Evidence for hydrogen abstraction from C1 of taurine by the high-spin Fe(IV) intermediate detected during oxygen activation by taurine:alpha-ketoglutarate dioxygenase (TauD)

The Fe(II)- and alpha-ketoglutarate-dependent dioxygenases catalyze hydroxylation reactions of considerable biomedical and environmental significance. Recently, the first oxidized iron intermediate in the reaction of a member of this family, taurine:alpha-ketoglutarate dioxygenase (TauD), was detected and shown to be a high-spin, formally Fe(IV) complex. The demonstration in this study that decay of the Fe(IV) complex is approximately 30-fold slower when it is formed in the presence of 1-[2H]2-taurine provides evidence that the intermediate abstracts hydrogen from C1, the site of hydroxylation, and suggests that quantum-mechanical tunneling may contribute to C1-H cleavage.     

Dioxygen activation and catalytic aerobic oxidation by a mononuclear nonheme iron(II) complex

We have used dioxygen, not artificial oxidants such as peracids, iodosylarenes, and hydroperoxides, in the generation of a mononuclear nonheme oxoiron(IV) complex, [Fe(IV)(TMC)(O)]2+ (TMC = 1,4,8,11-tetramethyl-1,4,8,11-tetraazacyclotetradecane), from its corresponding Fe(II) complex, [Fe(TMC)(CF3SO3)2]. The formation of oxoiron(IV) species by activating dioxygen was markedly dependent on iron(II) complexes and solvents, and this observation was interpreted with the electronic effect of iron(II) complexes on dioxygen activation to form oxoiron(IV) species. A catalytic aerobic oxidation of organic substrates was demonstrated in the presence of the [Fe(TMC)]2+ complex. By carrying out 18O-labeled water experiment, we were able to conclude that the oxidation of organic substrates was mediated by an oxoiron(IV) intermediate, not by a radical type of autoxidation process.     

Role of Fe(IV)-oxo intermediates in stoichiometric and catalytic oxidations mediated by iron pyridine-azamacrocycles

An iron(II) complex with a pyridine-containing 14-membered macrocyclic (PyMAC) ligand L1 (L1 = 2,7,12-trimethyl-3,7,11,17-tetra-azabicyclo[11.3.1]heptadeca-1(17),13,15-triene), 1, was prepared and characterized. Complex 1 contains low-spin iron(II) in a pseudo-octahedral geometry as determined by X-ray crystallography. Magnetic susceptibility measurements (298 K, Evans method) and M?ssbauer spectroscopy (90 K, δ = 0.50(2) mm/s, ΔE(Q) = 0.78(2) mm/s) confirmed that the low-spin configuration of Fe(II) is retained in liquid and frozen acetonitrile solutions. Cyclic voltammetry revealed a reversible one-electron oxidation/reduction of the iron center in 1, with E(1/2)(Fe(III)/Fe(II)) = 0.49 V vs Fc(+)/Fc, a value very similar to the half-wave potentials of related macrocyclic complexes. Complex 1 catalyzed the epoxidation of cyclooctene and other olefins with H(2)O(2). Low-temperature stopped-flow kinetic studies demonstrated the formation of an iron(IV)-oxo intermediate in the reaction of 1 with H(2)O(2) and concomitant partial ligand oxidation. A soluble iodine(V) oxidant, isopropyl 2-iodoxybenzoate, was found to be an excellent oxygen atom donor for generating Fe(IV)-oxo intermediates for additional spectroscopic (UV-vis in CH(3)CN: λ(max) = 705 nm, ε ≈ 240 M(-1) cm(-1); M?ssbauer: δ = 0.03(2) mm/s, ΔE(Q) = 2.00(2) mm/s) and kinetic studies. The electrophilic character of the (L1)Fe(IV)═O intermediate was established in rapid (k(2) = 26.5 M(-1) s(-1) for oxidation of PPh(3) at 0 °C), associative (ΔH(?) = 53 kJ/mol, ΔS(?) = -25 J/K mol) oxidation of substituted triarylphosphines (electron-donating substituents increased the reaction rate, with a negative value of Hammet's parameter ρ = -1.05). Similar double-mixing kinetic experiments demonstrated somewhat slower (k(2) = 0.17 M(-1) s(-1) at 0 °C), clean, second-order oxidation of cyclooctene into epoxide with preformed (L1)Fe(IV)═O that could be generated from (L1)Fe(II) and H(2)O(2) or isopropyl 2-iodoxybenzoate. Independently determined rates of ferryl(IV) formation and its subsequent reaction with cyclooctene confirmed that the Fe(IV)-oxo species, (L1)Fe(IV)═O, is a kinetically competent intermediate for cyclooctene epoxidation with H(2)O(2) at room temperature. Partial ligand oxidation of (L1)Fe(IV)═O occurs over time in oxidative media, reducing the oxidizing ability of the ferryl species; the macrocyclic nature of the ligand is retained, resulting in ferryl(IV) complexes with Schiff base PyMACs. NH-groups of the PyMAC ligand assist the oxygen atom transfer from ferryl(IV) intermediates to olefin substrates.     

