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Volume 149, Issue 6

Characterization of the expression and activity of the periplasmic nitrate reductase of in chemostat cultures Free

Abstract

The periplasmic nitrate reductase (Nap) from has a role in cellular redox balancing. Previously, transcription from the promoter in was shown to be responsive to the oxidation state of the carbon substrate. During batch culture, expression was higher during growth on reduced substrates such as butyrate compared to more oxidized substrates such as succinate. In the present study the effect of growth rate on expression in succinate-, acetate- and butyrate-limited chemostat cultures was investigated. In all three cases transcription from the promoter and Nap enzyme activity showed a strong correlation. At the fastest growth rates tested for the three substrates expression and Nap activity were highest when growth occurred on the most reduced substrate (butyrate > acetate > succinate). However, in all three cases a bell-shaped pattern of expression was observed as a function of growth rate, with the highest levels of expression and Nap activity being observed at intermediate growth rates. This effect was most pronounced on succinate, where an approximately fivefold variation was observed, and at intermediate dilution rates expression and Nap activity were comparable on all three carbon substrates. Analysis of mRNA prepared from the succinate-grown cultures revealed that different transcription initiation start sites for the operon were utilized as the growth rate changed. This study establishes a new regulatory feature of expression in that occurs at the level of transcription in response to growth rate in carbon-limited cultures.

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Keyword(s): MV, methylviologen and Nap, periplasmic nitrate reductase
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2003-06-01
2024-04-23
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References

