Pulse Radiolytic Investigations of Superoxide Catalyzed Disproportionation. Mechanism for Bovine Superoxide Dismutase

Bovine superoxide dismutase has been subjected to pulse radiolysis in the presence of 02, EDTA, and formate. The reaction mechanism was investigated by means of optical measurements at 650 and 300 nm, in the presence and in the absence of catalase. The results can be explained on the basis of several reactions involving the enzyme. These are:(a) E⁰ + O₂^- ⟶ E^- + O₂, k = (1.2 ± 0.2) × 10⁹ M^-1 sec^-1; (b) E^- + O₂^- (+2H⁺)⟶E° + H₂O₂, k = (2.2 ± 0.4) × 10₉ M^-1 sec^-1; (c) E^- + O₂^-⟶E^2- + O₂, k = (0-3) × 10⁸ M^-1 sec^-1; (d) E^2- + O₂ (+2H⁺)⟶E^- + H₂O₂, k = (1.2 ± 0.2) × 10⁹ M^-1 sec^-1; (e) E ° + H₂O₂ ⟶ E^2- + 2H⁺ + O₂; (f) E^- + O₂ ⟶E⁰ + O₂^-, k = 0.44 ± 0.12 M^-1 sec^-1. E⁰ refers to the native enzyme in which both copper atoms are oxidized while E^- and E^2- refer to the singly and doubly reduced enzyme, respectively. Reactions a and b account for the activity of the enzyme in the absence of H₂O₂, whereas reactions a, b, and d account for the activity of the modified form of the enzyme which was generated by reaction with H₂O₂, as in reaction e. Reaction f describes the slow reoxidation of reduced enzyme which was observed in the presence of O₂.