Publication

Towards Multiparametric Fluorescent Imaging of Amyloid Formation: Studies of a YFP Model of alpha-Synuclein Aggregation

van Ham, T. J., Esposito, A., Kumita, J. R., Hsu, S-T. D., Schierle, G. S. K., Kaminsk, C. F., Dobson, C. M., Nollen, E. A. A. & Bertoncini, C. W., 22-Jan-2010, In : Journal of Molecular Biology. 395, 3, p. 627-642 16 p.

Research output: Contribution to journalArticleAcademicpeer-review

APA

van Ham, T. J., Esposito, A., Kumita, J. R., Hsu, S-T. D., Schierle, G. S. K., Kaminsk, C. F., ... Bertoncini, C. W. (2010). Towards Multiparametric Fluorescent Imaging of Amyloid Formation: Studies of a YFP Model of alpha-Synuclein Aggregation. Journal of Molecular Biology, 395(3), 627-642. https://doi.org/10.1016/j.jmb.2009.10.066

Author

van Ham, Tjakko J. ; Esposito, Alessandro ; Kumita, Janet R. ; Hsu, Shang-Te D. ; Schierle, Gabriele S. Kaminski ; Kaminsk, Clemens F. ; Dobson, Christopher M. ; Nollen, Ellen A. A. ; Bertoncini, Carlos W. / Towards Multiparametric Fluorescent Imaging of Amyloid Formation : Studies of a YFP Model of alpha-Synuclein Aggregation. In: Journal of Molecular Biology. 2010 ; Vol. 395, No. 3. pp. 627-642.

Harvard

van Ham, TJ, Esposito, A, Kumita, JR, Hsu, S-TD, Schierle, GSK, Kaminsk, CF, Dobson, CM, Nollen, EAA & Bertoncini, CW 2010, 'Towards Multiparametric Fluorescent Imaging of Amyloid Formation: Studies of a YFP Model of alpha-Synuclein Aggregation', Journal of Molecular Biology, vol. 395, no. 3, pp. 627-642. https://doi.org/10.1016/j.jmb.2009.10.066

Standard

Towards Multiparametric Fluorescent Imaging of Amyloid Formation : Studies of a YFP Model of alpha-Synuclein Aggregation. / van Ham, Tjakko J.; Esposito, Alessandro; Kumita, Janet R.; Hsu, Shang-Te D.; Schierle, Gabriele S. Kaminski; Kaminsk, Clemens F.; Dobson, Christopher M.; Nollen, Ellen A. A.; Bertoncini, Carlos W.

In: Journal of Molecular Biology, Vol. 395, No. 3, 22.01.2010, p. 627-642.

Research output: Contribution to journalArticleAcademicpeer-review

Vancouver

van Ham TJ, Esposito A, Kumita JR, Hsu S-TD, Schierle GSK, Kaminsk CF et al. Towards Multiparametric Fluorescent Imaging of Amyloid Formation: Studies of a YFP Model of alpha-Synuclein Aggregation. Journal of Molecular Biology. 2010 Jan 22;395(3):627-642. https://doi.org/10.1016/j.jmb.2009.10.066


