Berlin 2018 – wissenschaftliches Programm
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BP: Fachverband Biologische Physik
BP 28: Single Molecule Biophysics
BP 28.12: Vortrag
Donnerstag, 15. März 2018, 12:45–13:00, H 1058
Reductive caging and photoactivation in single-molecule Förster resonance energy transfer experiments — •Atieh Aminian Jazi1, Evelyn Ploetz2, Muhamad Arizki1, Christine Ziegler4, Reinhard Krämer4, and Thorben Cordes1,3 — 1Zernike Institute for Advanced Materials, Groningen, The Netherlands — 2Department of Chemistry and CeNS, Ludwig Maximilians-Universität, Munich, Germany — 3Department Biology I, Ludwig-Maximilians-Universität München, Germany — 4Institute of Biophysics and Biophysical Chemistry, Universität Regensburg,Germany
Förster-resonance energy transfer(FRET), in combination with single-molecule detection, has become a powerful tool to investigate the structural dynamics of biomolecular systems. We used caging of fluorophores by reversible chemical deactivation of fluorescence.
The diffusing molecules can be reactivated by ultraviolet (UV) light. UV-reactivation allows retrieving both FRET-related distances and sorting of multiple intramolecular spices in solution-based smFRET. We employed caged FRET to investigate the structure of a membrane transporter BetP, as multi-subunit protein, and nucleic acids containing more than two fluorescent labels. The results revealed that chemical caging and photoactivation (uncaging) by UV light allows temporal uncoupling of convoluted fluorescence signals from multiple donors or acceptor molecules.Caged FRET also can be used in a further application to study the intermolecular details of low-affinity binding interactions with diffusion-based smFRET.