Disordered Motifs and Domains in Cell Control - October 11-15, 2014

Disordered Motifs and Domains in Cell Control

Poster Session I

3-POS Board 3 α-Synuclein D 2 : The Disorderly Disordered Parkinson’s Disease Amyloid Ashley S. Phillips 1 , Alexandre F. Gomes 2 , Rebecca Beveridge 1 , Jonas Gasparavicious 3 , Jay E. Gillam 3 , Fabio C. Gozzo 2 , Cait E. MacPhee 3 , Tilo Kunath 4 , Perdita E. Barran 1 . 1 University of Manchester, Manchester, United Kingdom, 2 State University of Campinas, São Paulo, Brazil, 3 University of Edinburgh, Edinburgh, United Kingdom, 4 MRC Centre for Regenerative Medicine, University of Edinburgh, Edinburgh, United Kingdom. Parkinson’s disease is the 2nd most common neurodegenerative disorder worldwide, affecting approximately 2% of the world population over 65. Current UK spending is ~£3.3billion and with those aged over 65 predicted to double by 2050, a treatment is quickly becoming of paramount importance. α-Synuclein is an amyloidogenic intrinsically disordered protein implicated in Parkinson’s disease aetiology, whose potential functions range from enzyme regulation to synaptic vesicle release. α-Synuclein co-exists in conformations ranging from disordered monomers to β-sheet rich fibrils. Our aim is to unravel the structures of the potentially druggable early aggregation species using multiple gas phase techniques, to inform a drug discovery process whose disordered subject limits the use of traditional techniques. Ion Mobility Mass Spectrometry (IMMS) is a hybrid mass spectrometry based gas phase electrophoretic technique which generates a rotationally averaged Collision Cross Section (CCS) for each conformer. Native Mass Spectrometry (MS) and IMMS have the ability to study the structure and conformational dynamics of complex protein samples. Under gentle Nano Electrospray Ionisation conditions (nESI) α-Synuclein presents ions, over a wide charge state range in positive and negative mode, characteristic of its intrinsically disordered nature. The CCS of α-Synuclein monomers range from ~1000Å 2 to ~3000Å 2 , again demonstrating its conformational flexibility. nESI also reveals small aggregate populations up to pentamer. MS and IMMS highlight the conformational plasticity of α-Synuclein, including its susceptibility to solution condition modification, genre-defining levels of day-to-day variation and the lack of a solution phase conformational response following in vitro aggregation. Crosslinking-IMMS has also been used to directly sample the abundance solution phase conformers. Our results are compared to established biophysical techniques. These results highlight the advantages of mass spectrometry-based approaches in determining transient structural forms and their application to studying the aggregation of amyloidogenic proteins.

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