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Structure of ADP·AIF4–-stabilized nitrogenase complex and its implications for signal transduction

Nature volume 387, pages 370–376 (1997)Cite this article

Abstract

The coupling of ATP hydrolysis to electron transfer by the enzyme nitrogenase during biological nitrogen fixation is an important example of a nucleotide-dependent transduction mechanism. The crystal structure has been determined for the complex between the Fe-protein and MoFe-protein components of nitrogenase stabilized by ADP·AIF4–, previously used as a nucleoside triphosphate analogue in nucleotide-switch proteins. The structure reveals that the dimeric Fe-protein has undergone substantial conformational changes. The β-phosphate and AIF4– groups are stabilized through intersubunit contacts that are critical for catalysis and the redox centre is repositioned to facilitate electron transfer. Interactions in the nitrogenase complex have broad implications for signal and energy transduction mechanisms in multiprotein complexes.

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References

  1. Stiefel, E. I.,, Coucouvanis, D. & Newton, W. E. (eds) Molybdenum Enzymes, Cofactors and Model Systems (Am. Chem. Soc., Washington, DC, 1993).

    Book  Google Scholar 

  2. Mortenson, L. E., Seefeldt, L. C., Morgan, T. V. & Bolin, J. T. The role of metal-clusters and MgATP in nitrogenase catalysis. Adv. Enzymol. Rel. Areas Mol. Biol. 67, 299–274 (1993).

    CAS  Google Scholar 

  3. Howard, J. B. & Rees, D. C. Nitrogenase: A nucleotide-dependent molecular switch. Annu. Rev. Biochem. 63, 235–264 (1994).

    Article  CAS  Google Scholar 

  4. Peters, J. W., Fisher, K. & Dean, D. R. Nitrogenase structure and function: A biochemical-genetic perspective. Annu. Rev. Microbiol. 49, 335–366 (1995).

    Article  CAS  Google Scholar 

  5. Burgess, B. K. & Lowe, D. J. Mechanism of molybdenum nitrogenase. Chem. Rev. 96, 2983–3011 (1996).

    Article  CAS  Google Scholar 

  6. Howard, J. B. & Rees, D. C. Structural basis of biological nitrogen fixation. Chem. Rev. 96, 2965–2982 (1996).

    Article  CAS  Google Scholar 

  7. Renner, K. A. & Howard, J. B. Aluminium fluoride inhibition of nitrogenase: stabilization of a nucleotide: Fe-protein : MoFe-protein complex. Biochemistry 35, 5353–5358 (1996).

    Article  CAS  Google Scholar 

  8. Duyvis, M., Wassink, H. & Haaker, H. Formation and characterization of a transition state complex of Azotobacter vinelandii nitrogenase. FEBS Lett. 380, 233–236 (1996).

    Article  CAS  Google Scholar 

  9. Fisher, A. J. et al. X-ray structures of the myosin motor domain of Dictyostelium discoideum complexed with MgADP·BeFx and MgADP·AlF−4 . Biochemistry 34, 8960–8972 (1995).

    Article  CAS  Google Scholar 

  10. Sondek, J., Lambright, D. G., Noel, J. P., Hamm, H. E. & Sigler, P. B. GTPase mechanisms of G proteins from the 1.7-Å crystal structure of transducin α·GDP·ALF−4 . Nature 372, 276–279 (1994).

    Article  ADS  CAS  Google Scholar 

  11. Coleman, D. E. et al. Structures of active conformers of Giα1 and the mechanism of GTP hydrolysis. Science 265, 1405–1412 (1994).

    Article  ADS  CAS  Google Scholar 

  12. Mittal, R., Ahmadian, M. R., Goody, R. S. & Wittinghofer, A. Formation of a transition-state analog of the Ras GTPase reaction by Ras-GDP, tetrafluoroaluminate, and GTPase-activating proteins. Science 273, 115–117 (1996).

    Article  ADS  CAS  Google Scholar 

  13. Kim, J. & Rees, D. C. Structural models for the metal centers in the nitrogenase molybdenum-iron protein. Science 257, 1677–1682 (1992).

    Article  ADS  CAS  Google Scholar 

  14. Kim, J. & Rees, D. C. Crystallographic structure and functional implications of the nitrogenase molybdenum-iron protein from Azotobacter vinelandii. Nature 360, 553–560 (1992).