Structural and electrochemical studies of novel bis(mu-methoxo)diiron complexes: observation of Fe(III)Fe(IV) and Fe(IV)Fe(IV) states resonating with phenoxyl radicals

Novel diiron complexes with an Fe2(mu-OMe)2 core were studied as models of the active site of nonheme iron-containing enzymes. X-ray crystal structures of the complexes showed the existence of two types of ligand folding-parallel and twisted-both of which have four virtually equivalent phenolato groups sticking out from the Fe2O2 rhombic plane. Cyclic voltammetry measurements revealed two or more distinct redox waves in a region of relatively high potential, in addition to known Fe(II)/Fe(III) redox waves in a region of lower potential. These new peaks were assigned to the high-valence state of iron atoms, that is, Fe(III)Fe(IV) and Fe(IV)Fe(IV), resonating with the phenoxyl radical(s). The split width of the redox waves ranged from 0.14 to 0.20 eV, which may be a measure of the electronic interaction of the phenolate groups through the Fe2(mu-OMe)2 core.     

The synthesis of organo-imido complexes of Cr(IV) and Fe(III)

The reaction of p-tolylazide with (5,10,15,20- tetraphenylporphyrinato) chromium(II) (Cr(TPP)) yields the high spin chromium(IV) organo-imido complex, CH₃C₆H₄N=Cr(TPP). N,N′-ethylene- bis-(salicylideneiminato)iron(II), (Fe(salen)), however reacts with arylazides to produce iron(III) organo-imido-bridged compounds of general formula, [Fe(salen)]₂NR showing magnetic coupling between the Fe(III) centres.     

CS‐Fe(II,III) complex as precursor for magnetite nanocrystal

Chitosan‐iron ions complex (CS‐Fe(II,III) complex) was used as precursor to synthesize magnetite nanocrystals and the mechanism was discussed. The magnetite nanocrystals have diameters of about 10 nm and clusters were formed due to slight aggregation of several magnetite nanocrystals. FT‐IR and X‐ray photoelectron spectrometer (XPS) investigations indicated that the Fe(II) and Fe(III) were chelated by ? NH₂ and ? OH groups of chitosan in CS‐Fe(II,III) complex, and the molar ratio of ? NH₂/Fe(II,III) was approximately 2. This chelation effect destroyed the hydrogen bonds of chitosan. In the following alkali treatment process, the chelated Fe(II) and Fe(III) provided nucleation site and formed the magnetite nanocrystals. After alkali treatment, the chelation effect between iron ions and ? NH₂ groups disappeared and some kind of weak interaction formed between magnetite and ? NH₂ groups. Moreover, the ? OH groups of chitosan have an interaction with the synthesized magnetite nanocrystals. Copyright © 2010 John Wiley & Sons, Ltd.     

