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Mechanism of Mo-Dependent Nitrogenase

  • Zhi-Yong Yang
  • Karamatullah Danyal
  • Lance C. SeefeldtEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 766)

Abstract

Nitrogenase is the enzyme responsible for biological reduction of dinitrogen (N2) to ammonia, a form usable for life. Playing a central role in the global biogeochemical nitrogen cycle, this enzyme has been the focus of intensive research for over 60 years. This chapter provides an overview of the features of nitrogenase as a background to the subsequent chapters of this volume that detail the many methods that have been applied in an attempt to gain a deeper understanding of this complex enzyme.

Key words

Nitrogen fixation Fe protein MoFe protein mechanism metalloenzyme MgATP 

Notes

Acknowledgments

The authors acknowledge the long collaboration with the Brian Hoffman and Dennis Dean laboratories in advancing understanding of nitrogenase. Work in the laboratory of the authors is supported by a generous grant from the National Institutes of Health (GM59087).

References

  1. 1.
    Smil V (2001) Enriching the Earth: Fritz Haber, Carl Bosch, and the Transformation of World Food Production. MIT Press, Cambridge, MAGoogle Scholar
  2. 2.
    Ferguson SJ (1998) Nitrogen cycle enzymology. Curr Opin Chem Biol 2:182–193PubMedGoogle Scholar
  3. 3.
    Fryzuk MD, MacKay BA (2004) Dinitrogen coordination chemistry: On the biomimetic borderlands. Chem Rev 104:385–401PubMedGoogle Scholar
  4. 4.
    Haber F (1922) The production of ammonia from nitrogen and hydrogen. Naturwissenschaften 10:1041–1049Google Scholar
  5. 5.
    Haber F (1923) The history of the ammonia process. Naturwissenschaften 11:339–340Google Scholar
  6. 6.
    Cheng Q (2008) Perspectives in biological nitrogen fixation research. J Integr Plant Biol 50:786–798PubMedGoogle Scholar
  7. 7.
    Smith BE (2002) Nitrogen reveals its inner secrets. Science 297:1654–1655PubMedGoogle Scholar
  8. 8.
    Raymond J, Siefert JL, Staples CR et al (2004) The natural history of nitrogen fixation. Mol Biol Evol 21:541–554PubMedGoogle Scholar
  9. 9.
    Burgess BK, Lowe DJ (1996) The mechanism of molybdenum nitrogenase. Chem Rev 96:2983–3011PubMedGoogle Scholar
  10. 10.
    Eady RR (1996) Structure-function relationships of alternative nitrogenases. Chem Rev 96:3013–3030PubMedGoogle Scholar
  11. 11.
    Bishop PE, Joerger RD (1990) Genetics and molecular biology of alternative nitrogen fixation systems. Annu Rev Plant Physiol Plant Mol Biol 41:109–125Google Scholar
  12. 12.
    Masepohl B, Schneider K, Drepper T et al (2002) Alternative nitrogenases. In: Leigh GJ (ed) Nitrogenase Fixation at the Millennium, pp. 191–222. Elsevier, AmsterdamGoogle Scholar
  13. 13.
    Barney BM, Lee HI, Dos Santos PC et al (2006) Breaking the N2 triple bond: Insights into the nitrogenase mechanism. Dalton Trans 19:2277–2284PubMedGoogle Scholar
  14. 14.
    Ribbe M, Gadkari D, Meyer O (1997) N2 fixation by Streptomyces thermoautotrophicus involves a molybdenum- dinitrogenase and a manganese-superoxide oxidoreductase that couple N2 reduction to the oxidation of superoxide produced from O2 by a molybdenum-CO dehydrogenase. J Biol Chem 272:26627–26633PubMedGoogle Scholar
  15. 15.
    Rees DC, Howard JB (2000) Nitrogenase: Standing at the crossroads. Curr Opin Chem Biol 4:559–566PubMedGoogle Scholar
  16. 16.
    Seefeldt LC, Dean DR (1997) Role of nucleotides in nitrogenase catalysis. Acc Chem Res 30:260–266Google Scholar
  17. 17.
    Seefeldt LC, Dance I, Dean DR (2004) Substrate interactions with nitrogenase: Fe versus Mo. Biochemistry 43:1401–1409PubMedGoogle Scholar
  18. 18.
    Dos Santos PC, Igarashi RY, Lee HI et al (2005) Substrate interactions with the nitrogenase active site. Acc Chem Res 38:208–214PubMedGoogle Scholar
  19. 19.
    Seefeldt LC, Hoffman BM, Dean DR (2009) Mechanism of Mo-dependent nitrogenase. Annu Rev Biochem 78:701–722PubMedGoogle Scholar
  20. 20.