  1. Baker S. C., Ferguson S. J., Ludwig B., Page M. D., Richter O.-M. H., Van Spanning R. J. M. 1998; Molecular genetics of the genus Paracoccus : metabolically versatile bacteria with bioenergetic flexibility. Microbiol Mol Biol Rev 62:1046–1078
     [Google Scholar]
  2. Bell L. C., Richardson D. J., Ferguson S. J. 1990; Periplasmic and membrane-bound respiratory nitrate reductases in Thiosphaera pantotropha . The periplasmic enzyme catalyses the first step in aerobic denitrification. FEBS Lett 265:85–87
     [Google Scholar]
  3. Bell L. C., Page M. D., Berks B. C., Richardson D. J., Ferguson S. J. 1993; Insertion of transposon Tn 5 into a structural gene of the membrane-bound nitrate reductase of Thiosphaera pantotropha results in anaerobic overexpression of periplasmic nitrate reductase activity. J Gen Microbiol 139:3205–3214
     [Google Scholar]
  4. Berks B. C., Ferguson S. J., Moir J. W., Richardson D. J. 1995a; Enzymes and electron transport systems that catalyse the respiratory reduction of nitrogen oxides and oxyanions. Biochim Biophys Acta 1232:97–173
     [Google Scholar]
  5. Berks B. C., Richardson D. J., Reilly A., Willis A. C., Ferguson S. J. 1995b; The napEDABC gene cluster encoding the periplasmic nitrate reductase system of Thiosphaera pantotropha . Biochem J 309:983–992
     [Google Scholar]
  6. Brondijk T. H., Fiegen D., Richardson D. J., Cole J. A. 2002; Roles of NapF, NapG and NapH, subunits of the Escherichia coli periplasmic nitrate reductase, in ubiquinol oxidation. Mol Microbiol 44:245–255
     [Google Scholar]
  7. Carter J. P., Hsaio Y. H., Spiro S., Richardson D. J. 1995; Soil and sediment bacteria capable of aerobic nitrate respiration. Appl Environ Microbiol 61:2852–2858
     [Google Scholar]
  8. Castillo F., Dobao M. M., Reyes F., Blasco R., Roldan M. D., Gavira M., Caballero F. J., Moreno-Vivian C., Martinez-Luque M. 1996; Molecular and regulatory properties of the nitrate reducing systems of Rhodobacter . Curr Microbiol 33:341–346
     [Google Scholar]
  9. Coleman K. J., Cornish-Bowden A., Cole J. A. 1978; Purification and properties of nitrite reductase from Escherichia coli K-12. Biochem J 175:483–493
     [Google Scholar]
  10. Darwin A. J., Ziegelhoffer E. C., Kiley P. J., Stewart V. 1998; Fnr, NarP and NarL regulation of Escherichia coli K-12 napF (periplasmic nitrate reductase) operon transcription in vitro . J Bacteriol 180:4192–4198
     [Google Scholar]
  11. Dobao M. M., Martinez-Luque M., Moreno V. C., Castillo F. 1994; Effect of carbon and nitrogen metabolism on nitrate reductase activity of Rhodobacter capsulatus E1F1. Can J Microbiol 40:645–650
     [Google Scholar]
  12. Ellington M. J. K., Bhakoo K. K., Sawers G., Richardson D. J., Ferguson S. J. 2002; Hierarchy of carbon source selection in Paracoccus pantotrophus : strict correlation between reduction state of the carbon substrate and aerobic expression of the nap operon. J Bacteriol 184:4767–4774
     [Google Scholar]
  13. Ellington M. J. K., Richardson D. J., Ferguson S. J. 2003; Rhodobacter capsulatus gains a competitive advantage from respiratory nitrate reduction during light–dark transitions. Microbiology 149:941–948
     [Google Scholar]
  14. Gavira M., Roldan M. D., Castillo F., Moreno-Vivian C. 2002; Regulation of nap gene expression and periplasmic nitrate reductase activity in the phototrophic bacterium Rhodobacter sphaeroides DSM158. J Bacteriol 184:1693–1702
     [Google Scholar]
  15. Harder W., Dijkhuzen L. 1982; Strategies of mixed substrate utilisation in microorganisms. Philos Trans R Soc Lond B 297:459–480
     [Google Scholar]
  16. Jones M. R., Richardson D. J., McEwan A. G., Jackson J. B., Fergsuon S. J. 1990; In vivo redox poising of the cyclic electron transport system of Rhodobacter capsulatus and the effects of auxiliary oxidants nitrate, nitrous oxide and trimethylamine- N -oxide as revealed by multiple short flash excitation. Biochim Biophys Acta 1017:209–216
     [Google Scholar]
  17. Liu H. P., Takio S., Satoh T., Yamamoto I. 1999; Involvement in denitrification of the napKEFDABC genes encoding the periplasmic nitrate reductase system in the denitrifying phototrophic bacterium Rhodobacter sphaeroides f.sp. denitrificans . Biosci Biotechnol Biochem 63:530–536
     [Google Scholar]
  18. Miller J. H. 1972 Experiments in Molecular Genetics pp  352–355 Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
     [Google Scholar]
  19. Moreno-Vivian C., Cabello P., Martinez-Luque M., Blasco R., Castillo F. 1999; Prokaryotic nitrate reduction: molecular properties and functional distinction among bacterial nitrate reductases. J Bacteriol 181:6573–6584
