Summary: A non-heme iron-sulfur protein isolated from Clostridium pasteurianum and other bacteria. It is a component of NITROGENASE, which is active in nitrogen fixation, and consists of two subunits with molecular weights of 59.5 kDa and 50.7 kDa, respectively.

Top Publications

  1. Georgiadis M, Komiya H, Chakrabarti P, Woo D, Kornuc J, Rees D. Crystallographic structure of the nitrogenase iron protein from Azotobacter vinelandii. Science. 1992;257:1653-9 pubmed
    ..The Fe-protein structure may be relevant to the functioning of other biochemical energy-transducing systems containing two nucleotide-binding sites, including membrane transport proteins...
  2. Lee C, Blank M, Fay A, Yoshizawa J, Hu Y, Hodgson K, et al. Stepwise formation of P-cluster in nitrogenase MoFe protein. Proc Natl Acad Sci U S A. 2009;106:18474-8 pubmed publisher
    ..e., one P-cluster is formed in one alphabeta dimer before the other in the second alphabeta dimer. ..
  3. Schindelin H, Kisker C, Schlessman J, Howard J, Rees D. Structure of ADP x AIF4(-)-stabilized nitrogenase complex and its implications for signal transduction. Nature. 1997;387:370-6 pubmed
    ..Interactions in the nitrogenase complex have broad implications for signal and energy transduction mechanisms in multiprotein complexes. ..
  4. Corbett M, Hu Y, Naderi F, Ribbe M, Hedman B, Hodgson K. Comparison of iron-molybdenum cofactor-deficient nitrogenase MoFe proteins by X-ray absorption spectroscopy: implications for P-cluster biosynthesis. J Biol Chem. 2004;279:28276-82 pubmed
    ..This type of model suggests that the P-cluster is formed by the condensation of two [Fe(4)S(4)] fragments, possibly concomitant with Fe protein (NifH)-induced conformational change. ..
  5. Dos Santos P, Dean D, Hu Y, Ribbe M. Formation and insertion of the nitrogenase iron-molybdenum cofactor. Chem Rev. 2004;104:1159-73 pubmed
  6. Hu Y, Corbett M, Fay A, Webber J, Hodgson K, Hedman B, et al. FeMo cofactor maturation on NifEN. Proc Natl Acad Sci U S A. 2006;103:17119-24 pubmed
    ..Our findings not only clarify the process of FeMoco maturation, but also provide useful insights into the other facets of nitrogenase chemistry. ..
  7. Igarashi R, Laryukhin M, Dos Santos P, Lee H, Dean D, Seefeldt L, et al. Trapping H- bound to the nitrogenase FeMo-cofactor active site during H2 evolution: characterization by ENDOR spectroscopy. J Am Chem Soc. 2005;127:6231-41 pubmed
    ..The data disfavors any model that involves protonation of sulfides, and thus suggests that the intermediate instead contains two chemically equivalent bound hydrides; it appears unlikely that these are terminal monohydrides. ..
  8. Kaiser J, Hu Y, Wiig J, Rees D, Ribbe M. Structure of precursor-bound NifEN: a nitrogenase FeMo cofactor maturase/insertase. Science. 2011;331:91-4 pubmed publisher
    ..6 angstrom resolution. From a structural comparison of NifEN with des-M-cluster NifDK and holo NifDK, we propose similar pathways of cluster insertion for the homologous NifEN and NifDK proteins. ..
  9. Han J, Newton W. Differentiation of acetylene-reduction sites by stereoselective proton addition during Azotobacter vinelandii nitrogenase-catalyzed C2D2 reduction. Biochemistry. 2004;43:2947-56 pubmed
    ..The observed nitrogenase-catalyzed production of ethylene-d(1) from C(2)D(2) implicates a reversible protonation step in the mechanistic pathway...

More Information


  1. Spatzal T, Aksoyoglu M, Zhang L, Andrade S, Schleicher E, Weber S, et al. Evidence for interstitial carbon in nitrogenase FeMo cofactor. Science. 2011;334:940 pubmed publisher
    ..Atomic-resolution x-ray diffraction data and an electron spin echo envelope modulation (ESEEM) analysis now provide direct evidence that the ligand is a carbon species...
  2. Peters J, Stowell M, Soltis S, Finnegan M, Johnson M, Rees D. Redox-dependent structural changes in the nitrogenase P-cluster. Biochemistry. 1997;36:1181-7 pubmed
    ..These observed redox-mediated structural changes of the P-cluster suggest a role for this cluster in coupling electron transfer and proton transfer in nitrogenase. ..
