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.

  • Article
  • Published:

The 2.8 Å structure of hydroxylamine oxidoreductase from a nitrifying chemoautotrophic bacterium, Nitrosomonas europaea

Nature Structural Biology volume 4, pages 276–284 (1997)Cite this article

Abstract

The 2.8 Å crystal structure of hydroxylamine oxidoreductase of a nitrifying chemoautotrophic bacterium, Nitrosomonas europaea, is described. Twenty-four haems lie in the centre bottom of the trimeric molecule, localized in four clusters within each monomer. The haem clusters within the trimer are aligned to form a ring that has inlet and outlet sites. The inlet is occupied by a novel haem, P460, and there are two possible outlet sites per monomer formed by paired haems lying within a cavity or cleft on the protein surface. The structure suggests pathways by which electron transfer may occur through the precisely arranged haems and provides a framework for the interpretation of previous and future biochemical and genetic observations.

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

Similar content being viewed by others

References

  1. Hooper, A.B. Biochemistry of the nitrifying lithoautotrophic bacteria in autotrophic bacteria (ed. Schlegel, H. G. & Bowein, B.) 239–265 (Science Tech Publishers, Madison, 1989).

    Google Scholar 

  2. Hollocher, T.C., Tate, M.E. & Nocholas, D.J.D. Oxidation of ammonia by Nitrosomonas europaea: definitive 180–tracer evidence that hydroxylamine formation involves a monooxygenase. J. Biol. Chem. 256, 10834–10836 (1981).

    CAS  PubMed  Google Scholar 

  3. Hyman, M.R. & Wood, P.M. Methan oxidation by Nitrosomonas europaea. Biochem. J. 212, 31–37 (1983).

    Article  CAS  Google Scholar 

  4. Hooper, A.B. & Nason, A. Characterization of hydroxylamine–cytochrome c reductase from the chemoautotophs Nitrosomonas europaea and Nitrosocystis oceanus. J. Biol. Chem. 240, 4044–4057 (1965).

    CAS  PubMed  Google Scholar 

  5. Yamanaka, T. & Shinra, M. Cytochrome c–552 and cytochrome c–554 derived from Nitrosomonas europaea. Purification, properties, and their function in hydroxylamine oxidation. J. Biochem. 75, 1265–1273 (1974).

    Article  CAS  Google Scholar 

  6. Yamazaki, T., Fukumori, Y. & Yamanaka, T. Cytochrome a1 of Nitrosomonas europaea resembles aa3–type cytochrome c oxidase in many respects. Biochim. Biophys. Acta 810, 174–183 (1985).

    Article  CAS  Google Scholar 

  7. DiSpirito, A.A., Lipscomb, J.D. & Hooper, A.B. Cytochrome aa3 from Nitrosomonas europaea. J. Biol. Chem. 261, 17048–17056 (1986).

    CAS  PubMed  Google Scholar 

  8. Suzuki, I. & Kwok, S.C. A partial resolution and reconstitution of the ammonia oxidizing system of Nitrosomonas europaea: role of cytochrome c554. Can. J. Biochem. 59, 484–488 (1981).

    Article  CAS  Google Scholar 

  9. Aleem, M.I.H. Generation of reducing power in chemosynthesis II. energy–linked reduction of pyridine nucleotides in the chemoautotroph, Nitrosomonas europaea. Biochim. Biophys. Acta 113, 216–224 (1966).

    Article  CAS  Google Scholar 

  10. Anderson, K.K., Kent, T.A., Lipscomb, J.D., Hooper, A.B. & Munck, E. Mossbauer, EPR, and optical studies of the P–460 center of hydroxylamine oxidoreductase from Nitrosomonas. J. Biol. Chem. 259, 6833–6840 (1984).

    Google Scholar 

  11. Arciero, D.M. & Hooper, A.B. Hydroxylamine oxidoreductase from Nitrosomonas europaea is a multimer of an octa–heme subunit. J. Biol. Chem. 268, 14645–14654 (1993).

