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Proceedings Paper

Step size of the rotary proton motor in single FoF1-ATP synthase from a thermoalkaliphilic bacterium by DCO-ALEX FRET
Author(s): Eva Hammann; Andrea Zappe; Stefanie Keis; Stefan Ernst; Doreen Matthies; Thomas Meier; Gregory M. Cook; Michael Börsch
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Paper Abstract

Thermophilic enzymes operate at high temperatures but show reduced activities at room temperature. They are in general more stable during preparation and, accordingly, are considered to be more rigid in structure. Crystallization is often easier compared to proteins from bacteria growing at ambient temperatures, especially for membrane proteins. The ATP-producing enzyme FoF1-ATP synthase from thermoalkaliphilic Caldalkalibacillus thermarum strain TA2.A1 is driven by a Fo motor consisting of a ring of 13 c-subunits. We applied a single-molecule Förster resonance energy transfer (FRET) approach using duty cycle-optimized alternating laser excitation (DCO-ALEX) to monitor the expected 13-stepped rotary Fo motor at work. New FRET transition histograms were developed to identify the smaller step sizes compared to the 10-stepped Fo motor of the Escherichia coli enzyme. Dwell time analysis revealed the temperature and the LDAO dependence of the Fo motor activity on the single molecule level. Back-and-forth stepping of the Fo motor occurs fast indicating a high flexibility in the membrane part of this thermophilic enzyme.

Paper Details

Date Published: 13 February 2012
PDF: 15 pages
Proc. SPIE 8228, Single Molecule Spectroscopy and Superresolution Imaging V, 82280A (13 February 2012); doi: 10.1117/12.907242
Show Author Affiliations
Eva Hammann, Univ. Stuttgart (Germany)
Andrea Zappe, Univ. Stuttgart (Germany)
Stefanie Keis, Univ. of Otago (New Zealand)
Stefan Ernst, Univ. of Stuttgart (Germany)
Jena Univ. Hospital (Germany)
Doreen Matthies, Max-Planck-Institute of Biophysics (Germany)
Thomas Meier, Max-Planck-Institute of Biophysics (Germany)
Gregory M. Cook, Univ. of Otago (New Zealand)
Michael Börsch, Univ. of Stuttgart (Germany)
Jena Univ. Hospital (Germany)

Published in SPIE Proceedings Vol. 8228:
Single Molecule Spectroscopy and Superresolution Imaging V
Jörg Enderlein; Zygmunt Karol Gryczynski; Rainer Erdmann; Felix Koberling; Ingo Gregor, Editor(s)

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