spFRET (single-pair fluorescence resonance energy transfer) with organic fluorophores has been used to demonstrate rotation of the subunits γ and ε in membrane-integrated FoF1 during proton transport-coupled ATP synthesis. Owing to the high light intensities used in single-molecule spectroscopy, organic fluorophores show a high probability for photobleaching. Luminescent CdSe/ZnS nanocrystals with a hydrophilic shell have been covalently bound to FoF1 either to the stator subunit b or to the rotor subunit c. TIRFM (total internal reflection microscopy) shows that covalent binding of the QD (quantum dot) via cysteine to FoF1 leads to a significant decrease in the blinking probability in the microsecond-to-second time range. This effect allows the observation of subunit movements in an extended time range. If the QD is bound to the rotor subunit c, the fluorescence anisotropy shows fluctuations in the presence of ATP, in contrast with the constant anisotropy observed in the absence of ATP.
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October 2008
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Conference Article|
September 19 2008
Quantum dots for single-pair fluorescence resonance energy transfer in membrane- integrated EFoF1
Eva Galvez;
Eva Galvez
*Instituto de Carboquimica (CSIC), Zaragoza, Spain
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Monika Düser;
Monika Düser
†Physikalisches Institut, Universität Stuttgart, Pfaffenwaldring 57, 70569 Stuttgart, Germany
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Michael Börsch;
Michael Börsch
†Physikalisches Institut, Universität Stuttgart, Pfaffenwaldring 57, 70569 Stuttgart, Germany
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Jörg Wrachtrup;
Jörg Wrachtrup
†Physikalisches Institut, Universität Stuttgart, Pfaffenwaldring 57, 70569 Stuttgart, Germany
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Peter Gräber
Peter Gräber
1
‡Institut für Physikalische Chemie, Universität Freiburg, Albertstrasse 23a, 79104 Freiburg, Germany
1To whom correspondence should be addressed (email [email protected]).
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Publisher: Portland Press Ltd
Received:
April 09 2008
Online ISSN: 1470-8752
Print ISSN: 0300-5127
© The Authors Journal compilation © 2008 Biochemical Society
2008
Biochem Soc Trans (2008) 36 (5): 1017–1021.
Article history
Received:
April 09 2008
Citation
Eva Galvez, Monika Düser, Michael Börsch, Jörg Wrachtrup, Peter Gräber; Quantum dots for single-pair fluorescence resonance energy transfer in membrane- integrated EFoF1. Biochem Soc Trans 1 October 2008; 36 (5): 1017–1021. doi: https://doi.org/10.1042/BST0361017
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