The effect of aggregation conditions on alpha-synuclein aggregation kinetics, fibril morphology and structural stability
MASTER project (published Januari 2019)
Parkinsons disease (PD), the second most frequently occurring neurodegenerative disease in the world, is mainly characterized by the loss of dopaminergic neurons and the presence of insoluble proteic complexes, so-called Lewy bodies (LB), in the brain . LB are largely composed of fibrillary aggregates (amyloid) of alpha-synuclein (aSyn) protein, which suggests that aSyn plays an important role in the pathogenesis of PD. In addition, aSyn gene mutations and duplication/triplication are associated with hereditary forms of PD .
The aSyn protein is small (14 kDa), intrinsically disordered, and in vivo mainly expressed in dopaminergic neurons in the brain . In vitro aggregation experiments often make use of recombinantly produced aSyn to produce amyloid fibrils at a variety of aggregation conditions. The aggregation of aSyn can lead to the formation of different polymorphs (fibrils with different morphology) depending on the aggregation condition used. An interesting fact is that the different polymorphs can, in vivo, induce different types of synucleopathies .
Description of the project:
The aim of this project is to make a systematic study of wild-type aSyn aggregation at different conditions (pH, salts, presence/absence of other proteins, etc.). This study will give mechanistical and structural information about the aSyn aggregation process. This type of information could be helpful for the comparison with aggregation of aSyn inside the brain or for the study of aggregation inhibitors.
The aggregation kinetics will be measured using Thioflavin T binding assays, the morphology of the formed fibrils will be analyzed using Atomic Force Microscopy and the structural stability of the fibrils will be characterized from proteinase K degradation patterns.
- In vitro aSyn aggregation reactions
- Thioflavin T fluorescence measurements
- Turbidity measurements
- Atomic force microscopy
- Circular Dichroism
Department: NanoBioPhysics (NBP) UTwente - https://www.utwente.nl/en/tnw/nbp/
Project leader: Prof.Dr. M.M.A.E. Claessens (Chair Nanobiophysics) - firstname.lastname@example.org