The influences of carbon donor ligands on biomimetic multi-iron complexes for N2 reduction†

The active site clusters of nitrogenase enzymes possess the only examples of carbides in biology. These are the only biological FeS clusters that are capable of reducing N2 to NH4+, implicating the central carbon and its interaction with Fe as important in the mechanism of N2 reduction. This biological question motivates study of the influence of carbon donors on the electronic structure and reactivity of unsaturated, high-spin iron centers. Here, we present functional and structural models that test the impacts of carbon donors and sulfide donors in simpler iron compounds. We report the first example of a diiron complex that is bridged by an alkylidene and a sulfide, which serves as a high-fidelity structural and spectroscopic model of a two-iron portion of the active-site cluster (FeMoco) in the resting state of Mo-nitrogenase. The model complexes have antiferromagnetically coupled pairs of high-spin iron centers, and sulfur K-edge X-ray absorption spectroscopy shows comparable covalency of the sulfide for C and S bridged species. The sulfur-bridged compound does not interact with N2 even upon reduction, but upon removal of the sulfide it becomes capable of reducing N2 to NH4+ with the addition of protons and electrons. This provides synthetic support for sulfide extrusion in the activation of nitrogenase cofactors.


Introduction
Nitrogenases are enzymes that accomplish the impressive feat of reducing N 2 to NH 4 + at ambient temperatures and pressures.
The active site of the most thoroughly studied nitrogenase is the iron-molybdenum cofactor (FeMoco), a unique iron-sulfur cluster composed of one molybdenum and seven iron atoms held together with a number of bridging atoms (Fig. 1a). 1 A range of kinetic, mutagenesis, and spectroscopic studies support N 2 binding at the iron atoms of FeMoco, 2-8 but the structures of intermediate species in the mechanism of N 2 reduction remain unclear. 1 A distinguishing feature of the FeMoco is the central carbide (formally C 4À ), which is bound to six iron atoms in the resting state. 9,10 Isotopic labeling studies show that the carbide is not exchanged during turnover. 11 X-ray emission spectroscopy has shown that the iron-carbide bonds are highly covalent. 12 The active-site clusters of nitrogenases are the only known examples of carbides in biological systems, and they are also the only catalysts for N 2 reduction in nature, implying the carbide serves an essential role in this transformation. However, the specic role of the carbide during the catalytic cycle for N 2 reduction remains obscure. One hypothesis for N 2 binding to FeMoco proposes cleavage of hemilabile Fe-S bonds during catalysis, in which case the carbide may function to preserve structural integrity by anchoring the core structure of the cofactor. 7,8,13,14 This idea is supported by several crystallographic studies showing that the belt sulde S2B (which bridges Fe2 and Fe6) can be reversibly displaced from the cluster (Fig. 1a). 7,8,14,15 These results suggest the Fe2/Fe6 locus as a primary substrate binding site in the cofactor. Another hypothesis proposes Fe-C bond cleavage during turnover to create an open coordination site on iron. 3,16 This could be accompanied by C-H bond formation, an idea that is supported by recent work on a synthetic diiron complex containing a bridging carbyne. 17 Other proposals involve direct interactions between the carbide and N 2 . [18][19][20] The wide variety of these proposals underscores the limited understanding of the structural and electronic contributions of bridging carbon ligands to reactivity in iron-sulfur clusters.
Synthetic complexes offer useful insights as structural or functional models of nitrogenase, but accessing species with both carbon and sulfur donors has been challenging. 21 In a recent study, Rauchfuss and coworkers reported the rst synthetic example of an iron cluster with both carbide and sulde (Fig. 1b), which have different bridging modes than FeMoco. 22 In this compound, the CO ligands lead to low-spin iron centers that contrast with the high-spin iron centers found in FeMoco.
