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Title: Folding, dimerisation and the interaction with Aβ₁₋₄₀ of human cystatin C
Author: Keeley, Emma
ISNI:       0000 0001 3596 2486
Awarding Body: University of Sheffield
Current Institution: University of Sheffield
Date of Award: 2008
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Amyloid formation is a predominant feature of many human diseases including Alzheimers' disease, Parkinsons' disease, type II diabetes and Creutzfeldt-Jacob disease. The process of amyloidogenesis involves the self-assembly of soluble protein into insoluble fibrous material. Amyloidogenic proteins, which share no common sequence, structure or function, form amyloid fibres which have a common morphology. Neither a detailed structure of mature amyloid or the mechanism by which it forms is fully understood. The work presented in this thesis uses the cystatins as a model system for probing the mechanism of amyloid formation. The formation of amyloid requires a refolding event as the protein involved refolds from its native structure to the cross-P structure common to amyloid. Characterisation of the folding pathway of human cystatin C indicates that it folds via a partially folded kinetic intermediate, in an analogous manner to chicken cystatin. Analysis of the early stages of amyloidogenesis in cystatins indicates domain-swapped dimers are the building block of cystatin amyloid. As is observed with chicken cystatin, the dimerisation of human cystatin C is a bimolecular process, m-value analysis indicates that the structure of the dimerisation transition state is very close to the structure of the unfolded state and is more unfolded than the kinetic intermediate identified in the human cystatin C folding pathway. No terameric species are observed in the amyloidogenesis of human cystatin C, supporting evidence that tetramers are an off-pathway intermediate in the amyloidogenesis of chicken cystatin. Following the formation of dimer, isomerisation of the proline conserved across the cystatins is required prior to the formation of amyloid fibres. A preliminary study of the interaction between human cystatin C and Aβ shows that there is no interaction between monomeric hCC and monomeric Aβ 1-40. Given that hCC has been shown to inhibit Aβ amyloid formation, hCC must interact with one of the oligomeric species of Aβ that is populated during amyloidogenesis. Further experimentation is required to determine the exact nature of the interaction between hCC and Aβ.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available