  3. Hu Y, Fay A, Lee C, Ribbe M. P-cluster maturation on nitrogenase MoFe protein. Proc Natl Acad Sci U S A. 2007;104:10424-9 pubmed
    ..Furthermore, we provide spectroscopic evidence that the [4Fe-4S] cluster-like fragments can be converted to P-clusters, thereby firmly establishing the physiological relevance of the previously identified P-cluster precursor...
  4. Einsle O, Tezcan F, Andrade S, Schmid B, Yoshida M, Howard J, et al. Nitrogenase MoFe-protein at 1.16 A resolution: a central ligand in the FeMo-cofactor. Science. 2002;297:1696-700 pubmed
    ..The presence of a nitrogen atom in the cofactor would have important implications for the mechanism of dinitrogen reduction by nitrogenase. ..
  5. Ribbe M, Burgess B. The chaperone GroEL is required for the final assembly of the molybdenum-iron protein of nitrogenase. Proc Natl Acad Sci U S A. 2001;98:5521-5 pubmed
    ..The involvement of the molecular chaperone GroEL in the insertion of a metal cluster into an apoprotein may have broad implications for the maturation of other metalloenzymes. ..
  6. Rupnik K, Lee C, Hu Y, Ribbe M, Hales B. [4Fe4S]2+ clusters exhibit ground-state paramagnetism. J Am Chem Soc. 2011;133:6871-3 pubmed publisher
    ..This unique characteristic suggests a specific coupling in these clusters necessary for nitrogen fixation and implies an evolutionary connection between the clusters in the two proteins. ..
  7. Goodwin P, Agar J, Roll J, Roberts G, Johnson M, Dean D. The Azotobacter vinelandii NifEN complex contains two identical [4Fe-4S] clusters. Biochemistry. 1998;37:10420-8 pubmed
  8. Hu Y, Corbett M, Fay A, Webber J, Hodgson K, Hedman B, et al. Nitrogenase Fe protein: A molybdate/homocitrate insertase. Proc Natl Acad Sci U S A. 2006;103:17125-30 pubmed
  9. Hu Y, Yoshizawa J, Fay A, Lee C, Wiig J, Ribbe M. Catalytic activities of NifEN: implications for nitrogenase evolution and mechanism. Proc Natl Acad Sci U S A. 2009;106:16962-6 pubmed publisher
    ..More importantly, the discrepancy between the two systems may provide useful insights into nitrogenase mechanism and allow reconstruction of a fully functional nitrogenase from the "skeleton" enzyme, NifEN. ..
  10. Hu Y, Fay A, Ribbe M. Identification of a nitrogenase FeMo cofactor precursor on NifEN complex. Proc Natl Acad Sci U S A. 2005;102:3236-41 pubmed
    ..The FeMoco-maturation assay described here will further address the remaining questions related to the assembly mechanism of the ever-intriguing FeMoco...
  11. Hoffman B, Dean D, Seefeldt L. Climbing nitrogenase: toward a mechanism of enzymatic nitrogen fixation. Acc Chem Res. 2009;42:609-19 pubmed publisher
    ..Although the summit remains distant, we look forward to continued progress in the ascent. ..
  12. Seefeldt L, Hoffman B, Dean D. Mechanism of Mo-dependent nitrogenase. Annu Rev Biochem. 2009;78:701-22 pubmed publisher
  13. Broach R, Rupnik K, Hu Y, Fay A, Cotton M, Ribbe M, et al. Variable-temperature, variable-field magnetic circular dichroism spectroscopic study of the metal clusters in the DeltanifB and DeltanifH mofe proteins of nitrogenase from Azotobacter vinelandii. Biochemistry. 2006;45:15039-48 pubmed
    ..These results suggest that the DeltanifH MoFe protein contains a pair of neighboring, unusual [4Fe-4S]-like clusters, which are paramagnetic in their oxidized state. ..
  14. Yoshizawa J, Blank M, Fay A, Lee C, Wiig J, Hu Y, et al. Optimization of FeMoco maturation on NifEN. J Am Chem Soc. 2009;131:9321-5 pubmed publisher
  15. Hu Y, Ribbe M. Decoding the nitrogenase mechanism: the homologue approach. Acc Chem Res. 2010;43:475-84 pubmed publisher
  16. Lancaster K, Roemelt M, Ettenhuber P, Hu Y, Ribbe M, Neese F, et al. X-ray emission spectroscopy evidences a central carbon in the nitrogenase iron-molybdenum cofactor. Science. 2011;334:974-7 pubmed publisher
    ..Identification of the central atom will drive further studies on its role in catalysis. ..