In functional synthetic models, mononuclear iron complexes have been used to gauge the N 2 -coordinating ability of compounds with Fe-C bonds. 16,17,[23][24][25][26][27][28] Of these complexes, only two systems produce NH 4 + (3.3-4.6 equiv. NH 4 + /Fe) upon treatment with acid and reductant (Fig. 1c). 16,27 A low-spin iron system, Fe(CP iPr 3 )N 2 , where CP iPr 3 is tris(2-(diisopropylphosphino)phenyl)methyl, displays lengthening of Fe-C bonds during reduction. The other, (CAAC) 2 Fe (CAAC ¼ cyclic (alkyl)(amino)carbene), is capable of mediating N 2 reduction to NH 4 + . 16,27,29 Studies of iron species with bridging carbon and sulfur ligands that are high-spin with greater electronic and structural delity to the Fe2/Fe6 site are needed to improve the understanding of how biological S-and C-based donors impact N 2 reactivity. Here, we present a new diiron complex that has both carbon and sulfur bridges between two high-spin iron centers, creating an Fe 2 CS diamond core that structurally overlays with a part of the core in FeMoco. [30][31][32] Further, we systematically evaluate the electronic structure and N 2 reducing ability of three related high-spin iron complexes with the carbon-based donors alkyl and alkylidene. Importantly, the high-spin iron alkyl and alkylidene complexes produce NH 4 + from N 2 .

Synthesis
We previously reported the rst high-spin iron complex with an unsupported alkylidene bridge, [  0.030(8) A longer than the average Fe-C bond in the diiron(II) complex 1 despite the higher oxidation state of iron. The sulde bridge causes the Fe-C-Fe bond angle to decrease from 95.6(3) in 1 to 81.74(6) in 2. The diamond core in 2 is contracted relative to the one in a previously reported bis(sulde) diiron(III) complex [L Me FeS] 2 , as in 2 the Fe-S bonds are 0.116(2) A shorter and the Fe/Fe distance is 0.641(2) A shorter. 34,35 Importantly, the core of 2 overlays well with the rhomb containing Fe2 and Fe6 in FeMoco, as the Fe-C and Fe-S average bond lengths in both structures are similar (Fe-C avg is 1.994(2) A in 2 vs. 2.00 A in FeMoco; Fe-S avg is 2.217(8) A in 2 and 2.25Å in FeMoco for the Fe2/Fe6 rhomb). 10 The Fe/Fe distance in 2 is 2.6027(6) A, which matches the distances between belt iron atoms in the crystal structure of FeMoco (2.61 A) quite well. The overall Fe/Fe/S/C core in 2 overlays with the Fe2/ Fe6/S2B/C core of the resting state FeMoco with an root-meansquare deviation (RMSD) of 0.08 A (Fig. 2b). These comparisons indicate that despite the difference in carbon coordination number, the alkylidene in 2 serves as an accurate structural model for the carbide bridge in FeMoco.
To compare the inuence of the alkylidene in 1 to a mononuclear alkyl analogue, we also prepared a threecoordinate iron(II) alkyl complex with diketiminate supporting ligands using a known method. 36,37 Adding 2.1 equiv. of MgBrCH 2 SiMe 3 to a solution of [L Me FeCl] 2 38 in THF led to the trimethylsilylmethyl iron(II) complex 3, which could be isolated in 56% yield (Scheme 2). The X-ray crystallographic data of 3 reveal a planar three-coordinate iron center featuring an Fe-C bond length of 2.017(3) A, which is comparable to the Fe-C bond lengths in other previously reported threecoordinate b-diketiminate iron(II) alkyl species. 33,36,37,39,40 Similar to these related compounds, 3 has a high-spin electronic conguration (S ¼ 2), as judged by the Evans method in solution (m eff in C 6 D 6 at 298 K is 5.4(2) m B ), and averaged C 2v symmetry evident in its 1 H NMR spectrum indicating rapid rotation around the Fe-C bond.
Reduction of N 2 to NH 4 + by C-bound iron complexes The conversion of N 2 to NH 4 + with the addition of reductant and acid was used to evaluate compounds 1-3 as functional models of nitrogenase. These studies used KC 8 27,41 In our work, the reductant (10 equiv.) was added rst to the complexes at 173 K and then the acid (10 equiv.) was added to the frozen mixture and stirred at 195 K (Scheme 3). This order of addition minimizes the potential for competitive protonation of the b-diketiminate ligand. 42 Using  Table S1 †). These control experiments demonstrate the NH 4 + formed by 1 and 3 is derived from N 2 , not from the diketiminate ligands or impurities. We also tested N 2 reduction in THF (right side of Table 1), but the NH 4 + yields were roughly ve times lower than the analogous experiments in Et 2 O. We hypothesize that this difference is a result of less favorable N 2 binding in THF (vide infra).
