Asymmetric binding of the high-affinity Q(H)(*)(-) ubisemiquinone in quinol oxidase (bo3) from Escherichia coli studied by multifrequency electron paramagnetic resonance spectroscopy.
Ubiquinone-2 (UQ-2) selectively labeled with (13)C (I =(1)/(2)) at either the position 1- or the 4-carbonyl carbon is incorporated into the ubiquinol oxidase bo(3) from Escherichia coli in which the native quinone (UQ-8) has been previously removed. The resulting stabilized anion radical in the high-affinity quinone-binding site (Q(H)(*)(-)) is investigated using multifrequency (9, 34, and 94 GHz) electron paramagnetic resonance (EPR) spectroscopy. The corresponding spectra reveal dramatic differences in (13)C hyperfine couplings indicating a strongly asymmetric spin density distribution over the quinone headgroup. By comparison with previous results on labeled ubisemiquinones in proteins as well as in organic solvents, it is concluded that Q(H)(*)(-) is most probably bound to the protein via a one-sided hydrogen bond or a strongly asymmetric hydrogen-bonding network. This observation is discussed with regard to the function of Q(H) in the enzyme and contrasted with the information available on other protein-bound semiquinone radicals.
Institut für Physikalische und Theoretische Chemie, J. W. Goethe Universität Frankfurt, Centre for Biological Magnetic Resonance, Frankfurt am Main, Germany.
S
S
Grimaldi
T
T
Ostermann
N
N
Weiden
T
T
Mogi
H
H
Miyoshi
B
B
Ludwig
H
H
Michel
T F
TF
Prisner
F
F
MacMillan
D001665:Binding Sites; D003067:Coenzymes; D004578:Electron Spin Resonance Spectroscopy; D004926:Escherichia coli / Q000201:enzymology; D006860:Hydrogen Bonding; D007700:Kinetics; D008958:Models, Molecular; D010088:Oxidoreductases / Q000737:chemistry* / Q000378:metabolism*; D055672:Static Electricity; D014451:Ubiquinone / Q000031:analogs & derivatives* / Q000737:chemistry* / Q000378:metabolism*
D003067:Coenzymes; D014451:Ubiquinone; D010088:Oxidoreductases; C020094:duroquinol oxidase; C024989:coenzyme Q10