RESUMO
Bradyrhizobium japonicum, a symbiotic nitrogen-fixing bacterium, has a complex respiratory electron-transport chain, capable of functioning throughout a wide range of oxygen tensions. It does so by synthesizing a number of terminal oxidases, each appropriate for different environmental conditions. We have previously described the cloning of the large catalytic subunit, coxX, from one of the terminal oxidases from B. japonicum [Surpin, M.A., Moshiri, F., Murphy, A.M. and Maier, R.J. (1994) Genetic evidence for a fourth terminal oxidase from Bradyrhizobium japonicum. Gene 143, 73-77]. In this work, we describe the remaining subunits of this terminal oxidase complex, which is encoded by the coxWXYZ operon. The polypeptide encoded by coxW does not contain any amino acid residues that are known to bind the CuA atom of cytochrome c terminal oxidases, but contains residues thought to be involved in ubiquinol binding. Terminal oxidase cyanide inhibition titration pattern comparisons of the wild type with a coxWXYZ insertion mutant indicated the new oxidase is expressed microaerobically. However analysis of hemes extracted from microaerobically incubated cells revealed the absence of heme O in this strain (from both the wild type and the mutant) of B. japonicum. Therefore, coxWXYZ most likely encodes a microaerobically-expressed bb3-type ubiquinol oxidase.
Assuntos
Proteínas de Membrana , Família Multigênica/genética , Oxirredutases/genética , Rhizobiaceae/genética , Sequência de Aminoácidos , Proteínas de Bactérias/genética , Genes Bacterianos/genética , Heme/análise , Resistência a Canamicina/genética , Dados de Sequência Molecular , Mutagênese Insercional , Fases de Leitura Aberta/genética , Óperon/genética , Oxirredutases/química , Mapeamento por Restrição , Homologia de Sequência de AminoácidosRESUMO
Bradyrhizobium japonicum, a symbiotic nitrogen-fixing bacterium, has a complex respiratory electron-transport chain, capable of functioning throughout a wide range of oxygen tensions. It does so by synthesizing a number of terminal oxidases, each appropriate for different environmental conditions. Several genes encoding terminal oxidases from B. japonicum have been cloned, but it is unknown what roles these individual oxidases play. In this paper, we describe the cloning and sequencing of the coxX gene encoding the large catalytic subunit for a fourth terminal oxidase from B. japonicum. The coxX gene encodes a 666-amino-acid (aa) protein (M(r) 74,527) that exhibits a high degree of homology to terminal oxidase proteins from a number of prokaryotic and eukaryotic species. This new oxidase exhibits greater homology to the Escherichia coli cytochrome o subunit I than any of the previously reported B. japonicum terminal oxidase genes.