To explore the species responsible for NH 4 + production from 1, we conducted low temperature 1 H NMR studies with smaller amounts of reductant and acid. Reduction of 1 with 1.6 equiv. of KC 8 in THF-d 8 at 203 K showed trace amounts (<10%) of 1 and 3, but most of the mixture consisted of unidentied species that we were unable to isolate due to thermal decomposition above 203 K. Further attempts to isolate reduced forms of 1 were not successful (see ESI †). As a result, we cannot condently attribute the N 2 reduction by 1 to any one active species. However, we reason that since the conversion of 1 to 3 upon reduction is relatively low, the N 2 reduction activity of 1 cannot be solely attributed to the formation of 3 under these conditions (see ESI † for further discussion). It is therefore evident that there is Scheme 2 Preparation of L Me FeCH 2 SiMe 3 (3). a Ammonium determined using the indophenol method. Error represented as a range of multiple trials; lack of error bar indicates a single trial. some reduced form of 1 (or a degradation product therefrom) that is capable of N 2 binding and reduction to NH 4 + .

Scheme 3 Conditions for N 2 reduction experiments.
In separate reduction experiment, treatment of 3 with 1.2 equiv. of KC 8 in the presence of 1.2 equiv. of 18-crown-6 in THF formed the iron(I) complex [L Me FeCH 2 SiMe 3 ][K(18-crown-6)] (4) in 55% yield (Scheme 4). Treatment of 4 with additional reductant did not result in further chemical changes. It was possible to characterize 4 in detail, including an X-ray crystal structure that showed separated cations (in which two THF molecules are bound to K + in addition to the 18-crown-6; see ESI †) and anions (in which the iron(I) ion is three-coordinate). The THF molecules are weakly bound to the K(18-crown-6) cation, as indicated by their absence in a low-quality crystal structure in which the K(18-crown-6) + unit was coordinated to the supporting ligand. Microanalysis also indicated the absence of the THF molecules in the isolated solid.
We investigated the ability of 4 to produce NH 4 + from N 2 under conditions similar to those described above, and found that it generates NH 4 + in comparable yields to 3 (Table 1). These experiments suggest that 4 is a feasible intermediate during the series of transformations leading to N 2 reduction by 3.

N 2 binding
Cooling 1, 2, or 3 under N 2 in THF-d 8 yielded no spectroscopic changes ( Fig. S8-S10 and S16 †). In contrast, cooling an Et 2 O solution of 4 under 1 atm N 2 resulted in a color change from green to red that was monitored using electronic absorption spectroscopy (Fig. 3a). Under 1 atm N 2 , a room-temperature solution of 4 in Et 2 O displayed prominent absorption bands at 450 and 750 nm whose intensity drastically decreased upon cooling. This marked change did not occur in upon cooling a sample under Ar (Fig. S18 †), indicating that the changes come from N 2 binding. Similar changes in the absorption spectrum were observed when cooling solutions of 4 in THF or 2-methyltetrahydrofuran (MeTHF) under N 2 ( Fig. S19 Due to decomposition at concentrations suitable for electronic absorption spectroscopy, we turned to 1 H NMR spectroscopy for more reproducible quantication. Variable temperature experiments in Et 2 O-d 10 allowed us to quantify the equilibrium between 4 and 4-N 2 , which are in slow exchange on the NMR time scale. Van't Hoff plots for N 2 binding to 4 gave DH ¼ À20 AE 1 kJ mol À1 and DS ¼ À57 AE 3 J mol À1 K À1 , where the large negative entropy is characteristic of binding a gas (Fig. S12 †). A parallel experiment performed in THF-d 8 yielded the thermodynamic parameters DH ¼ À26 AE 1 kJ mol À1 and DS ¼ À93 AE 5 J mol À1 K À1 (Fig. 3b and S14 †). These parameters are similar to those for N 2 binding to the related b-diketiminate iron(I) phenyl complex and other reported iron and cobalt complexes (Table S2 †). 