  17. Ribbe M, Hu Y, Guo M, Schmid B, Burgess B. The FeMoco-deficient MoFe protein produced by a nifH deletion strain of Azotobacter vinelandii shows unusual P-cluster features. J Biol Chem. 2002;277:23469-76 pubmed
    ..These unusual catalytic and spectroscopic properties might indicate the presence of a P-cluster precursor or a P-cluster trapped in an unusual conformation or oxidation state. ..
  18. Schmid B, Ribbe M, Einsle O, Yoshida M, Thomas L, Dean D, et al. Structure of a cofactor-deficient nitrogenase MoFe protein. Science. 2002;296:352-6 pubmed
    ..A predominantly positively charged funnel is revealed; this funnel is of sufficient size to accommodate insertion of the negatively charged cofactor...
  19. Schmid B, Einsle O, Chiu H, Willing A, Yoshida M, Howard J, et al. Biochemical and structural characterization of the cross-linked complex of nitrogenase: comparison to the ADP-AlF4(-)-stabilized structure. Biochemistry. 2002;41:15557-65 pubmed
  20. Hu Y, Fay A, Lee C, Yoshizawa J, Ribbe M. Assembly of nitrogenase MoFe protein. Biochemistry. 2008;47:3973-81 pubmed publisher
  21. Lee H, Igarashi R, Laryukhin M, Doan P, Dos Santos P, Dean D, et al. An organometallic intermediate during alkyne reduction by nitrogenase. J Am Chem Soc. 2004;126:9563-9 pubmed
    ..Of the two most attractive structures, one singly reduced at C3 (4), the other being the doubly reduced allyl alcohol product (6), we tentatively favor 6 because of the "natural" assignment it affords for H(b). ..
  22. Mills P, Footitt E, Ceyhan S, Waters P, Jakobs C, Clayton P, et al. Urinary AASA excretion is elevated in patients with molybdenum cofactor deficiency and isolated sulphite oxidase deficiency. J Inherit Metab Dis. 2012;35:1031-6 pubmed publisher
    ..This should be taken into account when interpreting the laboratory data. Sulphite was shown to inhibit ?-aminoadipic semialdehyde dehydrogenase in vitro. ..
  23. Afshar S, Johnson E, de Vries S, Schröder I. Properties of a thermostable nitrate reductase from the hyperthermophilic archaeon Pyrobaculum aerophilum. J Bacteriol. 2001;183:5491-5 pubmed
    ..When incubated at 100 degrees C, the purified nitrate reductase had a half-life of 1.5 h. This study constitutes the first description of a nitrate reductase from a hyperthermophilic archaeon...
  24. Igarashi R, Dos Santos P, Niehaus W, Dance I, Dean D, Seefeldt L. Localization of a catalytic intermediate bound to the FeMo-cofactor of nitrogenase. J Biol Chem. 2004;279:34770-5 pubmed
    ..Refinement of the binding mode and site was accomplished by the use of density functional and force field calculations pointing to an eta(2) coordination at Fe-6 of the FeMo-cofactor. ..
  25. Siemann S, Schneider K, Behrens K, Knöchel A, Klipp W, Muller A. FeMo cofactor biosynthesis in a nifE- mutant of Rhodobacter capsulatus. Eur J Biochem. 2001;268:1940-52 pubmed
    ..The implications of our findings for the biosynthesis of the FeMoco will be discussed. ..
  26. Zhang Y, Mei T, Yang D, Zhang Y, Wang B, Qu J. Synthesis and reactivity of thiolate-bridged multi-iron complexes supported by cyclic (alkyl)(amino)carbene. Dalton Trans. 2017;46:15888-15896 pubmed publisher
    ..Furthermore, complexes 3 and 4 exhibit good exchange reactivity toward the azide anion to give novel thiolate-bridged diiron complexes with two azido ligands in a trans arrangement...
  27. Chatelet C, Meyer J. Mapping the interaction of the [2Fe-2S] Clostridium pasteurianum ferredoxin with nitrogenase MoFe protein. Biochim Biophys Acta. 2001;1549:32-6 pubmed
    ..The positions on the ferredoxin molecule of the residues interacting with the MoFe protein were determined using the recently elucidated crystal structure of the homologous [2Fe-2S] ferredoxin from Aquifex aeolicus. ..