Spectroscopic and computational evaluation of the structure of the high-spin Fe(IV)-oxo intermediates in taurine: alpha-ketoglutarate dioxygenase from Escherichia coli and its His99Ala ligand variant

The Fe(II)- and alpha-ketoglutarate (alphaKG)-dependent dioxygenases activate O2 for cleavage of unactivated C-H bonds in their substrates. The key intermediate that abstracts hydrogen in the reaction of taurine:alphaKG dioxygenase (TauD), a member of this enzyme family, was recently characterized. The intermediate, denoted J, was shown to contain an iron(IV)-oxo unit. Other important structural features of J, such as the number, identity, and disposition of ligands in the Fe(IV) coordination sphere, are not yet understood. To probe these important structural features, a series of models for J with the Fe(IV) ion coordinated by the expected two imidazole (from His99 and His255), two carboxylate (succinate and Asp101), and oxo ligands have been generated by density functional theory (DFT) calculations, and spectroscopic parameters (M?ssbauer isomer shift, quadrupole splitting, and asymmetry parameter, 57Fe hyperfine coupling tensor, and zero field splitting parameters, D and E/D) have been calculated for each model. The calculated parameters of distorted octahedral models for J, in which one of the carboxylates serves as a monodentate ligand and the other as a bidentate ligand, and a trigonal bipyramidal model, in which both carboxylates serve as monodentate ligands, agree well with the experimental parameters, whereas the calculated parameters of a square pyramidal model, in which the oxo ligand is in the equatorial plane, are inconsistent with the data. Similar analysis of the Fe(IV) complex generated in the variant protein with His99, the residue that contributes the imidazole ligand cis to the oxo group, replaced by alanine suggests that the deleted imidazole is replaced by a water ligand. This work lends credence to the idea that the combination of M?ssbauer spectroscopy and DFT calculations can provide detailed structural information for reactive intermediates in the catalytic cycles of iron enzymes.     

A high field mössbauer study of a six coordinate iron II dithiocarbamate complex and comparison of the Fe(II)S6, Fe(III)S6 and Fe(IV)S6 chromophores in tris (diethyldithiocarbamate) complexation

Mössbauer spectroscopy shows that the quadruple interaction in tris (N, N′-diethyldithiocarbamate) iron II is negative, axially symmetric and that the compound is rapidly relaxing at 4.2 K. The results indicate a ⁵A ground term arising from trigonal compression of a Fe(II)S₆ chromophore and positive zero field splitting. A correlation is made with data for the Fe(III)S₆ and Fe(IV)S₆ tris-diethyldithiocarbamates.     

Dioxygen Reactivity of Biomimetic Iron-Catecholate and Iron-o-Aminophenolate Complexes of a Tris(2-pyridylthio)methanido Ligand: Aromatic CC Bond Cleavage of Catecholate versus o-Iminobenzosemiquinonate Radical Formation

An iron(III)-catecholate complex [L(1) Fe(III) (DBC)] (2) and an iron(II)-o-aminophenolate complex [L(1) Fe(II) (HAP)] (3; where L(1) =tris(2-pyridylthio)methanido anion, DBC=dianionic 3,5-di-tert-butylcatecholate, and HAP=monoanionic 4,6-di-tert-butyl-2-aminophenolate) have been synthesised from an iron(II)-acetonitrile complex [L(1) Fe(II) (CH(3) CN)(2) ](ClO(4) ) (1). Complex 2 reacts with dioxygen to oxidatively cleave the aromatic C?C bond of DBC giving rise to selective extradiol cleavage products. Controlled chemical or electrochemical oxidation of 2, on the other hand, forms an iron(III)-semiquinone radical complex [L(1) Fe(III) (SQ)](PF(6) ) (2(ox) -PF(6) ; SQ=3,5-di-tert-butylsemiquinonate). The iron(II)-o-aminophenolate complex (3) reacts with dioxygen to afford an iron(III)-o-iminosemiquinonato radical complex [L(1) Fe(III) (ISQ)](ClO(4) ) (3(ox) -ClO(4) ; ISQ=4,6-di-tert-butyl-o-iminobenzosemiquinonato radical) via an iron(III)-o-amidophenolate intermediate species. Structural characterisations of 1, 2, 2(ox) and 3(ox) reveal the presence of a strong iron?carbon bonding interaction in all the complexes. The bond parameters of 2(ox) and 3(ox) clearly establish the radical nature of catecholate- and o-aminophenolate-derived ligand, respectively. The effect of iron?carbon bonding interaction on the dioxygen reactivity of biomimetic iron-catecholate and iron-o-aminophenolate complexes is discussed.     

