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Metalloproteins with dinuclear cores are known to bind and activate dioxygen, with a subclass of these proteins having active sites containing FeMn cofactors and activities ranging from long-range proton-coupled electron transfer (PCET) to post-translational peptide modification. While mechanistic studies propose that these metallocofactors access FeIIIMnIV intermediates, there is a dearth of related synthetic analogs. Herein, the first well-characterized synthetic FeIII-(μ-O)-MnIV complex is reported; this complex shows similar spectroscopic features as the catalytically competent FeIIIMnIV intermediate X found in Class Ic ribonucleotide reductase and demonstrates PCET function towards phenolic substrates. This complex is prepared from the oxidation of the isolable FeIII-(μ-O)-MnIII species, whose stepwise assembly is facilitated by a tripodal ligand containing phosphinic amido groups. Structural and spectroscopic studies found proton movement involving the FeIIIMnIII core, whereby the initial bridging hydroxido ligand is converted to an oxido ligand with concomitant protonation of one phosphinic amido group. This series of FeMn complexes allowed us to address factors that may dictate the preference of an active site for a heterobimetallic cofactor over one that is homobimetallic: comparisons of the redox properties of our FeMn complexes with those of the di-Fe analogs suggested that the relative thermodynamic ease of accessing an FeIIIMnIV core can play an important role in determining the metal ion composition when the key catalytic steps do not require an overly potent oxidant. Moreover, these complexes allowed us to demonstrate the effect of the hyperfine interaction from non-Fe nuclei on 57Fe Mössbauer spectra which is relevant to MnFe intermediates in proteins.

Metalloproteins with active sites containing di-Fe cores exhibit diverse chemical reactivity that is linked to the precise transfer of protons and electrons which directly involve the di-Fe units. The redox conversions are commonly corroborated by spectroscopic methods, but the associated structural changes are often difficult to assess, particularly those related to proton movements. This report describes the development of di-Fe complexes in which the movements of protons and electrons are pinpointed during the stepwise oxidation of a di-FeII species to one with an FeIIIFeIV core. Complex formation was promoted using the phosphinic amido tripodal ligand [poat]3- (N,N',N″-[nitrilotris(ethane-2,1-diyl)]tris(P,P-diphenylphosphinic amido)) that provided dynamic coordination spheres that assisted in regulating both electron and proton transfer processes. Oxidation of an [FeII-(μ-OH)-FeIII] complex led to the corresponding di-FeIII species containing a hydroxido bridge that was not stable at room temperature and converted to a species containing an oxido bridging ligand and protonation of one phosphinic amido group to form [Hpoat]2-. Deprotonation led to a new species with an [FeIII-(μ-O)-FeIII] core that could be further oxidized to its FeIIIFeIV analogue. Reactions with phenols suggest homolytic cleavage of the O-H bond to give products that are consistent with the initial formation of a phenoxyl radical─spectroscopic studies indicated that the electron is transferred to the FeIV center, and the proton is initially transferred to the more sterically hindered oxido ligand but then relocates to [poat]3-. These findings offer new mechanistic insights related to the stability of and the reactions performed by di-Fe enzymes.

Bimetallic active sites are ubiquitous in metalloenzymes and have sparked investigations of synthetic models to aid in the establishment of structure-function relationship. We previously reported a series of discrete bimetallic complexes with [FeIII-(μ-OH)-MII] cores in which the ligand framework provides distinct binding sites for two metal centers. The formation of these complexes relied on a stepwise synthetic approach in which an FeIII-OH complex containing a sulfonamido tripodal ligand served as a synthon that promoted assembly. We have utilized this approach in the present study to produce a new series of bimetallic complexes with [FeIII-(μ-OH)-MII] cores (M = Ni, Cu, Zn) by using an ancillary ligand to the FeIII center that contains phosphinic amido groups. Assembly began with formation of an FeIII-OH that was subsequently used to bind the MII fragment that contained a triazacyclononane ligand. The series of bimetallic complexes were charactered structurally by X-ray diffraction methods, spectroscopically by absorption, vibrational, electron paramagnetic resonance spectroscopies, and electrochemically by cyclic voltammetry. A notable finding is that these new [FeIII-(μ-OH)-MII] complexes displayed significantly lower reduction potentials than their sulfonamido counterparts, which paves way for future studies on high valent bimetallic complexes in this scaffold.