27,46,47 Spin states and exchange coupling The electronic structures of 1 and 2 provide valuable insights into the inuences of S and C bridges in iron-sulfur clusters. The Mössbauer spectrum 48 of 1 at 80 K displays signals with an  isomer shi of 0.62 mm s À1 , consistent with high-spin iron(II) centers. 33 Inspection of the magnetic susceptibility of 1 reveals a sharp decrease in c M T with decreasing temperature, reaching a value of 0.12 cm 3 K mol À1 at 2 K (Fig. 4), indicating that the two iron(II) centers are antiferromagnetically coupled. The value of c M T depends linearly on T up to a value of 2.17 cm 3 K mol À1 at 225 K, which is consistent with two high-spin iron(II) centers that are antiferromagnetically coupled. To quantify the magnitude of the antiferromagnetic interaction, the data were t to the Van Vleck equation according to the spin Hamiltonian: . In this Hamiltonian, D and E are the axial and transverse zero-eld splitting parameters,Ŝ 1 andŜ 2 are the spin operators, g 1 and g 2 are the isotropic g-values, and J is the magnitude of the exchange interaction. The best t to the experimental data for 1 was accomplished with an exchange constant of J ¼ À34(2) cm À1 , and was relatively insensitive to the other parameters (see ESI †). Next, we examined the spin states in the diiron(III) complex 2, which has both C and S bridges. Its zero-eld Mössbauer spectrum at 80 K displays a doublet with d ¼ 0.26 mm s À1 and |DE Q | ¼ 1.95 mm s À1 (Fig. S22 †). The isomer shi is much lower than that of 1, consistent with more oxidized iron centers. To support this assignment, we used density-functional (DFT) calculations at the B3LYP/def2-TZVP(Fe,Si,S,N,CHSiMe 3 )/def2-SVP(C,H) level to give geometry optimized structures in all six possible spin states, and these were used to predict Mössbauer spectra for each spin state using the spectroscopic validation we established for b-diketiminate complexes. 49 A brokensymmetry model with two antiferromagnetically coupled highspin iron(III) centers gave the lowest energy structure, reproduced the crystallographic structure with a RMSD of 0.24 A, and predicted Mössbauer parameters (d 1 ¼ 0.35 mm s À1 , |DE Q | 1 ¼ 1.62 mm s À1 , d 2 ¼ 0.36 mm s À1 , |DE Q | 2 ¼ 1.67 mm s À1 ) that are within error of the experimental values; the other spin state possibilities gave isomer shi values that deviated from experiment by at least 0.20 mm s À1 or gave two distinct doublets (see ESI † for details). Other four-coordinate diiron(III) sulde complexes in the literature also have high-spin electronic congurations. [50][51][52][53] The magnetic susceptibility of 2 displays a c M T value of 0.91 cm 3 K mol À1 at 225 K that drops with decreasing temperature, supporting an antiferromagnetic exchange interaction between the iron(III) centers in 2 in agreement with the calculations. We modelled the dc susceptibility data with two high-spin iron(III) sites using the spin Hamiltonian described above with an exchange constant of J ¼ À120(10) cm À1 and isotropic g ¼ 2.0 (see ESI †).