Structural and magnetic investigations of the mixed-valence Fe(II,III) two-dimensional layer complex, [Fe2(II) Fe2(III)(HCOO)10(C6H7N)6]n.

The structure of the complex, [Fe2(II)Fe2(III)(HCOO)10(C6H7N6)n, (1) exhibits a neutral two-dimensional layer network of alternating iron(II) and iron(III) ions, bridged equatorially by formate groups. All iron atoms are octahedrally coordinated, with iron(III) coordinating axially to one gamma-picoline and one formate group, while the iron(II) centers interact axially with two gamma-picoline groups, above and below the layer plane. The complex crystallizes in the triclinic space group P1 at all studied temperatures [at 120 K, the cell dimensions are: a = 10.228(1), b = 12.071(1), c = 12.072(1) A, alpha = 89.801(2), beta = 71.149(2), gamma = 73.371(2) degrees]. An intralayer antiferromagnetic exchange interaction of J = -2.8 cm(-1) between iron(II) and iron(III) was observed in the magnetic studies. Decreasing the temperature to close to 20 K causes a magnetic-ordering phenomenon to occur and a low-temperature phase with a long-range antiferromagnetic spin orientation appears. The magnetic phase transition was confirmed by M?ssbauer spectroscopic studies at temperatures above and below the critical temperature. Structural information of 1 from synchrotron X-ray diffraction data collected at room temperature and 16 K suggests that the antiferromagnetic ordering is caused by an enhanced pi-pi interaction between chi-picoline groups from adjacent layers.     

N-O bond homolysis of an iron(II) TEMPO complex yields an iron(III) oxo intermediate

The reaction of TEMPO with the iron(I) synthon PhB(MesIm)(3)Fe(COE) leads to formation of the κ(1)-TEMPO complex PhB(MesIm)(3)Fe(TEMPO). Structural and spectroscopic data establish the complex contains divalent iron bound to a nitroxido anion and is isoelectronic to an iron(II) peroxo complex. Thermolysis of the complex results in N-O bond homolysis, leading to the formation of an iron(III) oxo intermediate. The oxo intermediate is active in oxygen atom transfer reactions and can be trapped by the triphenylmethyl radical to give the iron(II) alkoxo complex PhB(MesIm)(3)Fe(OCPh(3)).     

Mechanism of dioxygen cleavage in tetrahydrobiopterin-dependent amino acid hydroxylases

The reaction mechanism for the formation of the hydroxylating intermediate in aromatic amino acid hydroxylases (i.e., phenylalanine hydroxylase, tyrosine hydroxylase, tryptophan hydroxylase) was investigated by means of hybrid density functional theory. These enzymes use molecular oxygen to hydroxylate both the tetrahydrobiopterin cofactor and the aromatic amino acid. A mechanism is proposed in which dioxygen forms a bridging bond between the cofactor and iron. The product is an iron(II)-peroxy-pterin intermediate, and iron was found to be essential for the catalysis of this step. No stable intermediates involving a pterin radical cation and a superoxide ion O(2)(-) were found on the reaction pathway. Heterolysis of the O-O bond in the iron(II)-peroxy-pterin intermediate is promoted by one of the water molecules coordinated to iron and releases hydroxypterin and the high-valent iron oxo species Fe(IV)=O, which can carry out subsequent hydroxylation of aromatic rings. In the proposed mechanism, the formation of the bridging C-O bond is rate-limiting in the formation of Fe(IV)=O.     

