Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Letter
  • Published:

Nitration of a peptide phytotoxin by bacterial nitric oxide synthase

Nature volume 429, pages 79–82 (2004)Cite this article

Abstract

Nitric oxide (NO) is a potent intercellular signal in mammals that mediates key aspects of blood pressure, hormone release, nerve transmission and the immune response of higher organisms1,2,3,4. Proteins homologous to full-length mammalian nitric oxide synthases (NOSs) are found in lower multicellular organisms5. Recently, genome sequencing has shown that some bacteria contain genes coding for truncated NOS proteins; this is consistent with reports of NOS-like activities in bacterial extracts6,7. Biological functions for bacterial NOSs are unknown, but have been presumed to be analogous to their role in mammals. Here we describe a gene in the plant pathogen Streptomyces turgidiscabies that encodes a NOS homologue, and we reveal its role in nitrating a dipeptide phytotoxin required for plant pathogenicity8. High similarity between bacterial NOSs indicates a general function in biosynthetic nitration; thus, bacterial NOSs constitute a new class of enzymes9,10,11. Here we show that the primary function of Streptomyces NOS is radically different from that of mammalian NOS. Surprisingly, mammalian NO signalling and bacterial biosynthetic nitration share an evolutionary origin.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: The nos gene is upstream of characterized thaxtomin biosynthetic genes.
Figure 2: Deletion of nos decreases thaxtomin production by S. turgidiscabies.
Figure 3: Nitrite formation from Nω-hydroxy-l-arginine by recombinant stNOS, and sensitivity to inhibitors of mammalian NOSs.
Figure 4: The nitrate nitrogen of thaxtomin A derives from the guanidine nitrogen of l-Arg.

Similar content being viewed by others

References

  1. Lipton, S. Physiology. Nitric oxide and respiration. Nature 413, 118–119 (2001)

    Article  ADS  CAS  Google Scholar 

  2. Nathan, C. Inducible nitric oxide synthase: What difference does it make? J. Clin. Invest. 100, 2417–2423 (1997)

    Article  CAS  Google Scholar 

  3. Christopherson, K. S. & Bredt, D. S. Nitric oxide in excitable tissues: Physiological roles and disease. J. Clin. Invest. 100, 2424–2429 (1997)

    Article  CAS  Google Scholar 

  4. Garthwaite, J. & Boulton, C. L. Nitric-oxide signaling in the central-nervous system. Annu. Rev. Physiol. 57, 683–706 (1995)

    Article  CAS  Google Scholar 

  5. Torreilles, J. Nitric oxide: one of the more conserved and widespread signaling molecules. Front. Biosci. 6, D1161–D1172 (2001)

    CAS  PubMed  Google Scholar 

  6. Chen, Y. J. & Rosazza, J. P. N. Purification and characterization of nitric oxide synthase (NOSNoc) from a Nocardia species. J. Bacteriol. 177, 5122–5128 (1995)

    Article  CAS  Google Scholar 

  7. Choi, W. S., Chang, M. S., Han, J. W., Hong, S. Y. & Lee, H. W. Identification of nitric oxide synthase in Staphylococcus aureus. Biochem. Biophys. Res. Commun. 237, 554–558 (1997)

    Article  CAS  Google Scholar 

  8. Healy, F. G., Wach, M., Krasnoff, S. B., Gibson, D. M. & Loria, R. The txtAB genes of the plant pathogen Streptomyces acidiscabies encode a peptide synthetase required for phytotoxin thaxtomin A production and pathogenicity. Mol. Microbiol. 38, 794–804 (2000)

    Article  CAS  Google Scholar 

  9. Adak, S. et al. Cloning, expression, and characterization of a nitric oxide synthase protein from Deinococcus radiodurans. Proc. Natl Acad. Sci. USA 99, 107–112 (2002)

    Article  ADS  CAS  Google Scholar 

  10. Pant, K., Bilwes, A. M., Adak, S., Stuehr, D. J. & Crane, B. R. Structure of a nitric oxide synthase heme protein from Bacillus subtilis. Biochemistry 41, 11071–11079 (2002)

    Article  CAS  Google Scholar 

  11. Adak, S., Aulak, K. & Stuehr, D. J. Evidence for nitric oxide production by a nitric-oxide synthase-like protein from Bacillus subtilis. J. Biol. Chem. 277, 16167–16171 (2002)

    Article  CAS  Google Scholar 

  12. Alderton, W. K., Cooper, C. E. & Knowles, R. G. Nitric oxide synthases: structure, function and inhibition. Biochem. J. 357, 593–615 (2001)

    Article  CAS  Google Scholar 

  13. Pfeiffer, S., Mayer, B. & Hemmens, B. Nitric oxide: Chemical puzzles posed by a biological messenger. Angew. Chem. Int. Edn Engl. 38, 1714–1731 (1999)

    Article  Google Scholar 

  14. Stuehr, D. J. Mammalian nitric oxide synthases. Biochim. Biophys. Acta 1411, 217–230 (1999)

    Article  CAS  Google Scholar 

  15. Sun, J., Kelemen, G. H., Fernandez-Abalos, J. M. & Bibb, M. J. Green fluorescent protein as a reporter for spatial and temporal gene expression in Streptomyces coelicolor A3(2). Microbiology 145, 2221–2227 (1999)

