Cloning, sequencing and deletion from the chromosome of the gene encoding subunit I of the aa₃‐type cytochrome c oxidase of Rhodobacter sphaeroides

The ctaD gene encoding subunit I of the aa₃‐type cytochrome c oxidase from Rhodobacter sphaeroides has been cloned. The gene encodes a polypeptide of 565 residues which is highly homologous to the sequences of subunit I from other prokaryotic and eukaryotic sources, e.g. 51% identity with that from bovine, and 75% identity with that from Paracoccus denitrificans. The ctaD gene was deleted from the chromosome of R. sphaeroides, resulting in a strain that spectroscopically lacks cytochrome a. This strain maintains about 50% of the cytochrome c oxidase activity of the wild‐type strain owing to the presence of an alternate o‐type cytochrome c oxidase. The aa₃‐type oxidase was restored by complementing the chromosomal deletion with a plasmid‐borne copy of the CtaD gene. This system is well suited for site‐directed mutagenesis probing of the structure and function of cytochrome c oxidase.