EPR characterization of the iron-sulfur-containing NADH-ubiquinone oxidoreductase of the Escherichia coli aerobic respiratory chain.
The energy coupled NADH-ubiquinone (Q) oxidoreductase segment of the respiratory chain of Escherichia coli GR19N has been studied by EPR spectroscopy. Previously Matsushita et al. [(1987) Biochemistry 26, 7732-7737] have demonstrated the presence of two distinct NADH-Q oxidoreductases in E. coli membrane particles and designated them NADH dh I and NADH dh II. Although both enzymes oxidize NADH, only NADH dh I is coupled to the formation of the H+ electrochemical gradient. In addition to NADH, NADH dh I oxidizes nicotinamide hypoxanthine dinucleotide (deamino-NADH), while NADH dh II does not. In membrane particles we have detected EPR signals arising from four low-potential iron-sulfur clusters, one binuclear, one tetranuclear, and two fast spin relaxing g perpendicular = 1.94 type clusters (whose cluster structure has not yet been assigned). The binuclear cluster, temporarily designated [N-1]E, shows an EPR spectrum with gx,y,z = 1.92, 1.935, 2.03 and the Em7.4 value of -220 mV (n = 1). The tetranuclear cluster, [N-2]E, elicits a spectrum with gx,y,z = 1.90, 1.91, 2.05 and an Em7.4 of -240 mV (n = 1). These two clusters have been shown to be part of the NADH dh I complex by stability and inhibitor studies. When stored at 4 degrees C, both clusters are extremely labile as is the deamino-NADH-Q oxidoreductase activity. Addition of deamino-NADH in the presence of piericidin A results in nearly full reduction of [N-2]E within 17 s. In membrane particles pretreated with piericidin A, the cluster [N-1]E is only partly reducible by deamino-NADH and shows an altered line shape.(ABSTRACT TRUNCATED AT 250 WORDS)
Meinhardt
S W
SW
Department of Biochemistry and Biophysics, University of Pennsylvania, Philadelphia 19104.
Matsushita
K
K
Kaback
H R
HR
Ohnishi
T
T
D001426:Bacterial Proteins; D004578:Electron Spin Resonance Spectroscopy; D004579:Electron Transport; D004926:Escherichia coli / Q000201:enzymology; D007506:Iron-Sulfur Proteins; D008565:Membrane Proteins; D008667:Metalloproteins; D016660:NAD(P)H Dehydrogenase (Quinone); D010084:Oxidation-Reduction; D011725:Pyridines / Q000502:physiology; D011808:Quinone Reductases; D013816:Thermodynamics
D001426:Bacterial Proteins; D007506:Iron-Sulfur Proteins; D008565:Membrane Proteins; D008667:Metalloproteins; D011725:Pyridines; C100213:piericidin A; D016660:NAD(P)H Dehydrogenase (Quinone); D011808:Quinone Reductases