    CAS  PubMed  Google Scholar 

  12. Sayavedra–soto, L.A., Hommes, N.G. & Arp, D.J. Characterization of the gene encoding hydroxylamine oxidoreductase in Nitrosomonas europaea. J. Bateriol. 176, 504–510 (1994).

    Article  Google Scholar 

  13. Terry, K.R. & Hooper, A.B. Hydroxylamine oxidoreductase: A 20–Heme, 200000 molecular weight cytochrome c with unusual denaturation properties which forms a 63000 molecular weight monomer after heme removal. Biochemistry 20, 7026–7032 (1981).

    Article  CAS  Google Scholar 

  14. Arciero, D.M., Balny, C. & Hooper, A.B. Spectroscopic and rapid kinetic studies of reduction of cytochrome c554 by hydroxylamine oxidoreductase from Nitrosomonas europaea. Biochemistry 30, 11466–11472 (1991).

    Article  CAS  Google Scholar 

  15. Erickson, R.H. & Hooper, A.B. Preliminary characterization of a variant CO–binding heme protein from Nitrosomonas. Biochim. Biophys. Acta 275, 231–244 (1972).

    Article  CAS  Google Scholar 

  16. Lipscomb, J.D. & Hooper, A.B. Resolution of multiple heme centers of hydroxylamine oxidoreductase from Nitrosomonas. 1. electron paramagnetic redonance spectroscopy. Biochemistry 21, 3965–3972 (1982).

    Article  CAS  Google Scholar 

  17. Lipscomb, J.D., Anderson, K.K., Munck, E., Kent, T.A. & Hooper, A.B. Resolution of multiple heme centers of hydroxylamine oxidoreductase from Nitrosomonas. 2. Mosssbauer spectroscopy. Biochemistry 21, 3973–3976 (1982).

    Article  CAS  Google Scholar 

  18. Hooper, A.B., Debey, P., Anderson, K.K. & Balny, C. Heme P460 of hydroxylamine oxidoreductase of Nitrosomonas. reaction with CO and H2O2 . Eur. J. Biochem. 134, 83–87 (1983).

    Article  CAS  Google Scholar 

  19. Arciero, D.M., Hooper, A.B., Cai, M. & Timkovich, R. Evidence for the structure of the active site heme P460 in hydroxylamine oxidoreductase of Nitrosomonas. Biochemistry 32, 9370–9378 (1993).

    Article  CAS  Google Scholar 

  20. McTavish, H., LaQuier, F., Arciero, D., Logan, M., Mundfrom, G., Fuchs, J.A. & Hooper, A.B. Multiple copies of genes coding for electron transport proteins in the bacterium Nitrosomonas europaea. J. Bacteriol. 175, 2445–2447 (1993).

    Article  CAS  Google Scholar 

  21. Mikami, T. et al. Crystallization and preliminary X-ray studies of hydroxylamine oxidoreductase from Nitrosomonas europaea. J. Biochem. 110, 681–682 (1991).

    Article  CAS  Google Scholar 

  22. Igarashi, N., Moriyama, H., Mikami, T. & Tanaka, N. Detwinning of hemihedrally twinned crystals by the least squares method and its application to the crystal of hydroxylamine oxidoreductase from Nitrosomonas europaea. in the press.

  23. Lee, B.K. & Richards, F.M. The interpretation of protein structures: Estimation of static accessibility. J. Mole. Biol. 55, 379–400 (1971).

    Article  CAS  Google Scholar 

  24. Janin, J., Miller, S. & Chothia, C. Surface, subunit interfaces and interior of oligomeric protein. J. Molc. Biol. 204, 155–164 (1988).

    Article  CAS  Google Scholar 

  25. Prince, R.C., Larroque, C. & Hooper, A.B. Resolution of the hemes of hydroxylamine oxidoreductase by redox potentiometry and optical spectroscopy. FEBS Lett. 163, 25–27 (1983).

