Optical and magneto-optical activity of cytochrome bd from Geobacillus thermodenitrificans

Cytochromes bd are terminal oxidases in the respiratory chains of many prokaryotic organisms. They reduce O2 to 2H2O at the expense of electrons extracted from quinol. The oxidases can be divided into two subfamilies, L and S, based on the presence of either a long or a short hydrophilic connection between transmembrane helices 6 and 7 in subunit I designated as ‘Q-loop’. The L-subfamily members, e.g. the enzyme from Escherichia coli, are relatively well-studied and were shown to generate proton-motive force. The S-subfamily comprises the majority of cytochromes bd including the enzyme from Geobacillus thermodenitrificans but is very poor studied. We compared the properties of cytochromes bd from G. thermodenitrificans and E. coli at room temperature using a combination of absorption, CD and MCD spectroscopy. The G. thermodenitrificans enzyme does contain the high-spin heme bHS (“b595”) despite the fact that its characteristic Q00-band (“α”-band) at 595 nm is not seen in the absorption spectra; stoichiometry of hemes bLS, bHS and d per the enzyme complex is suggested to be 1:1:1. At 1 mM CO, 20–25% of ferrous heme bHS in the G. thermodenitrificans oxidase binds the ligand, while in case of the E. coli enzyme such a reaction is minor. In the G. thermodenitrificans oxidase, the excitonic interaction between ferrous hemes bHS and d decreased as compared to that in the E. coli bd. The latter may suggest that the two enzymes differ in the distance between heme d and heme bHS and/or in the angle between their porphyrin planes.

► G. thermodenitrificans cytochrome bd studied by absorption, CD and MCD spectroscopy.
► Purified enzyme retains high-spin heme bHS (previously called “b595”).
► At 1 mM CO, about 20–25% of high-spin ferroheme bHS binds the ligand.
► Compared to E. coli, excitonic interaction between ferrohemes bHS and d decreased.