56. Fay AW, Wiig JA, Lee CC, Hu Y (2015) Identification and characterization of functional
homologs of nitrogenase cofactor biosynthesis protein NifB from methanogens. Proc Natl
Acad Sci U S A 112(48):14829–14833
57. Boal AK, Grove TL, McLaughlin MI, Yennawar NH, Booker SJ, Rosenzweig AC (2011)
Structural basis for methyl transfer by a radical SAM enzyme. Science 332(6033):1089–
1092
58. Grove TL, Benner JS, Radle MI et al (2011) A radically different mechanism for
S-adenosylmethionine-dependent methyltransferases. Science 332(6029):604–607
59. Rettberg LA, Wilcoxen J, Lee CC et al (2018) Probing the coordination and function of
Fe 4 S 4 modules in nitrogenase assembly protein NifB. Nat Commun 9(1):2824
60. Rettberg LA, Wilcoxen J, Jasniewski AJ et al (2020) Identity and function of an essential
nitrogen ligand of the nitrogenase cofactor biosynthesis protein NifB. Nat Commun 11
(1):1757
61. Tanifuji K, Lee CC, Sickerman NS et al (2018) Tracing the ‘ninth sulfur’ of the nitrogenase
cofactor via a semi-synthetic approach. Nat Chem 10(5):568–572
62. Jasniewski AJ, Wilcoxen J, Tanifuji K et al (2019) Spectroscopic characterization of an
eight-iron nitrogenase cofactor precursor that lacks the “9th sulfur.” Angew Chem Int Ed
Engl 58(41):14703–14707
63. Fugate CJ, Jarrett JT (2012) Biotin synthase: insights into radical-mediated carbon-sulfur
bond formation. Biochim Biophys Acta 1824(11):1213–1222
64. Zheng L, White RH, Cash VL, Dean DR (1994) Mechanism for the desulfurization of
L-cysteine catalyzed by the nifS gene product. Biochemistry 33(15):4714–4720
65. Rupnik K, Tanifuji K, Rettberg L, Ribbe MW, Hu Y, Hales BJ (2019) Electron
paramagnetic resonance and magnetic circular dichroism spectra of the nitrogenase M cluster
precursor suggest sulfur migration upon oxidation: a proposal for substrate and inhibitor
binding. Chem BioChem. https://doi.org/10.1002/cbic.201900681
66. Goodwin PJ, Agar JN, Roll JT, Roberts GP, Johnson MK, Dean DR (1998) The Azotobacter
vinelandii NifEN complex contains two identical [4Fe-4S] clusters. Biochemistry 37
(29):10420–10428
67. Hu Y, Fay AW, Ribbe MW (2005) Identification of a nitrogenase FeMo cofactor precursor
on NifEN complex. Proc Natl Acad Sci U S A 102(9):3236–3241
68. Corbett MC, Hu Y, Fay AW, Ribbe MW, Hedman B, Hodgson KO (2006) Structural
insights into a protein-bound iron-molybdenum cofactor precursor. Proc Natl Acad Sci U S
A 103(5):1238–1243
69. Lancaster KM, Hu Y, Bergmann U, Ribbe MW, DeBeer S (2013) X-ray spectroscopic
observation of an interstitial carbide in NifEN-bound FeMoco precursor. J Am Chem Soc
135(2):610–612
70. Kaiser JT, Hu Y, Wiig JA, Rees DC, Ribbe MW (2011) Structure of precursor-bound
NifEN: a nitrogenase FeMo cofactor maturase/insertase. Science 331(6013):91–94
71. Hu Y, Corbett MC, Fay AW et al (2006a) Nitrogenase Fe protein: A molybdate/homocitrate
insertase. Proc Natl Acad Sci U S A 103(46):17125–17130
72. Hu Y, Corbett MC, Fay AW et al (2006b) FeMo cofactor maturation on NifEN. Proc Natl
Acad Sci U S A 103(46):17119–17124
73. Yoshizawa JM, Blank MA, Fay AW et al (2009) Optimization of FeMoco maturation on
NifEN. J Am Chem Soc 131(26):9321–9325
74. Fay AW, Blank MA, Lee CC et al (2011) Spectroscopic characterization of the isolated
iron-molybdenum cofactor (FeMoco) precursor from the protein NifEN. Angew Chem Int
Ed Engl 50(34):7787–7790
75. Fay AW, Blank MA, Rebelein JG et al (2016) Assembly scaffold NifEN: a structural and
functional homolog of the nitrogenase catalytic component. Proc Natl Acad Sci U S A 113
(34):9504–9508
182
C.-C. Lee et al.
