Electron spin relaxation enhancement measurements of interspin distances in human, porcine and Rhodobacter electron transfer flavoprotein-ubiquinone oxidoreductase (ETF-QO)

Fielding, Alistair J., Usselman, Robert J., Watmough, Nicholas J., Simkovic, Martin, Frerman, Frank E., Eaton, Gareth R. and Eaton, Sandra S. (2008) Electron spin relaxation enhancement measurements of interspin distances in human, porcine and Rhodobacter electron transfer flavoprotein-ubiquinone oxidoreductase (ETF-QO). Journal of Magnetic Resonance, 190 (2). pp. 222-232.

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Abstract

Electron transfer flavoprotein–ubiquinone oxidoreductase (ETF–QO) is a membrane-bound electron transfer protein that links primary flavoprotein dehydrogenases with the main respiratory chain. Human, porcine, and Rhodobacter sphaeroides ETF–QO each contain a single [4Fe–4S]2+,1+ cluster and one equivalent of FAD, which are diamagnetic in the isolated enzyme and become paramagnetic on reduction with the enzymatic electron donor or with dithionite. The anionic flavin semiquinone can be reduced further to diamagnetic hydroquinone. The redox potentials for the three redox couples are so similar that it is not possible to poise the proteins in a state where both the [4Fe–4S]+ cluster and the flavoquinone are fully in the paramagnetic form. Inversion recovery was used to measure the electron spin-lattice relaxation rates for the [4Fe–4S]+ between 8 and 18 K and for semiquinone between 25 and 65 K. At higher temperatures the spin-lattice relaxation rates for the [4Fe–4S]+ were calculated from the temperature-dependent contributions to the continuous wave linewidths. Although mixtures of the redox states are present, it was possible to analyze the enhancement of the electron spin relaxation of the FAD semiquinone signal due to dipolar interaction with the more rapidly relaxing [4Fe–4S]+ and obtain point-dipole interspin distances of 18.6 ± 1 Å for the three proteins. The point-dipole distances are within experimental uncertainty of the value calculated based on the crystal structure of porcine ETF–QO when spin delocalization is taken into account. The results demonstrate that electron spin relaxation enhancement can be used to measure distances in redox poised proteins even when several redox states are present.

Item Type:	Article
Faculty \ School:	Faculty of Science > School of Biological Sciences
UEA Research Groups:	Faculty of Science > Research Groups > Molecular Microbiology Faculty of Science > Research Centres > Centre for Molecular and Structural Biochemistry
Depositing User:	EPrints Services
Date Deposited:	01 Oct 2010 13:38
Last Modified:	24 Sep 2024 09:29
URI:	https://ueaeprints.uea.ac.uk/id/eprint/1597
DOI:	10.1016/j.jmr.2007.11.002

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