5 March 2015 Observing conformations of single FoF1-ATP synthases in a fast anti-Brownian electrokinetic trap
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Abstract
To monitor conformational changes of individual membrane transporters in liposomes in real time, we attach two fluorophores to selected domains of a protein. Sequential distance changes between the dyes are recorded and analyzed by Förster resonance energy transfer (FRET). Using freely diffusing membrane proteins reconstituted in liposomes, observation times are limited by Brownian motion through the confocal detection volume. A. E. Cohen and W. E. Moerner have invented and built microfluidic devices to actively counteract Brownian motion of single nanoparticles in electrokinetic traps (ABELtrap). Here we present a version of an ABELtrap with a laser focus pattern generated by electro-optical beam deflectors and controlled by a programmable FPGA. This ABELtrap could hold single fluorescent nanobeads for more than 100 seconds, increasing the observation times of a single particle more than 1000-fold. Conformational changes of single FRET-labeled membrane enzymes FoF1-ATP synthase can be detected in the ABELtrap.
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Bertram Su, Bertram Su, Monika G. Düser, Monika G. Düser, Nawid Zarrabi, Nawid Zarrabi, Thomas Heitkamp, Thomas Heitkamp, Ilka Starke, Ilka Starke, Michael Börsch, Michael Börsch, } "Observing conformations of single FoF1-ATP synthases in a fast anti-Brownian electrokinetic trap", Proc. SPIE 9329, Multiphoton Microscopy in the Biomedical Sciences XV, 93290A (5 March 2015); doi: 10.1117/12.2080975; https://doi.org/10.1117/12.2080975
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