Correlating Structure with Spectroscopy in Ascorbate Peroxidase Compound II.

Structural and spectroscopic investigations of compound II in ascorbate peroxidase (APX) have yielded conflicting conclusions regarding the protonation state of the crucial Fe(IV) intermediate. Neutron diffraction and crystallographic data support an iron(IV)-hydroxo formulation, whereas Mössbauer, X-ray absorption (XAS), and nuclear resonance vibrational spectroscopy (NRVS) studies appear consistent with an iron(IV)-oxo species. Here we examine APX with spectroscopy-oriented QM/MM calculations and extensive exploration of the conformational space for both possible formulations of compound II. We establish that irrespective of variations in the orientation of a vicinal arginine residue and potential reorganization of proximal water molecules and hydrogen bonding, the Fe-O distances for the oxo and hydroxo forms consistently fall within distinct, narrow, and nonoverlapping ranges. The accuracy of geometric parameters is validated by coupled-cluster calculations with the domain-based local pair natural orbital approach, DLPNO-CCSD(T). QM/MM calculations of spectroscopic properties are conducted for all structural variants, encompassing Mössbauer, optical, X-ray absorption, and X-ray emission spectroscopies and NRVS. All spectroscopic observations can be assigned uniquely to an Fe(IV)═O form. A terminal hydroxy group cannot be reconciled with the spectroscopic data. Under no conditions can the Fe(IV)═O distance be sufficiently elongated to approach the crystallographically reported Fe-O distance. The latter is consistent only with a hydroxo species, either Fe(IV) or Fe(III). Our findings strongly support the Fe(IV)═O formulation of APX-II and highlight unresolved discrepancies in the nature of samples used across different experimental studies.

Understanding Complex Interplay among Different Instabilities in Multiferroic BiMn7O12 Using 57Fe Probe Mössbauer Spectroscopy.

Here, we report the results of a Mössbauer study on hyperfine electrical and magnetic interactions in quadruple perovskite BiMn7O12 doped with 57Fe probes. Measurements were performed in the temperature range of 10 K < T < 670 K, wherein BiMn6.9657Fe0.04O12 undergoes a cascade of structural (T1 ≈ 590 K, T2 ≈ 442 K, and T3 ≈ 240 K) and magnetic (TN1 ≈ 57 K, TN2 ≈ 50 K, and TN3 ≈ 24 K) phase transitions. The analysis of the electric field gradient (EFG) parameters, including the dipole contribution from Bi3+ ions, confirmed the presence of the local dipole moments pBi, which are randomly oriented in the paraelectric cubic phase (T > T1). The unusual behavior of the parameters of hyperfine interactions between T1 and T2 was attributed to the dynamic Jahn-Teller effect that leads to the softening of the orbital mode of Mn3+ ions. The parameters of the hyperfine interactions of 57Fe in the phases with non-zero spontaneous electrical polarization (Ps), including the P1 ↔ Im transition at T3, were analyzed. On the basis of the structural data and the quadrupole splitting Δ(T) derived from the 57Fe Mössbauer spectra, the algorithm, based on the Born effective charge model, is proposed to describe Ps(T) dependence. The Ps(T) dependence around the Im ↔ I2/m phase transition at T2 is analyzed using the effective field approach. Possible reasons for the complex relaxation behavior of the spectra in the magnetically ordered states (T < TN1) are also discussed.

Characterizing the Biosynthesis of the [Fe(II)(CN)(CO)2(cysteinate)]- Organometallic Product of the Radical-SAM Enzyme HydG by EPR and Mössbauer Spectroscopy.

[FeFe]-hydrogenases employ a catalytic H-cluster, consisting of a [4Fe-4S]H cluster linked to a [2Fe]H subcluster with CO, CN- ligands, and an azadithiolate bridge, which mediates the rapid redox interconversion of H+ and H2. In the biosynthesis of this H-cluster active site, the radical S-adenosyl-l-methionine (radical SAM, RS) enzyme HydG plays the crucial role of generating an organometallic [Fe(II)(CN)(CO)2(cysteinate)]- product that is en route to forming the H-cluster. Here, we report direct observation of this diamagnetic organometallic Fe(II) complex through Mössbauer spectroscopy, revealing an isomer shift of δ = 0.10 mm s-1 and quadrupole splitting of ΔEQ = 0.66 mm s-1. These Mössbauer values are a change from the starting values of δ = 1.15 mm s-1 and ΔEQ = 3.23 mm s-1 for the ferrous "dangler" Fe in HydG. These values of the observed product complex B are in good agreement with Mössbauer parameters for the low-spin Fe2+ ions in synthetic analogues, such as 57Fe Syn-B, which we report here. These results highlight the essential role that HydG plays in converting a resting-state high-spin Fe(II) to a low-spin organometallic Fe(II) product that can be transferred to the downstream maturase enzymes.