BibTeX

@article{24e71db64d7e4117bb8d92f6682f4a24,
title = "Towards Multiparametric Fluorescent Imaging of Amyloid Formation: Studies of a YFP Model of alpha-Synuclein Aggregation",
abstract = "Misfolding and aggregation of proteins are characteristics of a range of increasingly prevalent neurodegenerative disorders including Alzheimer's and Parkinson's diseases. In Parkinson's disease and several closely related syndromes, the protein a-synuclein (AS) aggregates and forms amyloid-like deposits in specific regions of the brain. Fluorescence microscopy using fluorescent proteins, for instance the yellow fluorescent protein (YFP), is the method of choice to image molecular events such as protein aggregation in living organisms. The presence of a bulky fluorescent protein tag, however, may potentially affect significantly the properties of the protein of interest, for AS in particular, its relative small size and, as an intrinsically unfolded protein, its lack of defined secondary structure could challenge the usefulness of fluorescent-protein-based derivatives. Here, we subject a YFP fusion of AS to exhaustive studies in vitro designed to determine its potential as a means of probing amyloid formation in vivo. By employing a combination of biophysical and biochemical studies, we demonstrate that the conjugation of YFP does not significantly perturb the structure of AS in solution and find that the AS-YFP protein forms amyloid deposits in vitro that are essentially identical with those observed for wild-type AS, except that they are fluorescent. Of the several fluorescent properties of the YFP chimera that were assayed, we find that fluorescence anisotropy is a particularly useful parameter to follow the aggregation of AS-YFP, because of energy migration Forster resonance energy transfer (emFRET or homoFRET) between closely positioned YFP moieties occurring as a result of the high density of the fluorophore within the amyloid species. Fluorescence anisotropy imaging microscopy further demonstrates the ability of homoFRET to distinguish between soluble, pre-fibrillar aggregates and amyloid fibrils of AS-YFP. Our results validate the use of fluorescent protein chimeras of AS as representative models for studying protein aggregation and offer new opportunities for the investigation of amyloid aggregation in vivo using YFP-tagged proteins. (C) 2009 Elsevier Ltd. All rights reserved.",
keywords = "protein misfolding, protein aggregation, fluorescence anisotropy imaging microscopy, Parkinson's disease, fluorescence protein, PARKINSONS-DISEASE, LIVING CELLS, BETA-SYNUCLEIN, MOLECULAR-INTERACTIONS, PROTEIN AGGREGATION, CONTACT DYNAMICS, NMR-SPECTROSCOPY, FIBRIL FORMATION, FRET MICROSCOPY, LEWY BODIES",
author = "{van Ham}, {Tjakko J.} and Alessandro Esposito and Kumita, {Janet R.} and Hsu, {Shang-Te D.} and Schierle, {Gabriele S. Kaminski} and Kaminsk, {Clemens F.} and Dobson, {Christopher M.} and Nollen, {Ellen A. A.} and Bertoncini, {Carlos W.}",
year = "2010",
month = "1",
day = "22",
doi = "10.1016/j.jmb.2009.10.066",
language = "English",
volume = "395",
pages = "627--642",
journal = "Journal of Molecular Biology",
issn = "0022-2836",
publisher = "Academic Press",
number = "3",

}

RIS

TY - JOUR

T1 - Towards Multiparametric Fluorescent Imaging of Amyloid Formation

T2 - Studies of a YFP Model of alpha-Synuclein Aggregation

AU - van Ham, Tjakko J.

AU - Esposito, Alessandro

AU - Kumita, Janet R.

AU - Hsu, Shang-Te D.

AU - Schierle, Gabriele S. Kaminski

AU - Kaminsk, Clemens F.

AU - Dobson, Christopher M.

AU - Nollen, Ellen A. A.

AU - Bertoncini, Carlos W.

