Cytochrome aa₃ from heterocysts of the cyanobacterium Anabaena variabilis: Isolation and spectral characterization

Cytochrome c oxidase (ferrocytochrome-c:oxygen oxidoreductase, EC 1.9.3.1) has been prepared from heterocysts of Anabaena variabilis. The enzyme was solubilized with desoxycholate (0.2%) and octyl-β-d-thioglucoside (1%) followed by purification on two anion-exchange columns in the presence of genapol x-080. Purification factors of 145 with regard to protein and of 1373 with regard to chlorophyll were obtained. The γ-peak of the oxidized enzyme is at 420 nm. The difference spectra (oxidized/reduced) show absorption peaks at 440, 517 and 604 nm. The CO compound of the reduced enzyme exhibits maxima at 435 and 586 nm. The absorption characteristics including CO-difference spectra together with a high cyanide sensitivity (K_i = 0.5 μM) indicate that the oxidase is of the aa₃ type.