A New Approach for Investigating Iron Mineral Transformations in Soils and Sediments Using 57Fe-Labeled Minerals and 57Fe Mössbauer Spectroscopy.

Iron minerals in soils and sediments play important roles in many biogeochemical processes and therefore influence the cycling of major and trace elements and the fate of pollutants in the environment. However, the kinetics and pathways of Fe mineral recrystallization and transformation processes under environmentally relevant conditions are still elusive. Here, we present a novel approach enabling us to follow the transformations of Fe minerals added to soils or sediments in close spatial association with complex solid matrices including other minerals, organic matter, and microorganisms. Minerals enriched with the stable isotope 57Fe are mixed with soil or sediment, and changes in Fe speciation are subsequently studied by 57Fe Mössbauer spectroscopy, which exclusively detects 57Fe. In this study, 57Fe-labeled ferrihydrite was synthesized, mixed with four soils differing in chemical and physical properties, and incubated for 12+ weeks under anoxic conditions. Our results reveal that the formation of crystalline Fe(III)(oxyhydr)oxides such as lepidocrocite and goethite was strongly suppressed, and instead formation of a green rust-like phase was observed in all soils. These results contrast those from Fe(II)-catalyzed ferrihydrite transformation experiments, where formation of lepidocrocite, goethite, and/or magnetite often occurs. The presented approach allows control over the composition and crystallinity of the initial Fe mineral, and it can be easily adapted to other experimental setups or Fe minerals. It thus offers great potential for future investigations of Fe mineral transformations in situ under environmentally relevant conditions, in both the laboratory and the field.

Labile Iron Pool of Isolated Escherichia coli Cytosol Likely Includes Fe-ATP and Fe-Citrate but not Fe-Glutathione or Aqueous Fe.

The existence of labile iron pools (LFePs) in biological systems has been recognized for decades, but their chemical composition remains uncertain. Here, the LFeP in cytosol from Escherichia coli was investigated. Mössbauer spectra of whole vs lysed cells indicated significant degradation of iron-sulfur clusters (ISCs), even using an unusually gentle lysis procedure; this demonstrated the fragility of ISCs. Moreover, the released iron contributed to the non-heme high-spin Fe(II) species in the cell, which likely included the LFeP. Cytosol batches isolated from cells grown with different levels of iron supplementation were passed through a 3 kDa cutoff membrane, and resulting flow-through-solutions (FTSs) were subjected to SEC-ICP-MS. Mössbauer spectroscopy was used to evaluate the oxidation states of standards. FTSs exhibited iron-detected peaks likely due to different forms of Fe-citrate and Fe-nucleotide triphosphate complexes. Fe-Glutathione (GSH) complexes were not detected using physiological concentrations of GSH mixed with either Fe(II) or Fe(III); Fe(II)-GSH was concluded not to be a significant component of the LFeP in E. coli under physiological conditions. Aqueous iron was also not present in significant concentrations in isolated cytosol and is unlikely a major component of the pool. Fe appeared to bind ATP more tightly than citrate, but ATP also hydrolyzed on the timescale of tens of hours. Isolated cytosol contained excess ligands that coordinated the added Fe(II) and Fe(III). The LFeP in healthy metabolically active cells is undoubtedly dominated by the Fe(II) state, but the LFeP is redox-active such that a fraction might be present as stable and soluble Fe(III) complexes especially under oxidatively stressed cellular conditions.

Low-temperature Mössbauer spectroscopy of organs from 57Fe-enriched HFE(-/-) hemochromatosis mice: an iron-dependent threshold for generating hemosiderin.