    Article  ADS  CAS  Google Scholar 

  15. Georgiadis, M. M. et al. Crystallographic structure of the nitrogenase iron protein from Azotobacter vinelandii. Science 257, 1653–1659 (1992).

    Article  ADS  CAS  Google Scholar 

  16. Bolin, J. T., Campobasso, N., Muchmore, S. W., Morgan, T. V. & Mortenson, L. E. in Molybdenum Enzymes, Cofactors and Model Systems. ACS Symposium Series No. 535 (eds Stiefel, E. I., Coucouvanis, D. & Newton, W. E.) 186–195 (Am. Chem. Soc., Washington, 1993).

    Book  Google Scholar 

  17. Peters, J. P. et al. Redox-dependent structural changes in the nitrogenase P-cluster. Biochemistry 36, 1181–1187 (1997).

    Article  CAS  Google Scholar 

  18. Willing, A. & Howard, J. B. Cross-linking site in Azotobacter vinelandii complex. J. Biol. Chem. 265, 6596–6599 (1990).

    CAS  PubMed  Google Scholar 

  19. Howard, J. B. in Molybdenum Enzymes, Cofactors and Model Systems. ACS Symposium Series No. 535 (eds Stiefel, E. I., Coucouvanis, D. & Newton, W. E.) 271–289 (Am. Chem. Soc., Washington, 1993).

    Book  Google Scholar 

  20. Ludden, P. W. & Roberts, G. P. Regulation of nitrogenase activity by reversible ADP ribosylation. Curr. Top. Cell. Regul. 30, 23–56 (1989).

    Article  CAS  Google Scholar 

  21. Wolle, D., Kim, C., Dean, D. & Howard, J. B. Ionic interactions in the nitrogenase complex. J. Biol. Chem. 267, 3667–3673 (1992).

    CAS  PubMed  Google Scholar 

  22. Chen, L. et al. MgATP-induced conformational changes in the iron protein from A. vinelandii as studied by small-angle x-ray scattering. J. Biol. Chem. 269, 3290–3294 (1994).

    CAS  PubMed  Google Scholar 

  23. Pai, E. F. et al. Refined crystal structure of the triphosphate conformation of H-ras p21 at 1.35 Å resolution—implications for the mechanism of GTP hydrolysis. EMBO J. 9, 2351–2359 (1990).

    Article  CAS  Google Scholar 

  24. Noel, J. P., Hamm, H. E. & Sigler, P. B. The 2.2 Å crystal structure of transducin-α complexed with GTP-yS. Nature 366, 654–663 (1993).

    Article  ADS  CAS  Google Scholar 

  25. Lanzilotta, W. N., Ryle, M. J. & Seefeldt, L. C. Nucleotide hydrolysis and protein conformational changes in Azotobacter vinelandii nitrogenase iron protein: defining the function of aspartate 129. Biochemistry 34, 10712–10723 (1995).

    Google Scholar 

  26. Ryle, M. J. & Seefeldt, L. C. Elucidation of a MgATP signal transduction pathway in the nitrogenase iron protein: formation of a conformation resembling the MgATP-bound state by protein engineering. Biochemistry 35, 4766–4775 (1996).

    Article  CAS  Google Scholar 

  27. Seefeldt, I. C., Morgan, T. V., Dean, D. R. & Mortenson, L. E. Mapping the site(s) of MgATP and MgADP interaction with the nitrogenase of Azotobacter vinelandii. J. Biol. Chem. 267, 6680–6688 (1992).

    CAS  PubMed  Google Scholar 

  28. Ryle, M. J., Lanzilotta, W. N., Mortenson, L. E., Watt, G. D. & Seefeldt, L. C. Evidence for a central role of lysine 15 of Azotobacter vinelandii nitrogenase iron protein in nucleotide binding and protein conformational changes. J. Biol. Chem. 270, 13112–13117 (1995).

    Article  CAS  Google Scholar 

  29. Seefeldt, L. C. & Mortenson, L. E. Increasing nitrogenase catalytic efficiency for MgATP by changing serine 16 of its Fe protein to threonine: Use of Mn 2+ to show interaction of serine 16 with Mg2+. Prot. Sci. 2, 93–102 (1993).

