scavenging activity, thus lacking a potentially protective function. Our findings linking aS scavenging activity with brain lipid composition suggest a possible etiological mechanism in some neurodegenerative disorders.
Highlights
␣-Synuclein is a protein abundant in presynaptic nerve terminals in Parkinson disease (PD) and is a major component of intracellular Lewy bodies, the pathological hallmark of neurodegenerative disorders such as PD
To better understand the role of His-50, we analyzed the effect of docosahexaenoic acid (DHA) on aS-derived species: a naturally occurring variant, H50Q; an oxidized aS in which all methionines are sulfoxides; a fully lysine-alkylated aS; and aS fibrils, testing their ability to be chemically modified by DHA
Previous reports documented the ability of aS to interact with brain polyunsaturated fatty acids (PUFAs), which are critical for neuronal membranes, fluidity, and permeability, serving as an energy reservoir, and taking part in intra- and extracellular signaling as second messengers [15]
Summary
DHA and AA samples were incubated at pH 7 and 37 °C under shaking up to 24 h, in the absence and presence of aS (P/lipid molar ratio 1:50). It presents a specific pattern of several signals [37]: the most peculiar are those at mass/charge values of 329.2, 351.2, and 367.2 corresponding to a protonated molecular DHA (monoisotopic mass 328.2 Da), an adduct with sodium (328.2 Da ϩ 22), and an oxidized form of this adduct (328.2 Da ϩ 22 ϩ 16). After 24 h of incubation (Fig. 2B), the signals at m/z 329.2, 351.2, and 367.2, characteristics of DHA, disappear and new species are evident in the spectrum at m/z values of 381.3, 397.2, and 415.2 corresponding to hydroperoxide derivatives of DHA, which generate during the propagation phase of the fatty acids autoxidation (Scheme 1). Sodium adduct of DHA Sodium adduct of hydroxy DHA Sodium adduct of peroxy radical DHA or Sodium adduct of dialkoxy radical DHA Sodium adduct of alkoxy-peroxy diradical DHA Sodium adduct of dihydroperoxy DHA Arachidonic acid (monoisotopic mass 304.2) Sodium adduct of AA Sodium adduct of hydroxy AA Di-sodium adduct of AA Sodium adduct of dihydroperoxy AA dehydrated Sodium adduct of dihydroperoxy AA Di-sodium adduct of dihydroperoxy AA
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