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Molecules. 2019 Jan 30;24(3). pii: E504. doi: 10.3390/molecules24030504.

Structural Asymmetry and Kinetic Limping of Single Rotary F-ATP Synthases.

Author information

1
Single-Molecule Microscopy Group, Jena University Hospital, Friedrich Schiller University, 07743 Jena, Germany. hendrik.sielaff@med.uni-jena.de.
2
School of Life Sciences, Arizona State University, Tempe, Arizona, AZ 85287, USA. seiga@asu.edu.
3
School of Life Sciences, Arizona State University, Tempe, Arizona, AZ 85287, USA. frasch@asu.edu.
4
Department of Biology & Chemistry, University of Osnabrück, 49076 Osnabrück, Germany. junge@uos.de.
5
Single-Molecule Microscopy Group, Jena University Hospital, Friedrich Schiller University, 07743 Jena, Germany. michael.boersch@uni-jena.de.

Abstract

F-ATP synthases use proton flow through the FO domain to synthesize ATP in the F₁ domain. In Escherichia coli, the enzyme consists of rotor subunits γεc10 and stator subunits (αβ)₃δab₂. Subunits c10 or (αβ)₃ alone are rotationally symmetric. However, symmetry is broken by the b₂ homodimer, which together with subunit δa, forms a single eccentric stalk connecting the membrane embedded FO domain with the soluble F₁ domain, and the central rotating and curved stalk composed of subunit γε. Although each of the three catalytic binding sites in (αβ)₃ catalyzes the same set of partial reactions in the time average, they might not be fully equivalent at any moment, because the structural symmetry is broken by contact with b₂δ in F₁ and with ba in FO. We monitored the enzyme's rotary progression during ATP hydrolysis 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. We found that one dwell in the three-stepped rotary progression lasting longer than the other two by a factor of up to 1.6. This effect of the structural asymmetry is small due to the internal elastic coupling.

KEYWORDS:

Escherichia coli; FOF1 ATP synthase; cryo-EM structure; elasticity; single-molecule fluorescence; subunit rotation; symmetry

PMID:
30704145
PMCID:
PMC6384691
DOI:
10.3390/molecules24030504
[Indexed for MEDLINE]
Free PMC Article

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