| Many neurological diseases are linked to the aberrant folding of proteins and the production of amyloid fibrils. The misfolding and aggregation of the protein alpha-synuclein into Lewy bodies is thought to play a key role in the pathogenesis of Parkinson's disease. Despite extensive studies on this protein, very little is known about the structures and molecular mechanisms associated with the aggregation events. This work unravels detailed structural data about the aggregation pathways of alpha-synuclein, using an array of spectroscopic techniques, including circular dichroism, Raman spectroscopy, atomic force microscopy, nuclear magnetic resonance and thioflavin-T fluorescence.; Previous studies established that alpha-synuclein is predominantly unfolded in aqueous solution at neutral pH. My work revealed that under these conditions, alpha-synuclein adopts an ensemble of secondary structures (extended, alpha-helical and beta-sheet). This interpretation was based on the analysis of the Raman amide I band of the monomeric alpha-synuclein.; The morphology and the secondary structure of the soluble protein aggregates (spheroidal and chainlike oligomers) observed as probable intermediates that form during alpha-synuclein fibrillization, were characterized by Raman and atomic force microscopies. During the formation of alpha-synuclein oligomers under physiological conditions or in methanol-water solutions, a significant change in the ensemble of structures present took place; beta-sheet structure was enhanced and the extended structure was diminished. Importantly, these aggregates retained a significant amount of alpha-helical secondary structure. Moderate (15--20%) concentrations of methanol encouraged alpha-synuclein aggregation into chainlike oligomers that were indistinguishable from those formed in the absence of methanol.; In many sporadic neurodegenerative disorders, co-occurrence of alpha-synuclein and tau proteins as amyloid-like fibrils occurs, and it is thought that either protein may nucleate fibril deposition at the early stages that ultimately lead to the disease pathology. Despite the importance of these processes, the precise molecular changes and interactions have yet to be determined. In the last part of this work, we reveal by NMR that the major attractive forces between alpha-synuclein and tau are electrostatic and the interaction is localized in the C-terminal region (residues 119--140) of alpha-synuclein and the microtubule binding domain of tau protein. |
