Abstract
Dysregulation of the biosynthesis of cholesterol and other lipids has been implicated in various neurological diseases, including Parkinson’s disease, where the misfolding of membrane-associated α-Synuclein is a key molecular event. A mounting body of evidence suggests that α-Synuclein aggregation is strongly influenced by the lipid environment. The exact molecular mechanisms responsible for cholesterols effect on α-Synuclein binding to lipids and how this binding may affect α-Synuclein oligomerization and fibrillation remain elusive, as does the relative importance of cholesterol versus other lipid factors. We probed the interactions and fibrillation behaviour of α-Synuclein using styrene-maleic acid polymer nanodiscs, containing zwitterionic and anionic fluid lipid model systems with and without cholesterol. SPR and ThT fluorescence assays were then employed to monitor α-Synuclein binding, as well as fibrillation in the absence and presence of membrane models. 1H-15N correlated NMR was used to monitor the fold changes of α-Synuclein in response to nanodisc binding, and we determined individual residue apparent affinities for the lipid bilayers contained in the nanodiscs. Cholesterol inhibited α-Synuclein interaction with lipid bilayers. We also find that cholesterol, when present in the nanodiscs, significantly promotes α-Synuclein fibrillation, with more than a 20-fold reduction of lag-times before fibrillation onset. When α-Synuclein interaction was analysed for individual residues by solution-state NMR, we observed two different effects of cholesterol on α-Synuclein-membrane interaction. In nanodiscs made of DOPC, cholesterol modulated the NAC part of α-Synuclein, leading to stronger interaction of this region with the lipid bilayer. In contrast, in the nanodiscs comprising DOPC, DOPE and DOPG, the NAC part was mostly unaffected by cholesterol, while the binding of the N-terminal and the C-terminal are both inhibited.
Footnotes
Abbreviations α-Syn – α-Synuclein; AD – Alzheimer’s disease; DOPC – 1,2-dioleoyl-sn-glycero-3-phosphocholine; DOPE – 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine; DOPG – 1,2-dioleoyl-sn-glycero-3-phospho-(1’-rac-glycerol); DOSY – Diffusion-ordered spectroscopy; NAC – Non-Amyloid-Component; NMR – Nuclear magnetic resonance; PD – Parkinson’s disease; PM – plasma membrane; SMA – styrene-maleic acid; SPR – surface plasmon resonance; ThT – thioflavin T; TPE-TPP – tetraphenylethene tethered with triphenylphosphonium
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