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Effect of carbon substrate and aeration on nitrate reduction and expression of the periplasmic and membrane-bound nitrate reductases in carbon-limited continuous cultures of Paracoccus denitrificans Pd1222

Centre for Metalloprotein Spectroscopy and Biology, School of Biological Sciences, University of East Anglia, Norwich NR4 7TJ, UK

Author for correspondence: David J. Richardson. Tel: +44 1603 593250. Fax: 4-44 1603 592250. e-mail: d.richardson@uea.ac.uk

ABSTRACT

The expression and in situ activity of the membrane-bound and periplasmic nitrate reductases have been assayed in Paracoccus denitrificans Pd1222 grown under a range of aeration regimes in malate-limited or butyrate-limited chemostat cultures. In butyrate-limited cultures the expression of periplasmic nitrate reductase and the rate of in situ nitrate reduction were high at all oxygen concentrations measured between 0% and 100% air saturation. By contrast, in malate-limited cultures expression of the periplasmic nitrate reductase was low at 80-100% air saturation but increased to a maximum between 20% and 50% air saturation. Aerobic nitrate reduction was much higher in butyrate-limited than in malate-limited cultures, demonstrating a significant role for this process during butyrate metabolism. The rate of nitrate respiration increased in both the malate- and butyrate-limited cultures as aerobic metabolism switched completely to anaerobic metabolism. Expression of the membrane-bound nitrate reductase could be detected in butyrate-limited chemostat cultures maintained at an oxygen level of 100% air saturation. No membrane-bound nitrate reductase was detectable under similar conditions in malate-limited cultures but expression was detected at oxygen concentrations of 50% air saturation and below. Taken together, the results show that the nature of the carbon substrate and oxygen concentration can both influence expression of the periplasmic and membrane-bound nitrate reductases. The conditions under which expression of the periplasmic nitrate reductase and aerobic nitrate respiration are maximal can be rationalized in terms of a role for the periplasmic nitrate reductase in dissipating excess reductant generated during oxidative metabolism of reduced carbon substrates.

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