Abstract:
The aggregation of α-synuclein, a presynaptic protein, has an important role in the etiology of Parkinson's disease. Oligomers or protofibrils adopting the cross-β-sheet structure characteristic of fibrillating amyloid proteins are presumed to be the primary cytotoxic species. Current techniques for monitoring the kinetics of α-synuclein aggregation based on fluorescent dyes such as Thioflavin-T and Congo red detect only the terminal fibrillar species, are discontinuous and notoriously irreproducible. We have devised a new fluorescence aggregation assay that is continuous and provides a large set of fluorescence parameters sensitive to the presence of oligomeric intermediates as well as fibrils. The approach involves tagging functionally neutral Ala-to-Cys variants of α-synuclein with the long-lifetime fluorophore pyrene. Upon induction of aggregation at 37 °C, the entire family of steady-state descriptors of pyrene emission (monomer intensity, solvent polarity ratio (II/IIII), and anisotropy; and excimer intensity) change dramatically, particularly during the early stages in which oligomeric intermediates form and evolve. The pyrene probe senses a progressive decrease in polarity, an increase in molecular mass and close intermolecular association in a manner dependent on position in the sequence and the presence of point mutations. The time-resolved decays (0-160 ns) of intensity and anisotropy exhibited complex, characteristic features. The new assay constitutes a convenient platform for the high-throughput screening of agents useful in the diagnosis and therapy of Parkinson's disease as well as in basic investigations. © 2008 Elsevier Ltd. All rights reserved.
Registro:
Documento: |
Artículo
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Título: | Multiparametric Fluorescence Detection of Early Stages in the Amyloid Protein Aggregation of Pyrene-labeled α-Synuclein |
Autor: | Thirunavukkuarasu, S.; Jares-Erijman, E.A.; Jovin, T.M. |
Filiación: | Laboratory of Cellular Dynamics, Max Planck Institute for Biophysical Chemistry, 37077 Göttingen, Germany Departamento de Química Orgánica, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, 1428 Buenos Aires, Argentina
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Palabras clave: | fibrillization; flourescence anisotropy; neurodegenerative; oligomers; Parkinson's disease; alpha synuclein; pyrene; anisotropy; article; fluorescence; human; in vivo study; kinetics; Parkinson disease; point mutation; priority journal; protein aggregation; protein analysis; alpha-Synuclein; Amino Acid Sequence; Amyloid; Anisotropy; Fluorescent Dyes; Humans; Molecular Sequence Data; Mutagenesis, Site-Directed; Parkinson Disease; Protein Folding; Protein Structure, Quaternary; Pyrenes; Spectrometry, Fluorescence |
Año: | 2008
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Volumen: | 378
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Número: | 5
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Página de inicio: | 1064
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Página de fin: | 1073
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DOI: |
http://dx.doi.org/10.1016/j.jmb.2008.03.034 |
Título revista: | Journal of Molecular Biology
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Título revista abreviado: | J. Mol. Biol.
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ISSN: | 00222836
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CODEN: | JMOBA
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CAS: | alpha synuclein, 154040-18-3; pyrene, 129-00-0; alpha-Synuclein; Amyloid; Fluorescent Dyes; pyrene, 129-00-0; Pyrenes
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Registro: | https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_00222836_v378_n5_p1064_Thirunavukkuarasu |
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Citas:
---------- APA ----------
Thirunavukkuarasu, S., Jares-Erijman, E.A. & Jovin, T.M.
(2008)
. Multiparametric Fluorescence Detection of Early Stages in the Amyloid Protein Aggregation of Pyrene-labeled α-Synuclein. Journal of Molecular Biology, 378(5), 1064-1073.
http://dx.doi.org/10.1016/j.jmb.2008.03.034---------- CHICAGO ----------
Thirunavukkuarasu, S., Jares-Erijman, E.A., Jovin, T.M.
"Multiparametric Fluorescence Detection of Early Stages in the Amyloid Protein Aggregation of Pyrene-labeled α-Synuclein"
. Journal of Molecular Biology 378, no. 5
(2008) : 1064-1073.
http://dx.doi.org/10.1016/j.jmb.2008.03.034---------- MLA ----------
Thirunavukkuarasu, S., Jares-Erijman, E.A., Jovin, T.M.
"Multiparametric Fluorescence Detection of Early Stages in the Amyloid Protein Aggregation of Pyrene-labeled α-Synuclein"
. Journal of Molecular Biology, vol. 378, no. 5, 2008, pp. 1064-1073.
http://dx.doi.org/10.1016/j.jmb.2008.03.034---------- VANCOUVER ----------
Thirunavukkuarasu, S., Jares-Erijman, E.A., Jovin, T.M. Multiparametric Fluorescence Detection of Early Stages in the Amyloid Protein Aggregation of Pyrene-labeled α-Synuclein. J. Mol. Biol. 2008;378(5):1064-1073.
http://dx.doi.org/10.1016/j.jmb.2008.03.034