α-Synuclein induces both positive mean curvature and negative Gaussian curvature in membranes

Abstract

Using a combination of X-ray scattering, fluorescence correlation spectroscopy, coarse-grained molecular dynamics (MD) simulations and potential of mean force calculations, we have explored the membrane remodeling effects of monomeric α-synuclein (αS). Our initial findings from multiple approaches are that αS (1) causes a significant thinning of the bilayer and (2) stabilizes positive mean curvature, such that the maximum principle curvature matches that of synaptic vesicles, αS-induced tubules, and the synthetic lipid vesicles to which the protein binds most tightly. This suggests that αS binding to synaptic vesicles likely stabilizes their intrinsic curvature. We then show that αS induces local negative Gaussian curvature, an effect that occurs in regions of αS shown previously via NMR and corroborated by MD simulation to have significant conformational flexibility. The induction of negative Gaussian curvature, which has implications for all curvature-sensing and curvature-generating amphipathic α-helices, supports a hypothesis that connects helix insertion to fusion and fission of vesicles, processes that have recently been linked to αS function. Then, in an effort to explain these biophysical properties of αS, we promote an intrinsic curvature-field model that recasts long-range protein-protein interactions in terms of the interactions between the local curvature fields generated by lipid-protein complexes.

Figure 1

Exploring bilayer thickness. (A) LAXS (expt) and model-fit (SDP) bilayer form factors for pure lipid (black) and αS (red) samples. (B) ΔD_PP surfaces plotted versus local (relative to αS) x and y axes for asymmetric CGMD simulations with the average reoriented protein position in black (the white star indicates the N-terminus, purple squares indicate the linker region, and green tilted squares indicate GLY67-GLY68). Color map units are nanometers.

Figure 2

αS stabilizes positive mean curvature. (A) Representative height surfaces, h(x,y) = z, for the outer leaflet of the asymmetric system at two time points (18.2 and 32.8 μs scaled simulation times, left and right columns, respectively) showing both the outer leaflet (top row) and the inner leaflet (bottom row). αS (black), with the N-terminus (white star) highlighting the protein orientation, is correlated with regions of positive surface fluctuation (hot color map). (B) Time- and protein-averaged height surface for the asymmetric system. Panels A and B color map units are nanometers. (C) Snapshot of the asymmetric system showing positive mean curvature with two αS proteins (black) positioned at the apex. Mean (κ_m) and Gaussian (κ_g) curvatures are determined from the principle curvatures (κ₁ and κ₂), where κ₁ is the maximum curvature and κ₂ is the minimum curvature, with κ₁ ⊥ κ₂. (D) Local κ₁ and corresponding vesicle diameter, D = 2/〈κ₁〉, for the asymmetric system (the white star indicates the N-terminus, purple squares indicate the linker region, and green tilted squares indicate GLY67–GLY68).

Figure 3

αS induces negative Gaussian curvature. Time-averaged, reoriented 〈κ_g〉 determined from the continuous helix asymmetric system. Two regions of known increased flexibility, the linker region for the horseshoe helix conformation (purple) and GLY67–GLY68 (green), border a region of stabilized negative 〈κ_g〉. Color map scale units are inverse square nanometers.