Photoreactions of cytochrome c oxidase

The photoreduction of oxidized bovine heart cytochrome c oxidase (CcO) by visible and UV radiation was investigated in the absence and presence of external reagents. In the former case, the quantum yields for direct photoreduction of heme A (heme a + heme a(3)) were 2.6 +/- 0.5 x 10(-3), 4 +/- 1 x 10(-4), and 4 +/- 2 x 10(-6) with pulsed laser irradiation at 266, 355 and 532 run, respectively. Within experimental uncertainty, the quantum yields did not depend on pulse energy, implying that the mechanism is monophotonic. Irradiation with 355 rim light resulted in spectral changes similar to those produced independently by reduction with dithionite, whereby the low-spin heme a and Cu-A are reduced first. Extended illumination at 355 and 532 nm yielded substantial amounts of reduced heme a(3). Heme decomposition was noted with 266 nm light. In the presence of formate and cyanide ions, which bind at the binuclear heme a(3)/copper center in CcO, irradiation at 355 nm caused selective reduction of only the low-spin heme a and Cu-A. The addition of ferrioxalate ion dramatically increased the efficiency of cytochrome c oxidase photoreduction. The quantum efficiency for heme A reduction was found to be near unity, significantly greater than for other known methods of photoreduction. The active reductant is most likely ferrous iron, and its reduction of the enzyme is thermodynamically driven by the reformation of ferrioxalate in the presence of excess oxalate ion. Other metalloenzymes with redox potentials similar to those of cytochrome c oxidase should be amenable to indirect photoreduction by this method.