Quantification of Protein-Lipid Selectivity using FRET: Application to the M13 Major Coat Protein |
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| Authors: | Fernandes Fábio Loura Luís M S Koehorst Rob Spruijt Ruud B Hemminga Marcus A Fedorov Alexander Prieto Manuel |
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| Affiliation: | Centro de Química-Física Molecular, Instituto Superior Técnico, Lisbon, Portugal. |
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| Abstract: | Quantification of lipid selectivity by membrane proteins has been previously addressed mainly from electron spin resonance studies. We present here a new methodology for quantification of protein-lipid selectivity based on fluorescence resonance energy transfer. A mutant of M13 major coat protein was labeled with 7-diethylamino-3((4'iodoacetyl)amino)phenyl-4-methylcoumarin to be used as the donor in energy transfer studies. Phospholipids labeled with N-(7-nitro-2-1,3-benzoxadiazol-4-yl) were selected as the acceptors. The dependence of protein-lipid selectivity on both hydrophobic mismatch and headgroup family was determined. M13 major coat protein exhibited larger selectivity toward phospholipids which allow for a better hydrophobic matching. Increased selectivity was also observed for anionic phospholipids and the relative association constants agreed with the ones already presented in the literature and obtained through electron spin resonance studies. This result led us to conclude that fluorescence resonance energy transfer is a promising methodology in protein-lipid selectivity studies. |
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