Amyloid Proteins in Neurodegenerative Disease – Role of Extrinsic Modifiers

Abstract: ABSTRACT Self-assembly of disease-associated proteins into fibrillar homopolymers, so-called “amyloid fibrils” is a pathological hallmark of several debilitating human disorders, including Alzheimer’s disease (AD) and Parkinson’s disease (PD). AD and PD are associated with the formation of amyloid fibrils from the proteins amyloid-β (Aβ) and α-syn (α-syn), in the extracellular and intracellular space, respectively. In Aβ pathogenesis, there are different molecules such as metal ions and lipids that can interact with Aβ, and affect amyloid fibril formation but also other important pathological features such as cellular uptake, which contributes to the intracellular build-up of aggregates that precede the deposition of extracellular plaques. This thesis describes my research on the effects of different extrinsic modulators, mainly metals and lipids, in Aβ and α-syn aggregation, as well as in the cellular uptake of Aβ. I showed that Cu2+ inhibits Aβ(1-42) amyloid formation by impairment of the fibril elongation mechanism whereas Cu+ catalysed primary nucleation, indicating an important role of copper redox chemistry in AD. I further showed that Cu2+, and also Zn2+, whilst acting aggregation inhibitory, enhance Aβ(1-42) uptake and thereby contribute to other pathological effects, including prion-like cell-to-cell propagation. In a second part of my work, I explored the effect of lipid vesicles of biological and synthetic origin on the Aβ(1-42) aggregation process. I found that cell-derived extracellular vesicles impede the elongation process and induce the formation of amyloid fibrils with distinct morphology. Synthetic lipid vesicles, on the other hand, were found to have diverse effects on the Aβ(1-42) aggregation rate and mechanism and I showed that both lipid chemistry and the physical properties of the bilayers they participate in, contribute to the modulatory role of membrane in Aβ(1-42) aggregation. To analyse physical attributes of amyloid fibrils, I developed a nanofluidic-based method for visualization of single amyloid fibrils in solution and used it to compare persistence lengths of different fibril types as well as within samples, addressing the concept of polymorphism. Lastly, my work has contributed to the understanding of how graphene-based nanoparticles can modulate amyloid formation by interfering with both primary nucleation and secondary processes in aggregation. Keywords: Alzheimer’s disease, protein aggregation, amyloid fibril, amyloid-β, Aβ(1-42), α-synuclein, extracellular vesicles, lipid vesicle, kinetics, nanofluidics, fluorescence microscopy, atomic force microscopy