Aggregation and conformation of alpha-synuclein: effects of ligand binding and phosphomimetics

Parkinson’s disease (PD) affects 1 in 500 of the UK population. A critical step in disease aetiology is the formation of Lewy bodies (LBs), deposits of aggregated alpha-synuclein (αsyn) as amyloid inclusions, within surviving neurons. It is well established that αsyn within LBs has undergone a variety of post-translational modifications, including S129 phosphorylation, and LBs are known to be rich in metals such as copper. It has however, yet to be established how phosphorylation of αsyn and metal ions influence in the pathology of PD, and if the formation of LBs can be prevented by the use of small molecule inhibitors. Electrospray ionisation-ion mobility spectrometry- mass spectrometry (ESI-IMSMS) was used to investigate the conformational changes to compact states of αsyn known to be linked to amyloid formation, for WT and two αsyn mutants mimicking phosphorylation at S87 and S129, in the presence of copper, and their aggregation was monitored by thioflavin-T assays. Results demonstrate that the conformational state of αsyn can be modulated by interactions with copper, causing an increase in the compact state, leading to an increased rate of aggregation. S129D αsyn showed the highest affinity for copper, and the fastest aggregation rates overall. SH-SY5Y cell culture models were used to investigate intracellular aggregation and phosphorylation state upon copper exposure. Increased copper concentration correlated with increased formation of unmodified and phosphorylated intracellular aggregates, an increase in apoptosis, and decreased cell viability. ESI-IMS-MS demonstrated curcumin and its derivatives were able to disrupt amyloid assembly, via prevention of autofragmentation and dissociation of low-order oligomers, thus preventing the aggregation of αsyn. In SH-SY5Y model used however, curcumins were unable to prevent the metal-induced intracellular aggregation of αsyn. Together, results support the hypothesis that phosphorylation has a key role in PD progression, and demonstrated that modified curcumins may be a potential therapeutic for PD.