Skip to main content

Thank you for visiting 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.

Intermediates in the transformation of phosphonates to phosphate by bacteria


Phosphorus is an essential element for all known forms of life. In living systems, phosphorus is an integral component of nucleic acids, carbohydrates and phospholipids, where it is incorporated as a derivative of phosphate. However, most Gram-negative bacteria have the capability to use phosphonates as a nutritional source of phosphorus under conditions of phosphate starvation1. In these organisms, methylphosphonate is converted to phosphate and methane. In a formal sense, this transformation is a hydrolytic cleavage of a carbon–phosphorus (C–P) bond, but a general enzymatic mechanism for the activation and conversion of alkylphosphonates to phosphate and an alkane has not been elucidated despite much effort for more than two decades. The actual mechanism for C–P bond cleavage is likely to be a radical-based transformation2. In Escherichia coli, the catalytic machinery for the C–P lyase reaction has been localized to the phn gene cluster1. This operon consists of the 14 genes phnC, phnD, …, phnP. Genetic and biochemical experiments have demonstrated that the genes phnG, phnH, …, phnM encode proteins that are essential for the conversion of phosphonates to phosphate and that the proteins encoded by the other genes in the operon have auxiliary functions1,3,4,5,6. There are no functional annotations for any of the seven proteins considered essential for C–P bond cleavage. Here we show that methylphosphonate reacts with MgATP to form α-d-ribose-1-methylphosphonate-5-triphosphate (RPnTP) and adenine. The triphosphate moiety of RPnTP is hydrolysed to pyrophosphate and α-d-ribose-1-methylphosphonate-5-phosphate (PRPn). The C–P bond of PRPn is subsequently cleaved in a radical-based reaction producing α-d-ribose-1,2-cyclic-phosphate-5-phosphate and methane in the presence of S-adenosyl-l-methionine. Substantial quantities of phosphonates are produced worldwide for industrial processes, detergents, herbicides and pharmaceuticals7,8,9. Our elucidation of the chemical steps for the biodegradation of alkylphosphonates shows how these compounds can be metabolized and recycled to phosphate.

This is a preview of subscription content

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.

Figure 1: 31 P NMR spectra of the reaction products catalysed by PhnI, PhnM and PhnJ.
Figure 2: Ultraviolet–visible absorbance spectrum of PhnJ (31 μM) after anaerobic reconstitution of the Fe–S cluster (solid line).
Figure 3: Working model for the transformation of PRPn to PRcP.
Figure 4: Reaction pathway for the conversion of methylphosphonate to PRcP.


  1. 1

    Metcalf, W. W. & Wanner, B. L. Evidence for a fourteen-gene, phnC to phnP locus for phosphonate metabolism in Escherichia coli. Gene 129, 27–32 (1993)

    CAS  Article  Google Scholar 

  2. 2

    Ahn, Y., Ye, Q., Cho, H., Walsh, C. T. & Floss, H. G. Stereochemistry of carbon-phosphorus cleavage in ethylphosphonate catalyzed by C-P lyase from Escherichia coli. J. Am. Chem. Soc. 114, 7953–7954 (1992)

    CAS  Article  Google Scholar 

  3. 3

    Metcalf, W. W. & Wanner, B. L. Mutational analysis of an Escherichia coli fourteen-gene operon for phosphonate degradation, using TnphoA’ elements. J. Bacteriol. 175, 3430–3442 (1993)

    CAS  Article  Google Scholar 

  4. 4

    Hove-Jensen, B., Rosenkrantz, T. J., Haldimann, A. & Wanner, B. L. Escherichia coli phnN, encoding ribose 1,5-bisphosphokinase activity (phosphoribosyl diphosphate forming): dual role in phosphonate degradation and NAD biosynthesis pathways. J. Bacteriol. 185, 2793–2801 (2003)

    CAS  Article  Google Scholar 

  5. 5

    Errey, J. C. & Blanchard, J. S. Functional annotation and kinetic characterization of PhnO from Salmonella enterica. Biochemistry 45, 3033–3039 (2006)

    CAS  Article  Google Scholar 

  6. 6

    Hove-Jensen, B., McSorley, F. R. & Zechel, D. L. Physiological role of phnP-specified phosphoribosyl cyclic phosphodiesterase in catabolism of organophosphoric acids by the carbon-phosphorus lyase pathway. J. Am. Chem. Soc. 133, 3617–3624 (2011)

    CAS  Article  Google Scholar 

  7. 7

    Ternan, N. G., McGrath, J. W., McMullan, G. & Quinn, J. P. Organophosphonates: occurrence, synthesis and biodegradation by microorganisms. World J. Microbiol. Biotechnol. 14, 635–647 (1998)

    CAS  Article  Google Scholar 

  8. 8

    Kononova, S. V. & Nesmeyanova, M. A. Phosphonates and their degradation by microorganisms. Biochemistry (Mosc.) 67, 184–195 (2002)

    CAS  Article  Google Scholar 

  9. 9

    White, A. K. & Metcalf, W. W. Microbial metabolism of reduced phosphorus compounds. Annu. Rev. Microbiol. 61, 379–400 (2007)

    CAS  Article  Google Scholar 

  10. 10

    Avila, L. Z., Draths, K. M. & Frost, J. W. Metabolites associated with organophosphonate C-P bond cleavage: chemical synthesis and microbial degradation of [32P]-ethylphosphonic acid. Bioorg. Med. Chem. Lett. 1, 51–54 (1991)

