Succinate-dependent metabolism in Trypanosoma cruzi epimastigotes.
Trypanosoma cruzi epimastigotes permeabilized with digitonin (65 micrograms (mg protein)-1) to measure mitochondrial respiration were exposed to different substrates. Although none of the NADH-dependent substrates stimulated respiration, succinate supported not only oxygen consumption but also oxidative phosphorylation (respiratory control ratio of 1.9 +/- 0.3) indicating that the mitochondria were coupled. The rate of NADH-dependent oxygen consumption by membrane fractions (9.4 +/- 0.7 nmol min-1 (mg protein)-1) was reduced by 50% upon addition of catalase indicating that the electrons from NADH oxidation reduced oxygen to H2O2. NADH-dependent H2O2 production (16 +/- 1 nmol min-1 (mg protein)-1) was confirmed using cytochrome c peroxidase. This activity was inhibited by fumarate by 70%, suggesting a competition between fumarate and oxygen for the electrons from NADH, probably at the fumarate reductase level. The respiratory chain inhibitor antimycin blocked both respiration by intact cells and succinate-dependent cytochrome c by isolated membranes. No inhibition by antimycin was observed when NADH replaced succinate as an electron donor, indicating that the electrons from NADH oxidation reduced cytochrome c through a different route. Malonate blocked not only succinate-cytochrome c reductase and fumarate reductase, but also intact cell motility. These results suggest that succinate has a central role in the intermediate metabolism of i. cruzi, as it may be used for respiration or excreted to the extracellular space under anaerobic conditions. In addition, 2 potential sources of H2O2 were tentatively identified as: (a) the enzyme fumarate reductase; and (b) a succinate-dependent site, which may be the semiquinone form of Coenzyme Q9, as in mammalian mitochondria.
Denicola-Seoane
A
A
Department of Biochemistry, School of Medicine, Universidad de la República, Montevideo, Uruguay.
Rubbo
H
H
Prodanov
E
E
Turrens
J F
JF
D000818:Animals; D005650:Fumarates / Q000378:metabolism; D005947:Glucose / Q000378:metabolism; D006861:Hydrogen Peroxide / Q000378:metabolism*; D009243:NAD / Q000378:metabolism*; D010101:Oxygen Consumption / Q000502:physiology; D011392:Proline / Q000378:metabolism; D013386:Succinates / Q000378:metabolism*; D019802:Succinic Acid; D014349:Trypanosoma cruzi / Q000378:metabolism*
D005650:Fumarates; D013386:Succinates; D009243:NAD; C032005:fumaric acid; D011392:Proline; D019802:Succinic Acid; D006861:Hydrogen Peroxide; D005947:Glucose