    Hoffman BM, Dean DR, Seefeldt LC (2009) Climbing nitrogenase: Toward a mechanism of enzymatic nitrogen fixation. Acc Chem Res 42:609–619PubMedGoogle Scholar
  21. 21.
    Igarashi RY, Seefeldt LC (2003) Nitrogen fixation: The mechanism of the Mo-dependent nitrogenase. Crit Rev Biochem Mol Biol 38:351–384PubMedGoogle Scholar
  22. 22.
    Simpson FB, Burris RH (1984) A nitrogen pressure of 50 atmospheres does not prevent evolution of hydrogen by nitrogenase. Science 224:1095–1097PubMedGoogle Scholar
  23. 23.
    Kim J, Rees DC (1992) Crystallographic structure and functional implications of the nitrogenase molybdenum iron protein from Azotobacter vinelandii. Nature 360:553–560Google Scholar
  24. 24.
    Kim J, Rees DC (1992) Structural models for the metal centers in the nitrogenase molybdenum-iron protein. Science 257:1677–1682PubMedGoogle Scholar
  25. 25.
    Georgiadis MM, Komiya H, Chakrabarti P et al (1992) Crystallographic structure of the nitrogenase iron protein from Azotobacter vinelandii. Science 257:1653–1659PubMedGoogle Scholar
  26. 26.
    Kim J, Woo D, Rees DC (1993) X-ray crystal structure of the nitrogenase molybdenum-iron protein from Clostridium pasteurianum at 3.0-Å resolution. Biochemistry 32:7104–7115PubMedGoogle Scholar
  27. 27.
    Chan MK, Kim J, Rees DC (1993) The nitrogenase FeMo-cofactor and P-cluster pair: 2.2 Å resolution structures. Science 260:792–794PubMedGoogle Scholar
  28. 28.
    Einsle O, Tezcan FA, Andrade SLA et al (2002) Nitrogenase MoFe-protein at 1.16 Å resolution: A central ligand in the FeMo-cofactor. Science 297:1696–1700PubMedGoogle Scholar
  29. 29.
    Sørlie M, Christiansen J, Lemon BJ et al (2001) Mechanistic features and structure of the nitrogenase α-Gln195 MoFe protein. Biochemistry 40:1540–1549PubMedGoogle Scholar
  30. 30.
    Jang SB, Seefeldt LC, Peters JW (2000) Insights into nucleotide signal transduction in nitrogenase: Structure of an iron protein with MgADP bound. Biochemistry 39:14745–14752PubMedGoogle Scholar
  31. 31.
    Jang SB, Jeong MS, Seefeldt LC et al (2004) Structural and biochemical implications of single amino acid substitutions in the nucleotide-dependent switch regions of the nitrogenase Fe protein from Azotobacter vinelandii. J Biol Inorg Chem 9:1028–1033PubMedGoogle Scholar
  32. 32.
    Jang SB, Seefeldt LC, Peters JW (2000) Modulating the midpoint potential of the [4Fe-4S] cluster of the nitrogenase Fe protein. Biochemistry 39:641–648PubMedGoogle Scholar
  33. 33.
    Mayer SM, Lawson DM, Gormal CA et al (1999) New insights into structure-function relationships in nitrogenase: A 1.6 Å resolution X-ray crystallographic study of Klebsiella pneumoniae MoFe-protein. J Mol Biol 292:871–891PubMedGoogle Scholar
  34. 34.
    Mayer SM, Gormal CA, Smith BE et al (2002) Crystallographic analysis of the MoFe protein of nitrogenase from a nifV mutant of Klebsiella pneumoniae identifies citrate as a ligand to the molybdenum of iron molybdenum cofactor (FeMoco). J Biol Chem 277:35263–35266PubMedGoogle Scholar
  35. 35.
    Bolin JT, Ronco AE, Mortenson LE et al (1990) Structure of the nitrogenase MoFe protein: Spatial distribution of the intrinsic metal atoms determined by X-ray anomalous scattering. In: Gresshoff PM, Roth LE, Stacey G, Newton WE (eds) Nitrogen Fixation: Achievements and Objectives, pp. 117–124. Chapman and Hall, New York, NYGoogle Scholar
  36. 36.
    Bolin JT, Campobasso N, Muchmore SW et al (1993) Structure and environment of metal clusters in the nitrogenase molybdenum-iron protein form Clostridium pasteurianum. In: Stiefel EI, Coucouvanis D, Newton WE (eds) Molybdenum Enzymes, Cofactors and Model Systems, pp. 186–195. ACS, Washington, DCGoogle Scholar
  37. 37.
