The coordinate regulation of multiple terminal oxidases by the Pseudomonas putida ANR global regulator

Authors

  • Ana Ugidos,

    1. Departamento de Biotecnología Microbiana, Centro Nacional de Biotecnología, CSIC, Campus de la Universidad Autónoma de Madrid, Cantoblanco, 28049 – Madrid, Spain.
    2. Department of Life Sciences, Faculty of Natural Sciences, Imperial College London, Sir Alexander Fleming Building, London SW7 2AZ, UK.
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  • Gracia Morales,

    1. Departamento de Biotecnología Microbiana, Centro Nacional de Biotecnología, CSIC, Campus de la Universidad Autónoma de Madrid, Cantoblanco, 28049 – Madrid, Spain.
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  • Eduardo Rial,

    1. Centro de Investigaciones Biológicas, CSIC, Ramiro de Maeztu 9, 28040 – Madrid, Spain.
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  • Huw D. Williams,

    1. Department of Life Sciences, Faculty of Natural Sciences, Imperial College London, Sir Alexander Fleming Building, London SW7 2AZ, UK.
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  • Fernando Rojo

    Corresponding author
    1. Departamento de Biotecnología Microbiana, Centro Nacional de Biotecnología, CSIC, Campus de la Universidad Autónoma de Madrid, Cantoblanco, 28049 – Madrid, Spain.
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*E-mail frojo@cnb.csic.es; Tel. (+34) 91 585 45 39; Fax (+34) 91 585 45 06.

Summary

Pseudomonas putida KT2440 contains a branched aerobic respiratory chain with multiple terminal oxidases. Their relative proportion varies according to environmental conditions. The role of the oxygen-responsive ANR global regulator on expression of these terminal oxidases was analysed. During exponential growth in a highly aerated complete medium, ANR activated expression of the Cbb3-1 terminal oxidase (equivalent to Pseudomonas aeruginosa Cbb3-2), but had little role on expression of other terminal oxidases. In early stationary phase, or under oxygen limitation, inactivation of the anr gene led to increased expression of the bo3-type cytochrome (Cyo) and cyanide-insensitive (CIO) terminal oxidases, and to a much lower expression of Cbb3-1. DNase I footprints identified ANR binding sites at the promoters for these oxidases. Their location suggests that ANR is a transcriptional activator of Cbb3-1 genes and a repressor of CIO genes, consistent with expression data. ANR binding sites at the promoter for Cyo genes suggests a complex regulation in combination with other factors. Therefore, ANR coordinates expression of Cyo, CIO and Cbb3-1, but does not influence cytochrome aa3 and Cbb3-2 terminal oxidases under the conditions analysed. Functional assays showed that Cyo has a leading role during aerobic exponential growth, while Cbb3-1 becomes very important in stationary phase.

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