The mononuclear compounds display c M T values of 3.26 (3) and 1.93 (4) cm 3 K mol À1 at 225 K. These values are consistent with ground states of S ¼ 2 for 3 (high-spin iron(II)) and S ¼ 3/2 for 4 (high-spin iron(I)). The value of c M T decreases with decreasing temperature due to zero-eld splitting. We t the data with D values of À45.7(3) cm À1 for 3 and À14.9(2) cm À1 for 4. These large zero-eld splitting parameters are consistent with those observed in other three-coordinate iron(II) complexes. [54][55][56][57][58] The high-spin assignment is also consistent with the Mössbauer spectra of these mononuclear complexes, which display quadrupole doublets with d ¼ 0.43 mm s À1 and |DE Q | ¼ 1.28 mm s À1 for 3, and d ¼ 0.41 mm s À1 and |DE Q | ¼ 2.23 mm s À1 for 4 ( Fig. S23 and S24 †). These values are consistent with other high-spin iron(II) and iron(I) complexes in the literature, and DFT calculations similar to those described above validated these spin state assignments (see ESI †). 55,[59][60][61] We were interested in the spin state of 4-N 2 , but our inability to isolate it prevented characterization by magnetometry. However, the Mössbauer spectrum of 4 ash frozen in N 2saturated MeTHF was collected at 80 K and displays an additional doublet that was not observed in a control experiment under Ar. We attribute this new signal to 4-N 2 (Fig. 5a), for which the best t has d ¼ 0.64 mm s À1 and |DE Q | ¼ 2.55 mm s À1 . The isomer shi is consistent with reported values of highspin, four-coordinate iron(I) (S ¼ 3/2) yet too high for reported values for iron(I) in low spin congurations (S ¼ 1/2). 55,59-61 Geometry optimizations were performed for both possible spin states of iron(I) in 4-N 2 , and DFT calculations 49 were used to predict Mössbauer parameters for each model. The S ¼ 3/2 model predicted parameters (d ¼ 0.64 mm s À1 , |DE Q | ¼ 2.14 mm s À1 ) that are close to the experimental values, while the S ¼ 1/2 model (d ¼ 0.36 mm s À1 , |DE Q | ¼ 1.02 mm s À1 ) deviated substantially from the experimental data.
To corroborate our assignment of spin states for 4 and 4-N 2 as high-spin iron(I), we measured the X-band EPR spectrum of 4 frozen in MeTHF under N 2 and Ar at 77 K (Fig. 5b). The EPR spectrum under Ar displays three broad resonances with g eff values of 2.1, 3.8 and 5.6 and are consistent with a S ¼ 3/2 ground state. We simulated the EPR spectrum with the spin Hamiltonian employed to model the dc susceptibility data with |D| and |E| values of 12.9 cm À1 and 1.7 cm À1 , and g values of 2.36, 2.33, and 2.05. The EPR spectrum for a sample of 4 ash-frozen under N 2 displays an additional feature at g eff ¼ 5.4 that we attribute to 4-N 2 . The large effective g-value indicates that 4-N 2 has a high-spin conguration (S ¼ 3/2); however, due to the spectral convolution between 4 and 4-N 2 , we could not adequately model the EPR spectrum to extract the spin Hamiltonian parameters for 4-N 2 . The agreement with the DFT computations and the Mössbauer spectroscopy, however, supports this assignment of the iron(I) center as S ¼ 3/2 in 4-N 2 .

Covalency of bonds
To assess the effect of replacing a bridging sulde with an alkylidene on the Fe-S covalency, we compared sulfur K-edge Xray absorption spectra (XAS) for 2 to the related bis-sulde complex, [L Me FeS] 2 (Fig. 6a), which also has two high-spin iron(III) centers, but these centers are instead bridged by two sulde ligands. 34 We examined the pre-edge areas at 2470 eV determined by peak tting of the S K-edges, in order to quantify the S 3p character in the unoccupied metal d orbitals. 62 The contribution from the two bridging sulde ligands in [L Me FeS] 2 is 14% S 3p and the contribution from the single bridging sulde ligand in 2 is 6%. Because the 3p character in [L Me FeS] 2 is twice the value for 2 and reects the contributions from two suldes instead of one, it follows that the Fe-S covalency per bond is not signicantly perturbed by the substitution of an alkylidene for one of the sulde ligands. This interpretation of the XAS agrees with the similar isomer shis observed for 2 and [L Me FeS] 2 , which reect the electron density at the iron centers (Fig. S17 †).