    Article  CAS  Google Scholar 

  26. Prince, R.C. & Hooper, A.B. Resolution of the hemes of hydroxylamine oxidoreductase by redox potentiometry and electron spin resonance spectroscopy. Biochemistry 26, 970–974 (1987).

    Article  CAS  Google Scholar 

  27. Collins, M.J., Arciero, D.M. & Hooper, A.B. Optical spectropotentiometric resolution of the hemes of hydroxylamine oxidoreductase. Heme quantitation and pH dependence of Em. J. Biol. Chem. 268, 14655–14662 (1993).

    CAS  PubMed  Google Scholar 

  28. Hendrich, M.P. et al. The active site of hydroxylamine oxidoreductase from Nitrosomonas: Evidence for a new metal cluster in enzymes. J. Am. Chem. Soc. 116, 11961–11968 (1994).

    Article  CAS  Google Scholar 

  29. Deisenhofer, J., Epp, O., Miki, K., Huber, R. & Michel, H. X–ray structure analysis of a membrane protein complex. Electron density map at 3 Å resolution and a model of the chromophores of the photosynthetic reactioncenter from Rhodopseudomonas viridis. J. Mole. Biol. 180, 385–398 (1984).

    Article  CAS  Google Scholar 

  30. Deisenhofer, J., Epp, O., Miki, K., Huber, R. & Michel, H. Structure of the protein subunits in the photosynthetic reaction centre of Rhodoseudomonas viridis at 3 Å resolution. Nature 318, 618–624 (1985).

    Article  CAS  Google Scholar 

  31. Alien, J.P., Feher, G., Yeates, T.O., Komiya, H. & Rees, D.C. Structure of the reaction center from Rhodobacter sphaeroides R–26: the cofactors. Proc. Nat. Acad. Sci. U.S.A. 84, 5730–5734 (1987).

    Article  Google Scholar 

  32. McDermott, G. et al. Crystal structure of an integral membrane light–harvesting complex from photosynthetic bacteria. Nature 374, 517–521 (1995).

    Article  CAS  Google Scholar 

  33. Koepke, J., Hu, X., Muenke, C., Schulten, K. & Michel, H. The crystal structure of the light–harvesting complex II (B800–850). from Rhodospirillum molischianum. Structure 4, 581–597 (1996).

    Article  CAS  Google Scholar 

  34. Teske, A., Aim, E., Regan, J.M., Toze, S., Rittmann, B.E. & Stahl, D.A. Evolutionary relationships among ammonia- and nitrite-oxidizing bacteira. J. Bacteriol. 176, 6623–6630 (1994).

    Article  CAS  Google Scholar 

  35. Anderson, K.K. & Hooper, A.B. O2 and H2O are each the source of one O in NO2–: 15N–NMR evidence. FEBS Lett. 164, 236–240 (1983).

    Article  Google Scholar 

  36. Bergmann, D.J., Arciero, D.M. & Hooper, A.B. Organization of the hao gene cluster of Nitrosomonas europaea: Genes for two tetraheme c cytochromes. J. Bacteriol. 176, 3148–3153 (1994).

    Article  CAS  Google Scholar 

  37. Hommes, N.G., Sayavedra–soto, L.A. & Arp, D.J. Sequence of hey, a gene encoding cytochrome c–554 from Nitrosomonas europaea. Gene 146, 87–89 (1994).

    Article  CAS  Google Scholar 

  38. Olsen, T.C. & Hooper, A.B. Energy coupling in the bacterial oxidation of small molecules: an extracytoplasmic dehydrogenase in Nitrosomonas. FEMS, Microbiol. Lett. 19, 47–50 (1983).

    Article  Google Scholar 

  39. McTavish, H., Arciero, D.M. & Hooper, A.B. Interactions with membranes of cytochrome c554 from Nitrosomonas europaea. Arch. Biochem. Biophys. 324, 53–58 (1995).

