Structure and stability of amyloid fibrils studied by advanced vibrational spectroscopy

Protein misfolding often leads to the formation of refractory protein aggregates like amyloid fibrils. These fibrils possess a highly ordered structure and are implicated in over 25 severe diseases including Alzheimer's, Parkinson's and prion diseases. This work was focused on understanding the morphology and conformation of amyloid fibrils and their stability after formation. The deconstruction of fibrils as well as other aggregates like inclusion bodies under mild conditions was also investigated using Archaeal chaperones.;The process of spontaneous refolding is reported in HET-s (218-289) prion fibrils, which indicates the refolding of amyloid fibrils to be a general phenomenon. The mechanism of refolding was investigated and found to be associated with pH-dependent deprotonation of key amino acid residues along the fibril surface. Hence, a refolding mechanism is proposed for the first time. This is important because the ability to control fibril conformation can enable control of their biological activity. In addition, direct evidence has been obtained on the ability to probe two different levels of fibril chirality for the first time. Surface probe microscopy probes surface level chirality while vibrational circular dichroism probes filament level chirality. This is important because change in fibril chirality characterizes the refolding phenomenon and it shows that both techniques are required to understand the chirality of macromolecular assemblies.;This work also reports the ability of archaeal chaperones to deconstruct protein aggregates such as inclusion bodies and amyloid fibrils. The aggregates were deconstructed in terms of morphology, and their characteristic high ?-sheet content was found to be melted upon treatment with archaeal chaperones. This is important because protein aggregation poses a major problem in recombinant protein production and neurodegenerative diseases.;In summary, original contribution has been made through this work on amyloid fibril refolding, its mechanism and the ability of various techniques to reveal complementary information on this process, as well as on the use of archaeal chaperones to deconstruct inclusion bodies and amyloid fibrils.