    Sen S, Krishnakumar A, McClead J et al (2006) Insights into the role of nucleotide-dependent conformational change in nitrogenase catalysis: Structural characterization of the nitrogenase Fe protein Leu127 deletion variant with bound MgATP. J Inorg Biochem 100:1041–1052PubMedGoogle Scholar
  38. 38.
    Sen S, Igarashi R, Smith A et al (2004) A conformational mimic of the MgATP-bound “on state” of the nitrogenase iron protein. Biochemistry 43:1787–1797PubMedGoogle Scholar
  39. 39.
    Schmid B, Ribbe MW, Einsle O et al (2002) Structure of a cofactor-deficient nitrogenase MoFe protein. Science 296:352–356PubMedGoogle Scholar
  40. 40.
    Sarma R, Barney BM, Keable S et al (2009) Insights into substrate binding at FeMo-cofactor in nitrogenase from the structure of an α-70Ile MoFe protein variant. J Inorg Biochem 104:385–389PubMedGoogle Scholar
  41. 41.
    Jeong MS, Jang SB (2004) Structural basis for the changes in redox potential in the nitrogenase Phe135Trp Fe protein with MgADP bound. Mol Cells 18:374–382PubMedGoogle Scholar
  42. 42.
    Peters JW, Stowell MHB, Soltis SM et al (1997) Redox-dependent structural changes in the nitrogenase P-cluster. Biochemistry 36:1181–1187PubMedGoogle Scholar
  43. 43.
    Schindelin H, Kisker C, Schlessman JL et al (1997) Structure of ADP-AlF4 stabilized nitrogenase complex and its implications for signal transduction. Nature 387:370–376PubMedGoogle Scholar
  44. 44.
    Chiu HJ, Peters JW, Lanzilotta WN et al (2001) MgATP-bound and nucleotide-free structures of a nitrogenase protein complex between the Leu 127∆-Fe-protein and the MoFe-protein. Biochemistry 40:641–650PubMedGoogle Scholar
  45. 45.
    Schmid B, Einsle O, Chiu HJ et al (2002) Biochemical and structural characterization of the cross-linked complex of nitrogenase: Comparison to the ADP-AlF4 -stabilized structure. Biochemistry 41:15557–15565PubMedGoogle Scholar
  46. 46.
    Tezcan FA, Kaiser JT, Mustafi D et al (2005) Nitrogenase complexes: Multiple docking sites for a nucleotide switch protein. Science 309:1377–1380PubMedGoogle Scholar
  47. 47.
    Rees DC, Tezcan FA, Haynes CA et al (2005) Structural basis of biological nitrogen fixation. Philos Trans R Soc A 363:971–984Google Scholar
  48. 48.
    Howard JB, Rees DC (1994) Nitrogenase: A nucleotide-dependent molecular switch. Annu Rev Biochem 63:235–264PubMedGoogle Scholar
  49. 49.
    Rubio LM, Ludden PW (2008) Biosynthesis of the Iron-Molybdenum cofactor of nitrogenase. Annu Rev Microbiol 62:93–111PubMedGoogle Scholar
  50. 50.
    Hu Y, Fay AW, Lee CC et al (2008) Assembly of nitrogenase MoFe protein. Biochemistry 47:3973–3981PubMedGoogle Scholar
  51. 51.
    Martin AE, Burgess BK, Iismaa SE et al (1989) Construction and characterization of an Azotobacter vinelandii strain with mutations in the genes encoding flavodoxin and ferredoxin I. J Bacteriol 171:3162–3167PubMedGoogle Scholar
  52. 52.
    Mortenson LE (1964) Ferredoxin requirement for nitrogen fixation by extracts of Clostridium pasteurianum. Biochim Biophys Acta 81:473–478PubMedGoogle Scholar
  53. 53.
    Angove HC, Yoo SJ, Burgess BK et al (1997) Mössbauer and EPR evidence for an all-ferrous Fe4S4 cluster with S = 4 in the Fe protein of nitrogenase. J Am Chem Soc 119:8730–8731Google Scholar
  54. 54.
    Watt GD, Reddy KRN (1994) Formation of an all ferrous Fe4S4 cluster in the iron protein component of Azotobacter vinelandii nitrogenase. J Inorg Biochem 53:281–294Google Scholar
  55. 55.
    Musgrave KB, Angove HC, Burgess BK et al (1998) All-ferrous titanium(III) citrate reduced Fe protein of nitrogenase: An XAS study of electronic and metrical structure. J Am Chem Soc 120:5325–5326Google Scholar
  56. 56.