Dinuclear iron centers with a bridging hydroxido or oxido ligand form active sites within a variety of metalloproteins. A key feature of these sites is the ability of the protein to control the structures around the Fe centers, which leads to entatic states that are essential for function. To simulate this controlled environment, artificial proteins have been engineered using biotin-streptavidin (Sav) technology in which Fe complexes from adjacent subunits can assemble to form [FeIII-(μ-OH)-FeIII] cores. The assembly process is promoted by the site-specific localization of the Fe complexes within a subunit through the designed mutation of a tyrosinate side chain to coordinate the Fe centers. An important outcome is that the Sav host can regulate the Fe···Fe separation, which is known to be important for function in natural metalloproteins. Spectroscopic and structural studies from X-ray diffraction methods revealed uncommonly long Fe···Fe separations that change by less than 0.3 Å upon the binding of additional bridging ligands. The structural constraints imposed by the protein host on the di-Fe cores are unique and create examples of active sites having entatic states within engineered artificial metalloproteins.

High-valent nonheme FeIV-oxido species are key intermediates in biological oxidation, and their properties are proposed to be influenced by the unique microenvironments present in protein active sites. Microenvironments are regulated by noncovalent interactions, such as hydrogen bonds (H-bonds) and electrostatic interactions; however, there is little quantitative information about how these interactions affect crucial properties of high valent metal-oxido complexes. To address this knowledge gap, we introduced a series of FeIV-oxido complexes that have the same S = 2 spin ground state as those found in nature and then systematically probed the effects of noncovalent interactions on their electronic, structural, and vibrational properties. The key design feature that provides access to these complexes is the new tripodal ligand [poat]3-, which contains phosphinic amido groups. An important structural aspect of [FeIVpoat(O)]- is the inclusion of an auxiliary site capable of binding a Lewis acid (LAII); we used this unique feature to further modulate the electrostatic environment around the Fe-oxido unit. Experimentally, studies confirmed that H-bonds and LAII s can interact directly with the oxido ligand in FeIV-oxido complexes, which weakens the Fe═O bond and has an impact on the electronic structure. We found that relatively large vibrational changes in the Fe-oxido unit correlate with small structural changes that could be difficult to measure, especially within a protein active site. Our work demonstrates the important role of noncovalent interactions on the properties of metal complexes, and that these interactions need to be considered when developing effective oxidants.

The exchange-coupling constants (J) in a series of bimetallic complexes with an M2+(μ-OH)Fe3+ core (M = Mn, Fe, Ni, and Cu; series 1), which were reported in a recent study ( Sano et al. Inorg. Chem. 2017 , 56 , 14118 - 14128 ), have been analyzed with the help of density functional theory (DFT) calculations. The experimental J values of series 1 showed the remarkable property that they were virtually independent of metal M. This behavior contrasts with that observed for a related series of complexes with M2+Fe3+ cores reported by Chaudhuri and co-workers ( Biswas et al. Inorg. Chem. 2010 , 49 , 626 - 641 ) (series 2) in which J increases toward the upper end of the series. Broken symmetry DFT calculations for J, which yielded values in good agreement for the MnFe and NiFe complexes of series 1, gave for the CuFe complex a J value that was persistently much larger than that obtained from the experiment. Attempts to bridge the discrepancy by invoking various basis sets and corrections for hydrogen-bonding effects on J were not successful. The J values for series 1 were subsequently analyzed in the context of an exchange pathway model. From this analysis, it emerged that, in addition to the regular 2e-pathways, which contribute antiferromagnetic terms to J, there are also 3e-pathways that contribute ferromagnetic terms and have the propensity to keep J constant along series 1. It is shown that, while DFT evaluates the 2e-pathway terms reliably, this method seriously underestimates the 3e-pathway contributions, resulting in a too high J value for the CuFe complex of series 1. The pathway analysis of series 2 reveals that the 3e-pathway contributions to J are considerably smaller than those in series 1, resulting in J values that increase toward the upper end of the series, in accordance with the experiment.

