FIGURE 3.

A, optical absorption difference spectra of the reaction products by direct mixing of O₂ with CO-MV (mixed valence) Y167F aa₃ oxidase from P. denitrificans minus the oxidized form of the enzyme at the indicated times subsequent to mixing, pH 7.5. The enzyme concentration was 5 μm, and the path length of the cell was 0.5 cm. B, resonance Raman spectra of CO-MV Y167F at 0–5 min subsequent to mixing with ¹⁶O₂ (spectrum a) and ¹⁸O₂ (spectrum b). The inset shows the difference a-b spectrum. Several 0–5-min spectra were collected and added for the ¹⁶O₂ and ¹⁸O₂ experiments. The enzyme concentration was 50 μm, pH 7.5. The excitation wavelength was 428.7 nm, and the incident power was 1 milliwatt.