    Strop P, Takahara PM, Chiu HJ et al (2001) Crystal structure of the all-ferrous [4Fe-4S]0 form of the nitrogenase iron protein from Azotobacter vinelandii. Biochemistry 40:651–656PubMedGoogle Scholar
  57. 57.
    Yoo SJ, Angove HC, Burgess BK et al (1998) Magnetic circular dichroism study of the all-ferrous [4Fe-4S] cluster of the Fe-protein of Azotobacter vinelandii nitrogenase. J Am Chem Soc 120:9704–9705Google Scholar
  58. 58.
    Nyborg AC, Johnson JL, Gunn A et al (2000) Evidence for a two-electron transfer using the all-ferrous Fe protein during nitrogenase catalysis. J Biol Chem 275:39307–39312PubMedGoogle Scholar
  59. 59.
    Hagen WR, Dunham WR, Braaksma A et al (1985) On the prosthetic group(s) of component II from nitrogenase: EPR of the Fe-protein from Azotobacter vinelandii. FEBS Lett 187:146–150PubMedGoogle Scholar
  60. 60.
    Hagen WR, Eady RR, Dunham WR et al (1985) A novel S = 3/2 EPR signal associated with native Fe proteins of nitrogenase. FEBS Lett 189:250–254PubMedGoogle Scholar
  61. 61.
    Lindahl PA, Day EP, Kent TA et al (1985) Mössbauer, EPR, and magnetization studies of the Azotobacter vinelandii Fe protein. J Biol Chem 260:11160–11173PubMedGoogle Scholar
  62. 62.
    Thorneley RNF, Ashby GA (1989) Oxidation of nitrogenase iron protein by dioxygen without inactivation could contribute to high respiration rates of Azotobacter species and facilitate nitrogen fixation in other aerobic environments. Biochem J 261:181–187PubMedGoogle Scholar
  63. 63.
    Larsen C, Christensen S, Watt GD (1995) Reductant-independent ATP hydrolysis catalyzed by homologous nitrogenase proteins from Azotobacter vinelandii and heterologous crosses with Clostridium pasteurianum. Arch Biochem Biophys 323:215–222PubMedGoogle Scholar
  64. 64.
    Thorneley RNF, Lowe DJ (1985) Kinetics and mechanisms of the nitrogenase enzyme system. In: Spiro TG (ed) Molybdenum Enzymes, pp. 221–284. Wiley, New York, NYGoogle Scholar
  65. 65.
    Ryle MJ, Lanzilotta WN, Mortenson LE et al (1995) Evidence for a central role of lysine 15 of Azotobacter vinelandii nitrogenase iron protein in nucleotide binding and protein conformational changes. J Biol Chem 270:13112–13117PubMedGoogle Scholar
  66. 66.
    Watt GD (1979) An electrochemical method for measuring redox potentials of low potential proteins by microcoulometry at controlled potentials. Anal Biochem 99:399–407PubMedGoogle Scholar
  67. 67.
    Watt GD, Wang ZC, Knotts RR (1986) Redox reactions of and nucleotide binding to the iron protein of Azotobacter vinelandii. Biochemistry 25:8156–8162Google Scholar
  68. 68.
    Lanzilotta WN, Ryle MJ, Seefeldt LC (1995) Nucleotide hydrolysis and protein conformational changes in Azotobacter vinelandii nitrogenase iron protein: Defining the function of aspartate 129. Biochemistry 34:10713–10723PubMedGoogle Scholar
  69. 69.
    Peters JW, Szilagyi RK (2006) Exploring new frontiers of nitrogenase structure and mechanism. Curr Opin Chem Biol 10:101–108PubMedGoogle Scholar
  70. 70.
    Yates MG (1991) The enzymology of molybdenum-dependent nitrogen fixation. In: Stacey G, Burris RH, Evans HJ (eds) Biological Nitrogen Fixation, pp. 685–735. Chapman and Hall, New York, NYGoogle Scholar
  71. 71.
    Lanzilotta WN, Parker VD, Seefeldt LC (1999) Thermodynamics of nucleotide interactions with the Azotobacter vinelandii nitrogenase iron protein. Biochim Biophys Acta 1429:411–421PubMedGoogle Scholar
  72. 72.
    Cordewener J, Haaker H, Van Ewijk P et al (1985) Properties of the MgATP and MgADP binding sites on the Fe protein of nitrogenase from Azotobacter vinelandii. Eur J Biochem 148:499–508PubMedGoogle Scholar
  73. 73.
    Weston MF, Kotake S, Davis LC (1983) Interaction of nitrogenase with nucleotide analogs of ATP and ADP and the effect of metal ions on ADP inhibition. Arch Biochem Biophys 225:809–817PubMedGoogle Scholar
  74. 74.