    Article  CAS  Google Scholar 

  40. DiSpirito, A.A., Taaffe, L.R. & Hooper, A.B. Localization and concentration of hydroxylamine oxidoreductase and cytochrome c–552, c–554, cm–553, cm–552 and a in Nitrosomonas europaea. Biochim. Biophys. Acta 806, 320–330 (1985).

    Article  CAS  Google Scholar 

  41. Yamanaka, T., Shinra, M., Takahashi, K. & Shibasaka, M. Highly purified hydroxylamine oxidoreductase derived from Nitrosomonas europaea. Some physicochemical and enzymatic properties. J. Biochem. 86, 1101–1108 (1979).

    Article  CAS  Google Scholar 

  42. Otwinowski, Z. & Minor, W in Proc. CCP4 Study Weekend, 29–30 Jan. 1993, Data Collection and Processing (eds Sawyer, L., Isaacs, N. & Bailey, S.) 56–62 (SERC Daresbury Laboratory, Warrington, 1993).

    Google Scholar 

  43. Collabortive Computational project, Number 4 The CCP4 suite: programs for protein crystallography. Acta Crystallogr. D50, 760–763 (1994).

  44. Furey, W. & Swaminathan, S. PHASES – A program package for the processing and analysis of diffraction data from macromolecules. Am. Crystallogr. Assoc. Mtg Abstr.Ser.2 18, 73 (1990).

  45. Cowtan, K. in Joint CCP4 and ESF–EACBM Newsletter on Protein Crystallography 31, 34–38 (1994).

    Google Scholar 

  46. Jones, A.T. Interactive Computer Graphics: FRODO Meths. Enzymol. 115, 157–171 (1985).

    Article  CAS  Google Scholar 

  47. Brünger, A.T. XPLOR (version 3.1): A System for X–ray Crystallography and NMR (Yale University Press, 1993).

  48. Read, R.J. Improved fourier coefficients for maps using phases from partial structures with errors. Acta Crystallogr. A42, 140–149 (1986).

    Article  CAS  Google Scholar 

  49. Brünger, A.T. Free R-value: a novel statistical quantity for assessing the accuracy of crystal structures. Nature 355, 472–474 (1992).

    Article  Google Scholar 

  50. Jones, A.T. Improved methods for building protein models in electron density maps and the location of errors in these models. Acta Crystallogr. A47, 110–119 (1991).

    Article  CAS  Google Scholar 

  51. Kraulis, P. MOLSCRIPT: a program to produce both detailed and schematic plots of protein structures. J. Appl. Crystallogr. 24, 946–950 (1991).

    Article  Google Scholar 

  52. Merrit, E.A. & Murphy, M.E. Raster3D Version 2.0 – A program for photorealistic molecular graphics. Acta Crystallogr. D50, 869–873 (1994).

    Google Scholar 

  53. Nicholls, A., Sharp, K.A. & Honig, B. Protein folding and association: Insights from the interfacial and thermodynamic properties of hydrocarbons. Proteins Struct. Funct. Genet. 11, 281–296 (1991).

    Article  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Life Science, Faculty of Bioscience and Biotechnology, Tokyo Institute of Technology, 4259 Nagatsuta, Midori-ku, Yokohama, 226, Japan

    Noriyuki Igarashi, Hideaki Moriyama, Taketomo Fujiwara, Yoshihiro Fukumori & Nobuo Tanaka

Authors
  1. Noriyuki Igarashi
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Hideaki Moriyama
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Taketomo Fujiwara
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Yoshihiro Fukumori
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Nobuo Tanaka
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Igarashi, N., Moriyama, H., Fujiwara, T. et al. The 2.8 Å structure of hydroxylamine oxidoreductase from a nitrifying chemoautotrophic bacterium, Nitrosomonas europaea. Nat Struct Mol Biol 4, 276–284 (1997). https://doi.org/10.1038/nsb0497-276

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nsb0497-276

This article is cited by

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