    Article  CAS  Google Scholar 

  16. Southan, G. J. & Szabo, C. Selective pharmacological inhibition of distinct nitric oxide synthase isoforms. Biochem. Pharmacol. 51, 383–394 (1996)

    Article  CAS  Google Scholar 

  17. Carter, G. T. et al. Direct biochemical nitration in the biosynthesis of dioxapyrrolomycin: A unique mechanism for the introduction of nitro groups in microbial products. J. Chem. Soc. Chem. Commun. 17, 1271–1273 (1989)

    Article  Google Scholar 

  18. Koppenol, W. H. The basic chemistry of nitrogen monoxide and peroxynitrite. Free Radical Biol. Med. 25, 385–391 (1998)

    Article  CAS  Google Scholar 

  19. Hughes, M. N. Relationships between nitric oxide, nitroxyl ion, nitrosonium cation and peroxynitrite. Biochim. Biophys. Acta 1411, 263–272 (1999)

    Article  CAS  Google Scholar 

  20. Ridd, J. H. Some unconventional pathways in aromatic nitration. Acta Chem. Scand. A 52, 11–22 (1998)

    Article  CAS  Google Scholar 

  21. Packer, L. E. (ed.) Nitric oxide. Part B. Physiological and pathological processes. Methods Enzymol. 268 (1996).

  22. Ischiropoulos, H. (ed.) Serial review: Reactive nitrogen species, tyrosine nitration and cell signaling. Free Radical Biol. Med. 33 (2002).

  23. Alvarez, B. et al. Peroxynitrite-dependent tryptophan nitration. Chem. Res. Toxicol. 9, 390–396 (1996)

    Article  CAS  Google Scholar 

  24. Jalal, M. A. F., Hossain, M. B. & Van der Helm, D. Structure of anticancer antibiotic l-alanosine. Acta Crystallogr. C 42, 733–738 (1986)

    Article  Google Scholar 

  25. Ohba, K. et al. Nitropeptin, a new dipeptide antibiotic possessing a nitro group. J. Antibiot. 40, 709–713 (1987)

    Article  CAS  Google Scholar 

  26. Goyer, C., Vachon, J. & Beaulieu, C. Pathogenicity of Streptomyces scabies mutants altered in thaxtomin A production. Phytopathology 88, 442–445 (1998)

    Article  CAS  Google Scholar 

  27. Martin, G. E. & Hadden, C. E. Long-range 1H–15N heteronuclear shift correlation at natural abundance. J. Nat. Prod. 63, 543–585 (2000)

    Article  CAS  Google Scholar 

  28. Kieser, T., Bibb, M. J., Buttner, M. J., Chater, K. F. & Hopwood, D. A. Practical Streptomyces Genetics (John Innes Foundation, Norwich, 2000)

    Google Scholar 

  29. MacNeil, D. J. et al. Analysis of Streptomyces avermitilis genes required for avermectin biosynthesis utilizing a novel integration vector. Gene 111, 61–68 (1992)

    Article  CAS  Google Scholar 

  30. Healy, F. G., Krasnoff, S. B., Wach, M., Gibson, D. M. & Loria, R. Involvement of a cytochrome P450 monooxygenase in thaxtomin A biosynthesis by Streptomyces acidiscabies. J. Bacteriol. 184, 2019–2029 (2002)

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank A. Condo Jr and I. Keresztes of the NMR facility of the Department of Chemistry and Chemical Biology, Cornell University, and staff of the Mass Spectrometry Laboratory, School of Chemical Sciences, University of Illinois Urbana-Champaign, for assistance in acquiring spectra for isotopic labelling studies. We also thank R. Parry for helpful discussion, and K. Pant and Madhavan Buddha for help with the NOS assays. We acknowledge the support of the National Science Foundation and the United States Department of Agriculture National Research Initiative, Biology of Plant–Microbe Associations Program.

Author information

Author notes

  1. Raghida A. Bukhalid

    Present address: Bauer Center for Genomics Research, Harvard University, Cambridge, Massachusetts, 02138, USA

Authors and Affiliations

  1. Department of Plant Pathology, Cornell University, Ithaca, New York, 14853, USA

    Johan A. Kers, Michael J. Wach, Stuart B. Krasnoff, Kimberly D. Cameron, Raghida A. Bukhalid & Rosemary Loria

  2. Agricultural Research Service, United States Department of Agriculture, Ithaca, New York, 14853, USA

    Donna M. Gibson

  3. Department of Chemistry and Chemical Biology, Cornell University, Ithaca, New York, 14853, USA

    Joanne Widom & Brian R. Crane

Authors
  1. Johan A. Kers
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Michael J. Wach
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Stuart B. Krasnoff
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Joanne Widom
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Kimberly D. Cameron
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Raghida A. Bukhalid
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Donna M. Gibson
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Brian R. Crane
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Rosemary Loria
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Rosemary Loria.

Ethics declarations

Competing interests

The authors declare that they have no competing financial interests.

Supplementary information

Supplementary Figure 1

This figure contains an amino acid alignment of Streptomyces turgidiscabies NOS, Bacillus subtilis NOS, and murine inducible NOS. (DOC 1799 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Kers, J., Wach, M., Krasnoff, S. et al. Nitration of a peptide phytotoxin by bacterial nitric oxide synthase. Nature 429, 79–82 (2004). https://doi.org/10.1038/nature02504

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nature02504

This article is cited by

Comments

By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing