Investigating the impact of Cu on α-synuclein aggregation: A single-molecule approach.

Parkinson's disease (PD) is a prevalent neurodegenerative disorder characterized by the abnormal aggregation of α-synuclein. This study investigated the impact of Cu ions on α-synuclein aggregation in oligomer distribution using two single-molecule techniques. The aggregation of α-synuclein monomers with and without Cu revealed that Cu accelerated the formation of ThT-positive β-sheet structured aggregates. Nanopipettes of varying diameters from to 7 to 134 nm were employed to characterize the oligomers formed during the lag phase, demonstrating that Cu generated a wider range of oligomers from 10 nm to 20,000 nm over time. Confocal fluorescence spectroscopy analysis of ThT-positive fibrils in the plateau phase showed that Cu induces larger oligomers and fewer in number. The introduction of preformed seeds to the control and Cu-containing samples further accelerated the aggregation. The combination of seeds and Cu produced structurally distinct oligomers, with seeds catalyzing the formation of small oligomers that detach from the parent fibers and Cu, promoting the formation of larger oligomers. These results reveals that seeds and Cu act synergistically, with two different aggregation pathways coexisting in the early phase, leading to an intermediate composition of fibers and clusters at the end of aggregation.