A synthetic model of the putative Fe(II)-iminobenzosemiquinonate intermediate in the catalytic cycle of o-aminophenol dioxygenases

The oxidative ring cleavage of aromatic substrates by nonheme Fe dioxygenases is thought to involve formation of a ferrous-(substrate radical) intermediate. Here we describe the synthesis of the trigonal-bipyramdial complex Fe((Ph2)Tp)(ISQ(tBu)) (2), the first synthetic example of an iron(II) center bound to an iminobenzosemiquinonate (ISQ) radical. The unique electronic structure of this S = 3/2 complex and its one-electron oxidized derivative ([3](+)) have been established on the basis of crystallographic, spectroscopic, and computational analyses. These findings further demonstrate the viability of Fe(2+)-ISQ intermediates in the catalytic cycles of o-aminophenol dioxygenases.     

Hydrogenolysis of [PhBP3]Fe[triple bond]N-p-tolyl: probing the reactivity of an iron imide with H2

This paper describes the reductive hydrogenolysis of a low-spin (S = 1/2) iron(III) imide. Pseudotetrahedral [PhBP3]FeIIIN-p-tolyl is reduced by hydrogen at ambient temperature and pressure in benzene solution. The reduction appears to proceed in a stepwise fashion. An intermediate S = 2 iron(II) anilide, [PhBP3]Fe(N(H)-p-tolyl), is observed and has been independently generated and structurally characterized. Prolonged hydrogenolysis in benzene results in the complete hydrogenolysis of the Fe-N linkage to release H2N-p-tolyl. The major iron-containing product formed from this step is the diamagnetic cyclohexadienyl complex, [PhBP3]Fe(eta5-cyclohexadienyl), which has also been independently prepared and structurally characterized. Evidence is presented to suggest that the final [PhBP3]Fe(eta5-cyclohexadienyl) product is formed via benzene insertion into a reactive [PhBP3]FeII-H intermediate.     

Stereospecific alkane hydroxylation by non-heme iron catalysts: mechanistic evidence for an Fe(V)=O active species.

High-valent iron-oxo species have frequently been invoked in the oxidation of hydrocarbons by both heme and non-heme enzymes. Although a formally Fe(V)=O species, that is, [(Por(*))Fe(IV)=O](+), has been widely accepted as the key oxidant in stereospecific alkane hydroxylation by heme systems, it is not established that such a high-valent state can be accessed by a non-heme ligand environment. Herein we report a systematic study on alkane oxidations with H(2)O(2) catalyzed by a group of non-heme iron complexes, that is, [Fe(II)(TPA)(CH(3)CN)(2)](2+) (1, TPA = tris(2-pyridylmethyl)amine) and its alpha- and beta-substituted analogues. The reactivity patterns of this family of Fe(II)(TPA) catalysts can be modulated by the electronic and steric properties of the ligand environment, which affects the spin states of a common Fe(III)-OOH intermediate. Such an Fe(III)-peroxo species is high-spin when the TPA ligand has two or three alpha-substituents and is proposed to be directly responsible for the selective C-H bond cleavage of the alkane substrate. The thus-generated alkyl radicals, however, have relatively long lifetimes and are susceptible to radical epimerization and trapping by O(2). On the other hand, 1 and the beta-substituted Fe(II)(TPA) complexes catalyze stereospecific alkane hydroxylation by a mechanism involving both a low-spin Fe(III)-OOH intermediate and an Fe(V)=O species derived from O-O bond heterolysis. We propose that the heterolysis pathway is promoted by two factors: (a) the low-spin iron(III) center which weakens the O-O bond and (b) the binding of an adjacent water ligand that can hydrogen bond to the terminal oxygen of the hydroperoxo group and facilitate the departure of the hydroxide. Evidence for the Fe(V)=O species comes from isotope-labeling studies showing incorporation of (18)O from H(2)(18)O into the alcohol products. (18)O-incorporation occurs by H(2)(18)O binding to the low-spin Fe(III)-OOH intermediate, its conversion to a cis-H(18)O-Fe(V)=O species, and then oxo-hydroxo tautomerization. The relative contributions of the two pathways of this dual-oxidant mechanism are affected by both the electron donating ability of the TPA ligand and the strength of the C-H bond to be broken. These studies thus serve as a synthetic precedent for an Fe(V)=O species in the oxygen activation mechanisms postulated for non-heme iron enzymes such as methane monooxygenase and Rieske dioxygenases.     

