Peroxidatic activities of two superoxide scavengers: Cytochrome c and superoxide dismutase.

摘要

Both cytochrome c and superoxide dismutase are superoxide scavengers whose respective physiological functions are well established. Research on these enzymes has undergone intense revival following two discoveries: the role of cytochrome c in apoptosis, and that of superoxide dismutase in familial ALS. In neither case has the cytotoxic action been fully explained in terms of the enzyme's redox properties. We decided therefore to reinvestigate peroxidatic activities of cytochrome c and superoxide dismutase with relevance to positive feedback in apoptosis and ALS.; The data confirm that peroxidatic activity of native Cu,Zn-superoxide dismutase is minimal unless small molecules such as bicarbonate release this activity. Removing zinc increased H2O2-induced oxidation of TMPD by 8-fold compared with the native enzyme, and this enhanced peroxidatic activity was independent of the presence of bicarbonate. The data also confirm the strong peroxidatic activity of cytochrome c. Pre-incubating cytochrome c with H2O2 increased peroxidatic activity about thirty-fold within 10s followed by a sharp decline. The H2O2-induced release in peroxidatic activity is accompanied by aggregation the protein, without detectible scission that might reflect production of “microperoxidase”. Taking previous and current data together, cytochrome c clearly has a powerful but latent capability for positive feedback in the oxidative destruction of living cells.; In characterizing the peroxidatic activity of cytochrome c we report: (1) increasing reaction temperature increased peroxidatic activity of cytochrome c, (2) the reaction rate in HEPES buffer was 30% less than in phosphate buffer, (3) adding EDTA increased reaction rate by 76%, (4) optimal pH for peroxidatic activity was 8, (5) increasing ionic strength slowed the reaction, (6) adding halides increased peroxidatic activity, (7) calcium and zinc inhibited, while copper slowed the reaction, (8) trolox and the flavonoid anthocyanin inhibited by 20% and 85%, respectively; (9) ascorbate inhibited virtually completely until it was all oxidized, after which peroxidatic activity was greater than controls.; Pending further research it seems safe to say that under oxidative stress these novel capabilities for the positive feedback open new avenues for cytotoxic peroxidations. Some of these may well potentiate the apoptotic sequence or participate in the pathological processes of diseases.