α-synuclein's N-terminal region regulates amyloid fibril formation and liquid-liquid phase separation
Author(s)
Thrush, Rebecca
Type
Thesis
Abstract
Dementia and related neurodegenerative conditions pose a substantial and urgent challenge in today’s rapidly ageing population. Such diseases frequently exhibit a common pathological
feature: the aberrant accumulation of fibrillar protein aggregates in the nervous system, termed amyloids. In Parkinson’s disease, these amyloid fibrils are composed of α-synuclein, an
intrinsically disordered protein that is heavily post-translationally modified in vivo, including truncation at its N-terminus. Recently, α-synuclein has also been found to self-assemble
following liquid-liquid phase separation into immiscible liquid droplets called condensates, which can eventually mature into amyloids.
This thesis aimed to investigate the impact of N-terminal truncation on the amyloid and liquid-liquid phase separation aggregation pathways of α-synuclein. Employing a novel technique to
purify native N-terminally truncated variants, i.e., those lacking a starting methionine residue, this study first explored the effects of such modification on the amyloid aggregation of α-synuclein. The overall aggregation was characterised, before individual microscopic steps in the pathway were dissected. The research identified specific N-terminal residues crucial for interactions between α-synuclein monomers and pre-formed fibrils. It also highlighted N-terminal residues that are essential for fibril stability and the formation of mature amyloid fibrils.
This comprehensive analysis provided insight into the significance of the N-terminus in the aggregation of α-synuclein.
This thesis proceeded to explore the liquid-liquid phase separation induced aggregation of α-synuclein, and the involvement of the N-terminus in this pathway. An innovative technique was
developed using differential interference contrast imaging to study and quantify the formation of liquid-like condensates, and their liquid-to-solid transition. The investigation revealed that
the N-terminus influences, but is not critical for, the formation of liquid-like condensates. It was revealed that N-terminal truncation slows condensate growth and leads to an accelerated
liquid-to-solid transition, suggesting a condensate surface-dependent nucleation model for LLPS-induced α-synuclein aggregation.
feature: the aberrant accumulation of fibrillar protein aggregates in the nervous system, termed amyloids. In Parkinson’s disease, these amyloid fibrils are composed of α-synuclein, an
intrinsically disordered protein that is heavily post-translationally modified in vivo, including truncation at its N-terminus. Recently, α-synuclein has also been found to self-assemble
following liquid-liquid phase separation into immiscible liquid droplets called condensates, which can eventually mature into amyloids.
This thesis aimed to investigate the impact of N-terminal truncation on the amyloid and liquid-liquid phase separation aggregation pathways of α-synuclein. Employing a novel technique to
purify native N-terminally truncated variants, i.e., those lacking a starting methionine residue, this study first explored the effects of such modification on the amyloid aggregation of α-synuclein. The overall aggregation was characterised, before individual microscopic steps in the pathway were dissected. The research identified specific N-terminal residues crucial for interactions between α-synuclein monomers and pre-formed fibrils. It also highlighted N-terminal residues that are essential for fibril stability and the formation of mature amyloid fibrils.
This comprehensive analysis provided insight into the significance of the N-terminus in the aggregation of α-synuclein.
This thesis proceeded to explore the liquid-liquid phase separation induced aggregation of α-synuclein, and the involvement of the N-terminus in this pathway. An innovative technique was
developed using differential interference contrast imaging to study and quantify the formation of liquid-like condensates, and their liquid-to-solid transition. The investigation revealed that
the N-terminus influences, but is not critical for, the formation of liquid-like condensates. It was revealed that N-terminal truncation slows condensate growth and leads to an accelerated
liquid-to-solid transition, suggesting a condensate surface-dependent nucleation model for LLPS-induced α-synuclein aggregation.
Version
Open Access
Date Issued
2023-12-22
Date Awarded
2024-04-01
Copyright Statement
Creative Commons Attribution NonCommercial Licence
License URL
Advisor
Aprile, Francesco A.
Sponsor
Imperial College London
Publisher Department
Chemistry
Publisher Institution
Imperial College London
Qualification Level
Doctoral
Qualification Name
Doctor of Philosophy (PhD)