title: Fast flow microfluidics and single-molecule fluorescence for the rapid characterization of α-synuclein oligomers.
abstract: 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.
authors.affiliation: Department of Chemistry, University of Cambridge , Lensfield Road, Cambridge CB2 1EW, U.K.
authors.forename: Mathew H
authors.initials: MH
authors.lastname: Horrocks
authors.affiliation: Department of Chemistry, University of Cambridge , Lensfield Road, Cambridge CB2 1EW, U.K.|Istituto di Biofisica, CNR, U.O., Trento, Trentino, Italy.
authors.forename: Laura
authors.initials: L
authors.lastname: Tosatto
authors.affiliation: Department of Chemistry, University of Cambridge , Lensfield Road, Cambridge CB2 1EW, U.K.
authors.forename: Alexander J
authors.initials: AJ
authors.lastname: Dear
authors.affiliation: Department of Chemistry, University of Cambridge , Lensfield Road, Cambridge CB2 1EW, U.K.
authors.forename: Gonzalo A
authors.initials: GA
authors.lastname: Garcia
authors.affiliation: Department of Chemistry, University of Cambridge , Lensfield Road, Cambridge CB2 1EW, U.K.
authors.forename: Marija
authors.initials: M
authors.lastname: Iljina
authors.affiliation: Department of Chemistry, University of Cambridge , Lensfield Road, Cambridge CB2 1EW, U.K.
authors.forename: Nunilo
authors.initials: N
authors.lastname: Cremades
authors.affiliation: Istituto di Biofisica, CNR, U.O., Trento, Trentino, Italy.
authors.forename: Mauro
authors.initials: M
authors.lastname: Dalla Serra
authors.affiliation: Department of Chemistry, University of Cambridge , Lensfield Road, Cambridge CB2 1EW, U.K.
authors.forename: Tuomas P J
authors.initials: TP
authors.lastname: Knowles
authors.affiliation: Department of Chemistry, University of Cambridge , Lensfield Road, Cambridge CB2 1EW, U.K.
authors.forename: Christopher M
authors.initials: CM
authors.lastname: Dobson
authors.affiliation: Department of Chemistry, University of Cambridge , Lensfield Road, Cambridge CB2 1EW, U.K.
authors.forename: David
authors.initials: D
authors.lastname: Klenerman
mesh_terms: D005453:Fluorescence; D031541:Fluorescence Resonance Energy Transfer; D007834:Lasers; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation; D055672:Static Electricity; D051844:alpha-Synuclein / Q000032:analysis*
chemical_list: D051844:alpha-Synuclein