Cytochrome oxidase is a membrane protein complex that catalyses reduction of molecular oxygen to water and utilises the free energy of this reaction to generate a transmembrane proton gradient during respiration. The electron entry site in subunit II is a mixed-valence dinuclear copper centre in the enzymes which oxidize cytochrome c. This centre has been lost during the evolution of the quinol-oxidizing branch of cytochrome oxidases but can be restored by engineering (Van der Oost et al. (1992) EMBO J. 11, 3209-3217). Here we describe the crystal structures of the periplasmic fragment from the wild-type CyoA subunit of the Escherichia coli quinol oxidase and of the mutant with the engineered dinuclear copper centre ("purple CyoA") at high resolution. CyoA is folded as an 11-stranded, mostly antiparallel b-sandwich followed by three a-helices. The dinuclear copper centre is located at the loops between strands b5-b6 and b9-b10. The two coppers are at 2.5 Å distance and symmetrically coordinated to the main ligands which are two bridging cysteines and two terminal histidines. The residues that are distinct in cytochrome c and quinol oxidases are around the dinuclear copper centre. Structural comparison suggests a common ancestry for subunit II of cytochrome oxidase and blue copper-binding proteins.