     [Google Scholar]
  20. Philippot L., Hojberg O. 1999; Dissimilatory nitrate reductases in bacteria. Biochim Biophys Acta 1446:1–23
     [Google Scholar]
  21. Potter L. C., Cole J. A. 1999; Essential roles for the products of the napABCD genes, but not napFGH , in periplasmic nitrate reduction by Escherichia coli K-12. Biochem J 344:69–76
     [Google Scholar]
  22. Potter L. C., Angove H., Richardson D. J., Cole J. 2001; Nitrate reduction in the periplasm of gram-negative bacteria. Adv Microb Physiol 45:51–112
     [Google Scholar]
  23. Reyes F., Gavira M., Castillo F., Moreno-Vivian C. 1998; Periplasmic nitrate-reducing system of the phototrophic bacterium Rhodobacter sphaeroides DSM 158: transcriptional and mutational analysis of the napKEFDABC gene cluster. Biochem J 331:897–904
     [Google Scholar]
  24. Richardson D. J. 2000; Bacterial respiration: a flexible process for a changing environment. Microbiology 146:551–571
     [Google Scholar]
  25. Richardson D. J., Ferguson S. J. 1992; The influence of carbon substrate on the activity of the periplasmic nitrate reductase in aerobically grown Thiosphaera pantotropha . Arch Microbiol 157:535–537
     [Google Scholar]
  26. Richardson D. J., Berks B. C., Russell D. A., Spiro S., Taylor C. J. 2001; Functional, biochemical and genetic diversity of prokaryotic nitrate reductases. Cell Mol Life Sci 58:165–178
     [Google Scholar]
  27. Robertson L. A., Kuenen J. G. 1983; Thiosphaera pantotropha gen. nov. sp. nov., a facultatively anaerobic, facultatively autotrophic sulphur bacterium. J Gen Microbiol 129:2847–2855
     [Google Scholar]
  28. Robertson L. A., Kuenen J. G. 1984; Aerobic denitrification: a controversy revived. Arch Microbiol 139:351–354
     [Google Scholar]
  29. Robertson L. A., Kuenen J. G. 1990; Combined heterotrophic nitrification and aerobic denitrification in Thiosphaera pantotropha and other bacteria. Antonie Van Leeuwenhoek 57:139–152
     [Google Scholar]
  30. Robertson L. A., van Niel E. W. J., Torremans R. A. M., Kuenen J. G. 1988; Simultaneous nitrification and denitrification in aerobic chemostat cultures of Thiosphaera pantotropha . Microb Ecol 18:113–119
     [Google Scholar]
  31. Robertson L. A., Cornelisse R., De Vos P., Hadioetomo R., Kuenen J. G. 1989; Aerobic denitrification in various heterotrophic nitrifiers. Antonie Van Leeuwenhoek 56:289–299
     [Google Scholar]
  32. Sears H. J., Ferguson S. J., Richardson D. J., Spiro S. 1993; The identification of a periplasmic nitrate reductase in Paracoccus denitrificans . FEMS Microbiol Lett 113:107–112
     [Google Scholar]
  33. Sears H. J., Bennett B., Spiro S., Thomson A. J., Richardson D. J. 1995; Identification of periplasmic nitrate reductase Mo(V) EPR signals in intact cells of Paracoccus denitrificans . Biochem J 310:311–314
     [Google Scholar]
  34. Sears H. J., Spiro S., Richardson D. J. 1997; Effect of carbon substrate and aeration on nitrate reduction and expression of the periplasmic and membrane bound nitrate reductases in carbon limited cultures of Paracoccus denitrificans Pd1222. Microbiology 143:3767–3774
     [Google Scholar]
  35. Sears H. J., Sawers G., Berks B. C., Ferguson S. J., Richardson D. J. 2000; Control of periplasmic nitrate reductase gene expression ( napEDABC ) from Paracoccus pantotrophus in response to oxygen and carbon substrates. Microbiology 146:2977–2985
     [Google Scholar]
  36. Siddiqui R. A., Warnecke-Eberz U., Hengsberger A., Schneider B., Kostka S., Friedrich B. 1993; Structure and function of a periplasmic nitrate reductase in Alcaligenes eutrophus H16. J Bacteriol 175:5867–5876
     [Google Scholar]
  37. Stewart V., Lu Y., Darwin A. J. 2002; Periplasmic nitrate reductase (NapABC enzyme) supports anaerobic respiration by Escherichia coli K-12. J Bacteriol 184:1314–1323
     [Google Scholar]
  38. Wang H., Tseng C. P., Gunsalus R. P. 1999; The napF and narG nitrate reductase operons in E. coli are expressed in response to submicromolar concentrations of nitrate but not nitrite. J Bacteriol 181:5303–5308
     [Google Scholar]
  39. Zumft W. G. 1997; Cell biology and molecular basis of denitrification. Microbiol Mol Biol Rev 61:533–616
     [Google Scholar]
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Characterization of the expression and activity of the periplasmic nitrate reductase of Paracoccus pantotrophus in chemostat cultures
Microbiology 149, 1533 (2003); https://doi.org/10.1099/mic.0.26277-0
/content/journal/micro/10.1099/mic.0.26277-0
/content/journal/micro/10.1099/mic.0.26277-0
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