Biophysical properties and cellular toxicity of covalent crosslinked oligomers of α-synuclein formed by photoinduced side-chain tyrosyl radicals
Alpha-synuclein (αS), a 140 amino acid presynaptic protein, is the major component of the fibrillar aggregates (Lewy bodies) observed in dopaminergic neurons of patients affected by Parkinson's disease. It is currently believed that noncovalent oligomeric forms of αS, arising as intermediates i...
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paper:paper_08915849_v53_n4_p1004_Borsarelli2023-06-08T15:47:13Z Biophysical properties and cellular toxicity of covalent crosslinked oligomers of α-synuclein formed by photoinduced side-chain tyrosyl radicals Fauerbach, Jonathan Arturo Jares, Elizabeth Andrea Neurodegeneration Oxidative stress Parkinson's disease Photocrosslinking Protein aggregation Protein photooxidation SH-SY5Y alpha synuclein oligomer tris(2,2' bipyridine)ruthenium tyrosine anion exchange chromatography article biophysics controlled study cytotoxicity fluorescence human human cell light absorption light scattering oligomerization photosensitization polyacrylamide gel electrophoresis priority journal steady state Western blotting alpha-Synuclein Ammonium Sulfate Amyloid Cell Line Cell Survival Cross-Linking Reagents Free Radicals Humans Kinetics Organometallic Compounds Oxidative Stress Photochemical Processes Photosensitizing Agents Protein Stability Tyrosine Alpha-synuclein (αS), a 140 amino acid presynaptic protein, is the major component of the fibrillar aggregates (Lewy bodies) observed in dopaminergic neurons of patients affected by Parkinson's disease. It is currently believed that noncovalent oligomeric forms of αS, arising as intermediates in its aggregation, may constitute the major neurotoxic species. However, attempts to isolate and characterize such oligomers in vitro, and even more so in living cells, have been hampered by their transient nature, low concentration, polymorphism, and inherent instability. In this work, we describe the preparation and characterization of low molecular weight covalently bound oligomeric species of αS obtained by crosslinking via tyrosyl radicals generated by blue-light photosensitization of the metal coordination complex ruthenium (II) tris-bipyridine in the presence of ammonium persulfate. Numerous analytical techniques were used to characterize the αS oligomers: biochemical (anion-exchange chromatography, SDS-PAGE, and Western blotting); spectroscopic (optical: UV/Vis absorption, steady state, dynamic fluorescence, and dynamic light scattering); mass spectrometry; and electrochemical. Light-controlled protein oligomerization was mediated by formation of Tyr-Tyr (dityrosine) dimers through -C-C- bonds acting as covalent bridges, with a predominant involvement of residue Y39. The diverse oligomeric species exhibited a direct effect on the in vitro aggregation behavior of wild-type monomeric αS, decreasing the total yield of amyloid fibrils in aggregation assays monitored by thioflavin T (ThioT) fluorescence and light scattering, and by atomic force microscopy (AFM). Compared to the unmodified monomer, the photoinduced covalent oligomeric species demonstrated increased toxic effects on differentiated neuronal-like SH-SY5Y cells. The results highlight the importance of protein modification induced by oxidative stress in the initial molecular events leading to Parkinsons disease. © 2012 Elsevier Inc. All rights reserved. Fil:Fauerbach, J.A. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. Fil:Jares-Erijman, E.A. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. 2012 https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_08915849_v53_n4_p1004_Borsarelli http://hdl.handle.net/20.500.12110/paper_08915849_v53_n4_p1004_Borsarelli |
| institution |
Universidad de Buenos Aires |
| institution_str |
I-28 |
| repository_str |
R-134 |
| collection |
Biblioteca Digital - Facultad de Ciencias Exactas y Naturales (UBA) |
| topic |
Neurodegeneration Oxidative stress Parkinson's disease Photocrosslinking Protein aggregation Protein photooxidation SH-SY5Y alpha synuclein oligomer tris(2,2' bipyridine)ruthenium tyrosine anion exchange chromatography article biophysics controlled study cytotoxicity fluorescence human human cell light absorption light scattering oligomerization photosensitization polyacrylamide gel electrophoresis priority journal steady state Western blotting alpha-Synuclein Ammonium Sulfate Amyloid Cell Line Cell Survival Cross-Linking Reagents Free Radicals Humans Kinetics Organometallic Compounds Oxidative Stress Photochemical Processes Photosensitizing Agents Protein Stability Tyrosine |
| spellingShingle |
