Quantifying the charge dependences of alpha-synuclein's membrane interactions.

Abstract

Alpha-synuclein (aSyn) is a neuronal protein whose malfunction is responsible for the disease onset of alpha-synucleinopathies, such as Parkinson's Disease, Lewy body dementia, and multiple system atrophy. As part of its normal physiological function, aSyn is known to interact with the phospholipid membranes of synaptic vesicles. However, the binding interactions of aSyn with membranes are not well understood quantitatively. Using a tryptophan spectrofluorometry assay in combination with a new derivation of a binding equation, we have examined aSyn's membrane affinity. The binding equation yields a novel parameter σ, which can be understood as a binding site density. The binding site density informs on the spatial frequency of binding events, which corresponds to aSyn's overall propensity to interact with a particular membrane composition. In existing literature, aSyn has been shown to engage strongly with negatively charged membrane phospholipids, and calcium ions in solution are suggested to have conflicting impacts on aSyn's affinity depending on membrane charge. We investigate how incrementing proportions of the negatively charged phosphatidylserine (PS) in lipid vesicles affects their aSyn binding site densities, by examining responses in the σ parameter. Tryptophan fluorescence and σ also allow us to quantify the dependence of calcium ions’ ability to effect aSyn binding on the amount of overall membrane charge. This work furthers our understanding of aSyn's physiological binding preferences with respect to membrane and solution charges. Deviations from these preferences may prime us to identify mechanistic links to alpha-synucleinopathies.