Abstract
Membrane proteins exhibit different affinities for different lipid species, and protein–lipid selectivity regulates the membrane composition in close proximity to the protein, playing an important role in the formation of nanoscale membrane heterogeneities. Quantification of lipid selectivity by membrane proteins has been previously addressed mainly from Electron Spin Resonance studies (ESR). We present here a methodology for quantification of protein–lipid selectivity based on Förster Resonance Energy Transfer (FRET). Several FRET-based methods applied to the quantification of protein–lipid selectivity are presented, and different formalisms applied to the analysis of FRET data for particular geometries of donor–acceptor distribution are critically assessed. The FRET methodology has three interesting features: (1) by choosing donor–acceptor pairs with different Förster radii, it is possible to specifically study mainly the first-shell of lipids or also the outside shells; (2) the greater sensitivity of fluorescence requires much smaller amounts of material than ESR, (3) although this model leads to somewhat complex decay laws, it is actually not necessary to analyze the decay curves to recover the relevant parameters.
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Acknowledgments
F.F. acknowledges a research grant (SFRH/BPD/64320/2009) from Fundacão para a Ciência e Tecnologia (FCT). F.F., M.P., and L.M.S.L. acknowledge funding by FEDER (COMPETE program), and by FCT (Fundação para a Ciência e a Tecnologia), projects references PTDC/QUI-BIQ/112067/2009, PTDC/QUI-BIQ/099947/2008, and FCOMP-01-0124-FEDER-010787 (FCT PTDC/QUI-QUI/098198/2008).
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Fernandes, F., Prieto, M., Loura, L.M.S. (2012). Advanced FRET Methodologies: Protein–Lipid Selectivity Detection and Quantification. In: Sudhakaran, P., Surolia, A. (eds) Biochemical Roles of Eukaryotic Cell Surface Macromolecules. Advances in Experimental Medicine and Biology, vol 749. Springer, New York, NY. https://doi.org/10.1007/978-1-4614-3381-1_12
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DOI: https://doi.org/10.1007/978-1-4614-3381-1_12
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