Hereditary hemochromatosis is an iron-overload disease most often arising from a mutation in the Homeostatic Fe regulator (HFE) gene. HFE organs become overloaded with iron which causes damage. Iron-overload is commonly detected by NMR imaging, but the spectroscopic technique is insensitive to diamagnetic iron. Here, we used Mössbauer spectroscopy to examine the iron content of liver, spleen, kidney, heart, and brain of 57Fe-enriched HFE(-/-) mice of ages 3-52 wk. Overall, the iron contents of all investigated HFE organs were similar to the same healthy organ but from an older mouse. Livers and spleens were majorly overloaded, followed by kidneys. Excess iron was generally present as ferritin. Iron-sulfur clusters and low-spin FeII hemes (combined into the central quadrupole doublet) and nonheme high-spin FeII species were also observed. Spectra of young and middle-aged HFE kidneys were dominated by the central quadrupole doublet and were largely devoid of ferritin. Collecting and comparing spectra at 5 and 60 K allowed the presence of hemosiderin, a decomposition product of ferritin, to be quantified, and it also allowed the diamagnetic central doublet to be distinguished from ferritin. Hemosiderin was observed in spleens and livers from HFE mice, and in spleens from controls, but only when iron concentrations exceeded 2-3 mM. Even in those cases, hemosiderin represented only 10-20% of the iron in the sample. NMR imaging can identify iron-overload under non-invasive room-temperature conditions, but Mössbauer spectroscopy of 57Fe-enriched mice can detect all forms of iron and perhaps allow the process of iron-overloading to be probed in greater detail.

Electronic Configurations and the Effect of Peripheral Substituents of (Nitrosyl)iron Corroles.

There have been debates on the electronic configurations of (nitrosyl)iron corroles for decades. In this work, pentacoordinate [Fe(TPC)(NO)], [Fe(TTC)(NO)], and [Fe(TpFC)(NO)] with different para-substituted phenyl groups (TPC, TTC, and TpFC = tris(phenyl, 4-tolyl, or 4-fluorophenyl)corrole, respectively) have been isolated and investigated by various techniques including single-crystal X-ray diffraction, UV-vis spectroscopy, cyclic voltammetry, Fourier transform infrared, NMR, and absorption fine structure spectroscopy. Multitemperature and high-magnetic-field (3, 6, and 9 T) Mössbauer spectroscopy was also applied on all three complexes, which determined the S = 0 diamagnetic states, consistent with the magnetic susceptibility and electron paramagnetic resonance measurements. Density functional theory predictions by different functionals were compared, and the new calculation strategy, which gave remarkable agreement of the experimental Mössbauer parameters (ΔEQ and δ), allowed further assignment on the electronic configuration of {FeNO}6-(corrole3-) with antiferromagnetically coupled (S = 1/2, FeIII) and (S = 1/2, NO). Correlated sequences between the electronic donating/withdrawing capability of para substituents and the reduction/oxidation potentials, metal out-of-plane displacements (Δ4 and Δ23), and Mössbauer parameters (Vzz and ΔEQ) were also established, which suggests the strong effects of peripheral substituents.

Mössbauer-based molecular-level decomposition of the Saccharomyces cerevisiae ironome, and preliminary characterization of isolated nuclei.

One hundred proteins in Saccharomyces cerevisiae are known to contain iron. These proteins are found mainly in mitochondria, cytosol, nuclei, endoplasmic reticula, and vacuoles. Cells also contain non-proteinaceous low-molecular-mass labile iron pools (LFePs). How each molecular iron species interacts on the cellular or systems' level is underdeveloped as doing so would require considering the entire iron content of the cell-the ironome. In this paper, Mössbauer (MB) spectroscopy was used to probe the ironome of yeast. MB spectra of whole cells and isolated organelles were predicted by summing the spectral contribution of each iron-containing species in the cell. Simulations required input from published proteomics and microscopy data, as well as from previous spectroscopic and redox characterization of individual iron-containing proteins. Composite simulations were compared to experimentally determined spectra. Simulated MB spectra of non-proteinaceous iron pools in the cell were assumed to account for major differences between simulated and experimental spectra of whole cells and isolated mitochondria and vacuoles. Nuclei were predicted to contain ∼30 µM iron, mostly in the form of [Fe4S4] clusters. This was experimentally confirmed by isolating nuclei from 57Fe-enriched cells and obtaining the first MB spectra of the organelle. This study provides the first semi-quantitative estimate of all concentrations of iron-containing proteins and non-proteinaceous species in yeast, as well as a novel approach to spectroscopically characterizing LFePs.