    Ryle MJ, Seefeldt LC (2000) Hydrolysis of nucleoside triphosphates other than ATP by nitrogenase. J Biol Chem 275:6214–6219PubMedGoogle Scholar
  75. 75.
    Sarma R, Mulder DW, Brecht E et al (2007) Probing the MgATP-bound conformation of the nitrogenase Fe protein by solution small-angle X-ray scattering. Biochemistry 46:14058–14066PubMedGoogle Scholar
  76. 76.
    Shah VK, Brill WJ (1977) Isolation of an iron-molybdenum cofactor from nitrogenase. Proc Natl Acad Sci USA 74:3249–3253PubMedGoogle Scholar
  77. 77.
    Zimmermann R, Münck E, Brill WJ et al (1978) Nitrogenase X: Mössbauer and EPR studies on reversibly oxidized MoFe protein from Azotobacter vinelandii OP: Nature of the iron centers. Biochim Biophys Acta 537:185–207PubMedGoogle Scholar
  78. 78.
    Lindahl PA, Papaefthymiou V, Orme-Johnson WH et al (1988) Mössbauer studies of solid thionin-oxidized MoFe protein of nitrogenase. J Biol Chem 263:19412–19418PubMedGoogle Scholar
  79. 79.
    Hagen WR, Wassink H, Eady RR et al (1987) Quantitative EPR of an S = 7/2 system in thionine-oxidized MoFe proteins of nitrogenase: A redefinition of the P-cluster concept. Eur J Biochem 169:457–465PubMedGoogle Scholar
  80. 80.
    Pierik AJ, Wassink H, Haaker H et al (1993) Redox properties and EPR spectroscopy of the P-clusters of Azotobacter vinelandii MoFe protein. Eur J Biochem 212:51–61PubMedGoogle Scholar
  81. 81.
    Tittsworth RC, Hales BJ (1993) Detection of EPR signals assigned to the 1-equiv-oxidized P-clusters of the nitrogenase MoFe protein from Azotobacter vinelandii. J Am Chem Soc 115:9763–9767Google Scholar
  82. 82.
    Lanzilotta WN, Fisher K, Seefeldt LC (1997) Evidence for electron transfer-dependent formation of a nitrogenase iron protein-molybdenum-iron protein tight complex: The role of aspartate 39. J Biol Chem 272:4157–4165PubMedGoogle Scholar
  83. 83.
    Morgan TV, Mortenson LE, McDonald JW et al (1988) Comparison of redox and EPR properties of the molybdenum iron proteins of Clostridium pasteurianum and Azotobacter vinelandii nitrogenases. J Inorg Biochem 33:111–120PubMedGoogle Scholar
  84. 84.
    Lanzilotta WN, Christiansen J, Dean DR et al (1998) Evidence for coupled electron and proton transfer in the [8Fe-7S] cluster of nitrogenase. Biochemistry 37:11376–11384PubMedGoogle Scholar
  85. 85.
    Hoover TR, Robertson AD, Cerny RL et al (1987) Identification of the V factor needed for the synthesis of the iron-molybdenum cofactor of nitrogenase as homocitrate. Nature 329:855–857PubMedGoogle Scholar
  86. 86.
    Lee HI, Benton PM, Laryukhin M et al (2003) The interstitial atom of the nitrogenase FeMo-cofactor: ENDOR and ESEEM show it is not an exchangeable nitrogen. J Am Chem Soc 125:5604–5605PubMedGoogle Scholar
  87. 87.
    Yang TC, Maeser NK, Laryukhin M et al (2005) The interstitial atom of the nitrogenase FeMo-cofactor: ENDOR and ESEEM evidence that it is not a nitrogen. J Am Chem Soc 127:12804–12805PubMedGoogle Scholar
  88. 88.
    Lukoyanov D, Pelmenschikov V, Maeser N et al (2007) Testing if the interstitial atom, X, of the nitrogenase molybdenum-iron cofactor is N or C: ENDOR, ESEEM, and DFT studies of the S = 3/2 resting state in multiple environments. Inorg Chem 46:11437–11449PubMedGoogle Scholar
  89. 89.
    George SJ, Igarashi RY, Xiao Y et al (2008) Extended X-ray absorption fine structure and nuclear resonance vibrational spectroscopy reveal that NifB-co, a FeMo-co precursor, comprises a 6Fe core with an interstitial light atom. J Am Chem Soc 130:5673–5680PubMedGoogle Scholar
  90. 90.
    Lovell T, Li J, Liu T et al (2001) FeMo cofactor of nitrogenase: A density functional study of states MN, MOX, MR, and MI. J Am Chem Soc 123:12392–12410PubMedGoogle Scholar
  91. 91.
