alpha-synuclein and dimyristoylphosphatidylglycerol

Parkinson's disease is an incurable neurodegenerative disorder caused by the aggregation of α-synuclein (AS). This amyloid protein contains a 12-residue-long segment, AS

Topics: alpha-Synuclein; Anions; Circular Dichroism; Hydrogen-Ion Concentration; Macromolecular Substances; Phosphatidylglycerols; Protein Aggregates; Protein Structure, Secondary; Vibration

Alpha-synuclein (aSN) is a presynaptic protein with a pathological role in Parkinson's disease (PD). The mutants A30P, E46K and A53T are involved in PD early-onset forms. aSN is natively unfolded but can self-assemble to oligomers and fibrils and binds anionic membranes in a helical conformation. We study the influence of wild-type (wt) aSN and familial variants on the chain order and thermotropic phase behavior of anionic dimyristoylphosphatidylglycerol (DMPG) bilayers by using electron spin resonance and calorimetry, respectively. The alpha-helical conformation of the proteins in the membrane-bound state is assessed by circular dichroism thermal scans. wt and mutated aSN upon binding to fluid DMPG vesicles progressively increase chain order. Lipid:protein molar binding stoichiometries correspond to 50 for A30P, 35-36 for aSN and A53T, 30 for E46K. The temperature range over which the variants assume the α-helical fold correlates directly with the density of proteins on vesicle surfaces. All variants preserve the characteristic chain flexibility gradient and impart motional restriction in the lipid chain. This is evident at the first CH2 segments and is markedly reduced at the chain termini, disappearing completely for A30P. The proteins slightly reduce DMPG main transition temperature, revealing preferential affinity for the fluid phase, and broaden the transition, promoting gel-fluid phase coexistence. The overall results are consistent with protein surface association in which the degree of binding correlates with the degree of folding and perturbation of the membrane bilayer. However, the degree of binding of monomer to membrane does not correlate directly with aSN toxicity in vivo.

Topics: alpha-Synuclein; Amino Acid Substitution; Gene Expression; Humans; Lipid Bilayers; Mutation; Phase Transition; Phosphatidylcholines; Phosphatidylglycerols; Protein Binding; Protein Conformation, alpha-Helical; Protein Conformation, beta-Strand; Protein Folding; Protein Interaction Domains and Motifs; Recombinant Proteins; Solutions; Structure-Activity Relationship; Thermodynamics

Alpha-synuclein is a small presynaptic protein, which is linked to the development of Parkinson's disease. Alpha-synuclein partitions between cytosolic and vesicle-bound states, where membrane binding is accompanied by the formation of an amphipathic helix in the N-terminal section of the otherwise unstructured protein. The impact on alpha-synuclein of binding to vesicle-like liposomes has been studied extensively, but far less is known about the impact of alpha-synuclein on the membrane. The interactions of alpha-synuclein with phosphatidylglycerol membranes are studied here by using spin-labeled lipid species and electron spin resonance (ESR) spectroscopy to allow a detailed analysis of the effect on the membrane lipids. Membrane association of alpha-synuclein perturbs the ESR spectra of spin-labeled lipids in bilayers of phosphatidylglycerol but not of phosphatidylcholine. The interaction is inhibited at high ionic strength. The segmental motion is hindered at all positions of spin labeling in the phosphatidylglycerol sn-2 chain, while still preserving the chain flexibility gradient characteristic of fluid phospholipid membranes. Direct motional restriction of the lipid chains, resulting from penetration of the protein into the hydrophobic interior of the membrane, is not observed. Saturation occurs at a protein/lipid ratio corresponding to approximately 36 lipids/protein added. Alpha-synuclein exhibits a selectivity of interaction with different phospholipid spin labels when bound to phosphatidylglycerol membranes in the following order: stearic acid > cardiolipin > phosphatidylcholine > phosphatidylglycerol approximately phosphatidylethanolamine > phosphatidic acid approximately phosphatidylserine > N-acyl phosphatidylethanolamine > diglyceride. Accordingly, membrane-bound alpha-synuclein associates at the interfacial region of the bilayer where it may favor a local concentration of certain phospholipids.

Topics: alpha-Synuclein; Electron Spin Resonance Spectroscopy; Humans; Hydrophobic and Hydrophilic Interactions; Lipid Bilayers; Models, Chemical; Nerve Tissue Proteins; Phosphatidylcholines; Phosphatidylglycerols; Phosphoproteins; Protein Binding; Recombinant Proteins; Spin Labels; Static Electricity; Synucleins; Titrimetry