Anti-amyloid compounds inhibit alpha-synuclein aggregation induced by Protein Misfolding Cyclic Amplification (PMCA). [Protein Structure and Folding]

February 28th, 2014 by Herva, M. E., Zibaee, S., Fraser, G., Barker, R. A., Goedert, M., Spillantini, M. G.

Filaments made of alpha-synuclein form the characteristic Lewy pathology in Parkinson′s and other diseases. The formation of alpha-synuclein filaments can be reproduced in vitro by incubation of recombinant protein, but the filament growth is very slow and highly variable and so unsuitable for fast high throughput anti-aggregation drug screening. To overcome this obstacle we have investigated whether the protein misfolding cyclic amplification (PMCA) technique, used for fast amplification of prion protein aggregates, could be adapted for growing alpha-synuclein aggregates and thus suitable for screening of drugs to affect alpha-synuclein aggregation for the treatment of the yet incurable alpha-synucleinopathies. Circular dichroism, electron microscopy, native and SDS-PAGE gels were used to demonstrate alpha-synuclein aggregate formation by PMCA and the strain imprint of the alpha-synuclein fibrils was studied by proteinase K digestion. We also demonstrated that alpha-synuclein fibrils are able to seed new alpha-synuclein PMCA reactions and to enter and aggregate in cells in culture. In particular, we have generated a line of ″chronically infected″ cells, which transmit alpha-synuclein aggregates even after multiple passages. To evaluate the sensitivity of the PMCA system as an alpha-synuclein anti-aggregating drug screening assay a panel of 10 drugs was tested. Anti-amyloid compounds proved efficient in inhibiting alpha-synuclein fibril formation induced by PMCA. Our results show that alpha-synuclein PMCA is a fast and reproducible system that could be used as a high throughput screening method for finding new alpha-synuclein anti-aggregating compounds.
  • Posted in Journal of Biological Chemistry, Publications
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