PY - 2010/1/22

Y1 - 2010/1/22

N2 - Misfolding and aggregation of proteins are characteristics of a range of increasingly prevalent neurodegenerative disorders including Alzheimer's and Parkinson's diseases. In Parkinson's disease and several closely related syndromes, the protein a-synuclein (AS) aggregates and forms amyloid-like deposits in specific regions of the brain. Fluorescence microscopy using fluorescent proteins, for instance the yellow fluorescent protein (YFP), is the method of choice to image molecular events such as protein aggregation in living organisms. The presence of a bulky fluorescent protein tag, however, may potentially affect significantly the properties of the protein of interest, for AS in particular, its relative small size and, as an intrinsically unfolded protein, its lack of defined secondary structure could challenge the usefulness of fluorescent-protein-based derivatives. Here, we subject a YFP fusion of AS to exhaustive studies in vitro designed to determine its potential as a means of probing amyloid formation in vivo. By employing a combination of biophysical and biochemical studies, we demonstrate that the conjugation of YFP does not significantly perturb the structure of AS in solution and find that the AS-YFP protein forms amyloid deposits in vitro that are essentially identical with those observed for wild-type AS, except that they are fluorescent. Of the several fluorescent properties of the YFP chimera that were assayed, we find that fluorescence anisotropy is a particularly useful parameter to follow the aggregation of AS-YFP, because of energy migration Forster resonance energy transfer (emFRET or homoFRET) between closely positioned YFP moieties occurring as a result of the high density of the fluorophore within the amyloid species. Fluorescence anisotropy imaging microscopy further demonstrates the ability of homoFRET to distinguish between soluble, pre-fibrillar aggregates and amyloid fibrils of AS-YFP. Our results validate the use of fluorescent protein chimeras of AS as representative models for studying protein aggregation and offer new opportunities for the investigation of amyloid aggregation in vivo using YFP-tagged proteins. (C) 2009 Elsevier Ltd. All rights reserved.

AB - Misfolding and aggregation of proteins are characteristics of a range of increasingly prevalent neurodegenerative disorders including Alzheimer's and Parkinson's diseases. In Parkinson's disease and several closely related syndromes, the protein a-synuclein (AS) aggregates and forms amyloid-like deposits in specific regions of the brain. Fluorescence microscopy using fluorescent proteins, for instance the yellow fluorescent protein (YFP), is the method of choice to image molecular events such as protein aggregation in living organisms. The presence of a bulky fluorescent protein tag, however, may potentially affect significantly the properties of the protein of interest, for AS in particular, its relative small size and, as an intrinsically unfolded protein, its lack of defined secondary structure could challenge the usefulness of fluorescent-protein-based derivatives. Here, we subject a YFP fusion of AS to exhaustive studies in vitro designed to determine its potential as a means of probing amyloid formation in vivo. By employing a combination of biophysical and biochemical studies, we demonstrate that the conjugation of YFP does not significantly perturb the structure of AS in solution and find that the AS-YFP protein forms amyloid deposits in vitro that are essentially identical with those observed for wild-type AS, except that they are fluorescent. Of the several fluorescent properties of the YFP chimera that were assayed, we find that fluorescence anisotropy is a particularly useful parameter to follow the aggregation of AS-YFP, because of energy migration Forster resonance energy transfer (emFRET or homoFRET) between closely positioned YFP moieties occurring as a result of the high density of the fluorophore within the amyloid species. Fluorescence anisotropy imaging microscopy further demonstrates the ability of homoFRET to distinguish between soluble, pre-fibrillar aggregates and amyloid fibrils of AS-YFP. Our results validate the use of fluorescent protein chimeras of AS as representative models for studying protein aggregation and offer new opportunities for the investigation of amyloid aggregation in vivo using YFP-tagged proteins. (C) 2009 Elsevier Ltd. All rights reserved.

KW - protein misfolding

KW - protein aggregation

KW - fluorescence anisotropy imaging microscopy

KW - Parkinson's disease

KW - fluorescence protein

KW - PARKINSONS-DISEASE

KW - LIVING CELLS

KW - BETA-SYNUCLEIN

KW - MOLECULAR-INTERACTIONS

KW - PROTEIN AGGREGATION

KW - CONTACT DYNAMICS

KW - NMR-SPECTROSCOPY

KW - FIBRIL FORMATION

KW - FRET MICROSCOPY

KW - LEWY BODIES

U2 - 10.1016/j.jmb.2009.10.066

DO - 10.1016/j.jmb.2009.10.066

M3 - Article

VL - 395

SP - 627

EP - 642

JO - Journal of Molecular Biology

JF - Journal of Molecular Biology

SN - 0022-2836

IS - 3

ER -

ID: 5022999