The intermediates formed in the reaction of oxygen with mixed-valence cytochrome c oxidase were followed as a function of time by monitoring the resonance Raman and optical absorption spectra with a flow-flash-probe technique. At early times (50 μs), the optical absorption spectra confirm the presence of the primary intermediate (Im). This intermediate has an Fe-O2 stretching mode in the resonance Raman spectrum at 568 cm-1 that was used as a marker for its population. By following the change in intensity of this mode, the first-order rate constant for the decay of Im, was measured to be 4.5 × 103 s-1. The formation of the next intermediate (IIm) of the enzyme with a 607-nm peak in the optical absorption difference spectrum was monitored by its optical absorption spectrum and displays a first-order rate constant for its formation, which is the same as the rate constant for the decay Im. The resonance Raman spectrum of Im and IIm are nearly identical, indicating that the π-electron depletion of the porphyrin macrocycle in cytochrome a3 is the same for these two intermediates. The absorption spectrum and the resonance Raman spectrum of an intermediate formed by mixing oxygen with CO-bound mixed-valence cytochrome c oxidase and allowing O2 to spontaneously replace CO are the same as those of IIm formed in the photolysis experiments. IIm, formed in this way, decays biphasically to the resting form of the enzyme.
|Original language||English (US)|
|Journal||Journal of the American Chemical Society|
|Publication status||Published - Dec 19 1990|
ASJC Scopus subject areas
- Colloid and Surface Chemistry