  28. Hallenbeck P, George G, Prince R, Thorneley R. Characterization of a modified nitrogenase Fe protein from Klebsiella pneumoniae in which the 4Fe4S cluster has been replaced by a 4Fe4Se cluster. J Biol Inorg Chem. 2009;14:673-82 pubmed publisher
  29. Chen X, Zhang W, Duncan J, Lee S. Iron-amide-sulfide and iron-imide-sulfide clusters: heteroligated core environments relevant to the nitrogenase FeMo cofactor. Inorg Chem. 2012;51:12891-904 pubmed publisher
    ..Cluster 10 presents an [Fe(4)NS(3)] core framework virtually isometric with the isostructural [Fe(4)S(3)X] subunit of the FeMo cofactor, thus providing a synthetic structural representation for this portion of the cofactor core. ..
  30. Paustian T, Shah V, Roberts G. Apodinitrogenase: purification, association with a 20-kilodalton protein, and activation by the iron-molybdenum cofactor in the absence of dinitrogenase reductase. Biochemistry. 1990;29:3515-22 pubmed
    ..This suggests that sulfhydryl(s) is (are) important for the FeMoco-activation reaction. ..
  31. Hu Y, Ribbe M. Biosynthesis of the iron-molybdenum cofactor of nitrogenase. J Biol Chem. 2013;288:13173-7 pubmed publisher
    ..Here, we review the recent progress in the research area of M-cluster assembly, with an emphasis on our work that provides useful insights into the mechanistic details of this process. ..
  32. Rubio L, Ludden P. Biosynthesis of the iron-molybdenum cofactor of nitrogenase. Annu Rev Microbiol. 2008;62:93-111 pubmed publisher
    ..This effort is starting to provide insights into the structures of the FeMo-co biosynthetic intermediates and into the biochemical details of FeMo-co synthesis. ..
  33. Kastner J, Blöchl P. Model for acetylene reduction by nitrogenase derived from density functional theory. Inorg Chem. 2005;44:4568-75 pubmed
    ..Our proposed mechanism is consistent with experimentally observed stereoselectivity and the ability of C2H2 to suppress H2 production by nitrogenase. ..
  34. Cui Z, Dunford A, Durrant M, Henderson R, Smith B. Binding sites of nitrogenase: kinetic and theoretical studies of cyanide binding to extracted FeMo-cofactor derivatives. Inorg Chem. 2003;42:6252-64 pubmed
  35. Fisher K, Newton W, Lowe D. Electron paramagnetic resonance analysis of different Azotobacter vinelandii nitrogenase MoFe-protein conformations generated during enzyme turnover: evidence for S = 3/2 spin states from reduced MoFe-protein intermediates. Biochemistry. 2001;40:3333-9 pubmed
    ..J., and Thorneley, R. N. F. (1984) Biochem. J. 224, 887-909] and propose that they arise from reduced states of the MoFe protein and reflect different conformations of the FeMo cofactor with different protonation states. ..
  36. Hu Y, Fay A, Schmid B, Makar B, Ribbe M. Molecular insights into nitrogenase FeMoco insertion: TRP-444 of MoFe protein alpha-subunit locks FeMoco in its binding site. J Biol Chem. 2006;281:30534-41 pubmed
    ..Our results provide some of the initial molecular insights into the FeMoco insertion process and, moreover, have useful implications for the overall scheme of nitrogenase assembly. ..
  37. Hu Y, Corbett M, Fay A, Webber J, Hedman B, Hodgson K, et al. Nitrogenase reactivity with P-cluster variants. Proc Natl Acad Sci U S A. 2005;102:13825-30 pubmed
  38. Dance I. New insights into the reaction capabilities of His195 adjacent to the active site of nitrogenase. J Inorg Biochem. 2017;169:32-43 pubmed publisher
    ..The accumulated results suggest that protonated His195 could be the agent for the first, most difficult, transfer of H to bound substrate N2. ..
  39. Reiss J, Hahnewald R. Molybdenum cofactor deficiency: Mutations in GPHN, MOCS1, and MOCS2. Hum Mutat. 2011;32:10-8 pubmed publisher
    ..For type A deficiency an effective substitution therapy has been described recently. ..
  40. Dance I. The mechanism of nitrogenase. Computed details of the site and geometry of binding of alkyne and alkene substrates and intermediates. J Am Chem Soc. 2004;126:11852-63 pubmed
    ..Alternative binding sites and geometries for ethyne and ethene, relevant to the wild-type protein, are described. This model defines the location and scene for detailed investigation of the mechanism of nitrogenase. ..
