The misfolding and amyloid formation of α-synuclein, a 140 amino acid neuronal protein, is central in Parkinson’s disease etiology. Critical to discovering the relationship of α-synuclein to disease is the understanding of how environmental factors modulate the protein folding landscape. Membranes are of particular interest as they alter α-synuclein conformation and can both stimulate and inhibit amyloidogenesis <i>in vitro</i>. In order to examine structures that promote fibrillation we used time-resolved Förster resonance energy transfer (trFRET) measurements, which afford a nanosecond snapshot of polypeptide conformations. Using an intramolecular dansyl/rhodamine pair incorporated at positions 40 and 90, respectively, trFRET results reveal a heterogeneous collection of compact and extended subpopulations. Distinct distance distributions of α-synuclein have been obtained in its soluble, fibrillar, and membrane-bound forms at both acidic and neutral pH. At acidic pH, fibrillation is significantly accelerated and a larger proportion of compact population is observed at pH 5.5 than at pH 7. Importantly, trFRET has detected membrane-bound states at aggregation promoting lipid concentrations which are not apparent from other conformation-sensitive techniques such as circular dichroism and site-specific Trp fluorescence spectroscopy. Moreover, both compact (~27-35Å) and extended (≥70 Å) structures are present on the lipid membrane, hinting at the formation of aggregation prone structures on the membrane surface. By monitoring trFRET as a function of aggregation time, mechanistic insights will be gained into the α-synuclein amyloid formation process in the presence of phospholipid membranes.

Examining the folding landscape of α-synuclein using time-resolved FRET

249th National Meeting (2015)

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