Exotic FeII/FeIII Local Environments in the Hexagonal Channels of Hydroxyapatite.

In this fundamental solid-state chemistry study, two sample series were investigated in depth: iron(III)-doped hydroxyapatite (HA) compounds obtained from a co-sintering process of hematite and pure HA under air and iron(III)-doped HA compounds obtained from a co-sintering process from iron(II) acetate and pure HA under an argon atmosphere. X-ray diffraction, UV-visible, Fourier transform infrared, 1H and 31P NMR, electron paramagnetic resonance (EPR,) and Mössbauer spectroscopy methods were coupled to unravel the Fe valence states, the interactions with other anionic species (OH- and PO43-), and finally the complex local environments in hexagonal channels in both the series. In particular, we highlighted the associated mechanism to ensure electroneutrality with a focus on deprotonation versus calcium substitution. By diverging mechanisms, Fe3+ and Fe2+ ions were found to be located in different coordinated sites: 4(+1) coordinated site for Fe3+ and 2(+3) coordinated site for Fe2+ and clearly associated with very different Mössbauer and EPR signatures as various absorption bands (leading to different sample colors).

Mössbauer studies of the ferryl, ferrous and ferric states of dehaloperoxidase from A. ornata.

Dehaloperoxidase (DHP) is a multi-functional catalytic globin from the marine worm A. ornata, whose physiological functions include oxygen transport and oxidation of toxic substrates present in its habitat. In the Fe(III) state, DHPA has an isomer shift of 0.42 mm/s, characteristic for high-spin heme proteins. Changes in pH have subtle effects on the electronic structure of DHP in the Fe(III) state detectable in the high-field spectra, which show a pH-dependent mixture of species with different zero-field splittings between 5 and 18 cm-1. The short-lived intermediate obtained by direct reaction of the Fe(III) enzyme with H2O2 has an isomer shift of 0.10 mm/s, indicative of an Fe(IV)-oxo state and of an S = 1 electronic ground state confirmed by variable field studies. The O2-bound state of DHP has an isomer shift of 0.28 mm/s and a high-field spectrum characteristic for diamagnetic heme complexes, similarly to other haemoglobins. Overall, the isomer shift and quadrupole splitting of DHP in the four states studied are expectedly similar to both peroxidases and to myoglobin. The differences in electronic structure between DHP and other heme proteins and enzyme are observed in the high-field Mössbauer spectra of the ferric state, which show pH-dependent zero-field splittings suggesting a heme site in which the ligand field strength at the iron ion is tuned by pH. This tunability is correlated with variable electron-donating properties of the iron, which can perform multiple functions.

Yeast cells depleted of the frataxin homolog Yfh1 redistribute cellular iron: Studies using Mössbauer spectroscopy and mathematical modeling.

The neurodegenerative disease Friedreich's ataxia arises from a deficiency of frataxin, a protein that promotes iron-sulfur cluster (ISC) assembly in mitochondria. Here, primarily using Mössbauer spectroscopy, we investigated the iron content of a yeast strain in which expression of yeast frataxin homolog 1 (Yfh1), oxygenation conditions, iron concentrations, and metabolic modes were varied. We found that aerobic fermenting Yfh1-depleted cells grew slowly and accumulated FeIII nanoparticles, unlike WT cells. Under hypoxic conditions, the same mutant cells grew at rates similar to WT cells, had similar iron content, and were dominated by FeII rather than FeIII nanoparticles. Furthermore, mitochondria from mutant hypoxic cells contained approximately the same levels of ISCs as WT cells, confirming that Yfh1 is not required for ISC assembly. These cells also did not accumulate excessive iron, indicating that iron accumulation into yfh1-deficient mitochondria is stimulated by O2. In addition, in aerobic WT cells, we found that vacuoles stored FeIII, whereas under hypoxic fermenting conditions, vacuolar iron was reduced to FeII. Under respiring conditions, vacuoles of Yfh1-deficient cells contained FeIII, and nanoparticles accumulated only under aerobic conditions. Taken together, these results informed a mathematical model of iron trafficking and regulation in cells that could semiquantitatively simulate the Yfh1-deficiency phenotype. Simulations suggested partially independent regulation in which cellular iron import is regulated by ISC activity in mitochondria, mitochondrial iron import is regulated by a mitochondrial FeII pool, and vacuolar iron import is regulated by cytosolic FeII and mitochondrial ISC activity.

