N-Alpha-Acetylation of α-Synuclein Increases Its Helical Folding Propensity, GM1 Binding Specificity and Resistance to Aggregation

A switch in the conformational properties of alpha-synuclein (alpha S) is hypothesized to be a key step in the pathogenic mechanism of Parkinson's disease (PD). Whereas the beta-sheet-rich state of alpha S has long been associated with its pathological aggregation in PD, a partially alpha-helical state was found to be related to physiological lipid binding; this suggests a potential role of the alpha-helical state in controlling synaptic vesicle cycling and resistance to beta-sheet rich aggregation. N-terminal acetylation is the predominant post-translational modification of mammalian alpha S. Using circular dichroism, isothermal titration calorimetry, and fluorescence spectroscopy, we have analyzed the effects of N-terminal acetylation on the propensity of recombinant human alpha S to form the two conformational states in interaction with lipid membranes. Small unilamellar vesicles of negatively charged lipids served as model membranes. Consistent with previous NMR studies using phosphatidylserine, we found that membrane-induced alpha-helical folding was enhanced by N-terminal acetylation and that greater exothermic heat could be measured upon vesicle binding of the modified protein. Interestingly, the folding and lipid binding enhancements with phosphatidylserine in vitro were weak when compared to that of alpha S with GM1, a lipid enriched in presynaptic membranes. The resultant increase in helical folding propensity of N-acetylated alpha S enhanced its resistance to aggregation. Our findings demonstrate the significance of the extreme N-terminus for folding nucleation, for relative GM1 specificity of alpha S-membrane interaction, and for a protective function of N-terminal-acetylation against alpha S aggregation mediated by GM1.