First Synthesis of a Binuclear [Mn(II)(bipy)-Fe(III)(porphyrin)] Complex: Spectroscopic Characterization and First Evidence of Reversible Formation of Manganese(III) as Manganese Peroxidase

A [(P)Fe(III)-Mn(II)] bimetallic complex, mimicking the active site of manganese peroxidase, has been synthesized. A modified highly fluorinated porphyrin, 5,10,15-tris(pentafluorophenyl)-20-(o-aminophenyl)porphyrin, has been used to introduce, through a short spacer linked to the amino function, a manganese auxiliary ligand, 6-aminomethyl-2,2'-bipyridine. Two successive metalations by FeCl(2) and MnCl(2) afforded the [(P)Fe(III)-Mn(II)] bimetallic complex that has been characterized by elemental analysis and FAB(+) mass spectrometry. X-band EPR spectroscopy and magnetic susceptibility measurements were in agreement with two high spin Fe(III) and Mn(II) centers without magnetic exchange interaction. Moreover, there is no higher intermolecular association through &mgr;-chloro bridging as observed by EPR with a simpler chloromanganese complex, Mn(bipy)(2)Cl(2), at high concentration. Addition of pentafluoroiodosobenzene in methanol at 0 degrees C led to the progressive and complete disappearance of the EPR Mn(II) signals, that were recovered after addition of a phenol. This result is consistent with Mn(III) formation. This production of Mn(III) requires the presence of the iron porphyrin and is proposed to occur through the intermediate formation of a Fe(IV) dimethoxide species which can be related to the oxidation of Mn(II) catalyzed by manganese peroxidase compound II.     

Interfacial aggregate growth process of Fe(II) and Fe(III) complexes with pyridylazophenol in solvent extraction system

The complexation mechanism and aggregate formation of bis[2-(5-bromo-2-pyridylazo)-5-diethylaminophenolate] iron(II) and iron(III) complexes at the heptane-water interface were studied spectrophotometrically by the high-speed stirring method and the centrifugal liquid membrane method. Furthermore, the reduction process of the Fe(III) complex with ascorbic acid at the interface was spectrophotometrically observed. The chemical compositions of the interfacial aggregate of complexes have been proved by the X-ray photoelectron spectroscopy. The aggregation of the complex at the interface was observed as a red-shifted, very strong and narrower absorption band with respect to the absorption band of the monomer complex. The aggregate of Fe(III) complex showed more shifted spectrum than that of Fe(II) complex, which proposed the larger aggregation number of Fe(III) aggregate (n = 8) than that of Fe(II) aggregate (n = 3). The obtained rate constants of interfacial aggregation were smaller than rate constants of interfacial monomer complexation, because the formation of aggregate required the assembly of the monomers.     

