Alpha Synuclein is a cytosolic neuronal protein associated with a group of neurodegenerative diseases, called synucleinopathies. A feature of synucleinopathies is the presence of Lewy bodies, which are intracellular inclusions composed mainly by alpha synuclein aggregates complexed with iron. Alpha synuclein is able to bind cellular membranes, and these interactions are considered critical for at least some normal functions of the protein, e.g. promoting protein aggregation and toxicity. Recent findings suggest that oligomers represent the main toxic aggregate species but little is known about the mechanisms that induce aggregation and about the onset of the pathology. Another feature of synucleinopathies is the amount of brain iron accumulation which correlates with disease stage and severity.
We correlated iron content with alpha synuclein propensity to form aggregates and to interact with biological membranes. By atomic force microscopy, we investigated the role of iron (II) in the interaction of the wild-type and the mutated A53T alpha synuclein species with artificial lipid bilayers composed by dioleoylglycerophosphocholine (DOPC), sphingomyelin and cholesterol, mimicking lipid raft domains. We tested monomeric species of the protein in presence and absence of ferrous chloride salt.
Co-incubation of the mutated A53T monomer with iron on the membrane, leads to the formation of globular protein aggregates which preferentially interact with lipid rafts (liquid-ordered phase), assembling ring-shape structures of 0.2-1 μm in diameter. On the contrary, the monomer in absence of metal interacts primary with the DOPC (liquid-disordered phase) leading to the formation of defects without any aggregation. Similar results are obtained with the wild-type form, which forms protein clusters in presence of iron, but the effect seems to be less strong compared to the mutated species.
In absence of iron, the wild-type monomer has different behaviour compared to the mutated form, interacting with both lipid phases. The interaction of the protein seems stronger on lipid raft domains, which appear to have pore-like defects and some aggregation structures.
Our work showed that iron has a huge effect in promoting aggregation of alpha synuclein on artificial membranes, highlighting some of the molecular mechanisms which are at the basis of neurodegenerative diseases development.