Neurodegeneration Oxidative stress Parkinson's disease Photocrosslinking Protein aggregation Protein photooxidation SH-SY5Y alpha synuclein oligomer tris(2,2' bipyridine)ruthenium tyrosine anion exchange chromatography article biophysics controlled study cytotoxicity fluorescence human human cell light absorption light scattering oligomerization photosensitization polyacrylamide gel electrophoresis priority journal steady state Western blotting alpha-Synuclein Ammonium Sulfate Amyloid Cell Line Cell Survival Cross-Linking Reagents Free Radicals Humans Kinetics Organometallic Compounds Oxidative Stress Photochemical Processes Photosensitizing Agents Protein Stability Tyrosine Fauerbach, Jonathan Arturo Jares, Elizabeth Andrea Biophysical properties and cellular toxicity of covalent crosslinked oligomers of α-synuclein formed by photoinduced side-chain tyrosyl radicals |
| topic_facet |
Neurodegeneration Oxidative stress Parkinson's disease Photocrosslinking Protein aggregation Protein photooxidation SH-SY5Y alpha synuclein oligomer tris(2,2' bipyridine)ruthenium tyrosine anion exchange chromatography article biophysics controlled study cytotoxicity fluorescence human human cell light absorption light scattering oligomerization photosensitization polyacrylamide gel electrophoresis priority journal steady state Western blotting alpha-Synuclein Ammonium Sulfate Amyloid Cell Line Cell Survival Cross-Linking Reagents Free Radicals Humans Kinetics Organometallic Compounds Oxidative Stress Photochemical Processes Photosensitizing Agents Protein Stability Tyrosine |
| description |
Alpha-synuclein (αS), a 140 amino acid presynaptic protein, is the major component of the fibrillar aggregates (Lewy bodies) observed in dopaminergic neurons of patients affected by Parkinson's disease. It is currently believed that noncovalent oligomeric forms of αS, arising as intermediates in its aggregation, may constitute the major neurotoxic species. However, attempts to isolate and characterize such oligomers in vitro, and even more so in living cells, have been hampered by their transient nature, low concentration, polymorphism, and inherent instability. In this work, we describe the preparation and characterization of low molecular weight covalently bound oligomeric species of αS obtained by crosslinking via tyrosyl radicals generated by blue-light photosensitization of the metal coordination complex ruthenium (II) tris-bipyridine in the presence of ammonium persulfate. Numerous analytical techniques were used to characterize the αS oligomers: biochemical (anion-exchange chromatography, SDS-PAGE, and Western blotting); spectroscopic (optical: UV/Vis absorption, steady state, dynamic fluorescence, and dynamic light scattering); mass spectrometry; and electrochemical. Light-controlled protein oligomerization was mediated by formation of Tyr-Tyr (dityrosine) dimers through -C-C- bonds acting as covalent bridges, with a predominant involvement of residue Y39. The diverse oligomeric species exhibited a direct effect on the in vitro aggregation behavior of wild-type monomeric αS, decreasing the total yield of amyloid fibrils in aggregation assays monitored by thioflavin T (ThioT) fluorescence and light scattering, and by atomic force microscopy (AFM). Compared to the unmodified monomer, the photoinduced covalent oligomeric species demonstrated increased toxic effects on differentiated neuronal-like SH-SY5Y cells. The results highlight the importance of protein modification induced by oxidative stress in the initial molecular events leading to Parkinsons disease. © 2012 Elsevier Inc. All rights reserved. |
| author |
Fauerbach, Jonathan Arturo Jares, Elizabeth Andrea |
| author_facet |
Fauerbach, Jonathan Arturo Jares, Elizabeth Andrea |
| author_sort |
Fauerbach, Jonathan Arturo |
| title |
Biophysical properties and cellular toxicity of covalent crosslinked oligomers of α-synuclein formed by photoinduced side-chain tyrosyl radicals |
| title_short |
Biophysical properties and cellular toxicity of covalent crosslinked oligomers of α-synuclein formed by photoinduced side-chain tyrosyl radicals |
| title_full |
Biophysical properties and cellular toxicity of covalent crosslinked oligomers of α-synuclein formed by photoinduced side-chain tyrosyl radicals |
| title_fullStr |
Biophysical properties and cellular toxicity of covalent crosslinked oligomers of α-synuclein formed by photoinduced side-chain tyrosyl radicals |
| title_full_unstemmed |
Biophysical properties and cellular toxicity of covalent crosslinked oligomers of α-synuclein formed by photoinduced side-chain tyrosyl radicals |
| title_sort |
biophysical properties and cellular toxicity of covalent crosslinked oligomers of α-synuclein formed by photoinduced side-chain tyrosyl radicals |
| publishDate |
2012 |
| url |
https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_08915849_v53_n4_p1004_Borsarelli http://hdl.handle.net/20.500.12110/paper_08915849_v53_n4_p1004_Borsarelli |
| work_keys_str_mv |
AT fauerbachjonathanarturo biophysicalpropertiesandcellulartoxicityofcovalentcrosslinkedoligomersofasynucleinformedbyphotoinducedsidechaintyrosylradicals AT jareselizabethandrea biophysicalpropertiesandcellulartoxicityofcovalentcrosslinkedoligomersofasynucleinformedbyphotoinducedsidechaintyrosylradicals |
| _version_ |
1768542419232489472 |