  41. Chow T, Tabita F. Reciprocal light-dark transcriptional control of nif and rbc expression and light-dependent posttranslational control of nitrogenase activity in Synechococcus sp. strain RF-1. J Bacteriol. 1994;176:6281-5 pubmed
    ..By contrast, transcription of the rbcL gene, which encodes the catalytic subunit of the key enzyme of CO2 fixation (a light-dependent process), was enhanced in the light. ..
  42. Scott K, Rolfe B, Shine J. Biological nitrogen fixation: primary structure of the Rhizobium trifolii iron protein gene. DNA. 1983;2:149-55 pubmed
    ..The nif structural genes are preceded by a DNA sequence which is repeated at least three times in the Rhizobium trifolii genome. ..
  43. Cotton M, Rupnik K, Broach R, Hu Y, Fay A, Ribbe M, et al. VTVH-MCD study of the Delta nifB Delta nifZ MoFe protein from Azotobacter vinelandii. J Am Chem Soc. 2009;131:4558-9 pubmed publisher
    ..These results suggest that the MoFe protein is synthesized in a stepwise fashion where NifZ is specifically required for the formation of the second P-cluster. ..
  44. Rittle J, Peters J. Fe-N2/CO complexes that model a possible role for the interstitial C atom of FeMo-cofactor (FeMoco). Proc Natl Acad Sci U S A. 2013;110:15898-903 pubmed publisher
    ..The data presented are contextualized in support of a hypothesis wherein modulation of a belt Fe-C interaction in the FeMo-cofactor facilitates substrate binding and reduction. ..
  45. Zhou Z, Yan W, Wan H, Tsai K. Synthesis and characterization of homochiral polymeric S-malato molybdate(VI): toward the potentially stereospecific formation and absolute configuration of iron-molybdenum cofactor in nitrogenase. J Inorg Biochem. 2002;90:137-43 pubmed
    ..The absolute configuration of wild-type FeMo-cofactor is assigned as Delta(R). ..
  46. Vijayakumar K, Gunny R, Grunewald S, Carr L, Chong K, Devile C, et al. Clinical neuroimaging features and outcome in molybdenum cofactor deficiency. Pediatr Neurol. 2011;45:246-52 pubmed publisher
    ..Imaging pattern on early brain MRI (<1 week) may prompt the diagnosis, potentially allowing early treatment and disease modifications. ..
  47. Sarma R, Barney B, Keable S, Dean D, Seefeldt L, Peters J. Insights into substrate binding at FeMo-cofactor in nitrogenase from the structure of an alpha-70(Ile) MoFe protein variant. J Inorg Biochem. 2010;104:385-9 pubmed publisher
  48. Pickett C, Vincent K, Ibrahim S, Gormal C, Smith B, Best S. Electron-transfer chemistry of the iron-molybdenum cofactor of nitrogenase: delocalized and localized reduced states of FeMoco which allow binding of carbon monoxide to iron and molybdenum. Chemistry. 2003;9:76-87 pubmed
  49. Hitzert M, Bos A, Bergman K, Veldman A, Schwarz G, Santamaria Araujo J, et al. Favorable outcome in a newborn with molybdenum cofactor type A deficiency treated with cPMP. Pediatrics. 2012;130:e1005-10 pubmed publisher
    ..We strongly recommend starting cPMP treatment as soon as possible after birth in infants diagnosed with MoCD type A. ..
  50. Hinnemann B, Nørskov J. Modeling a central ligand in the nitrogenase FeMo cofactor. J Am Chem Soc. 2003;125:1466-7 pubmed
    ..By comparison of bond geometries with the crystallographically determined values, we show that the central ligand is most likely nitrogen. ..
  51. Hales B. Magnetic circular dichroism spectroscopy. Methods Mol Biol. 2011;766:207-19 pubmed publisher
    ..The basics of this technique are discussed along with examples of how MCD spectroscopy has been successfully used to elucidate the metal clusters of Nif proteins from nitrogen-fixing bacteria. ..
  52. Rupnik K, Lee C, Wiig J, Hu Y, Ribbe M, Hales B. Nonenzymatic synthesis of the P-cluster in the nitrogenase MoFe protein: evidence of the involvement of all-ferrous [Fe4S4](0) intermediates. Biochemistry. 2014;53:1108-16 pubmed publisher
    ..These results demonstrate that the nitrogenase P-cluster can be generated in the absence of NifH and MgATP. ..
  53. Tolland J, Thorneley R. Stopped-flow Fourier transform infrared spectroscopy allows continuous monitoring of azide reduction, carbon monoxide inhibition, and ATP hydrolysis by nitrogenase. Biochemistry. 2005;44:9520-7 pubmed