Hydrogen bonds (H-bonds) have been shown to modulate the chemical reactivities of iron centers in iron-containing dioxygen-activating enzymes and model complexes. However, few examples are available that investigate how systematic changes in intramolecular H-bonds within the secondary coordination sphere influence specific properties of iron intermediates, such as iron-oxido/hydroxido species. Here, we used 57 Fe nuclear resonance vibrational spectroscopy (NRVS) to probe the Fe-O/OH vibrations in a series of FeIII -hydroxido and FeIV/III -oxido complexes with varying H-bonding networks but having similar trigonal bipyramidal primary coordination spheres. The data show that even subtle changes in the H-bonds to the Fe-O/OH units result in significant changes in their vibrational frequencies, thus demonstrating the utility of NRVS in studying the effect of the secondary coordination sphere to the reactivities of iron complexes.

Hydrogen bonds (H-bonds) within the secondary coordination sphere are often invoked as essential noncovalent interactions that lead to productive chemistry in metalloproteins. Incorporating these types of effects within synthetic systems has proven a challenge in molecular design that often requires the use of rigid organic scaffolds to support H-bond donors or acceptors. We describe the preparation and characterization of a new hybrid tripodal ligand ([H2pout]3-) that contains two monodeprotonated urea groups and one phosphinic amide. The urea groups serve as H-bond donors, while the phosphinic amide group serves as a single H-bond acceptor. The [H2pout]3- ligand was utilized to stabilize a series of Mn-hydroxido complexes in which the oxidation state of the metal center ranges from 2+ to 4+. The molecular structure of the MnIII-OH complex demonstrates that three intramolecular H-bonds involving the hydroxido ligand are formed. Additional evidence for the formation of intramolecular H-bonds was provided by vibrational spectroscopy in which the energy of the O-H vibration supports its assignment as an H-bond donor. The stepwise oxidation of [MnIIH2pout(OH)]2- to its higher oxidized analogs was further substantiated by electrochemical measurements and results from electronic absorbance and electron paramagnetic resonance spectroscopies. Our findings illustrate the utility of controlling both the primary and secondary coordination spheres to achieve structurally similar Mn-OH complexes with varying oxidation states.

The design of molecular complexes of earth-abundant first-row transition metals that can catalyze multi-electron C-H bond activation processes is of interest for achieving efficient, low-cost syntheses of target molecules. To overcome the propensity of these metals to perform single-electron processes, redox-active ligands have been utilized to provide additional electron equivalents. Herein, we report the synthesis of a novel redox active ligand, [ibaps]3-, which binds to transition metals such as FeII and CoII in a meridional fashion through the three anionic nitrogen atoms and provides additional coordination sites for other ligands. In this study, the neutral bidentate ligand 2,2'-bipyridine (bpy) was used to complete the coordination spheres of the metal ions and form NEt4[MII(ibaps)bpy] (M = Fe (1) or Co (1-Co)) salts. The FeII salt exhibited rich electrochemical properties and could be chemically oxidized by 1 and 2 equiv. of ferrocenium to form singly and doubly oxidized species, respectively. The reactivity of 1 towards intramolecular C-H bond amination of aryl azides at benzylic and aliphatic carbon centers was explored, and moderate to good yields of the resulting indoline products were obtained.

Bimetallic complexes are important sites in metalloproteins but are often difficult to prepare synthetically. We have previously introduced an approach to form discrete bimetallic complexes with MII-(μ-OH)-FeIII (MII = Mn, Fe) cores using the tripodal ligand N,N',N″-[2,2',2″-nitrilotris(ethane-2,1-diyl)]tris(2,4,6-trimethylbenzenesulfonamido) ([MST]3-). This series is extended to include the rest of the late 3d transition metal ions (MII = Co, Ni, Cu, Zn). All of the bimetallic complexes have similar spectroscopic and structural properties that reflect little change despite varying the MII centers. Magnetic studies performed on the complexes in solution using electron paramagnetic resonance spectroscopy showed that the observed spin states varied incrementally from S = 0 through S = 5/2; these results are consistent with antiferromagnetic coupling between the high-spin MII and FeIII centers. However, the difference in the MII ion occupancy yielded only slight changes in the magnetic exchange coupling strength, and all complexes had J values ranging from +26(4) to +35(3) cm-1.

