Fast Flow Microfluidics and Single-Molecule Fluorescence for the Rapid Characterization of α-Synuclein Oligomers
Dear, Alexander J
Garcia, Gonzalo A
Serra, Mauro Dalla
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Horrocks, M., Tosatto, L., Dear, A. J., Garcia, G. A., Iljina, M., Cremades Casasin, N., Serra, M. D., et al. (2015). Fast Flow Microfluidics and Single-Molecule Fluorescence for the Rapid Characterization of α-Synuclein Oligomers. Analytical Chemistry, 87 8818-8826. https://doi.org/10.1021/acs.analchem.5b01811
[Image - see article] α-Synuclein oligomers can be toxic to cells and may be responsible for cell death in Parkinson’s disease. Their typically low abundance and highly heterogeneous nature, however, make such species challenging to study using traditional biochemical techniques. By combining fast-flow microfluidics with single-molecule fluorescence, we are able to rapidly follow the process by which oligomers of αS are formed and to characterize the species themselves. We have used the technique to show that populations of oligomers with different FRET efficiencies have varying stabilities when diluted into low ionic strength solutions. Interestingly, we have found that oligomers formed early in the aggregation pathway have electrostatic repulsions that are shielded in the high ionic strength buffer and therefore dissociate when diluted into lower ionic strength solutions. This property can be used to isolate different structural groups of αS oligomers and can help to rationalize some aspects of αS amyloid fibril formation.
M.H.H. thanks the Royal Society of Chemistry (Analytical Chemistry Trust Fund) for his studentship. L.T. has been the recipient of a grant PAT Post Doc Outgoing 2009 – 7th Framework Program Marie Curie COFUND actions. A.J.D. is funded by the Schiff Foundation.
External DOI: https://doi.org/10.1021/acs.analchem.5b01811
This record's URL: https://www.repository.cam.ac.uk/handle/1810/250289