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Cloning, sequencing and deletion from the chromosome of the gene encoding subunit I of the aa 3 ‐type cytochrome c oxidase of Rhodobacter sphaeroides
Author(s) -
Shapleigh James P.,
Gennis Robert B.
Publication year - 1992
Publication title -
molecular microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.857
H-Index - 247
eISSN - 1365-2958
pISSN - 0950-382X
DOI - 10.1111/j.1365-2958.1992.tb01511.x
Subject(s) - rhodobacter sphaeroides , cytochrome c oxidase , biology , oxidase test , gene , microbiology and biotechnology , cytochrome , protein subunit , electron transport complex iv , mutagenesis , cytochrome b , site directed mutagenesis , genetics , biochemistry , mutation , mutant , enzyme , mitochondrial dna , mitochondrion , bacteria
Summary The ctaD gene encoding subunit I of the aa 3 ‐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 aa 3 ‐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.