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• Paredes, J
• Casares, S
• Ruedas-Rama, M
• Fernandez, E
• Castello, F
• Varela, L
• Orte, A
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• Paredes, J
• Casares, S
• Ruedas-Rama, M
• Fernandez, E
• Castello, F
• Varela, L
• Orte, A
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• Paredes, J
• Casares, S
• Ruedas-Rama, M
• Fernandez, E
• Castello, F
• Varela, L
• Orte, A

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Int. J. Mol. Sci. 2012, 13(8), 9400-9418; doi:10.3390/ijms13089400

Article

Early Amyloidogenic Oligomerization Studied through Fluorescence Lifetime Correlation Spectroscopy

Jose M. Paredes ¹ , Salvador Casares ² , Maria J. Ruedas-Rama ¹ , Elena Fernandez ² , Fabio Castello ¹ , Lorena Varela ^2,†  and Angel Orte ^1,*

¹ Department of Physical Chemistry, Faculty of Pharmacy, Campus Cartuja, Granada, 18071, Spain ² Department of Physical Chemistry, Faculty of Sciences, Campus Fuentenueva, Granada, 18071, Spain ^† Present address: Department of Biochemistry, University of Oxford, South Parks Road, Oxford, OX1 3QU, UK.

* Author to whom correspondence should be addressed.

Received: 19 June 2012; in revised form: 13 July 2012 / Accepted: 19 July 2012 / Published: 25 July 2012

(This article belongs to the Special Issue Advances in Single Molecule Spectroscopy)

Download PDF Full-Text [1372 KB, Updated Version, uploaded 30 July 2012 12:04 CEST]

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Abstract: Amyloidogenic protein aggregation is a persistent biomedical problem. Despite active research in disease-related aggregation, the need for multidisciplinary approaches to the problem is evident. Recent advances in single-molecule fluorescence spectroscopy are valuable for examining heterogenic biomolecular systems. In this work, we have explored the initial stages of amyloidogenic aggregation by employing fluorescence lifetime correlation spectroscopy (FLCS), an advanced modification of conventional fluorescence correlation spectroscopy (FCS) that utilizes time-resolved information. FLCS provides size distributions and kinetics for the oligomer growth of the SH3 domain of α-spectrin, whose N47A mutant forms amyloid fibrils at pH 3.2 and 37 °C in the presence of salt. The combination of FCS with additional fluorescence lifetime information provides an exciting approach to focus on the initial aggregation stages, allowing a better understanding of the fibrillization process, by providing multidimensional information, valuable in combination with other conventional methodologies.

Keywords: amyloids; protein aggregation; pulsed interleaved excitation; protein oligomers; single-molecule fluorescence

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