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Proceedings Paper

The regulatory switch of F1-ATPase studied by single-molecule FRET in the ABEL trap
Author(s): Samuel D. Bockenhauer; Thomas M. Duncan; W. E. Moerner; Michael Börsch
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Paper Abstract

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 motor region of F1. We investigate this proposed mechanism by labeling F1 specifically with two fluorophores to monitor the C-terminus of the ε subunit by Förster resonance energy transfer. Single F1 molecules are trapped in solution by an Anti-Brownian electrokinetic trap which keeps the FRET-labeled F1 in place for extended observation times of several hundreds of milliseconds, limited by photobleaching. FRET changes in single F1 and FRET histograms for different biochemical conditions are compared to evaluate the proposed regulatory mechanism.

Paper Details

Date Published: 1 April 2014
PDF: 14 pages
Proc. SPIE 8950, Single Molecule Spectroscopy and Superresolution Imaging VII, 89500H (1 April 2014); doi: 10.1117/12.2042688
Show Author Affiliations
Samuel D. Bockenhauer, Stanford Univ. (United States)
Thomas M. Duncan, SUNY Upstate Medical Univ. (United States)
W. E. Moerner, Stanford Univ. (United States)
Michael Börsch, Friedrich-Schiller-Univ. Jena (Germany)


Published in SPIE Proceedings Vol. 8950:
Single Molecule Spectroscopy and Superresolution Imaging VII
Jörg Enderlein; Ingo Gregor; Zygmunt Karol Gryczynski; Rainer Erdmann; Felix Koberling, Editor(s)

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