A New Look at Molecular and Electronic Structure of Homoleptic Diiron(II,II) Complexes with N,N-Bidentate Ligands: Combined Experimental and Theoretical Study.

Paddlewheel-type binuclear complexes featuring metal-metal bonding have been the subject of widespread interest due to fundamental concern in their electronic structures and potential applications. Here, we explore the molecular and electronic structures of diiron(II,II) complexes with N,N'-diarylformamidinate ligands. While a paddlewheel-type diiron(II,II) complex with N,N'-diphenylformamidinate ligands (DPhF) exhibits the centrosymmetric [Fe2 (µ-DPhF)4 ] structure, a minor alteration in the ligand system, i. e., switching from phenyl to p-tolyl N-substituted formamidinate ligand (DTolF), resulted in the isolation of an unprecedented non-centrosymmetric [Fe(µ-DTolF)3 Fe(κ2 -DTolF)] complex. Both complexes were characterized using single-crystal X-ray diffraction, magnetic measurements, 57 Fe Mössbauer spectroscopy, and cyclic voltammetry along with high-level ab-initio calculations. The results provide a new view on a range of factors controlling the ground-state electronic configuration and structural diversity of homoleptic diiron(II,II) complexes. Model calculations determined that the Mayer bond orders for Fe-Fe interactions are significantly lower than 1 and equal to 0.15 and 0.28 for [Fe2 (µ-DPhF)4 ] and [Fe(µ-DTolF)3 Fe(κ2 -DTolF)], respectively.

Stabilization of intermediate spin states in mixed-valent diiron dichalcogenide complexes.

The electronic structure and ground spin states, S, observed for mixed-valent iron-sulfur dimers (FeII-FeIII) are typically determined by the Heisenberg exchange interaction, J, that couples the magnetic interaction of the two metal centres either ferromagnetically (J > 0, S = 9/2) or antiferromagnetically (J < 0, S = 1/2). In the case of antiferromagnetically coupled iron centres, stabilization of the high-spin S = 9/2 ground state is also feasible through a Heisenberg double-exchange interaction, B, which lifts the degeneracy of the Heisenberg spin states. This theorem also predicts intermediate spin states for mixed-valent dimers, but those have so far remained elusive. Herein, we describe the structural, electron paramagnetic resonance and Mössbauer spectroscopic, and magnetic characterization of a series of mixed-valent complexes featuring [Fe2Q2]+ (Q = S2-, Se2-, Te2-), where the Se and Te complexes favour S = 3/2 spin states. The incorporation of heavier chalcogenides in this series reveals a delicate balance of antiferromagnetic coupling, Heisenberg double-exchange and vibronic coupling.

Versatile Fe-Sn Bonding Interactions in a Metallostannylene System: Multiple Bonding and C-H Bond Activation.

The metallostannylene Cp*(iPr2MeP)(H)2Fe-SnDMP (1; Cp* = η5-C5Me5; DMP = 2,6-dimesitylphenyl), formed by hydrogen migration in a putative Cp*(iPr2MeP)HFe[Sn(H)DMP] intermediate, serves as a robust platform for exploration of transition-metal main-group element bonding and reactivity. Upon one-electron oxidation, 1 expels H2 to generate the coordinatively unsaturated [Cp*(iPr2MeP)Fe═SnDMP][B(C6F5)4] (3), which possesses a highly polarized Fe-Sn multiple bond that involves interaction of the tin lone pair with iron. Evidence from EPR and 57Fe Mössbauer spectroscopy, along with DFT studies, shows that 3 is primarily an iron-based radical with charge localization at tin. Upon reduction of 3, C-H bond activation of the phosphine ligand was observed to produce Cp*HFe(κ2-(P,Sn)═Sn(DMP)CH2CHMePMeiPr) (5). Complex 5 was also accessed via thermolysis of 1, and kinetics studies of this thermolytic pathway indicate that the reductive elimination of H2 from 1 to produce a stannylyne intermediate, Cp*(iPr2MeP)Fe[SnDMP] (A), is likely rate-determining. Evidence indicates that the production of 5 proceeds through a concerted C-H bond activation. DFT investigations suggest that the transition state for this transformation involves C-H cleavage across the Fe-Sn bond and that a related transition state where C-H bond activation occurs exclusively at the tin center is disfavored, illustrating an effect of iron-tin cooperativity in this system.

