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Facile stop codon method reveals elevated neuronal toxicity by discrete S87p-α-synuclein oligomers
Facile "stop codon" method reveals elevated neuronal toxicity by discrete S87p-alpha-synuclein oligomers

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Issue Date
2014-01
Publisher
Academic Press
Citation
Biochemical and Biophysical Research Communications, Vol.443 No.3, pp.1085-1091
Abstract
Herein, a new method for preparing phosphorylated proteins at specific sites has been applied to alpha-synuclein (alpha-Syn). Three different alpha-Syn species phosphorylated at Serine 87 (S87p-alpha-Syn), Serine 129 (S129p-alpha-Syn) and Serine 87/129 (S87p,129p-alpha-Syn) were prepared through the 'stop codon' method and verified by LC/MS/MS and immunoblotting. Each type of phosphorylated alpha-Syn was tested for oligomerization trends and cellular toxicity with dopamine (DA), Cu2+ ions and pyridoxal 5'-phosphate. Aggregation trends induced by DA or DA/Cu2+ were similar between phosphorylated and non-phosphorylated alpha-Syn in SDS-PAGE. However, except for the monomer, phosphorylated oligomers showed higher toxicity than the non-phosphorylated alpha-Syn (Np-alpha-Syn) oligomers via WST-1 assays when tested on SH-SY5Y human neuroblastoma cells. In particular, S87p-alpha-Syn and S87p,129p-alpha-Syn oligomers induced by DA/Cu2+, showed higher toxicity than did S129p-alpha-Syn. When alpha-Syn was treated with pyridoxal 5'-phosphate in the presence of DA or Cu2+ to determine aggregation effects, high inhibition effects were shown in both non-phosphorylated and phosphorylated versions. alpha-Syn co-incubated with DA or DA/Cu2+ showed less cellular toxicity upon pyridoxal 5'-phosphate treatment, especially in the case of DA-induced Np-alpha-Syn. This study supports that phosphorylated oligomers of alpha-Syn at residue 87 can contribute to neuronal toxicity and the pyridoxal 5'-phosphate can be used as an inhibitor for alpha-Syn aggregation. (C) 2013 Elsevier Inc. All rights reserved.
ISSN
0006-291X
URI
https://hdl.handle.net/10371/177949
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College of Medicine/School of Medicine (의과대학/대학원)Dept. of Medicine (의학과)Journal Papers (저널논문_의학과)
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