title: Free radical intermediates in the reaction of pyruvate:ferredoxin oxidoreductase in Tritrichomonas foetus hydrogenosomes.
abstract: Aerobic incubations of the Tritrichomonas foetus hydrogenosomal fraction containing pyruvate, CoA, and the spin trap 5,5-dimethyl-1-pyrroline N-oxide (DMPO) gave spectra of two radical adducts. One was a carbon-centered radical adduct of DMPO. This radical was centered at C-3 of pyruvate as determined in experiments using [13C]pyruvate. The other radical detected was identified as the CoA radical adduct of DMPO by comparison with an adduct obtained by incubating CoA with DMPO, H2O2 and horseradish peroxidase. Deletion of CoA led to an increased stability of the carbon-centered radical adduct of DMPO, disappearance of the thiyl radical adduct of DMPO, and appearance of a hydroxyl radical adduct of DMPO. Superoxide dismutase suppressed the appearance of the DMPO-hydroxyl radical adduct but did not have any inhibitory effect on the appearance of the other adducts. Catalase had no significant effect on any of the adducts. Addition of pyruvate to these hydrogenosomal preparations stimulated oxygen consumption. Addition of CoA led to a further increase in the rate of O2 uptake but had no effect in the absence of pyruvate. The formation of two substrate free radicals as intermediates in the generation of acetyl-CoA represents a novel mechanism for this enzymatic reaction and indicates that the pyruvate:ferredoxin oxidoreductase from T. foetus differs significantly from the pyridine nucleotide-dependent pyruvate dehydrogenase complex of other eukaryotic cells in its catalytic mechanism.
mesh_terms: D004578:Electron Spin Resonance Spectroscopy; D005609:Free Radicals; D007658:Ketone Oxidoreductases / Q000378:metabolism*; D008830:Microbodies / Q000201:enzymology; D010084:Oxidation-Reduction; D050743:Pyruvate Synthase; D013113:Spin Labels; D014318:Tritrichomonas / Q000201:enzymology*
chemical_list: D005609:Free Radicals; D013113:Spin Labels; D007658:Ketone Oxidoreductases; D050743:Pyruvate Synthase