STRUCTURE OF SUBUNIT II OF A QUINOL OXIDASE WITH REENGINEERED CuA SITE. Matthias Wilmanns, Kristina Djinovic, Pekka Lappalainen, Mark Kelly, Elisabeth Sauer-Deriksson & Matti Saraste, EMBL Heidelberg, Postfach 102209, D-69012 Heidelberg, Germany

The crystal structures of the periplasmic fragment from the wild-type CyoA subunit II of the Escherichia coli quinol oxidase and of a mutant with a reengineered dinuclear copper centre ("purple CyoA") have been solved at 2.3 and 2.5 Å, respectively. Quinol oxidases belong to the superfamily of cytochrome oxidases. This enzyme is a member of the protein complex that catalyses reduction of molecular oxygen to water and utilizes 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 in the enzymes which oxidize cytochrome c. This centre has been lost during the evolution of the quinol-oxidizing branch of cytochrome oxidases.

CyoA is folded as a 11-stranded, mostly antiparallel ß-sandwich followed by three [[alpha]]-helices. The dinuclear copper centre is located at the loops between strands ß5-ß6 and ß9-ß10. 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. A recent structure of CyoA with reduced dinuclear copper centre shows a virtual identical arrangement of the two coppers except for increased distances between the two terminal histidines and the copper ions. Structural comparison suggests a common ancestry for subunit II of cytochrome oxidase and blue copper proteins.