α-Synuclein Has a High Affinity for Packing Defects in a Bilayer Membrane

Abstract

A number of neurodegenerative disorders, including Parkinson's disease, dementia with Lewy bodies, and multiple system atrophy, are characterized by the intracellular deposition of fibrillar aggregates that contain a high proportion of ␣-synuclein (␣S). The interaction with the membrane-water interface strongly modulates folding and aggregation of the protein. The present study investigates the lipid binding and the coil-helix transition of ␣S, using titration calorimetry, differential scanning calorimetry, and circular dichroism spectroscopy. Titration of the protein with small unilamellar vesicles composed of zwitterionic phospholipids below the chain melting temperature of the lipids yielded exceptionally large exothermic heat values. The sigmoidal titration curves were evaluated in terms of a simple model that assumes saturable binding sites at the vesicle surface. The cumulative heat release and the ellipticity were linearly correlated as a result of simultaneous binding and helix folding. There was no heat release and folding of ␣S in the presence of large unilamellar vesicles, indicating that a small radius of curvature is necessary for the ␣S-membrane interaction. The heat release and the negative heat capacity of the proteinvesicle interaction could not be attributed to the coilhelix transition of the protein alone. We speculate that binding and helix folding of ␣S depends on the presence of defect structures in the membrane-water interface, which in turn results in lipid ordering in the highly curved vesicular membranes. This will be discussed with regard to a possible role of the protein for the stabilization of synaptic vesicle membranes. ␣-Synuclein (␣S)1 is a protein of 140 amino acids that has been identified as a major component of the intracytoplasmic fibrillar deposits (Lewy bodies) associated with idiopathic and inherited forms of Parkinson's disease (1). The majority of cases are idiopathic, whereas mutations in the ␣S gene are known to be responsible for rare inherited, early onset variants of Parkinson's disease (2-4). Although the molecular mode of action of ␣S and of its homologs is as yet unknown, it was assumed that the protein modulates the dopamine neurotransmission by regulating synaptic vesicle (SV) mobilization from the presynaptic reserve pool (5, 6) or directly regulating the dopamine metabolism (7-9). However, attempts to identify a specific SV-binding protein, e.g. by protein cross-linking, have been unsuccessful so far.At a certain threshold concentration, ␣S tends to aggregate into amyloid fibrils (10), whereas the homolog ␤S, which lacks a stretch of amino acids within the central portion of ␣S, has a much lower fibrillization propensity (11, 12) and may even inhibit ␣S fibrillization (13-15), indicating that the hydrophobic central part of ␣S is essential for its fibrillar aggregation (16). A recent study on the structure of mature fibrillar aggregates, using site-directed spin labeling, indicates that the N terminus of ␣S is less ordered than the central portion and that t...

19F NMR studies of α‐synuclein‐membrane interactions

“…26,27 In that model, a-synuclein prefers SUV defects, which are affected by head group size, charge, and the hydrophobic thickness. The reduced binding of a-synuclein to DMPC/DMPG SUVs is due to the increase in the curvature that arises as a consequence of the acyl chain, which agree with the conclusion of Nuscher et al 45 As we observe, these properties affect a-synuclein-membrane interactions.…”

Anle138b and related compounds are aggregation specific fluorescence markers and reveal high affinity binding to α-synuclein aggregates

“…Expression and purification were performed as described previously [23]. Briefly, pET-5a/α-synuclein (136TAT) plasmid (wt-plasmid by Philipp Kahle, LMU Munich; 136-TAC/TAT-Mutation by Matthias Habeck) was used to transform Escherichia coli BL21(DE3)pLys (Novagen, Madison, WI, USA), and expression was induced with isopropyl ß-D-1-thiogalactopyranoside (IPTG, Peqlab, Erlangen, Germany).…”

α-Synuclein assembles into higher-order multimers upon membrane binding to promote SNARE complex formation

“…Moreover, α-synuclein is a major component of intracellular protein aggregates called Lewy bodies, which are pathological hallmarks of neurodegenerative disorders such as PD, Lewy body dementia, and multiple system atrophy (11-14). Strikingly, neurotoxic α-synuclein aggregates propagate between neurons during neurodegeneration, suggesting that such α-synuclein aggregates are not only intrinsically neurotoxic but also nucleate additional fibrillization (15-18).α-Synuclein is highly concentrated in presynaptic terminals where α-synuclein exists in an equilibrium between a soluble and a membrane-bound state, and is associated with synaptic vesicles (19)(20)(21)(22). The labile association of α-synuclein with membranes (23, 24) suggests that binding of α-synuclein to synaptic vesicles, and its dissociation from these vesicles, may regulate its physiological function.…”