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Evidence for a key role of cytochrome bo3 oxidase in respiratory energy metabolism of Gluconobacter oxydans

ABSTRACT: The obligatory aerobic acetic acid bacterium Gluconobacter oxydans oxidizes a variety of substrates in the periplasm by membrane-bound dehydrogenases, which transfer the reducing equivalents to ubiquinone. Two quinol oxidases, cytochrome bo3 and cytochrome bd, then catalyze transfer of the electrons from ubiquinol to molecular oxygen. In this study, mutants lacking either of these terminal oxidases were characterized. Deletion of the cydAB genes for cytochrome bd had no obvious influence on growth, whereas the lack of the cyoBACD genes for cytochrome bo3 severely reduced the growth rate and the cell yield. Using a respiration activity monitoring system and adjusting different levels of oxygen availability, hints for a low oxygen affinity of cytochrome bd oxidase were obtained, which were supported by measurements of oxygen consumption in a respirometer. The H+/O ratio of the ΔcyoBACD mutant with mannitol as substrate was 0.56 ± 0.11 and more than 50% lower than that of the reference strain (1.26 ± 0.06) and the delta-cydAB mutant (1.31 ± 0.16), indicating that cytochrome bo3 oxidase is the main component for proton extrusion via the respiratory chain. Plasmid-based overexpression of cyoBACD led to increased growth rates and growth yields, both in the wild type and the delta-cyoBACD mutant, suggesting that cytochrome bo3 might be the rate-limiting factor of the respiratory chain.

ORGANISM(S): Gluconobacter oxydans 621H

PROVIDER: GSE47782 | GEO | 2013/07/26

SECONDARY ACCESSION(S): PRJNA207800

REPOSITORIES: GEO

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Project description:The genome-wide transcriptional responses of the strictly aerobic -proteobacterium Gluconobacter oxydans 621H to oxygen limitation, to the absence of the cytochrome bc1 complex, and to low pH were studied using DNA microarray analyses. Oxygen limitation caused expression changes of 486 genes, representing 20% of the chromosomal genes. Genes with an increased mRNA level included those for terminal oxidases, the cytochrome bc1 complex, transhydrogenase, two alcohol dehydrogenases, heme biosynthesis, PTS proteins, proteins involved in cyclic diGMP synthesis and degradation, two sigma factors, flagella and chemotaxis proteins, several stress proteins, and a putative exporter protein. The downregulated genes comprised those for respiratory dehydrogenases, enzymes of central metabolism, PQQ biosynthesis, outer membrane receptors, Sec proteins, and proteins involved in transcription and translation. A M-NM-^TqrcABC mutant of G. oxydans showed a growth defect during cultivation on mannitol at pH 4 under oxygen saturation. Comparison of the transcriptomes of this mutant versus the wild type under these conditions revealed 51 differentially expressed genes. Interestingly, almost all of the 45 genes with increased expression in the M-NM-^TqrcABC mutant at pH 4 were also upregulated in the wild type grown at pH 6 under oxygen limitation. These results support an active role of the cytochrome bc1 complex in G. oxydans respiration. The transcriptome comparison of G. oxydans wild type at pH 4 versus pH 6 in mannitol medium under oxygen-saturated conditions uncovered only 72 differentially expressed genes. The 35 upregulated genes included those for cytochrome bd oxidase, major polyol dehydrogenase, iron storage and oxidative stress proteins. Among the 37 downregulated genes were some encoding enzymes dealing with carbon dioxide, such as biotin carboxylase, biotin carboxyl carrier protein, and carboanhydrase. These results give first insights into global transcriptional responses of G. oxydans. DNA microarray experiments were repeated independently three times for G. oxydans M-NM-^TqrcABC versus wild type and G. oxydans grown at pH 4 versus pH 6 and four times for G. oxydans grown at oxygen limitation versus pH oxygen saturation in biological replicates.

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Evidence for a key role of cytochrome bo3 oxidase in respiratory energy metabolism of Gluconobacter oxydans

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