Broken symmetry DFT calculations indicated strong antiferromagnetic coupling between iron centers in both [L Me FeS] 2  and 2 (Fig. 7). The overlap (J) was À84 cm À1 for [L Me FeS] 2 and À313 cm À1 for 2, but these values from a single reference DFT calculation relative to experiment are commonly overestimated. [63][64][65][66] Calculated spectra (Fig. S41-43 †) were thus obtained using the lowest energy solution with S ¼ 0. It should be noted that the differences in intensities of the calculated S Kedges can be attributed to the difference in number of absorbers. Calculated S 3p character in the unoccupied dorbitals was 11.94% for [L Me FeS] 2 and 5.94% for 2. These calculations agree remarkably well with the experimental data described in the previous paragraph, and conrm that the Fe-S covalency is the same for both compounds. These results contrast with those presented in a study by Pollock et al., in which the replacement of a sulde with an imido (N t Bu 2À ) in [Fe 2 S 2 Cl 4 ] 2À decreased the iron-sulfur covalency. 50 The Fe K-edge XAS obtained for 1, 2 and [L Me FeS] 2 are presented in Fig. 6b. These data reveal a signicant shi in the rising edges between 1 (7115 eV) and 2 (7118 eV), as expected for more oxidized iron sites. The overlaying pre-edge and edge features of 2 and [L Me FeS] 2 in Fe K-edge XAS also reect similar electronic structures at the iron centers, in agreement with the S K-edge XAS data described above, despite the substitution of the sulde for an alkylidene.

Electronic structure
To further understand the nature of the bonding in these complexes, we analyzed the localized orbitals of the brokensymmetry DFT model using the intrinsic atomic orbitalintrinsic bond order (IAOIBO) method. 67 From this analysis, the Mayer bond order 68 (MBO) provides a convenient method to sum all of the contributions to the bond; it has been applied to FeMoco and related systems to understand the magnitude of bonding between two atoms. 68,69 The MBOs for the Fe-C interactions are similar between 3 (0.87) and 1 (0.90). Upon the introduction of a sulde ligand and oxidation of the iron centers, the Fe-C interactions of the bridging alkylidene in 2 display a similar MBO of 0.86. This MBO analysis demonstrates the similarity of Fe-C bonding across 1, 2, and 3, despite the differences in metal nuclearity, oxidation state, and carbon donor identity.
Despite the comparable Fe-C MBO in 1 and 2, we sought to understand the electronic implications of the Fe-C-Fe angle contraction from 95.6(3) in 1 to 81.74(6) in 2. There is also a substantial distortion of the alkylidene carbon geometry away from tetrahedral moving from 1 (s 4 ¼ 0.85) to 2 (s 4 ¼ 0.65, see ESI †). 70 Analysis of the localized Fe-C orbitals shows that in 1 the electron density in each orbital is more localized on one discrete Fe-C bond, while in 2 there is a delocalized bonding orbital with electron density between both iron centers and the alkylidene carbon (Fig. 8). The IAOIBO analysis of 2 shows the alkylidene exclusively forms s bonds with the iron centers, while the sulde forms one s bond with each iron as well as pbonding interactions. The covalency of the Fe-S bonds can be measured by summing the Fe-S bonding orbitals in 2, giving 23% Fe character and 77% S character. In the bis-sulde complex [L Me FeS] 2 these are similar (24% Fe, 76% S), consistent with the S K-edge XAS data described above that show similar Fe-S covalency for the compounds. The MBOs of the Fe-S bonds are also identical (1.1) between the two compounds, despite the 0.116 (2)     underscore the electronic similarities between the model complex and the biological cluster site.
In contrast to the copious literature study of Fe-S bonding in FeS clusters, 71-74 the Fe-C bonding interactions in high-spin complexes remain poorly understood. The incorporation of both a bridging alkylidene and sulde into 2 allows us to assess the relative covalency of Fe-S and Fe-C bonds. The IAOIBO analysis reveals the average Fe-C electron distribution in the Fe-C bonds of 2 to be 43% Fe and 57% C in character, suggesting that the Fe-C bonds are more covalent than the Fe-S bonds (Fig. 9). Though the limited number of complexes in our studies prevented us from clearly distinguishing the oxidationstate dependence of covalency of Fe-C bonds, the diiron(II) alkylidene complex 1 reveals a similar total Fe/C electron distribution of 40% Fe and 60% C, which is consistent with the comparable Fe-C MBO values in 1 and 2. This orbital analysis provides insight into Fe-C bonding in high-spin iron complexes relevant to FeMoco.

