Kazuki Tanifuji

Synthetic analogues of [Fe4S4(Cys)3(His)] in hydrogenases and [Fe4S4(Cys)4] in HiPIP derived from all-ferric [Fe4S4{N(SiMe3)2}4]

The all-ferric [Fe4S4]4+ cluster [Fe4S4{N(SiMe3)2}4] 1 and its one-electron reduced form [1]- serve as convenient precursors for the synthesis of 3∶1-site differentiated [Fe4S4] clusters and high-potential iron-sulfur protein (HiPIP) model clusters. The reaction of 1 with four equivalents (equiv) of the bulky thiol HSDmp (Dmp = 2,6-(mesityl)2C6H3, mesityl = 2,4,6-Me3C6H2) followed by treatment with tetrahydrofuran (THF) resulted in the isolation of [Fe4S4(SDmp)3(THF)3] 2. Cluster 2 contains an octahedral iron atom with three THF ligands, and its Fe(S)3(O)3 coordination environment is relevant to that in the active site of substrate-bound aconitase. An analogous reaction of [1]- with four equiv of HSDmp gave [Fe4S4(SDmp)4]- 3, which models the oxidized form of HiPIP. The THF ligands in 2 can be replaced by tetramethyl-imidazole (Me4Im) to give [Fe4S4(SDmp)3(Me4Im)] 4 modeling the [Fe4S4(Cys)3(His)] cluster in hydrogenases, and its one-electron reduced form [4]- was synthesized from the reaction of 3 with Me4Im. The reversible redox couple between 3 and [3]- was observed at E1/2 = -820 mV vs. Ag/Ag+, and the corresponding reversible couple for 4 and [4]- is positively shifted by +440 mV. The cyclic voltammogram of 3 also exhibited a reversible oxidation couple, which indicates generation of the all-ferric [Fe4S4]4+ cluster, [Fe4S4(SDmp)4].

Formation of a Nitrogenase P-cluster [Fe8S7] Core via Reductive Fusion of Two All-Ferric [Fe4S4] Clusters

Iron–sulfur clusters are ubiquitous in electron-transfer proteins, and the cuboidal [Fe4S4] core is thermodynamically robust, exhibiting multiple oxidation states with only minor geometry change. Although the removal of an iron or sulfur atom from an [Fe4S4] core is chemically difficult, [2–4] the reversible transformation between [Fe4S4] and [Fe3S4] clusters has been suggested to occur under physiological conditions. Further, the nitrogenase P-cluster core [Fe8S7] (Figure 1), which plays an important role in the electron-

Combining a Nitrogenase Scaffold and a Synthetic Compound into an Artificial Enzyme

Nitrogenase catalyzes substrate reduction at its cofactor center ([(Cit)MoFe7S9C](n-); designated M-cluster). Here, we report the formation of an artificial, nitrogenase-mimicking enzyme upon insertion of a synthetic model complex ([Fe6S9(SEt)2](4-); designated Fe6(RHH)) into the catalytic component of nitrogenase (designated NifDK(apo)). Two Fe6(RHH) clusters were inserted into NifDK(apo), rendering the conformation of the resultant protein (designated NifDK(Fe)) similar to the one upon insertion of native M-clusters. NifDK(Fe) can work together with the reductase component of nitrogenase to reduce C2H2 in an ATP-dependent reaction. It can also act as an enzyme on its own in the presence of Eu(II)DTPA, displaying a strong activity in C2H2 reduction while demonstrating an ability to reduce CN(-) to C1-C3 hydrocarbons in an ATP-independent manner. The successful outcome of this work provides the proof of concept and underlying principles for continued search of novel enzymatic activities based on this approach.