    Pelmenshikov V, Case DA, Noodleman L (2008) Ligand-bound S = 1/2 FeMo-cofactor of nitrogenase: Hyperfine interaction analysis and implication for the central ligand X identity. Inorg Chem 47:6162–6172Google Scholar
  92. 92.
    Dance I (2003) The consequences of an interstitial N atom in the FeMo cofactor of nitrogenase. Chem Commun 3:324–325Google Scholar
  93. 93.
    Dance I (2007) The mechanistically significant coordination chemistry of dinitrogen at FeMo-co, the catalytic site of nitrogenase. J Am Chem Soc 129:1076–1088PubMedGoogle Scholar
  94. 94.
    Orme-Johnson WH, Hamilton WD, Jones TL et al (1972) Electron paramagnetic resonance of nitrogenase and nitrogenase components from Clostridium pasteurianum W5 and Azotobacter vinelandii OP. Proc Natl Acad Sci USA 69:3142–3145PubMedGoogle Scholar
  95. 95.
    Schultz FA, Gheller SF, Newton WE (1988) Iron molybdenum cofactor of nitrogenase: Electrochemical determination of the electron stoichiometry of the oxidized/semi-reduced couple. Biochem Biophys Res Commun 152:629–635PubMedGoogle Scholar
  96. 96.
    Huynh BH, Henzl MT, Christner JA et al (1980) Nitrogenase XII: Mössbauer studies of the MoFe protein from Clostridium pasteurianum W5. Biochim Biophys Acta 623:124–138PubMedGoogle Scholar
  97. 97.
    Yoo SJ, Angove HC, Papaefthymiou V et al (2000) Mössbauer study of the MoFe protein of nitrogenase from Azotobacter vinelandii using selective 57Fe enrichment of the M-centers. J Am Chem Soc 122:4926–4936Google Scholar
  98. 98.
    Watt GD, Burns A, Lough S et al (1980) Redox and spectroscopic properties of oxidized MoFe protein from Azotobacter vinelandii. Biochemistry 19:4926–4932PubMedGoogle Scholar
  99. 99.
    Lovell T, Liu T, Case DA et al (2003) Structural, spectroscopic, and redox consequences of a central ligand in the FeMoco of nitrogenase: A density functional theoretical study. J Am Chem Soc 125:8377–8383PubMedGoogle Scholar
  100. 100.
    Lee HI, Hales BJ, Hoffman BM (1997) Metal-ion valencies of the FeMo cofactor in CO-inhibited and resting state nitrogenase by 57Fe Q-band ENDOR. J Am Chem Soc 119:11395–11400Google Scholar
  101. 101.
    Burgess BK (1990) The iron-molybdenum cofactor of nitrogenase. Chem Rev 90:1377–1406Google Scholar
  102. 102.
    Smith BE, Durrant MC, Fairhurst SA et al (1999) Exploring the reactivity of the isolated iron-molybdenum cofactor of nitrogenase. Coord Chem Rev 185:669–687Google Scholar
  103. 103.
    Igarashi RY, Laryukhin M, Dos Santos PC et al (2005) Trapping H- bound to the nitrogenase FeMo-cofactor active site during H2 evolution: Characterization by ENDOR spectroscopy. J Am Chem Soc 127:6231–6241PubMedGoogle Scholar
  104. 104.
    Benton PMC, Laryukhin M, Mayer SM et al (2003) Localization of a substrate binding site on FeMo-cofactor in nitrogenase: Trapping propargyl alcohol with an α-70-substituted MoFe protein. Biochemistry 42:9102–9109PubMedGoogle Scholar
  105. 105.
    Lee HI, Igarashi RY, Laryukhin M et al (2004) An organometallic intermediate during alkyne reduction by nitrogenase. J Am Chem Soc 126:9563–9569PubMedGoogle Scholar
  106. 106.
    Barney BM, Laryukhin M, Igarashi RY et al (2005) Trapping a hydrazine reduction intermediate on the nitrogenase active site. Biochemistry 44:8030–8037PubMedGoogle Scholar
  107. 107.
    Barney BM, Yang TC, Igarashi RY et al (2005) Intermediates trapped during nitrogenase reduction of N2, CH3-N=NH, and H2N-NH2. J Am Chem Soc 127:14960–14961PubMedGoogle Scholar
  108. 108.
    Barney BM, Lukoyanov D, Yang TC et al (2006) A methyldiazene (HN=N-CH3) derived species bound to the nitrogenase active site FeMo-cofactor: Implications for mechanism. Proc Natl Acad Sci USA 103:17113–17118PubMedGoogle Scholar
  109. 109.
