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FoF1-ATP synthase is the essential membrane enzyme maintaining the cellular level of adenosine triphosphate (ATP) and comprises two rotary motors. We measure subunit rotation in FoF1-ATP synthase by intramolecular Foerster resonance energy…

Quantitative Methods · Quantitative Biology 2015-05-27 Torsten Rendler , Marc Renz , Eva Hammann , Stefan Ernst , Nawid Zarrabi , Michael Boersch

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 Forster…

Quantitative Methods · Quantitative Biology 2015-06-11 Bertram Su , Monika G. Dueser , Nawid Zarrabi , Thomas Heitkamp , Ilka Starke , Michael Börsch

FoF1-ATP synthase is the membrane protein catalyzing the synthesis of the 'biological energy currency' adenosine triphosphate (ATP). The enzyme uses internal subunit rotation for the mechanochemical conversion of a proton motive force to…

Biomolecules · Quantitative Biology 2015-06-15 Thomas Heitkamp , Hendrik Sielaff , Anja Korn , Marc Renz , Nawid Zarrabi , Michael Boersch

Observation times of freely diffusing single molecules in solution are limited by the photophysics of the attached fluorescence markers and by a small observation volume in the femtolitre range that is required for a sufficient…

Conformational changes of individual fluorescently labeled proteins can be followed in solution using a confocal microscope. Two fluorophores attached to selected domains of the protein report fluctuating conformations. Based on F\"orster…

Quantitative Methods · Quantitative Biology 2015-06-12 Nawid Zarrabi , Caterina Clausen , Monika G. Dueser , Michael Boersch

Trapping nanoscopic objects to observe their dynamic behaviour for extended periods of time is an ongoing quest. Particularly, sub-100nm transparent objects are hard to catch and most techniques rely on immobilisation or transient diffusion…

Quantitative Methods · Quantitative Biology 2017-01-13 Maria Dienerowitz , Thomas Heitkamp , Thomas Gottschall , Jens Limpert , Michael Borsch

FoF1-ATP synthases are ubiquitous membrane-bound, rotary motor enzymes that can catalyze ATP synthesis and hydrolysis. Their enzyme kinetics are controlled by internal subunit rotation, by substrate and product concentrations, by mechanical…

Biomolecules · Quantitative Biology 2021-06-29 Thomas Heitkamp , Michael Börsch

FoF1-ATP synthase catalyzes the synthesis of adenosine triphosphate (ATP). The F1 portion can be stripped from the membrane-embedded Fo portion of the enzyme. F1 acts as an ATP hydrolyzing enzyme, and ATP hydrolysis is associated with…

Biomolecules · Quantitative Biology 2018-02-14 Hendrik Sielaff , Thomas Heitkamp , Andrea Zappe , Nawid Zarrabi , Michael Boersch

Single molecule FRET (fluorescence resonance energy transfer) is a powerful technique for detecting real-time conformational changes and molecular interactions during biological reactions. In this review, we examine different techniques of…

Biomolecules · Quantitative Biology 2007-05-23 Ivan Rasnik , Sean A. McKinney , Taekjip Ha

Single-molecule F\"orster Resonance Energy Transfer (smFRET) is ideally suited to resolve the dynamics of biomolecules. A significant challenge to date is capturing and quantifying the exchange between multiple conformational states, mainly…

Single molecule time traces reveal the time evolution of unsynchronized kinetic systems. Especially single molecule F\"orster resonance energy transfer (smFRET) provides access to enzymatically important timescales, combined with molecular…

Quantitative Methods · Quantitative Biology 2018-01-17 Sonja Schmid , Thorsten Hugel

Confocal time resolved single-molecule spectroscopy using pulsed laser excitation and synchronized multi channel time correlated single photon counting (TCSPC) provides detailed information about the conformational changes of a biological…

Biological Physics · Physics 2009-11-13 N. Zarrabi , M. G. Dueser , S. Ernst , R. Reuter , G. D. Glick , S. D. Dunn , J. Wrachtrup , M. Boersch

Multiplexed, real-time fluorescence detection at the single-molecule level is highly desirable to reveal the stoichiometry, dynamics, and interactions of individual molecular species within complex systems. However, traditionally…

F1-ATPase is the soluble portion of the membrane-embedded enzyme FoF1-ATP synthase that catalyzes the production of adenosine triphosphate in eukaryotic and eubacterial cells. In reverse, the F1 part can also hydrolyze ATP quickly at three…

Biomolecules · Quantitative Biology 2015-06-18 Samuel D. Bockenhauer , Thomas M. Duncan , W. E. Moerner , Michael Boersch

Conformational dynamics of biomolecules are of fundamental importance for their function. Single-molecule F\"orster Resonance Energy Transfer (smFRET) is a powerful approach to inform on the structure and the dynamics of labeled molecules.…

Anti-Brownian traps confine single particles in free solution by closed-loop feedback forces that directly counteract Brownian motion. The extended-duration measurement of trapped objects allows detailed characterization of photophysical…

Instrumentation and Detectors · Physics 2023-05-15 Allison H. Squires , Abhijit A. Lavania , Peter D. Dahlberg , W. E. Moerner

Recent genome and transcriptome sequencing projects have unveiled a plethora of highly structured RNA molecules as central mediators of cellular function. Single molecule Forster Resonance Energy Transfer (smFRET) is a powerful tool for…

ATP synthases utilize a proton motive force to synthesize ATP. In reverse, these membrane-embedded enzymes can also hydrolyze ATP to pump protons over the membrane. To prevent wasteful ATP hydrolysis, distinct control mechanisms exist for…

Biomolecules · Quantitative Biology 2018-01-16 Hendrik Sielaff , Dhirendra Singh , Gerhard Grueber , Michael Börsch

The enzyme FoF1-ATP synthase provides the 'chemical energy currency' adenosine triphosphate (ATP) for living cells. Catalysis is driven by mechanochemical coupling of subunit rotation within the enzyme with conformational changes in the…

Biomolecules · Quantitative Biology 2015-06-04 Stefan Ernst , Monika G. Dueser , Nawid Zarrabi , Michael Boersch
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