Structure of the binuclear heme iron-copper site in the quinol-oxidizing cytochrome aa₃ from Bacillus subtilis

Cytochrome aa₃-600 is a terminal quinol oxidase of Bacillus subtilis, belonging to the large family of structurally and functionally related respiratory enzymes to which the mitochondrial cytochrome c oxidase also belongs. However, the Cu_A center typical of the cytochrome c oxidases is lacking from cytochrome aa₃-600. The presence of only one copper, viz. Cu_B of the binuclear heme iron-copper site, makes cytochrome aa₃-600 especially suitable for XAS analysis of this structure. Cu and Fe XAS data for fully oxidized cytochrome aa₃-600 indicate a structure for the binuclear site similar to that previously reported for mitochondrial cytochrome c oxidase (see Powers et al. (1981) Biophys. J. 34, 465-468). Heme Fe_a3 has a proximal histidine nitrogen ligand 2.10 ± 0.02 A ̊ from the iron, and a distal S or Cl ligand at 2.36 ± 0.03 A ̊. The latter is also a ligand of Cu_B (2.21 ± 0.02 A ̊), and apparently forms a bridge between the two metals which are 3.70 ± 0.06 A ̊ apart. Cu_B has two more close-lying ligands at 1.95 ± 0.02 A ̊, which are likely histidine nitrogens. The similarity between EXAFS of Cu_B and type 1 'blue' copper is contrasted to EPR and optical spectroscopic properties of Cu_B, and the nature of the bridging ligand is discussed.