Year: 2016
Type: dissertation
Abstract: Amyloids are insoluble aggregates rich in β-structure that result from the self-assembly of polypeptides in a non-native structural fold. The deposition of these aggregates onto organs has been traditionally linked to many human diseases, however the discovery of functional amyloids in nature suggests there may also be beneficial roles for this self-assembly process. Pathogenic and functional amyloids both contain a cross-β backbone composed of β-sheets that run perpendicular to the fibril axis, but there lacks clear molecular details on the structural differences to explain their divergent biological behaviours. Solid-state NMR (SSNMR) has become an invaluable tool for structural determination of amyloids, as poor solubility of these assemblies make them challenging for traditional X-ray crystallography and solution NMR methods. This thesis aimed to identify key structural features between cytotoxic and non-cytotoxic amyloid fibrils using SSNMR, and to examine the effects of protein-lipid interactions on membrane disruption and amyloid formation that can reflect amyloid cytotoxicity. First, three amyloid fibrils formed from fragments of the human prion protein were examined, with the findings that a parallel β-sheet arrangement and packing of hydrophobic residues into the fibril core defined the basic structure in these assemblies. Next, these three peptide amyloids were sonicated to explore fragmentation-associated cytotoxicity of these amyloids. The results here highlighted the importance of solvent-exposed hydrophobicity on amyloid-membrane interaction, linking the increased surface hydrophobicity of fragmented fibrillar species to greater membrane disruption and cytotoxicity. Finally, SSNMR experiments were applied to characterize the structure of human apolipoprotein serum amyloid A when associated with high-density lipoprotein (HDL)-like particles and when misfolded into amyloid fibrils. The results led to a preliminary model that can explain functional differences between lipid-free and HDL-associated SAA. Together, these findings highlight the differing effects of lipid interaction on the pathogenicity of amyloid fibrils.
Source: TSpace (University of Toronto)
Author: Jason Yau
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: green
OpenAlex ID: W3088520593