Spectroscopic evidence for a heme-superoxide/Cu(I) intermediate in a functional model of cytochrome c oxidase

A superstructured tetraphenylporphyrin with a covalently attached proximal imidazole axial base and three distal imidazole pickets has been developed as a model for the active site of terminal oxidases such as cytochrome c oxidase. The oxygen adduct of the Fe-only heme (at low temperature) has a diamagnetic NMR and is EPR silent, which taken together with a resonance Raman oxygen isotope sensitive band (nuFe-O) at 575/554 cm-1 (16O2/18O2) indicates formation of a six-coordinate heme-superoxide complex. Unexpectedly, the Fe/Cu complex, where the copper is in a trisimidazole environment approximately 5 A above the heme plane, displays similar characteristics: a diamagnetic NMR, EPR silence, and nuFe-O at 570/544 cm-1. This indicates the dioxygen adduct of this Fe/Cu system is also a superoxide. This contrasts with previously characterized partially reduced dioxygen intermediates of binuclear heme/copper complexes that form Fe/Cu mu-peroxo complexes.