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Journal of Bacteriology, November 2003, p. 6308-6315, Vol. 185, No. 21
0021-9193/03/$08.00+0     DOI: 10.1128/JB.185.21.6308-6315.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

A Mutant of Paracoccus denitrificans with Disrupted Genes Coding for Cytochrome c550 and Pseudoazurin Establishes These Two Proteins as the In Vivo Electron Donors to Cytochrome cd1 Nitrite Reductase

Isobel V. Pearson,1 M. Dudley Page,1 Rob J. M. van Spanning,2 and Stuart J. Ferguson1*

Department of Biochemistry, University of Oxford, Oxford OX1 3QU, United Kingdom,1 Department of Molecular Cell Physiology, Faculty of Biology, Biocentrum Amsterdam, Free University, 1081 HV Amsterdam, The Netherlands2

Received 9 April 2003/ Accepted 8 August 2003

In Paracoccus denitrificans, electrons pass from the membrane-bound cytochrome bc1 complex to the periplasmic nitrite reductase, cytochrome cd1. The periplasmic protein cytochrome c550 has often been implicated in this electron transfer, but its absence, as a consequence of mutation, has previously been shown to result in almost no attenuation in the ability of the nitrite reductase to function in intact cells. Here, the hypothesis that cytochrome c550 and pseudoazurin are alternative electron carriers from the cytochrome bc1 complex to the nitrite reductase was tested by construction of mutants of P. denitrificans that are deficient in either pseudoazurin or both pseudoazurin and cytochrome c550. The latter organism, but not the former (which is almost indistinguishable in this respect from the wild type), grows poorly under anaerobic conditions with nitrate as an added electron acceptor and accumulates nitrite in the medium. Growth under aerobic conditions with either succinate or methanol as the carbon source is not significantly affected in mutants lacking either pseudoazurin or cytochrome c550 or both these proteins. We concluded that pseudoazurin and cytochrome c550 are the alternative electron mediator proteins between the cytochrome bc1 complex and the cytochrome cd1-type nitrite reductase. We also concluded that expression of pseudoazurin is mainly controlled by the transcriptional activator FnrP.

* Corresponding author. Mailing address: Department of Biochemistry, University of Oxford, South Parks Road, Oxford, OX1 3QU, United Kingdom. Phone: 44 1865 275240. Fax: 44 1865 275259. E-mail: stuart.ferguson{at}bioch.ox.ac.uk.

Journal of Bacteriology, November 2003, p. 6308-6315, Vol. 185, No. 21
0021-9193/03/$08.00+0     DOI: 10.1128/JB.185.21.6308-6315.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



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