1887

Abstract

GlnR is the global transcriptional regulator of nitrogen assimilation in . Under nitrogen starvation, GlnR controls the transcription of at least nine genes associated with nitrogen metabolism. In this study, we identified a new GlnR target gene, , named (itrate/itrite ssimilation egulator). analysis of NnaR revealed the presence of two distinct domains: an N-terminal uroporphyrinogen-III synthase (HemD)-like enzymatic domain and a C-terminal DNA binding domain. Complementation experiments with a haemin auxotroph Δ mutant strain revealed that NnaR has no HemD activity. Physiological studies of an  : : Tn mutant showed that NnaR is involved in regulating nitrite reduction. By electrophoretic mobility shift assays the functionality of the NnaR DNA binding domain was confirmed, and it was found that NnaR binds in front of the genes (putative nitrate extrusion protein), (nitrite reductase), (putative nitrite/sulphite reductase) and (putative nitrate reductase), which are associated with nitrate/nitrite assimilation. Furthermore, a cooperative binding of NnaR together with GlnR to the promoter was observed, suggesting that NnaR may act as a GlnR co-activator.

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2012-05-01
2024-03-29
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References

  1. Ahmad M., Roberts J. N., Hardiman E. M., Singh R., Eltis L. D., Bugg T. D. ( 2011). Identification of DypB from Rhodococcus jostii RHA1 as a lignin peroxidase. Biochemistry 50:5096–5107 [View Article][PubMed]
    [Google Scholar]
  2. Arcondéguy T., Jack R., Merrick M. ( 2001). PII signal transduction proteins, pivotal players in microbial nitrogen control. Microbiol Mol Biol Rev 65:80–105 [View Article][PubMed]
    [Google Scholar]
  3. Bradford M. M. ( 1976). A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein–dye binding. Anal Biochem 72:248–254 [View Article][PubMed]
    [Google Scholar]
  4. Browning D. F., Busby S. J. ( 2004). The regulation of bacterial transcription initiation. Nat Rev Microbiol 2:57–65 [View Article][PubMed]
    [Google Scholar]
  5. Bullock W. O., Fernandez J. M., Short J. M. ( 1987). XL1-Blue: a high efficiency plasmid transforming recA Escherichia coli strain with beta-galactosidase selection. Biotechniques 5:376–379
    [Google Scholar]
  6. Chartrand P., Tardif D., Săsărman A. ( 1979). Uroporphyrin- and coproporphyrin I-accumulating mutant of Escherichia coli K12. J Gen Microbiol 110:61–66[PubMed] [CrossRef]
    [Google Scholar]
  7. Commichau F. M., Stülke J. ( 2008). Trigger enzymes: bifunctional proteins active in metabolism and in controlling gene expression. Mol Microbiol 67:692–702 [View Article][PubMed]
    [Google Scholar]
  8. Darie S., Gunsalus R. P. ( 1994). Effect of heme and oxygen availability on hemA gene expression in Escherichia coli: role of the fnr, arcA, and himA gene products. J Bacteriol 176:5270–5276[PubMed]
    [Google Scholar]
  9. Darwin A. J., Tyson K. L., Busby S. J. W., Stewart V. ( 1997). Differential regulation by the homologous response regulators NarL and NarP of Escherichia coli K-12 depends on DNA binding site arrangement. Mol Microbiol 25:583–595 [View Article][PubMed]
    [Google Scholar]
  10. Eiglmeier K., Honoré N., Iuchi S., Lin E. C. C., Cole S. T. ( 1989). Molecular genetic analysis of FNR-dependent promoters. Mol Microbiol 3:869–878 [View Article][PubMed]
    [Google Scholar]
  11. Fink D., Weißschuh N., Reuther J., Wohlleben W., Engels A. ( 2002). Two transcriptional regulators GlnR and GlnRII are involved in regulation of nitrogen metabolism in Streptomyces coelicolor A3(2). Mol Microbiol 46:331–347 [View Article][PubMed]
    [Google Scholar]
  12. Fu H. A., Iuchi S., Lin E. C. ( 1991). The requirement of ArcA and Fnr for peak expression of the cyd operon in Escherichia coli under microaerobic conditions. Mol Gen Genet 226:209–213 [View Article][PubMed]
    [Google Scholar]
  13. Hopwood D. A., Bibb M. J., Chater K. F., Kieser T., Bruton C. J., Kieser H. M., Lydiate D. J., Smith C. P., Ward J. M., Schrempf H. ( 1985). Genetic Manipulation of Streptomyces: a Laboratory Manual Norwich: John Innes Foundation;
    [Google Scholar]
  14. Hopwood D. A., Chater K. F., Bibb M. J. ( 1995). Genetics of antibiotic production in Streptomyces coelicolor A3(2), a model streptomycete. Biotechnology 28:65–102[PubMed]
    [Google Scholar]
  15. Hutchings M. I., Hoskisson P. A., Chandra G., Buttner M. J. ( 2004). Sensing and responding to diverse extracellular signals? Analysis of the sensor kinases and response regulators of Streptomyces coelicolor A3(2). Microbiology 150:2795–2806 [View Article][PubMed]
    [Google Scholar]
  16. Kieser T., Bibb M. J., Buttner M. J., Chater K. F., Hopwood D. A. ( 2000). Practical Streptomyces Genetics Norwich: John Innes Foundation;