    CAS  Article  Google Scholar 

  11. 11

    Frost, J. W., Loo, S., Cordeiro, M. L. & Li, D. Radical-based dephosphorylation and organophosphonate biodegradation. J. Am. Chem. Soc. 109, 2166–2171 (1987)

    CAS  Article  Google Scholar 

  12. 12

    Parker, G. F., Higgins, T. P., Hawkes, T. & Robson, R. L. Rhizobium (Sinorhizobium) meliloti phn genes: characterization and identification of their protein products. J. Bacteriol. 181, 389–395 (1999)

    CAS  PubMed  PubMed Central  Google Scholar 

  13. 13

    Kamat, S. S. et al. Catalytic mechanism and three-dimensional structure of adenine deaminase. Biochemistry 50, 1917–1927 (2011)

    MathSciNet  CAS  Article  Google Scholar 

  14. 14

    Maynes, J. T., Yuan, R. G. & Snyder, F. F. Identification, expression and characterization of the Escherichia coli guanine deaminase. J. Bacteriol. 182, 4658–4660 (2000)

    CAS  Article  Google Scholar 

  15. 15

    Hall, R. S. et al. Three dimensional structure and catalytic mechanism of cytosine deaminase. Biochemistry 50, 5077–5085 (2011)

    CAS  Article  Google Scholar 

  16. 16

    Krenitsky, T. A., Neil, S. M., Elion, G. B. & Hitchings, G. H. A comparison of the specificities of xanthine oxidase and aldehyde oxidase. Arch. Biochem. Biophys. 150, 585–599 (1972)

    CAS  Article  Google Scholar 

  17. 17

    La . Vallie, E. R., McCoy, J. M., Smith, D. B. & Riggs, P . Enzymatic and chemical cleavage of fusion proteins. Curr. Protocols Mol. Biol. Unit 16.4B. (1994)

  18. 18

    Jochimsen, B. et al. Five phosphonate operon gene products as components of a multi-enzyme complex of the carbon-phosphorus lyase pathway. Proc. Natl Acad. Sci. USA 108, 11393–11398 (2011)

    ADS  CAS  Article  Google Scholar 

  19. 19

    Seibert, C. M. &. Raushel, F. M. Structural and catalytic diversity within the amidohydrolase superfamily. Biochemistry 44, 6383–6391 (2005)

    CAS  Article  Google Scholar 

  20. 20

    Frey, P. A., Hegeman, A. D. & Ruzicka, F. J. The radical SAM superfamily. Crit. Rev. Biochem. Mol. Biol. 43, 63–88 (2008)

    CAS  Article  Google Scholar 

  21. 21

    Chatterjee, A. et al. Reconstitution of ThiC in thiamine pyrimidine biosynthesis expands the radical SAM superfamily. Nature Chem. Biol. 4, 758–765 (2008)

    CAS  Article  Google Scholar 

  22. 22

    McGlynn, S. E. et al. Identification and characterization of a novel member of the radical AdoMet enzyme superfamily and implications for the biosynthesis of the Hmd hydrogenase active site cofactor. J. Bacteriol. 192, 595–598 (2010)

    CAS  Article  Google Scholar 

  23. 23

    Zhang, Y. et al. Diphthamide biosynthesis requires an organic radical generated by iron-sulphur enzyme. Nature 465, 891–896 (2010)

    ADS  CAS  Article  Google Scholar 

  24. 24

    Parast, C. V., Wong, K. K., Lewisch, S. A. & Kozarich, J. W. Hydrogen exchange of the glycyl radical of pyruvate formate-lyase is catalyzed by cysteine 419. Biochemistry 34, 2393–2399 (1995)

    CAS  Article  Google Scholar 

  25. 25

    Buis, J. M. & Broderick, J. B. Pyruvate formate-lyase activating enzyme: elucidation of a novel mechanism for glycyl radical formation. Arch. Biochem. Biophys. 433, 288–296 (2005)

    CAS  Article  Google Scholar 

  26. 26

    Thauer, R. K. & Shima, S. Methane as fuel for anaerobic microorganisms. Ann. NY Acad. Sci. 1125, 158–170 (2008)

    ADS  CAS  Article  Google Scholar 

Download references


We thank D. Barondeau and his laboratory for use of the anaerobic chamber and for advice on the assembly of Fe–S clusters in radical SAM enzymes. We also thank C. Xu for help with some of the 31P NMR spectra and S. Burrows for help with cloning phnI and phnH. This work was supported in part by the National Institutes of Health (GM93342, GM71790) and the Robert A. Welch Foundation (A-840). The cryoprobe for the NMR spectrometer was purchased with funds from the National Science Foundation (0840464)

Author information




S.S.K., H.J.W. and F.M.R. designed the experiments. S.S.K. and H.J.W. performed the experiments. S.S.K., H.J.W. and F.M.R. wrote the manuscript.

Corresponding author

Correspondence to Frank M. Raushel.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Information

The file contains Supplementary Figures 1-24 with legends, Supplementary Table 1, Supplementary Text and additional references. (PDF 1937 kb)

PowerPoint slides

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Kamat, S., Williams, H. & Raushel, F. Intermediates in the transformation of phosphonates to phosphate by bacteria. Nature 480, 570–573 (2011).

Download citation

Further reading


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.


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