Stabilization of α-Synuclein Secondary Structure upon Binding to Synthetic Membranes

Abstract

␣-Synuclein is a highly conserved presynaptic protein of unknown function. A mutation in the protein has been causally linked to Parkinson's disease in humans, and the normal protein is an abundant component of the intraneuronal inclusions (Lewy bodies) characteristic of the disease. ␣-Synuclein is also the precursor to an intrinsic component of extracellular plaques in Alzheimer's disease. The ␣-synuclein sequence is largely composed of degenerate 11-residue repeats reminiscent of the amphipathic ␣-helical domains of the exchangeable apolipoproteins. We hypothesized that ␣-synuclein should associate with phospholipid bilayers and that this lipid association should stabilize an ␣-helical secondary structure in the protein. We report that ␣-synuclein binds to small unilamellar phospholipid vesicles containing acidic phospholipids, but not to vesicles with a net neutral charge. We further show that the protein associates preferentially with vesicles of smaller diameter (20 -25 nm) as opposed to larger (ϳ125 nm) vesicles. Lipid binding is accompanied by an increase in ␣-helicity from 3% to approximately 80%. These observations are consistent with a role in vesicle function at the presynaptic terminal.

19F NMR studies of α‐synuclein‐membrane interactions

“…28 Our data [ Fig. 2(B)] are consistent both with this conclusion and with conclusions based on 15 N NMR data, which show that the N-terminal region of the protein binds SDS while the C-terminal region remains disordered. 13,20,24,28,31,32 Solution NMR is a powerful tool for accessing processes that occur over a range of timescales.…”

“…[8][9][10][11][12] The protein binds lipids and anionic detergents through the seven imperfect, cationic, 11-amino acid repeats located in its N-terminal and hydrophobic regions. [6][7][8][13][14][15][16][17][18][19][20] Electron paramagnetic resonance (EPR) data on its complex with small unilamellar vesicles (SUVs) suggest that the first $100 residues of the monomeric protein adopt an a-helical conformation that lies on the membrane surface. [21][22][23][24][25] The last $40 residues lack defined structure and do not appear to be involved in membrane interactions.…”

“…13,21 Recent solution NMR data, however, appear incompatible with this model. More specifically, 15 N intensity and relaxation data from titration of SUVs sugget there exists several binding modes in which the first 25 residues adopt a helical state that anchor the interaction with SUVs. 26,27 A variety of other techniques, including circular dichroism spectropolarimetry, 12,13,19,28 fluorescence spectroscopy, 21,[29][30][31] and EPR [21][22][23][24][25] have been used to study how the phospholipid composition of vesicles affects a-synuclein affinity.…”

Acyl-CoA Synthetase Activity Links Wild-Type but Not Mutant α-Synuclein to Brain Arachidonate Metabolism

“…In astrocytes lacking α-synuclein palmitic (16:0) and arachidonic (20:4n-6) acid uptake and metabolism is depressed through an unknown mechanism (11). Lack of α-synuclein in brain depresses 16:0 uptake and significantly alters its metabolism (29), although the impact on brain 20:4n-6 uptake and metabolism is unknown.In addition, α-synuclein modulates phospholipase C and D activities in vitro (22,(31)(32)(33), suggesting a role in lipid-mediated signal transduction. Studies in vivo also demonstrate that α-synuclein affects enzymes involved in lipid metabolism, because α-synuclein overexpression leads to phospholipase D inhibition in yeast (34) and down regulates expression of phospholipase A 2 and of long chain fatty acid CoA synthetase in Drosophila (35).…”

“…In addition, α-synuclein modulates phospholipase C and D activities in vitro (22,(31)(32)(33), suggesting a role in lipid-mediated signal transduction. Studies in vivo also demonstrate that α-synuclein affects enzymes involved in lipid metabolism, because α-synuclein overexpression leads to phospholipase D inhibition in yeast (34) and down regulates expression of phospholipase A 2 and of long chain fatty acid CoA synthetase in Drosophila (35).…”