Aggregation Kinetics and Characteristics of Truncated α-synuclein Variants 1-119 and 72-140 Related to Parkinson’s Disease A biophysical investigation of truncated forms of the Parkinson’s protein α-synuclein using kinetic assays, atomic force microscopy, and cell viability assays

Abstract

Parkinson’s disease is a neurodegenerative disorder associated with the pathological aggregation of the protein α-synuclein (α-syn) into amyloid fibrils. While extensive research has focused on the structure and aggregation kinetics of full length α-syn (FL), recent findings suggest that truncated variants are common in the brain and may exhibit distinct aggregation behaviors and structural properties. This master thesis aims to investigate the α-syn truncations 1-119 and 72-140 and compare them to the FL protein. Specifically, the study examines their aggregation kinetics, the morphologies and secondary structure of the resulting aggregates and finally their cytotoxic effects on human neuroblastoma SH-SY5Y cells. The results show that there are distinct differences in aggregation behaviors of these truncated variants compared to FL α-syn. The 72-140 variant aggregates rapidly but does not form fibrils, instead it produces large globular oligomers which lack β-sheet structure. These aggregates are not cytotoxic to SH-SY5Y cells, possibly due to the absence of an ordered fibrillar structure and/or their size. Although 72–140 does not appear to co-form fibrils with other variants, it was found to have some catalytic effect on their aggregation. The 1-119 variant aggregates faster than FL α-syn. The kinetic curves are characterized by a reduced lag time and they form shorter fibrils, suggesting that enhanced nucleation may explain the faster aggregation rate. The 1-119 fibrils are toxic to cells, but less toxic than FL fibrils. When co-aggregated with FL or 72-140 the aggregation kinetics are predominantly governed by the 1-119 variant. FL and 1-119 do not appear to co-form fibrils, but rather form fibrils independently while influencing each other’s aggregation kinetics. These results furthers the understanding of truncated α-syn variants and their possible involvement in Parkinson’s disease initiation and progression. This project also creates a base for further research in this area, especially for further investigations of 72-140 and other C-terminal fragments.