Sanghwa Han

Vibrational structure of the formyl group on heme a. Implications on the properties of cytochrome c oxidase

Resonance Raman spectra have been recorded for heme a derivatives in which the oxygen atom of the formyl group has been isotopically labeled and for Schiff base derivatives of heme a in which the Schiff base nitrogen has been isotopically labeled. The 14N-15N isotope shift in the C = N stretching mode of the Schiff base is close to the theoretically predicted shift for an isolated C = N group for both the ferric and ferrous oxidation states and in both aqueous and nonaqueous solutions. In contrast, the 16O-18O isotope shift of the C = O stretching mode of the formyl group is significantly smaller than that predicted for an isolated C = O group and is also dependent on whether the environment is aqueous or nonaqueous. This differences between the theoretically predicted shifts and the observed shifts are attributed to coupling of the C = O stretching mode to as yet unidentified modes of the heme. The complex behavior of the C = O stretching vibration precludes the possibility of making simple interpretations of frequency shifts of this mode in cytochrome c oxidase.

Time-resolved resonance Raman spectroscopy of intermediates in cytochrome oxidase.

This chapter focuses on time-resolved resonance Raman spectroscopy of intermediates in cytochrome oxidase. Cytochrome oxidase is the membrane-bound terminal enzyme in the electron transfer chain. It serves the dual role of generating a proton gradient by coupling redox events to proton translocation and maintaining continued electron flow for oxidative phosphorylation by catalyzing the four-electron reduction of O 2 to H 2 0. The reaction of this enzyme is very complex. However, great progress in its understanding has been made by resonance Raman studies. Most of the intermediates in the reaction scheme have been identified and their kinetics determined. The reaction scheme of the enzyme is divided into two phases. In the oxidative phase, the four-electron reduced enzyme reacts with oxygen and becomes fully oxidized. In the reductive phase, the enzyme is rereduced. The first intermediate in the reaction is labeled as compound A and is considered to be a ferric-superoxide species. In the Raman spectrum, this species has an iron-oxygen stretching mode at 568 cm -1 .

Intermediates Formed in the Reaction of Cytochrome c Oxidase with Oxygen

Cytochrome c oxidase is the terminal enzyme in the electron transport chain. In this role, it catalyzes the transfer of four electrons from cytochrome c to di-oxygen, the physiological substrate. Oxygen binds to cytochrome c oxidase on the microsecond time scale and is fully reduced to water in milliseconds. An understanding of the molecular basis for the catalytic mechanism of the oxygen reduction has been sought after for many years but has remained elusive due to the difficulty in establishing the identity of each intermediate on this time scale by optical spectroscopic techniques. Recently, we [1–5] and others [6–11] have applied resonance Raman spectroscopy to the study of the oxygen reduction process and achieved considerable success. In this paper, we discuss the findings from the resonance Raman scattering experiments and based on these results present a model for the catalytic pathway.