Abstract
The S100 proteins are a large family of 10–12 kDa EF-hand signaling proteins that bind calcium, and in some cases zinc and copper, functioning as central regulators in a diversity of cellular processes. These proteins have tissue, cell, and subcellular-specific expression patterns, and many have an extracellular function. Altogether, these properties underlie their functional diversity and involvement in several pathological conditions including cancer, inflammation, and neurodegeneration. S100 proteins exhibit considerable structural plasticity, being able to exist as monomers or assemble into dimers, higher oligomers, and amyloids, frequently in a metal-dependent manner. Many of these oligomers are functionally relevant, and S100 amyloids have been recently found in prostatic inclusions. Here, we report experimental procedures for the isolation and quantitation of S100 oligomers from tissues, purification of recombinant human S100 protein for assays and use as standards, and an amyloidogenesis assay that allows monitoring the formation of S100 β-oligomers and amyloids in apo- and metal-bound S100 proteins.
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Abbreviations
- ATR FT-IR:
-
Attenuated total reflectance Fourier transform infrared spectroscopy
- DLS:
-
Dynamic light scattering
- KPi:
-
Potassium phosphate
- OD:
-
Optical density
- ThT:
-
Thioflavin T
- SEC:
-
Size exclusion chromatography
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Acknowledgments
This work was supported by grants POCTI/QUI/45758 and PTDC/QUI/70101 (to CMG) from the Fundação para a Ciência e a Tecnologia (FCT/MCTES, Portugal), by grants FR 1488/3-1 and FR 1488/5-1 from the Deutsche Forschungsgemeinschaft (DFG) (to GF). CMG and GF are recipients of a CRUP/DAAD collaborative grant A-15/08. HMB was a recipient of a Ph. D. fellowship (SFRH/BD/31126/2006) from Fundação para a Ciência e a Tecnologia (FCT/MCTES, Portugal). Ludmilla Morozova-Roche (Umeå University) is gratefully acknowledged for AFM imaging.
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Botelho, H.M., Fritz, G., Gomes, C.M. (2012). Analysis of S100 Oligomers and Amyloids. In: Sigurdsson, E., Calero, M., Gasset, M. (eds) Amyloid Proteins. Methods in Molecular Biology, vol 849. Humana Press. https://doi.org/10.1007/978-1-61779-551-0_25
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DOI: https://doi.org/10.1007/978-1-61779-551-0_25
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