BesC Initiates C-C Cleavage through a Substrate-Triggered and Reactive Diferric-Peroxo Intermediate.

BesC catalyzes the iron- and O2-dependent cleavage of 4-chloro-l-lysine to form 4-chloro-l-allylglycine, formaldehyde, and ammonia. This process is a critical step for a biosynthetic pathway that generates a terminal alkyne amino acid which can be leveraged as a useful bio-orthogonal handle for protein labeling. As a member of an emerging family of diiron enzymes that are typified by their heme oxygenase-like fold and a very similar set of coordinating ligands, recently termed HDOs, BesC performs an unusual type of carbon-carbon cleavage reaction that is a significant departure from reactions catalyzed by canonical dinuclear-iron enzymes. Here, we show that BesC activates O2 in a substrate-gated manner to generate a diferric-peroxo intermediate. Examination of the reactivity of the peroxo intermediate with a series of lysine derivatives demonstrates that BesC initiates this unique reaction trajectory via cleavage of the C4-H bond; this process represents the rate-limiting step in a single turnover reaction. The observed reactivity of BesC represents the first example of a dinuclear-iron enzyme that utilizes a diferric-peroxo intermediate to capably cleave a C-H bond as part of its native function, thus circumventing the formation of a high-valent intermediate more commonly associated with substrate monooxygenations.

Spontaneous assembly of redox-active iron-sulfur clusters at low concentrations of cysteine.

Iron-sulfur (FeS) proteins are ancient and fundamental to life, being involved in electron transfer and CO2 fixation. FeS clusters have structures similar to the unit-cell of FeS minerals such as greigite, found in hydrothermal systems linked with the origin of life. However, the prebiotic pathway from mineral surfaces to biological clusters is unknown. Here we show that FeS clusters form spontaneously through interactions of inorganic Fe2+/Fe3+ and S2- with micromolar concentrations of the amino acid cysteine in water at alkaline pH. Bicarbonate ions stabilize the clusters and even promote cluster formation alone at concentrations >10 mM, probably through salting-out effects. We demonstrate robust, concentration-dependent formation of [4Fe4S], [2Fe2S] and mononuclear iron clusters using UV-Vis spectroscopy, 57Fe-Mössbauer spectroscopy and 1H-NMR. Cyclic voltammetry shows that the clusters are redox-active. Our findings reveal that the structures responsible for biological electron transfer and CO2 reduction could have formed spontaneously from monomers at the origin of life.

The Effect of High Selenite and Selenate Concentrations on Ferric Oxyhydroxides Transformation under Alkaline Conditions.

Iron-based nanomaterials have high technological impacts on various pro-environmental applications, including wastewater treatment using the co-precipitation method. The purpose of this research was to identify the changes of iron nanomaterial's structure caused by the presence of selenium, a typical water contaminant, which might affect the removal when the iron co-precipitation method is used. Therefore, we have investigated the maturation of co-precipitated nanosized ferric oxyhydroxides under alkaline conditions and their thermal transformation into hematite in the presence of selenite and selenate with high concentrations. Since the association of selenium with precipitates surfaces has been proven to be weak, the mineralogy of the system was affected insignificantly, and the goethite was identified as an only ferric phase in all treatments. However, the morphology and the crystallinity of ferric oxyhydroxides was slightly altered. Selenium affected the structural order of precipitates, especially at the initial phase of co-precipitation. Still, the crystal integrity and homogeneity increased with time almost constantly, regardless of the treatment. The thermal transformation into well crystalized hematite was more pronounced in the presence of selenite, while selenate-treated and selenium-free samples indicated the presence of highly disordered fraction. This highlights that the aftermath of selenium release does not result in destabilization of ferric phases; however, since weak interactions of selenium are dominant at alkaline conditions with goethite's surfaces, it still poses a high risk for the environment. The findings of this study should be applicable in waters affected by mining and metallurgical operations.