Carotenoid cleavage oxygenases (CCOs) are non-heme iron enzymes that catalyze scission of alkene groups in carotenoids and stilbenoids to form biologically important products. CCOs possess a rare four-His iron center whose resting-state structure and interaction with substrates are incompletely understood. Here, we address this knowledge gap through a comprehensive structural and spectroscopic study of three phyletically diverse CCOs. The crystal structure of a fungal stilbenoid-cleaving CCO, CAO1, reveals strong similarity between its iron center and those of carotenoid-cleaving CCOs, but with a markedly different substrate-binding cleft. These enzymes all possess a five-coordinate high-spin Fe(II) center with resting-state Fe-His bond lengths of ∼2.15 Å. This ligand set generates an iron environment more electropositive than those of other non-heme iron dioxygenases as observed by Mössbauer isomer shifts. Dioxygen (O2) does not coordinate iron in the absence of substrate. Substrates bind away (∼4.7 Å) from the iron and have little impact on its electronic structure, thus excluding coordination-triggered O2 binding. However, substrate binding does perturb the spectral properties of CCO Fe-NO derivatives, indicating proximate organic substrate and O2-binding sites, which might influence Fe-O2 interactions. Together, these data provide a robust description of the CCO iron center and its interactions with substrates and substrate mimetics that illuminates commonalities as well as subtle and profound structural differences within the CCO family.

High-valent Fe-OH species are often invoked as key intermediates but have only been observed in Compound II of cytochrome P450s. To further address the properties of non-heme FeIV-OH complexes, we demonstrate the reversible protonation of a synthetic FeIV-oxo species containing a tris-urea tripodal ligand. The same protonated FeIV-oxo species can be prepared via oxidation, suggesting that a putative FeV-oxo species was initially generated. Computational, Mössbauer, XAS, and NRVS studies indicate that protonation of the FeIV-oxo complex most likely occurs on the tripodal ligand, which undergoes a structural change that results in the formation of a new intramolecular H-bond with the oxido ligand that aids in stabilizing the protonated adduct. We suggest that similar protonated high-valent Fe-oxo species may occur in the active sites of proteins. This finding further argues for caution when assigning unverified high-valent Fe-OH species to mechanisms.

Cupredoxins are electron-transfer proteins that have active sites containing a mononuclear Cu center with an unusual trigonal monopyramidal structure (Type 1 Cu). A single Cu-Scys bond is present within the trigonal plane that is responsible for its unique physical properties. We demonstrate that a cysteine-containing variant of streptavidin (Sav) can serve as a protein host to model the structure and properties of Type 1 Cu sites. A series of artificial Cu proteins are described that rely on Sav and a series of biotinylated synthetic Cu complexes. Optical and EPR measurements highlight the presence of a Cu-Scys bond, and XRD analysis provides structural evidence. We further provide evidence that changes in the linker between the biotin and Cu complex within the synthetic constructs allows for small changes in the placement of Cu centers within Sav that have dramatic effects on the structural and physical properties of the resulting artificial metalloproteins. These findings highlight the utility of the biotin-Sav technology as an approach for simulating active sites of metalloproteins.

The structural and electronic properties of a series of manganese complexes with terminal oxido ligands are described. The complexes span three different oxidation states at the manganese center (III-V), have similar molecular structures, and contain intramolecular hydrogen-bonding networks surrounding the Mn-oxo unit. Structural studies using X-ray absorption methods indicated that each complex is mononuclear and that oxidation occurs at the manganese centers, which is also supported by electron paramagnetic resonance (EPR) studies. This gives a high-spin Mn(V)-oxo complex and not a Mn(IV)-oxy radical as the most oxidized species. In addition, the EPR findings demonstrated that the Fermi contact term could experimentally substantiate the oxidation states at the manganese centers and the covalency in the metal-ligand bonding. Oxygen-17-labeled samples were used to determine spin density within the Mn-oxo unit, with the greatest delocalization occurring within the Mn(V)-oxo species (0.45 spins on the oxido ligand). The experimental results coupled with density functional theory studies show a large amount of covalency within the Mn-oxo bonds. Finally, these results are examined within the context of possible mechanisms associated with photosynthetic water oxidation; specifically, the possible identity of the proposed high valent Mn-oxo species that is postulated to form during turnover is discussed.

