Electron and proton transfer in the ba3 oxidase from Thermus thermophilus
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- Smirnova, I.A., Zaslavsky, D., Fee, J.A. et al. J Bioenerg Biomembr (2008) 40: 281. doi:10.1007/s10863-008-9157-7
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The ba3-type cytochrome c oxidase from Thermus thermophilus is phylogenetically very distant from the aa3–type cytochrome c oxidases. Nevertheless, both types of oxidases have the same number of redox-active metal sites and the reduction of O2 to water is catalysed at a haem a3-CuB catalytic site. The three-dimensional structure of the ba3 oxidase reveals three possible proton-conducting pathways showing very low homology compared to those of the mitochondrial, Rhodobacter sphaeroides and Paracoccus denitrificans aa3 oxidases. In this study we investigated the oxidative part of the catalytic cycle of the ba3-cytochrome c oxidase using the flow-flash method. After flash-induced dissociation of CO from the fully reduced enzyme in the presence of oxygen we observed rapid oxidation of cytochrome b (k≅6.8 × 104 s−1) and formation of the peroxy (PR) intermediate. In the next step a proton was taken up from solution with a rate constant of ∼1.7 × 104 s−1, associated with formation of the ferryl (F) intermediate, simultaneous with transient reduction of haem b. Finally, the enzyme was oxidized with a rate constant of ∼1,100 s−1, accompanied by additional proton uptake. The total proton uptake stoichiometry in the oxidative part of the catalytic cycle was ~1.5 protons per enzyme molecule. The results support the earlier proposal that the PR and F intermediate spectra are similar (Siletsky et al. Biochim Biophys Acta 1767:138, 2007) and show that even though the architecture of the proton-conducting pathways is different in the ba3 oxidases, the proton-uptake reactions occur over the same time scales as in the aa3-type oxidases.