Structure and Reactivity of an Asymmetric Synthetic Mimic of Nitrogenase Cofactor

The Mo nitrogenase catalyzes the ambient reduction of N2 to NH3 at its M-cluster site. A complex metallocofactor with a core composition of [MoFe7 S9 C], the M-cluster, can be extracted from the protein scaffold and used to facilitate the catalytic reduction of CN- , CO, and CO2 into hydrocarbons in the isolated state. Herein, we report the synthesis, structure, and reactivity of an asymmetric M-cluster analogue with a core composition of [MoFe5 S9 ]. This analogue, referred to as the Mo-cluster, is the first synthetic example of an M-cluster mimic with Fe and Mo positioned at opposite ends of the cluster. Moreover, the ability of the Mo-cluster to reduce C1 substrates to hydrocarbons suggests the feasibility of developing nitrogenase-based biomimetic approaches to recycle C1  waste into fuel products.

A Convenient Route to Synthetic Analogues of the Oxidized Form of High-Potential Iron–Sulfur Proteins

An amide-bound [Fe4S4](3+) cluster, [Fe4S4{N(SiMe3)2}4](-) (1), was found to serve as a convenient precursor for synthetic analogues of the oxidized form of high-potential iron-sulfur proteins. Treatment of 1 with 4 equiv of bulky thiols led to replacement of the amide ligands with thiolates, giving rise to a series of [Fe4S4(SR)4](-) clusters (R = Dmp (2a), Tbt (2b), Eind (2c), Dxp (2d), Dpp (2e); Dmp = 2,6-di(mesityl)phenyl, Tbt = 2,4,6-tris[bis(trimethylsilyl)methyl]phenyl, Eind = 1,1,3,3,5,5,7,7-octaethyl-s-hydrindacen-4-yl, Dxp = 2,6-di(m-xylyl)phenyl, Dpp = 2,6-diphenylphenyl). These clusters were characterized by the mass spectrum, the EPR spectrum, and X-ray crystallography. The redox potentials of the [Fe4S4](3+/2+) couple, -0.82 V (2a), -0.86 V (2b), -0.84 V (2c), -0.74 V (2d), and -0.63 V (2e) vs Ag/Ag(+) in THF, are significantly more negative than that of [Fe4S4(SPh)4](-/2-) (-0.21 V).

Non-centrosymmetric coordination polymer with a highly hindered octahedral copper center bridged by mandelate.

A novel chiral coordination polymer, [Cu(C(6)H(5)CH(OH)COO)(μ-C(6)H(5)CH(OH)COO)] (1-L and 1-D), was synthesized through a reaction of copper acetate with L-mandelic acid at room temperature. Although previously reported copper mandelate prepared by hydrothermal reaction was a centrosymmetric coordination polymer because of the racemization of mandelic acid, the current coordination polymer shows noncentrosymmetry and a completely different structure from that previously reported. The X-ray crystallography for 1-L revealed that the copper center of the compound showed a highly distorted octahedral structure bridged by a chiral mandelate ligand in the unusual coordination mode to construct a one-dimensional (1D) zigzag chain structure. These 1D chains interdigitated each other to give a layered structure as a result of the formation of multiple aromatic interactions and hydrogen bonds between hydroxyl and carboxylate moieties at mandelate ligands. The coordination polymer 1-L belongs to the noncentrosymmetric space group of C2 to show piezoelectric properties and second harmonic generation (SHG) activity.

Synthetic Analogues of Nitrogenase Metallocofactors: Challenges and Developments

Nitrogenase is the only known biological system capable of reducing N2 to NH3 , which is a critical component of bioavailable nitrogen fixation. Since the discovery of discrete iron-sulfur metalloclusters within the nitrogenase MoFe protein, synthetic inorganic chemists have sought to reproduce the structural features of these clusters in order to understand how they facilitate the binding, activation and hydrogenation of N2 . Through the decades following the initial identification of these clusters, significant progress has been made to synthetically replicate certain compositional and functional aspects of the biogenic clusters. Although much work remains to generate synthetic iron-sulfur clusters that can reduce N2 to NH3 , the insights borne from past and recent developments are discussed in this concept article.