    Barney BM, McClead J, Lukoyanov D et al (2007) Diazene (HN=NH) is a substrate for nitrogenase: Insights into the pathway of N2 reduction. Biochemistry 46:6784–6794PubMedGoogle Scholar
  110. 110.
    Barney BM, Lukoyanov D, Igarashi RY et al (2009) Trapping an intermediate of dinitrogen (N2) reduction on nitrogenase. Biochemistry 48:9094–9102PubMedGoogle Scholar
  111. 111.
    Kim CH, Newton WE, Dean DR (1995) Role of the MoFe protein α-subunit histidine-195 residue in FeMo-cofactor binding and nitrogenase catalysis. Biochemistry 34:2798–2808PubMedGoogle Scholar
  112. 112.
    Thomann H, Bernardo M, Newton WE et al (1991) N coordination of MoFe cofactor requires His-195 of the MoFe protein α-subunit and is essential for biological nitrogen fixation. Proc Natl Acad Sci USA 88:6620–6623PubMedGoogle Scholar
  113. 113.
    Scott DJ, May HD, Newton WE et al (1990) Role for the nitrogenase MoFe protein α-subunit in FeMo-cofactor binding and catalysis. Nature 343:188–190PubMedGoogle Scholar
  114. 114.
    Dilworth MJ, Fisher K, Kim CH et al (1998) Effects on substrate reduction of substitution of histidine-195 by glutamine in the alpha-subunit of the MoFe protein of Azotobacter vinelandii nitrogenase. Biochemistry 37:17495–17505PubMedGoogle Scholar
  115. 115.
    Fisher K, Dilworth MJ, Newton WE (2000) Differential effects on N2 binding and reduction, HD formation, and azide reduction with α-195His- and α-191Gln-substituted MoFe proteins of Azotobacter vinelandii nitrogenase. Biochemistry 39:15570–15577PubMedGoogle Scholar
  116. 116.
    Burgess BK (1985) Substrate reactions of nitrogenase. In: Spiro TG (ed) Metal Ions in Biology: Molybdenum Enzymes, pp. 161–219. Wiley, New York, NYGoogle Scholar
  117. 117.
    Willing AH, Georgiadis MM, Rees DC et al (1989) Cross-linking of nitrogenase components. J Biol Chem 264:8499–8503PubMedGoogle Scholar
  118. 118.
    Willing A, Howard JB (1990) Crosslinking site in Azotobacter vinelandii complex. J Biol Chem 265:6596–6599PubMedGoogle Scholar
  119. 119.
    Emerich DW, Burris RH (1976) Interactions of heterologous nitrogenase components that generate catalytically inactive complexes. Proc Natl Acad Sci USA 73:4369–4373PubMedGoogle Scholar
  120. 120.
    Emerich DW, Burris RH (1978) Complementary functioning of the component proteins of nitrogenase from several bacteria. J Bacteriol 134:936–943PubMedGoogle Scholar
  121. 121.
    Emerich DW, Ljones T, Burris RH (1978) Nitrogenase: Properties of the catalytically inactive complex between the Azotobacter vinelandii MoFe protein and the Clostridium pasteurianum Fe protein. Biochim Biophys Acta 527:359–369PubMedGoogle Scholar
  122. 122.
    Chan JM, Ryle MJ, Seefeldt LC (1999) Evidence that MgATP accelerates primary electron transfer in a Clostridium pasteurianum Fe protein-Azotobacter vinelandii MoFe protein nitrogenase tight complex. J Biol Chem 274:17593–17598PubMedGoogle Scholar
  123. 123.
    Lanzilotta WN, Seefeldt LC (1997) Changes in the midpoint potentials of the nitrogenase metal centers as a result of iron protein-molybdenum-iron protein complex formation. Biochemistry 36:12976–12983PubMedGoogle Scholar
  124. 124.
    Clarke TA, Yousafzai FK, Eady RR (1999) Klebsiella pneumoniae nitrogenase: Formation and stability of putative beryllium fluoride-ADP transition state complexes. Biochemistry 38:9906–9913PubMedGoogle Scholar
  125. 125.
    Duyvis MG, Wassink H, Haaker H (1996) Formation and characterization of a transition state complex of Azotobacter vinelandii nitrogenase. FEBS Lett 380:233–236PubMedGoogle Scholar
  126. 126.
    Renner KA, Howard JB (1996) Aluminum fluoride inhibition of nitrogenase: Stabilization of a nucleotide-Fe-protein-MoFe protein complex. Biochemistry 35:5353–5358PubMedGoogle Scholar
  127. 127.