Di-tert-butyldiphosphatetrahedrane as a Source of 1,2-Diphosphacyclobutadiene Ligands.

Reactions of di-tert-butyldiphosphatetrahedrane (1) with cycloocta-1,5-diene- or anthracene-stabilised metalate anions of iron and cobalt consistently afford complexes of the rarely encountered 1,2-diphosphacyclobutadiene ligand, which have previously been very challenging synthetic targets. The subsequent reactivity of 1,2-diphosphacyclobutadiene cobaltates toward various electrophiles has also been investigated and is compared to reactions of related 1,3-diphosphacyclobutadiene complexes. The results highlight the distinct reactivity of such isomeric species, showing that the 1,2-isomers can act as precursors for previously unknown triphospholium ligands. The electronic structures of the new complexes were investigated by several methods, including NMR, EPR and Mößbauer spectroscopies as well as quantum chemical calculations.

Heme-Heme Interactions in Diheme Cytochromes: Effect of Mixed-Axial Ligation on the Electronic Structure and Electrochemical Properties.

Diheme cytochromes, the simplest members in the multiheme family, play substantial biochemical roles in enzymatic catalysis as well as in electron transfer. A series of diiron(III) porphyrin dimers have been synthesized as active site analogues of diheme cytochromes. The complexes contain six-coordinated iron(III) having thiophenol and imidazole at the fifth and sixth coordination sites, respectively. The iron centers in the complexes have been found to be in a low-spin state, as confirmed through solid-state Mössbauer and electron paramagnetic resonance (EPR) spectroscopic investigations. Mössbauer quadrupole splitting of complexes having mixed ligands is substantially larger than that observed when both axial ligands are the same. Rhombic types of EPR spectra with narrow separation between gx, gy, and gz clearly distinguish heme thiolate coordination compared to bis(imidazole)-ligated low-spin heme centers. The redox potential in diheme cytochromes has been found to be tuned by interheme interactions along with the nature of axial ligands. The effect of mixed-axial ligation within the diiron(III) porphyrin dimers is demonstrated by a positive shift in the Fe(III)/Fe(II) redox couple upon thiophenolate coordination compared to their bis(imidazole) analogues. The pKa of the imidazole also decides the extent of the shift for the Fe(III)/Fe(II) couple, while the potential of the mixed-ligated diiron(III) porphyrin dimer is more positive compared to their monomeric analogue. A variation of around 1.1 V for the Fe(III)/Fe(II) redox potential in the diiron(III) porphyrin dimer can be achieved with the combined effect of axial ligation and a metal spin state, while such a large variation in the redox potential, compared to their monomeric analogues, is attributed to the heme-heme interactions observed in dihemes. Moreover, theoretical calculations also support the experimental shifts in the redox potential values.

Electron inventory of the iron-sulfur scaffold complex HypCD essential in [NiFe]-hydrogenase cofactor assembly.

The [4Fe-4S] cluster containing scaffold complex HypCD is the central construction site for the assembly of the [Fe](CN)2CO cofactor precursor of [NiFe]-hydrogenase. While the importance of the HypCD complex is well established, not much is known about the mechanism by which the CN- and CO ligands are transferred and attached to the iron ion. We report an efficient expression and purification system producing the HypCD complex from E. coli with complete metal content. This enabled in-depth spectroscopic characterizations. The results obtained by EPR and Mössbauer spectroscopy demonstrate that the [Fe](CN)2CO cofactor and the [4Fe-4S] cluster of the HypCD complex are redox active. The data indicate a potential-dependent interconversion of the [Fe]2+/3+ and [4Fe-4S]2+/+ couple, respectively. Moreover, ATR FTIR spectroscopy reveals potential-dependent disulfide formation, which hints at an electron confurcation step between the metal centers. MicroScale thermophoresis indicates preferable binding between the HypCD complex and its in vivo interaction partner HypE under reducing conditions. Together, these results provide comprehensive evidence for an electron inventory fit to drive multi-electron redox reactions required for the assembly of the CN- and CO ligands on the scaffold complex HypCD.