Photosynthetic water oxidation is catalyzed by a Mn4O5Ca cluster with an unprecedented arrangement of metal ions in which a single manganese center is bonded to a distorted Mn3O4Ca cubane-like structure. Several mechanistic proposals describe the unique manganese center as a site for water binding and subsequent formation of a high valent Mn-oxo center that reacts with a M-OH unit (M = Mn or CaII) to form the O-O bond. The conversion of low valent Mn-OHn (n = 1,2) to a Mn-oxo species requires that a single manganese site be able to accommodate several oxidation states as the water ligand is deprotonated. To study these processes, the preparation and characterization of a new monomeric MnIV-OH complex is described. The MnIV-OH complex completes a series of well characterized Mn-OH and Mn-oxo complexes containing the same primary and secondary coordination spheres; this work thus demonstrates that a single ligand can support mononuclear Mn complexes spanning four different oxidation states (II through V) with oxo and hydroxo ligands that are derived from water. Moreover, we have completed a thermodynamic analysis based on this series of manganese complexes to predict the formation of high valent Mn-oxo species; we demonstrated that the conversion of a MnIV-OH species to a MnV-oxo complex would likely occur via a stepwise proton transfer-electron transfer mechanism. The large dissociation energy for the MnIVO-H bond (~95 kcal/mol) diminished the likelihood that other pathways are operative within a biological context. Furthermore, these studies showed that reactions between Mn-OH and Mn-oxo complexes lead to non-productive, one-electron processes suggesting that initial O-O bond formation with the OEC does not involve an Mn-OH unit.

Manganese-hydroxo species have been implicated in C-H bond activation performed by metalloenzymes, but the electronic properties of many of these intermediates are not well characterized. The present work presents a detailed characterization of three Mn(n)-OH complexes (where n = II, III, and IV) of the tris[(N'-tert-butylureaylato)-N-ethylene]aminato ([H3buea](3-)) ligand using X- and Q-band dual mode electron paramagnetic resonance (EPR). Quantitative simulations for the [Mn(II)H3buea(OH)](2-) complex demonstrated the ability to characterize similar Mn(II) species commonly present in the resting states of manganese-containing enzymes. The spin states of the Mn(III) and Mn(IV) complexes determined from EPR spectroscopy are S = 2 and 3/2, respectively, as expected for the C3 symmetry imposed by the [H3buea](3-) ligand. Simulations of the spectra indicated the constant presence of two Mn(IV) species in solutions of [Mn(IV)H3buea(OH)] complex. The simulations of perpendicular- and parallel-mode EPR spectra allow determination of zero-field splitting and hyperfine parameters for all complexes. For the Mn(III) and Mn(IV) complexes, density functional theory calculations are used to determine the isotropic Mn hyperfine values, to compare the excited electronic state energies, and to give theoretical estimates of the zero-field energy.

Heterobimetallic cores are important units within the active sites of metalloproteins but are often difficult to duplicate in synthetic systems. We have developed a synthetic approach for the preparation of a complex with a Mn(II)-(μ-OH)-Fe(III) core, in which the metal centers have different coordination environments. Structural and physical data support the assignment of this complex as a heterobimetallic system. A comparison with analogous homobimetallic complexes, Mn(II)-(μ-OH)-Mn(III) and Fe(II)-(μ-OH)-Fe(III) cores, further supports this assignment.

Many peroxy-containing secondary metabolites (1,2) have been isolated and shown to provide beneficial effects to human health (3-5). Yet, the mechanisms of most endoperoxide biosyntheses are not well understood. Although [...]