homologs of nitrogenase cofactor biosynthesis protein NifB from methanogens. Proc Natl
Acad Sci U S A 112(48):14829–14833
57. Boal AK, Grove TL, McLaughlin MI, Yennawar NH, Booker SJ, Rosenzweig AC (2011)
Structural basis for methyl transfer by a radical SAM enzyme. Science 332(6033):1089–
1092
58. Grove TL, Benner JS, Radle MI et al (2011) A radically different mechanism for
S-adenosylmethionine-dependent methyltransferases. Science 332(6029):604–607
59. Rettberg LA, Wilcoxen J, Lee CC et al (2018) Probing the coordination and function of
Fe 4 S 4 modules in nitrogenase assembly protein NifB. Nat Commun 9(1):2824
60. Rettberg LA, Wilcoxen J, Jasniewski AJ et al (2020) Identity and function of an essential
nitrogen ligand of the nitrogenase cofactor biosynthesis protein NifB. Nat Commun 11
(1):1757
61. Tanifuji K, Lee CC, Sickerman NS et al (2018) Tracing the ‘ninth sulfur’ of the nitrogenase
cofactor via a semi-synthetic approach. Nat Chem 10(5):568–572
62. Jasniewski AJ, Wilcoxen J, Tanifuji K et al (2019) Spectroscopic characterization of an
eight-iron nitrogenase cofactor precursor that lacks the “9th sulfur.” Angew Chem Int Ed
Engl 58(41):14703–14707
63. Fugate CJ, Jarrett JT (2012) Biotin synthase: insights into radical-mediated carbon-sulfur
bond formation. Biochim Biophys Acta 1824(11):1213–1222
64. Zheng L, White RH, Cash VL, Dean DR (1994) Mechanism for the desulfurization of
L-cysteine catalyzed by the nifS gene product. Biochemistry 33(15):4714–4720
65. Rupnik K, Tanifuji K, Rettberg L, Ribbe MW, Hu Y, Hales BJ (2019) Electron
paramagnetic resonance and magnetic circular dichroism spectra of the nitrogenase M cluster
precursor suggest sulfur migration upon oxidation: a proposal for substrate and inhibitor
binding. Chem BioChem. https://doi.org/10.1002/cbic.201900681
66. Goodwin PJ, Agar JN, Roll JT, Roberts GP, Johnson MK, Dean DR (1998) The Azotobacter
vinelandii NifEN complex contains two identical [4Fe-4S] clusters. Biochemistry 37
(29):10420–10428
67. Hu Y, Fay AW, Ribbe MW (2005) Identification of a nitrogenase FeMo cofactor precursor
on NifEN complex. Proc Natl Acad Sci U S A 102(9):3236–3241
68. Corbett MC, Hu Y, Fay AW, Ribbe MW, Hedman B, Hodgson KO (2006) Structural
insights into a protein-bound iron-molybdenum cofactor precursor. Proc Natl Acad Sci U S
A 103(5):1238–1243
69. Lancaster KM, Hu Y, Bergmann U, Ribbe MW, DeBeer S (2013) X-ray spectroscopic
observation of an interstitial carbide in NifEN-bound FeMoco precursor. J Am Chem Soc
135(2):610–612
70. Kaiser JT, Hu Y, Wiig JA, Rees DC, Ribbe MW (2011) Structure of precursor-bound
NifEN: a nitrogenase FeMo cofactor maturase/insertase. Science 331(6013):91–94
71. Hu Y, Corbett MC, Fay AW et al (2006a) Nitrogenase Fe protein: A molybdate/homocitrate
insertase. Proc Natl Acad Sci U S A 103(46):17125–17130
72. Hu Y, Corbett MC, Fay AW et al (2006b) FeMo cofactor maturation on NifEN. Proc Natl
Acad Sci U S A 103(46):17119–17124
73. Yoshizawa JM, Blank MA, Fay AW et al (2009) Optimization of FeMoco maturation on
NifEN. J Am Chem Soc 131(26):9321–9325
74. Fay AW, Blank MA, Lee CC et al (2011) Spectroscopic characterization of the isolated
iron-molybdenum cofactor (FeMoco) precursor from the protein NifEN. Angew Chem Int
Ed Engl 50(34):7787–7790
75. Fay AW, Blank MA, Rebelein JG et al (2016) Assembly scaffold NifEN: a structural and
functional homolog of the nitrogenase catalytic component. Proc Natl Acad Sci U S A 113
(34):9504–9508
182
C.-C. Lee et al.