Subunit inequivalence in superoxide anion formation during photooxidation of human oxyhemoglobin.

Subunit inequivalence in the photooxidation of human oxyhemoglobin was investigated by quantitative analysis of the fraction of alpha and beta chains oxidized after low intensity flash photolysis using white light in quartz cuvettes. This reaction was previously shown to generate methemoglobin and superoxide anion as photoproducts (Demma, L.S., and Salhany, J.M. (1977) J. Biol. Chem. 252, 1226-1230). The present results indicate that superoxide anion photodissociates from the alpha chain about 3 to 4 times more extensively than from beta. This difference was observed in the presence of superoxide dismutase and catalase at both pH 7 and 9, suggesting that photolysis is directly responsible. The reaction of superoxide anion with oxyhemoglobin was also studied with the same analytical methods and no chain differences could be observed. If the electronic structure of the oxyheme complex is viewed as a spin equilibrium between a singlet ground state and a charge transfer configuration, our results may indicate that the uv component of white light perturbs this equilibrium to a greater extent in the oxygenated alpha chain, implying that the separation of energy levels may be smaller in that chain as compared with beta.