Oxygen sensitivity of mitochondrial reactive oxygen species generation depends on metabolic conditions.
The mitochondrial generation of reactive oxygen species (ROS) plays a central role in many cell signaling pathways, but debate still surrounds its regulation by factors, such as substrate availability, [O2] and metabolic state. Previously, we showed that in isolated mitochondria respiring on succinate, ROS generation was a hyperbolic function of [O2]. In the current study, we used a wide variety of substrates and inhibitors to probe the O2 sensitivity of mitochondrial ROS generation under different metabolic conditions. From such data, the apparent Km for O2 of putative ROS-generating sites within mitochondria was estimated as follows: 0.2, 0.9, 2.0, and 5.0 microM O2 for the complex I flavin site, complex I electron backflow, complex III QO site, and electron transfer flavoprotein quinone oxidoreductase of beta-oxidation, respectively. Differential effects of respiratory inhibitors on ROS generation were also observed at varying [O2]. Based on these data, we hypothesize that at physiological [O2], complex I is a significant source of ROS, whereas the electron transfer flavoprotein quinone oxidoreductase may only contribute to ROS generation at very high [O2]. Furthermore, we suggest that previous discrepancies in the assignment of effects of inhibitors on ROS may be due to differences in experimental [O2]. Finally, the data set (see supplemental material) may be useful in the mathematical modeling of mitochondrial metabolism.
From the Departments of Biochemistry, Rochester, New York 14642.
David L
DL
Hoffman
Anesthesiology, University of Rochester Medical Center, Rochester, New York 14642. Electronic address: paul_brookes@urmc.rochester.edu.
Paul S
PS
Brookes
D000818:Animals; D042967:Electron Transport Complex I / Q000378:metabolism; D042963:Electron Transport Complex II / Q000378:metabolism; D014450:Electron Transport Complex III / Q000378:metabolism; D004734:Energy Metabolism / Q000502:physiology*; D018698:Glutamic Acid / Q000494:pharmacology; D008293:Malates / Q000494:pharmacology; D008297:Male; D008314:Malonates / Q000494:pharmacology; D008928:Mitochondria / Q000378:metabolism*; D010100:Oxygen / Q000494:pharmacology*; D010101:Oxygen Consumption / Q000502:physiology*; D010172:Palmitoylcarnitine / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D017382:Reactive Oxygen Species / Q000378:metabolism*; D012402:Rotenone / Q000494:pharmacology; D019802:Succinic Acid / Q000494:pharmacology; D014475:Uncoupling Agents / Q000494:pharmacology
D008293:Malates; D008314:Malonates; D017382:Reactive Oxygen Species; D014475:Uncoupling Agents; D012402:Rotenone; D010172:Palmitoylcarnitine; D018698:Glutamic Acid; C030298:malic acid; C030290:malonic acid; D019802:Succinic Acid; D042963:Electron Transport Complex II; D042967:Electron Transport Complex I; D014450:Electron Transport Complex III; D010100:Oxygen