NADH oxidation and NAD+ reduction catalysed by tightly coupled inside-out vesicles from Paracoccus denitrificans.
Tightly coupled inside-out vesicles were prepared from Paracoccus denitrificans cells (SPP, sub-Paracoccus particles) and characterized kinetically. The rate of NADH oxidation, catalysed by SPP, increases 6-8 times on addition of gramicidin. The vesicles are capable of catalysing Delta micro H+-dependent reverse electron transfer from quinol to NAD+. The kinetic parameters of the NADH-oxidase and the reverse electron transfer carried out by membrane-bound P. denitrificans complex I were estimated and compared with those of the mitochondrial enzyme. The data demonstrate that catalytic properties of the dinucleotide-binding site of the bacterial and mitochondrial complex I are almost identical, pointing out similar organization of the site in mammals and P. denitrificans. Inhibition of the bacterial complex I by a specific inhibitor of Q reduction, rotenone, is very different from that of the mitochondrial enzyme. The inhibitor is capable of suppressing the NADH oxidation reaction only at micromolar concentrations, while the activity of mitochondrial enzyme is suppressed by nanomolar concentrations of rotenone. In contrast to the mitochondrial enzyme, rotenone, even at concentrations as high as 10 micro m, does not inhibit the reverse, Delta micro H+-dependent NAD+-reductase reaction on SPP.
Kotlyar
Alexander B
AB
Department of Biochemistry, George S. Wise Faculty of Life Sciences, Tel Aviv University, Ramat Aviv, Israel. s2shak@post.tau.ac.it
Borovok
Natalia
N
D000818:Animals; D001426:Bacterial Proteins / Q000378:metabolism*; D001665:Binding Sites; D002384:Catalysis; D004579:Electron Transport; D042967:Electron Transport Complex I; D006096:Gramicidin / Q000494:pharmacology; D007700:Kinetics; D046911:Macromolecular Substances; D008928:Mitochondria / Q000201:enzymology; D009243:NAD / Q000378:metabolism*; D009247:NADH, NADPH Oxidoreductases / Q000378:metabolism*; D010084:Oxidation-Reduction; D010231:Paracoccus denitrificans / Q000737:chemistry*; D018892:Proton-Motive Force; D012402:Rotenone / Q000494:pharmacology; D013386:Succinates / Q000494:pharmacology; D014451:Ubiquinone / Q000378:metabolism
D001426:Bacterial Proteins; D046911:Macromolecular Substances; D013386:Succinates; D012402:Rotenone; D009243:NAD; D014451:Ubiquinone; D006096:Gramicidin; D009247:NADH, NADPH Oxidoreductases; D042967:Electron Transport Complex I