Interconversion between [Fe4S4] and [Fe2S2] Clusters Bearing Amide Ligands

Structural conversion of [Fe4S4] clusters into [Fe2S2] clusters has been suggested to be a fundamental process for some O2-sensing proteins. While the formation of [Fe2S2] clusters from synthetic [Fe4S4] clusters has been unprecedented, an all-ferric [Fe4S4](4+) cluster Fe4S4{N(SiMe3)2}4 (1) was found to split in the presence of pyridines, giving [Fe2S2] clusters Fe2S2{N(SiMe3)2}2(L)2 (2, L = pyridines). The structural conversion between [Fe4S4] and [Fe2S2] clusters appeared to be reversible, and the thermodynamic parameters for the equilibrium reactions between 1 + L and 2 were determined. Assembly of two [Fe2S2] clusters was also induced by chemical reductions of Fe2S2{N(SiMe3)2}2(Py)2 (Py = pyridine), and the resultant [Fe4S4] clusters [1](-) and [1](2-) were found to split into two [Fe2S2] clusters by oxidation with [Cp2Fe](+) in the presence of pyridine.

Tracing the ‘ninth sulfur’ of the nitrogenase cofactor via a semi-synthetic approach

AbstractThe M-cluster is the [(homocitrate)MoFe7S9C] active site of nitrogenase that is derived from an 8Fe core assembled viacoupling and rearrangement of two [Fe4S4] clusters concomitant with the insertion of an interstitial carbon and a ‘ninth sulfur’. Combining synthetic [Fe4S4] clusters with an assembly protein template, here we show that sulfite can give rise to the ninth sulfur that is incorporated in the catalytically important belt region of the cofactor after the radical S-adenosyl-l-methionine-dependent carbide insertion and the concurrent 8Fe-core rearrangement have already taken place. Based on the differential reactivity of the formed cluster species, we also propose a new [Fe8S8C] cluster intermediate, the L*-cluster, which is similar to the [Fe8S9C] L-cluster, but lacks the ninth sulfur from sulfite. This work provides a semi-synthetic tool for protein reconstitution that could be widely applicable for the functional analysis of other FeS systems.The M-cluster in the active site of nitrogenase is derived from an 8Fe core assembled via coupling and rearrangement of two [Fe4S4] clusters concomitant with the insertion of an interstitial carbon and a ninth sulfur. Now, by combining synthetic [Fe4S4] clusters and assembly with a protein template, it has been shown that sulfite gives rise to the ninth sulfur that is inserted into the nitrogenase cofactor after the radical SAM-dependent carbide insertion and cofactor core rearrangement.

Ambient conversion of CO 2 to hydrocarbons by biogenic and synthetic [Fe 4 S 4 ] clusters

The Fe protein of nitrogenase contains a redox active [Fe4S4] cluster that plays a key role in electron transfer and substrate reduction. Here we show that the Fe protein of Methanosarcina acetivorans can reduce CO2 and CO to hydrocarbons under ambient conditions. Further, we demonstrate that this reactivity is inherent to [Fe4S4] clusters, showing the ability of a synthetic [Fe4S4] compound to catalyse the same ambient reaction in solutions. Theoretical calculations suggest a reaction mechanism involving an aldehyde-like intermediate that gives rise to hydrocarbon products upon proton-coupled electron transfer and concomitant removal of water molecules. These results provide a framework for mechanistic investigations of FeS-based activation and reduction of CO2 and CO while facilitating potential development of FeS catalysts capable of ambient conversion of CO2 and CO into fuel products.The Fe protein of nitrogenase contains a redox-active [Fe4S4] cluster that plays a key role in electron transfer and substrate reduction. Here, Hu and co-workers show that the Fe protein of Methanosarcina acetivorans can reduce CO2 and CO to hydrocarbons under ambient conditions.