The regulatory switch of F1-ATPase studied by single-molecule FRET in the ABEL trap

@inproceedings{Bockenhauer2014TheRS,
  title={The regulatory switch of F1-ATPase studied by single-molecule FRET in the ABEL trap},
  author={Samuel D Bockenhauer and Thomas M. Duncan and W. E. Moerner and Michael B{\"o}rsch},
  booktitle={Photonics West - Biomedical Optics},
  year={2014}
}
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 catalytic binding sites. Therefore, catalysis of 'non-productive' ATP hydrolysis by F1 (or FoF1) must be minimized in the cell. In bacteria, the ε subunit is thought to control and block ATP hydrolysis by mechanically inserting its C-terminus into the rotary… 
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30 Citations
Background Citations
5
Methods Citations
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30 Citations

Regulatory conformational changes of the Ɛ subunit in single FRET-labeled F0F1-ATP synthase
TLDR
An experimental system is developed that can reveal conditions under which ε inhibits the holoenzyme FoF1-ATP synthase in vitro and labels the C-terminal domain of ε and the γ subunit specifically with two different fluorophores for single-molecule Förster resonance energy transfer (smFRET).
Regulatory conformational changes of the epsilon subunit in single FRET-labeled FoF1-ATP synthase
TLDR
Labeling the C-terminal domain of epsilon and the gamma subunit specifically with two different fluorophores for single-molecule Foerster resonance energy transfer (smFRET) allowed monitoring of the conformation of ePSilon in the reconstituted enzyme in real time.
Structural Asymmetry and Kinetic Limping of Single Rotary F-ATP Synthases
TLDR
The enzyme’s rotary progression during ATP hydrolysis is monitored by three single-molecule techniques: fluorescence video-microscopy with attached actin filaments, Förster resonance energy transfer between pairs of fluorescence probes, and a polarization assay using gold nanorods, and it is found that one dwell in the three-steppedrotary progression lasting longer than the other two by a factor of up to 1.6.
Observing single FoF1-ATP synthase at work using an improved fluorescent protein mNeonGreen as FRET donor
TLDR
The novel FRET donor mNeonGreen is evaluated as a fusion to FoF1-ATP synthase and compare it to the previously used fluorophore EGFP to evaluate the biochemical purification procedures and activity measurements of the fully functional mutant enzyme.
Analyzing conformational changes in single FRET-labeled A1 parts of archaeal A1AO-ATP synthase
TLDR
The lifetimes of fluorescence donor and acceptor dyes are analyzed to distinguish between smFRET signals of conformational changes and potential artefacts to prevent wasteful ATP hydrolysis.
The regulatory subunit ε in Escherichia coli FOF1-ATP synthase.
Fast ATP-Dependent Subunit Rotation in Reconstituted FoF1-ATP Synthase Trapped in Solution.
TLDR
It is reported that kinetic monitoring of functional rotation can be prolonged from milliseconds to seconds by utilizing an anti-Brownian electrokinetic trap (ABEL trap), and broad distributions of ATP-dependent catalytic rates were revealed.
ATP synthase from Escherichia coli: Mechanism of rotational catalysis, and inhibition with the ε subunit and phytopolyphenols.
Observing conformations of single FoF1-ATP synthases in a fast anti-Brownian electrokinetic trap
TLDR
A version of an ABELtrap with a laser focus pattern generated by electro-optical beam deflectors and controlled by a programmable FPGA is presented, which could hold single fluorescent nanobeads for more than 100 seconds and increase the observation times of a single particle more than 1000-fold.
Insights into the regulatory function of the ɛ subunit from bacterial F-type ATP synthases: a comparison of structural, biochemical and biophysical data
TLDR
It is concluded that the ɛ subunit from the bacterial F-type ATP synthases is indeed capable of regulating ATP hydrolysis activity in a wide variety of bacteria, making it a potentially valuable drug target, but its exact role is still under debate.
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References

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TLDR
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TLDR
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TLDR
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TLDR
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