Discussion
The diiron alkylidene sulde complex 2 presented here incorporates structural elements relevant to nitrogenase, and the Fe/Fe/ S2B/C diamond core overlays extremely well with the Fe2/Fe6/S/C rhomb in the resting state of FeMoco (Fig. 2b). Enzymatic studies implicate the Fe2 and Fe6 centers as a primary substrate binding site, 7,8,14,15,75 and therefore it is particularly important that the core of our synthetic model structurally resembles these centers.
In addition to the local structural similarity, the electronic structure of 2 has signicant similarities to this site in FeMoco. First, the oxidation states are identical, based on the diiron(III) assignment for the Fe2/Fe6 sites in FeMoco based on SpReAD analysis. 30 Se Ka-HERFD XAS studies on FeMoco with selective substitution of S2B with Se 31 assign these iron centers as an antiferromagnetically-coupled diferric pair, which is consistent with QM/MM studies. 32 In 2, the iron sites have high-spin electronic congurations and display antiferromagnetic coupling between the iron centers, which agrees with calculations indicating that the belt irons Fe2 and Fe6 in the resting state of FeMoco are antiferromagnetically coupled pairs. [30][31][32] The comparison of XAS data between 2 and the analogous doubly sulde-bridged complex [L Me Fe(m-S)] 2 enables us to evaluate the inuence of carbon donors within iron-sulfur clusters. This is signicant because there are few other examples of highspin iron-sulfur clusters with any carbon-based ligands. 46,76,77 The sulfur pre-edge intensities from the S K-edge XAS data indicate that the marker sulde does not change its covalency from the addition of the carbon-based bridge. The similarity in Mössbauer isomer shis of the iron(III) sites in 2 and its bis-sulde analogue further emphasizes the similarity of C and S bridges in terms of their inuences on the iron centers. Within 2, though, the Fe-C bonds are more covalent than the Fe-S bonds and form exclusively s-interactions in contrast to the p interactions contributed by suldes (see ESI, Fig. S33-S35 †). The contracted Fe-C-Fe angle in 2 gives a delocalized orbital that stretches over both iron atoms and the carbon atom (Fig. 8 above), contributing to the greater antiferromagnetic exchange coupling observed in 2. An analogous superexchange interaction between the iron centers facilitated by the carbon bridge could help to rationalize the different coupling present in FeMoco compared to other FeS clusters. 12,78,79 Naturally, there are differences between the carbon bridges in these synthetic complexes compared to FeMoco as well. For example, the average Fe-C MBO in complexes 1 and 2 (0.9) are higher than the average Fe-C MBO in FeMoco (0.32). 69 The variation in the Fe-C MBO between complexes 1 and 2 and the FeMoco may be attributed to the coordination of the carbide to six iron centers (m 6 ) rather than the two iron centers in 1 and 2 (m 2 ).
The ability of the complexes to reduce N 2 was assessed by adding KC 8  . We ascribe the difference to the open coordination sites in 1 and 3, while the iron centers in 2 are four-coordinate. The differences in oxidation state are less likely to be important because our N 2 reduction experiments used a 10-fold excess of reductant relative to the complex. The necessity for a coordinatively unsaturated iron center in these N 2 reduction studies parallels N 2 reduction in nitrogenase, as enzymatic studies imply S2B dissociation upon binding substrates. 7,8,14 It has been hypothesized that dissociation of S2B in the initial stages of FeMoco reduction could be the "trigger" that brings about N 2 binding and reduction, and in this context we see that the formal loss of an S atom moving from 2 to 1 causes a change from very little N 2 reduction activity (2) to signicant N 2 conversion to NH 4 + (1). Thus, these synthetic complexes support the importance of sulfur dissociation for bringing about N 2 reducing ability in iron-carbon-sulfur clusters. It is informative to compare the reactivity of 1 and 3 with other diketiminate-iron complexes that lack C and S ligands. We previously reported that addition of four or more equivalents of reductant to [L Me Fe(m-Cl)] 2 under N 2 forms complexes M 2 [L Me Fe(m-N 2 )] 3 (M ¼ K, Rb, Cs). 