    Miller RW, Eady RR, Fairhurst SA et al (2001) Transition state complexes of the Klebsiella pneumoniae nitrogenase proteins: Spectroscopic properties of aluminum fluoride-stabilized and beryllium fluoride-stabilized MgADP complexes reveal conformational differences of the Fe protein. Eur J Biochem 268:809–818PubMedGoogle Scholar
  128. 128.
    Spee JH, Arendsen AF, Wassink H et al (1998) Redox properties and electron paramagnetic resonance spectroscopy of the transition state complex of Azotobacter vinelandii nitrogenase. FEBS Lett 432:55–58PubMedGoogle Scholar
  129. 129.
    Duyvais MG, Wassink H, Haaker H (1998) Nitrogenase of Azotobacter vinelandii: Kinetic analysis of the Fe protein redox cycle. Biochemistry 37:17345–17354Google Scholar
  130. 130.
    Christiansen J, Dean DR, Seefeldt LC (2001) Mechanistic features of the Mo-containing nitrogenase. Annu Rev Plant Physiol Plant Mol Biol 52:269–295PubMedGoogle Scholar
  131. 131.
    Hageman RV, Burris RH (1978) Nitrogenase and nitrogenase reductase associate and dissociate with each catalytic cycle. Proc Natl Acad Sci USA 75:2699–2702PubMedGoogle Scholar
  132. 132.
    Lowe DJ, Thorneley RNF (1984) The mechanism of Klebsiella pneumoniae nitrogenase action: The determination of rate constants required for the simulation of kinetics of N2 reduction and H2 evolution. Biochem J 224:895–901PubMedGoogle Scholar
  133. 133.
    Thorneley RNF, Lowe DJ (1984) The mechanism of Klebsiella pneumoniae nitrogenase action: Pre-steady-state kinetics of an enzyme-bound intermediate in N2 reduction and of NH3 formation. Biochem J 224:887–894PubMedGoogle Scholar
  134. 134.
    Thorneley RNF, Lowe DJ (1984) The mechanism of Klebsiella pneumoniae nitrogenase action: Stimulation of the dependences of H2-evolution rate on component-protein concentration and ratio and sodium dithionite concentration. Biochem J 224:903–909PubMedGoogle Scholar
  135. 135.
    Liang J, Burris RH (1988) Hydrogen burst associated with nitrogenase-catalyzed reactions. Proc Natl Acad Sci USA 85:9446–9450PubMedGoogle Scholar
  136. 136.
    Rivera-Ortiz JM, Burris RH (1975) Interactions among substrates and inhibitors of nitrogenase. J Bacteriol 123:537–545PubMedGoogle Scholar
  137. 137.
    Davis LC, Shah VK, Brill WJ (1975) Nitrogenase: Effect of component ratio, ATP and H2 on the distribution of electrons to alternative substrates. Biochim Biophys Acta 403:67–78PubMedGoogle Scholar
  138. 138.
    Lee HI, Cameron LM, Hales BJ et al (1997) CO binding to the FeMo cofactor of CO-inhibited nitrogenase: 13CO and 1H Q-band ENDOR investigation. J Am Chem Soc 119:10121–10126Google Scholar
  139. 139.
    Pollock CR, Lee HI, Cameron LM et al (1995) Investigation of CO bound to inhibited forms of nitrogenase MoFe protein by 13C ENDOR. J Am Chem Soc 117:8686–8687Google Scholar
  140. 140.
    Christie PD, Lee HI, Cameron LM et al (1996) Identification of the CO-binding cluster in nitrogenase MoFe protein by ENDOR of 57Fe isotopomers. J Am Chem Soc 118:8707–8709Google Scholar
  141. 141.
    Maskos Z, Hales BJ (2003) Photo-lability of CO bound to Mo-nitrogenase from Azotobacter vinelandii. J Inorg Biochem 93:11–17PubMedGoogle Scholar
  142. 142.
    Maskos Z, Fisher K, Sørlie M et al (2005) Variant MoFe proteins of Azotobacter vinelandii: Effects of carbon monoxide on electron paramagnetic resonance spectra generated during enzyme turnover. J Biol Inorg Chem 10:394–406PubMedGoogle Scholar
  143. 143.
    Dos Santos PC, Mayer SM, Barney BM et al (2007) Alkyne substrate interaction within the nitrogenase MoFe protein. J Inorg Biochem 101:1642–1648PubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2011

Authors and Affiliations

  • Zhi-Yong Yang
    • 1
  • Karamatullah Danyal
    • 1
  • Lance C. Seefeldt
    • 1
    Email author
  1. 1.Department of Chemistry and BiochemistryUtah State UniversityLoganUSA

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