Amyloidogenic peptides or proteins self-assemble to form oligomers and fibrils in many neurodegenerative diseases such as Alzheimer’s disease, Parkinson’s disease, and type-2 diabetes. Fibrils formed by amyloidogenic peptides or proteins are observed as plaques in patients, and the build-up of these plaques are the hallmark of amyloid diseases. The amyloid fibrils were believed to be the cause of neurodegenerative diseases because they were observed in patients’ brains. Amyloid fibrils are stable, and their structures are extensively characterized over the years through solid-state NMR and cryogenic electron microscopy.
In the last few decades, the intermediate form, soluble amyloid oligomers, have emerged as the neurotoxic form of amyloids rather than amyloid fibrils. Amyloid oligomers are short-lived and inherently heterogeneous, forming various species of many different sizes and morphologies The heterogeneous nature of amyloid oligomers makes them difficult to elucidate their structures by common structural techniques such as NMR, X-ray crystallography, and cryo-EM. While not much is known about amyloid oligomers, many studies have found that amyloid oligomers are comprised of amyloid monomers adapting an antiparallel β-hairpin conformation.
Effort to gain insights into structures of elusive amyloid oligomers, the Nowick laboratory has developed a chemical model system which mimics β-hairpin of amyloid peptides or proteins. In the last few years, structural studies of these model systems have shed light on how amyloid oligomers may assemble in disease pathology. Among many amyloid peptides and proteins, amyloid beta (Aβ) is the most intensively studied amyloid peptide and is central to the pathology of Alzheimer’s disease. Thus, advances in amyloid research rely on improved access to Aβ. In the first two chapters of my thesis, I describe an efficient method of expression and purification of Aβ(M1-42) and its mutants, by combining protein expression and peptide purification method, as well as of efficient expression and labeling N-terminal cysteine Aβ with fluorophores or biotin using maleimide conjugation reagents. In the Chapters 3 and 4, I describe my effort in studying amyloid oligomers by stabilizing β-hairpin region of Aβ and by a chemical model system derived from transthyretin. Lastly, in the epilogue, I describe my collaborative efforts during my time in the Nowick laboratory.