MUTAGENESIS OF A GENE ENCODING A CYTOCHROME-O-LIKE TERMINAL OXIDASE OF AZOTOBACTER-VINELANDII - A CYTOCHROME-O MUTANT IS AERO-TOLERANT DURING NITROGEN-FIXATION
D. Leung et al., MUTAGENESIS OF A GENE ENCODING A CYTOCHROME-O-LIKE TERMINAL OXIDASE OF AZOTOBACTER-VINELANDII - A CYTOCHROME-O MUTANT IS AERO-TOLERANT DURING NITROGEN-FIXATION, FEMS microbiology letters, 119(3), 1994, pp. 351-357
The amino acid sequence obtained by translating the nucleotide sequenc
e of a 0.55 kb fragment, amplified from Azotobacter vinelandii chromos
omal DNA by PCR, was 57% identical to part, of the Escherichia coli cy
oB gene, encoding subunit I of the cytochrome bo-type quinol oxidase.
This fragment was mutated in vitro by insertion of a kanamycin-resista
nce cassette and introduced into the chromosome of A. vinelandii by ho
mologous recombination. The mutant contained no spectrally detectable
cytochrome o. However, in the stationary phase of growth, the level of
the alternative oxidase (cytochrome bd) was 11-fold higher than in th
e wild-type strain. Respiration of the mutant was insensitive to chlor
promazine, an inhibitor thought to act specifically on cytochrome o. C
ytochrome o-deficient mutants fixed nitrogen in air, clearly distingui
shing the role of this oxidase from that of cytochrome bd, which is re
quired for respiratory protection of oxygen-labile nitrogenase.