Subunit rotation in a single FoF1-ATP synthase in a living bacterium monitored by FRET

K. Seyfert; T. Oosaka; H. Yaginuma; S. Ernst; H. Noji; R. Iino; M. Börsch

doi:10.1117/12.873066

28 February 2011 Subunit rotation in a single F_oF₁-ATP synthase in a living bacterium monitored by FRET

K. Seyfert, T. Oosaka, H. Yaginuma, S. Ernst, H. Noji, R. Iino, M. Börsch

Proceedings Volume 7905, Single Molecule Spectroscopy and Imaging IV; 79050K (2011) https://doi.org/10.1117/12.873066
Event: SPIE BiOS, 2011, San Francisco, California, United States

Abstract

F_oF₁-ATP synthase is the ubiquitous membrane-bound enzyme in mitochondria, chloroplasts and bacteria which provides the 'chemical energy currency' adenosine triphosphate (ATP) for cellular processes. In Escherichia coli ATP synthesis is driven by a proton motive force (PMF) comprising a proton concentration difference ΔpH plus an electric potential ΔΨ across the lipid membrane. Single-molecule in vitro experiments have confirmed that proton-driven subunit rotation within F_oF₁-ATP synthase is associated with ATP synthesis. Based on intramolecular distance measurements by single-molecule fluorescence resonance energy transfer (FRET) the kinetics of subunit rotation and the step sizes of the different rotor parts have been unraveled. However, these experiments were accomplished in the presence of a PMF consisting of a maximum ΔpH ~ 4 and an unknown ΔΨ. In contrast, in living bacteria the maximum ΔpH across the plasma membrane is likely 0.75, and ΔΨ has been measured between -80 and -140 mV. Thus the problem of in vivo catalytic turnover rates, or the in vivo rotational speed in single F_oF₁-ATP synthases, respectively, has to be solved. In addition, the absolute number of functional enzymes in a single bacterium required to maintain the high ATP levels has to be determined. We report our progress of measuring subunit rotation in single F_oF₁-ATP synthases in vitro and in vivo, which was enabled by a new labeling approach for single-molecule FRET measurements.

Citation Download Citation

K. Seyfert, T. Oosaka, H. Yaginuma, S. Ernst, H. Noji, R. Iino, and M. Börsch "Subunit rotation in a single F_oF₁-ATP synthase in a living bacterium monitored by FRET", Proc. SPIE 7905, Single Molecule Spectroscopy and Imaging IV, 79050K (28 February 2011); https://doi.org/10.1117/12.873066

ACCESS THE FULL ARTICLE

INSTITUTIONAL
Select your institution to access the SPIE Digital Library.

SELECT YOUR INSTITUTION

PERSONAL
Sign in with your SPIE account to access your personal subscriptions or to use specific features such as save to my library, sign up for alerts, save searches, etc.

PERSONAL SIGN IN

No SPIE Account? Create one

PURCHASE THIS CONTENT

SUBSCRIBE TO DIGITAL LIBRARY

50 downloads per 1-year subscription

Members: $195

Non-members: $335 ADD TO CART

25 downloads per 1 - year subscription

Members: $145

Non-members: $250 ADD TO CART

PURCHASE SINGLE ARTICLE

Includes PDF, HTML & Video, when available