[Fe.sup.III]-[O.sub.2.sup.*-] intermediates are well known in heme enzymes, but none have been characterized in the nonheme mononuclear [Fe.sup.II] enzyme family. Many steps in the [O.sub.2] activation and reaction cycle of [Fe.sup.II]-containing homoprotocatechuate 2,3-dioxygenase are made detectable by using the alternative substrate 4-nitrocatechol (4NC) and mutation of the active site His200 to Asn (H200N). Here, the first intermediate (Int-1) observed after adding [O.sub.2] to the H200N-4NC complex is trapped and characterized using EPR and Mossbauer (MB) spectroscopies. Int-1 is a high-spin ([S.sub.1] = 5/2) [Fe.sup.III] antiferromagnetically (AF) coupled to an [S.sub.2] = 1/2 radical (J [approximately equal to] 6 [cm.sup.-1] in H = [JS.sub.1]*[S.sub.2]). It exhibits parallel-mode EPR signals at g = 8.17 from the S = 2 multiplet, and g = 8.8 and 11.6 from the S = 3 multiplet. These signals are broadened significantly by [sup.17][O.sub.2] hyperfine interactions ([A17.sub.O] [approximately equal to] 180 MHz). Thus, Int-1 is an AF-coupled [Fe.sup.III]-[O.sub.2.sup.*-] species. The experimental observations are supported by density functional theory calculations that show nearly complete transfer of spin density to the bound [O.sub.2]. Int-1 decays to form a second intermediate (Int-2). MB spectra show that it is also an AF-coupled [Fe.sup.III]-radical complex. Int-2 exhibits an EPR signal at g = 8.05 arising from an S = 2 state. The signal is only slightly broadened by [sup.17][O.sub.2] ( oxygen activation | oxygenase | spectroscopy | superoxide doi/ 10.1073/pnas.1010015107

PNA oligomers that contain up to three adjacent bipyridine ligands are synthesized and the interaction with [Ni.sup.2+] and [Cu.sup.2+] of these oligomers and of duplexes formed from them are examined. It is indicated that even when two [Cu.sup.2+] ions coordinate to a PNA duplex in which two bipyridine pairs are next to each other, the two metal-ligand complexes that form in the duplex are far enough from each other that the dipolar coupling is very weak.

The electronic properties of an unusually redox-rich iron system, [Ph[Bp.sup.R.sub.3]]Fe--[N.sub.x] (where [Ph[Bp.sup.R.sub.3]] is [PhB([CH.sub.2]P[R.sub.2).sub.3]]-), are explored by Mossbauer, EPR, magnetization, and density-functional methods to gain a detailed picture regarding their oxidation states and electronic structures. The complexes of primary interest in this article are the two terminal iron(IV) nitride species, [[PhBp.sup.iPr].sub.3]]Fe [equivalent to] N (3a) and [PhBpCH2[Cy.sub.3]]Fe [equivalent to] N (3b), and the formally diiron(I) bridged-Fe([mu]-[N.sub.2])Fe species, {[PhBp[iPr.sub.3]][Fe}.sub.2]([mu]-[N.sub.2]) (4). Complex 4 is chemically related to 3a via a spontaneous nitride coupling reaction. The diamagnetic iron(IV) nitrides 3a and 3b exhibit unique electronic environments that are reflected in their unusual Mossbauer parameters, including quadrupole-splitting values of 6.01(1) mm/s and isomer shift values of -0.34(1) mm/s. The data for 4 suggest that this complex can be described by a weak ferromagnetic interaction (J/D < 1) between two iron(I) centers. For comparison, four other relevant complexes also are characterized: a diamagnetic iron(IV) trihydride [Ph[BP.sup.iPr.sub.3]]Fe[(H).sub.3](P[Me.sub.3]) (5), an S = 3/2 iron(I) phosphine adduct [Ph[Bp.sup.iPr.sub.3]]FeP[Me.sub.3] (6), and the S = 2 iron(II) precursors to 3a, [Ph[Bp.sup.iPr.sub.3]]Fe-Cl and [Ph[BP.sup.iPr.sub.3]]Fe-2,3:5,6-dibenzo-7-aza bicyclo[2.2.1]hepta-2,5-diene (dbabh). The electronic properties of these respective complexes also have been explored by density-functional methods to help corroborate our spectral assignments and to probe their electronic structures further. dinitrogen activation | high-valent iron | nitrogenase | high-valent iron nitride | spectroscopy

The tetraheme cytochrome c(sub 554) [cyt c(sub 554)] from Nitrosomonas europae is believed to function as an electron-transfer protein from hydroxylamine oxidoreductase. The NO reductase activity of cyt c(sub 554) may be important during ammonia oxidation in N and europaea at low oxygen concentrations to detoxify NO produced by reduction of nitrite or incomplete oxidation of hydroxylamine.

The electrochemical properties of cyt c(sub 554) using EPR and Mossbauer spectroscopy on chemically reduced and electrochemically poised samples are studied. A comparison of the reduction potentials between homologous hemes of cyt c(sub 554) and other members of the new class of multiheme proteins is discussed.