    [Google Scholar]
  17. Kiley P. J., Beinert H. ( 2003). The role of Fe–S proteins in sensing and regulation in bacteria. Curr Opin Microbiol 6:181–185 [View Article][PubMed]
    [Google Scholar]
  18. Ling M., Allen S. W., Wood J. M. ( 1994). Sequence analysis identifies the proline dehydrogenase and Δ1-pyrroline-5-carboxylate dehydrogenase domains of the multifunctional Escherichia coli PutA protein. J Mol Biol 243:950–956 [View Article][PubMed]
    [Google Scholar]
  19. Magasanik B. ( 1982). Genetic control of nitrogen assimilation in bacteria. Annu Rev Genet 16:135–168 [View Article][PubMed]
    [Google Scholar]
  20. Merrick M. J., Edwards R. A. ( 1995). Nitrogen control in bacteria. Microbiol Rev 59:604–622[PubMed]
    [Google Scholar]
  21. Moir J. W. B., Wood N. J. ( 2001). Nitrate and nitrite transport in bacteria. Cell Mol Life Sci 58:215–224 [View Article][PubMed]
    [Google Scholar]
  22. Okanishi M., Suzuki K., Umezawa H. ( 1974). Formation and reversion of Streptomycete protoplasts: cultural condition and morphological study. J Gen Microbiol 80:389–400[PubMed] [CrossRef]
    [Google Scholar]
  23. Pullan S. T., Chandra G., Bibb M. J., Merrick M. ( 2011). Genome-wide analysis of the role of GlnR in Streptomyces venezuelae provides new insights into global nitrogen regulation in actinomycetes. BMC Genomics 12:175 [View Article][PubMed]
    [Google Scholar]
  24. Reuther J., Wohlleben W. ( 2007). Nitrogen metabolism in Streptomyces coelicolor: transcriptional and post-translational regulation. J Mol Microbiol Biotechnol 12:139–146 [View Article][PubMed]
    [Google Scholar]
  25. Sambrook J., Fritsch E. F., Maniatis T. ( 1989). Molecular Cloning: a Laboratory Manual , 2nd edn. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory Press;
    [Google Scholar]
  26. Schäfer A., Tauch A., Jäger W., Kalinowski J., Thierbach G., Pühler A. ( 1994). Small mobilizable multi-purpose cloning vectors derived from the Escherichia coli plasmids pK18 and pK19: selection of defined deletions in the chromosome of Corynebacterium glutamicum . Gene 145:69–73 [View Article][PubMed]
    [Google Scholar]
  27. Shao Z., Gao J., Ding X., Wang J., Chiao J., Zhao G. ( 2011). Identification and functional analysis of a nitrate assimilation operon nasACKBDEF from Amycolatopsis mediterranei U32. Arch Microbiol 193:463–477 [View Article][PubMed]
    [Google Scholar]
  28. Simon R., Priefer V., Pühler A. ( 1983). A broad host range mobilization system for in vivo genetic engineering: transposon mutagenesis in Gram negative bacteria. Nat Biotechnol 1:784–791 [View Article]
    [Google Scholar]
  29. Southern E. M. ( 1975). Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol 98:503–517 [View Article][PubMed]
    [Google Scholar]
  30. Stamford N. P., Capretta A., Battersby A. R. ( 1995). Expression, purification and characterisation of the product from the Bacillus subtilis hemD gene, uroporphyrinogen III synthase. Eur J Biochem 231:236–241 [View Article][PubMed]
    [Google Scholar]
  31. Tiffert Y., Supra P., Wurm R., Wohlleben W., Wagner R., Reuther J. ( 2008). The Streptomyces coelicolor GlnR regulon: identification of new GlnR targets and evidence for a central role of GlnR in nitrogen metabolism in actinomycetes. Mol Microbiol 67:861–880 [View Article][PubMed]
    [Google Scholar]
  32. Tiffert Y., Franz-Wachtel M., Fladerer C., Nordheim A., Reuther J., Wohlleben W., Mast Y. ( 2011). Proteomic analysis of the GlnR-mediated response to nitrogen limitation in Streptomyces coelicolor M145. Appl Microbiol Biotechnol 89:1149–1159 [View Article][PubMed]
    [Google Scholar]
  33. Volff J.-N., Eichenseer C., Viell P., Piendl W., Altenbuchner J. ( 1996). Nucleotide sequence and role in DNA amplification of the direct repeats composing the amplifiable element AUD1 of Streptomyces lividans 66. Mol Microbiol 21:1037–1047 [View Article][PubMed]
    [Google Scholar]
  34. Wang J., Zhao G.-P. ( 2009). GlnR positively regulates nasA transcription in Streptomyces coelicolor . Biochem Biophys Res Commun 386:77–81 [View Article][PubMed]
    [Google Scholar]
  35. Wohlleben W., Stegmann E., Süßmuth R. D. ( 2009). Chapter 18. Molecular genetic approaches to analyze glycopeptide biosynthesis. Methods Enzymol 458:459–486 [View Article][PubMed]
    [Google Scholar]
  36. Wray L. V. Jr, Atkinson M. R., Fisher S. H. ( 1991). Identification and cloning of the glnR locus, which is required for transcription of the glnA gene in Streptomyces coelicolor A3(2). J Bacteriol 173:7351–7360[PubMed]
    [Google Scholar]
  37. Zhu W., Becker D. F. ( 2003). Flavin redox state triggers conformational changes in the PutA protein from Escherichia coli . Biochemistry 42:5469–5477 [View Article][PubMed]
    [Google Scholar]
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