82 These clusters, which lack C and S ligands, possess iron centers in lower oxidation states (Fe 0 2 Fe 1+ ) yet they do not form measurable amounts of NH 4 + upon the addition of acid. Similarly, previously reported diketiminate-supported iron complexes containing bridging FeNNFe cores do not react with acid to give NH 4 + . [83][84][85] In contrast, we see here that the incorporation of C-based ligands and an open coordination site leads to the ability to reduce N 2 to NH 4 + . Though we were unable to deconvolute the inuence of the nuclearity and the carbon ligand identity on N 2 reduction ability in these studies, it appears that the presence of an Fe-C bond is benecial for N 2 reduction. We note that two previous carbon-ligated iron systems from Peters and coworkers yield NH 4 + (3.3-4.6 equiv. NH 4 + /Fe) as well. 16,27 Though we isolated the carbon-ligated low-valent species relevant to the N 2 reduction studies from 3, the mechanism of N 2 reduction by the diiron alkylidene complex 1 remains unclear. However, a recent article by Agapie and co-workers described a diiron m-alkylidyne m-hydride complex with a Fe/ Fe/H/C core, which undergoes Fe-C bond cleavage and C-H bond formation to give various products including iron(II) alkyl and alkylidene species with N 2 bound. 17 These may bear resemblance to the intermediates during N 2 reduction by 1. This gains signicance because the Fe/Fe/H/C diamond core in the alkylidyne complex is in a more reduced state (closer to the level of N 2 -binding FeMoco intermediates) than the diiron alkylidene sulde 2. However, the mechanisms may be different since the strong-eld phosphine ligand sphere in the alkylidyne complex is less electronically similar to the FeMoco than the weak-eld ligand sphere present in 1, 2, and 3 that gives rise to high-spin iron centers.

Conclusions
This paper has presented a series of complexes that helps to understand the inuence of Fe-C bonding on the electronic structure and N 2 reactivity of high-spin iron sites like those in the FeMoco of nitrogenase. The mononuclear iron(II) alkyl 3 and the diiron(II) alkylidene 1 can reduce N 2 to NH 4 + upon addition of acid and reductant, suggesting that Fe-C bonds are benecial for N 2 reduction. Importantly, the lack of a sulfur bridge is essential for N 2 reduction activity, supporting the idea that sulfur dissociation is a reasonable step toward N 2 binding in FeMoco.
The structural relevance of 1 to the resting state of FeMoco was extended by incorporating a sulde ligand to give 2, which has both a carbon donor and a sulde as bridges. Complex 2 has nearly identical metrical parameters as the Fe2/Fe6/S2B/C rhomb within the FeMoco resting state structure, and the antiferromagnetic coupling of the iron(III) centers in 2 aligns well with the analogous coupling of Fe2 and Fe6 in FeMoco. In both dinuclear complexes 1 and 2, the bridging ligands facilitate electronic communication between the iron centers, giving rise to antiferromagnetic coupling. The addition of a bridging sulde ligand in 2 enhances the antiferromagnetic coupling interaction. The IAOIBO picture of the Fe-C interactions in 1 and 2 depicts highly covalent Fe-C bonds which can mediate superexchange, suggesting that the presence of a carbon ligand may contribute to the different exchange interactions observed in FeMoco compared to other FeS clusters.
The study of the bonding interactions in these simplied structural models also shows a surprisingly strong similarity between Fe-C bonds and Fe-S bonds. First, S K-edge XAS experiments show that the substitution of an alkylidene for a bridging sulde minimally inuences the Fe-S covalency in the other bridge. Further, the similar Fe-C Mayer bond orders in 1-3 are similar despite differences in the identity of the carbon ligand, the iron oxidation state, and the complex nuclearity. This property of the Fe-C bonds is reminiscent of the well-known similarity of Fe-S interactions in different oxidation states of iron-sulfur clusters. 86 The insights provided by these model complexes improve our understanding of the Fe-C interactions at high spin iron sites, which helps rene our hypotheses about the structural and electronic implications of the carbide in FeMoco, and the key determinants of N 2 reduction by the enzyme.

Conflicts of interest
There are no conicts to declare.