    Article  CAS  Google Scholar 

  30. Wolle, D., Dean, D. R. & Howard, J. B. Nucleotide-iron-sulfur cluster signal transduction in the nitrogenase iron-protein: the role of Asp125. Science 258, 992–995 (1992).

    Article  ADS  CAS  Google Scholar 

  31. Frech, M. et al. Role of glutamine-61 in the hydrolysis of GTP by p21H-ras. An experimental and theoretical study. Biochemistry 33, 3237–3244 (1994).

    Article  CAS  Google Scholar 

  32. Maegley, K. A., Admiraal, S. J. & Herschlag, D. Ras-catalyzed hydrolysis of GTP: A new perspective from model studies. Proc. Natl Acad. Sci. USA 93, 8160–8166 (1996).

    Article  ADS  CAS  Google Scholar 

  33. Brownbridge, G. G., Lowe, P. N., Moore, K. J. M., Skinner, R. H. & Webb, M. R. Interaction of GTPase activating proteins (GAPs) with p21ras measured by a novel fluorescence anisotropy method: Essential role of Arg-903 of GAP in activation of GTP hydrolysis on p21ras. J. Biol. Chem. 268, 10914–10919 (1993).

    CAS  PubMed  Google Scholar 

  34. Otwinowski, Z. in Data Collection and Processing (eds Sawyer, L., Issacs, N. & Bailey, S.) 56–62 (SERC Daresbury Laboratory, UK, 1993).

    Google Scholar 

  35. Bailey, S. The CCP4 suite—programs for protein crystallography. Acta Crystallogr. D 50, 760–763 (1994).

    Google Scholar 

  36. Navaza, J. AMORE—an automated package for molecular replacement. Acta Crystallogr. A 50, 157–163 (1994).

    Article  Google Scholar 

  37. Kleywegt, G. J. & Jones, T. A. in First Map to Final Model (eds Bailey, S., Hubbard, R. & Waller, D.) 59–66 (SERC Daresbury Laboratory, UK, 1994).

    Google Scholar 

  38. Jones, T. A., Zhou, J. Y., Cowan, S. W. & Kjeldgaard, M. Improved methods for building protein models in electron density maps and the location of errors in these models. Acta Crystallogr. A 47, 110–119 (1991).

    Article  Google Scholar 

  39. Brünger, A. T. X-PLOR version 3.1—A system for X-ray crystallography and NMR (Yale University Press, New Haven and London, 1992).

    Google Scholar 

  40. Laskowski, R. A., McArthur, M. W., Moss, D. S. & Thornton, J. M. PROCHECK—a program to check the stereochemical quality of protein structures. J. Appl. Crystallogr. 26, 283–291 (1993).

    Article  CAS  Google Scholar 

  41. Kabsch, W. A solution for the best rotation to relate two sets of vectors. Acta Crystallogr. A 32, 922–923 (1976).

    Article  Google Scholar 

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

    Article  Google Scholar 

  43. Bacon, D. J. & Anderson, W. F. A fast algorithm for rendering space-filling molecule pictures. J. Mol. Graph. 6, 219–220 (1988).

    Article  Google Scholar 

  44. Merritt, E. A. & Murphy, M. E. P. Raster3D Version 2.0-a program for photorealistic molecular graphics. Acta Crystallogr. D 50, 869–873 (1994).

    CAS  Google Scholar 

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Authors and Affiliations

  1. Division of Chemistry and Chemical Engineering, 147-75CH, California Institute of Technology, Pasadena, California, 91125, USA

    Hermann Schindelin, Caroline Kisker, Jamie L. Schlessman & Douglas C. Rees

  2. Department of Biochemistry, University of Minnesota, Minneapolis, Minnesota, 55455, USA

    James B. Howard

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  1. Hermann Schindelin
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  2. Caroline Kisker
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  3. Jamie L. Schlessman
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  4. James B. Howard
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  5. Douglas C. Rees
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Schindelin, H., Kisker, C., Schlessman, J. et al. Structure of ADP·AIF4–-stabilized nitrogenase complex and its implications for signal transduction. Nature 387, 370–376 (1997). https://doi.org/10.1038/387370a0

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