Olefin cis-dihydroxylation with bio-inspired iron catalysts. evidence for an Fe(II)/Fe(IV) catalytic cycle

Iron(II) complexes of a series of N-acylated dipyridin-2-ylmethylamine ligands (R-DPAH) have been investigated as catalysts for the cis-dihydroxylation of olefins to model the action of Rieske dioxygenases that catalyze arene cis-dihydroxylation. The Rieske dioxygenases have a mononuclear iron active site coordinated to a 2-histidine-1-carboxylate facial triad motif. The R-DPAH ligands are designed to provide a facial N,N,O-ligand set that mimics the enzyme active site. The iron(II) complexes of the R-DPAH ligands activate H(2)O(2) to effect the oxidation of olefin substrates into cis-diol products. As much as 90% of the H(2)O(2) oxidant is converted into cis-diol, but a large excess of olefin is required to achieve the high conversion efficiency. Reactivity and mechanistic comparisons with the previously characterized Fe(TPA)/H(2)O(2) catalyst/oxidant combination (TPA = tris(pyridin-2-ylmethyl)amine) lead us to postulate an Fe(II)/Fe(IV) redox cycle for the Fe(R-DPAH) catalysts in which an Fe(IV)(OH)(2) oxidant carries out the cis-hydroxylation of olefins. This hypothesis is supported by three sets of observations: (a) the absence of a lag phase in the conversion of the H(2)O(2) oxidant into a cis-diol product, thereby excluding the prior oxidation of the Fe(II) catalyst to an Fe(III) derivative as established for the Fe(TPA) catalyst; (b) the incorporation of H(2)(18)O into the cis-diol product, thereby requiring O-O bond cleavage to occur prior to cis-diol formation; and (c) the formation of cis-diol as the major product of cyclohexene oxidation, rather than the epoxide or allylic alcohol products more commonly observed in metal-catalyzed oxidations of cyclohexene, implicating an oxidant less prone to oxo transfer or H-atom abstraction.     

Origin of the unusual kinetics of iron deposition in human H-chain ferritin

From microorganisms to humans, ferritin plays a central role in the biological management of iron. The ferritins function as iron storage and detoxification proteins by oxidatively depositing iron as a hydrous ferric hydroxide mineral core within their shell-like structures. The mechanism by which the mineral core is formed has been the subject of intense investigation for many years. A diiron ferroxidase site located on the H-chain subunit of vertebrate ferritins catalyzes the oxidation of Fe(II) to Fe(III) by molecular oxygen. A previous stopped-flow kinetics study of a transient mu-peroxodiFe(III) intermediate formed at this site revealed very unusual kinetics curves, the shape of which depended markedly on the amount of iron presented to the protein. In the present work, a mathematical model for catalysis is developed that explains the observed kinetics. The model consists of two sequential mechanisms. In the first mechanism, turnover of iron at the ferroxidase site is rapid, resulting in steady-state production of the peroxo intermediate with continual formation of the mineral core until the available Fe(II) in solution is consumed. At this point, the second mechanism comes into play whereby the peroxo intermediate decays and the ferroxidase site is postulated to vacate its complement of iron. The kinetic data reveal for the first time that Fe(II) in excess of that required to saturate the ferroxidase site promotes rapid turnover of Fe(III) at this site and that the ferroxidase site plays a role in catalysis at all levels of iron loading of the protein (48-800 Fe/protein). The data also provide evidence for a second intermediate, a putative hydroperoxodiFe(III) complex, that is a decay product of the peroxo intermediate.     

Volume profile analysis for the reversible binding of superoxide to form iron(II)-superoxo/Iron(III)-peroxo porphyrin complexes

The one-electron reduced iron(II)-dioxygen adduct, {Fe(II)-O(2)}(-), is known to be an important intermediate in the catalytic cycle of heme (mono)oxygenases. The same type of species, considered as Fe(III)-peroxo, can be formed in a direct reaction between a Fe(II) center and superoxide. In a unique high-pressure study of the reaction between superoxide and the Fe(II) complex of a crown ether porphyrin conjugate in dimethylsulfoxide (DMSO), the overall Fe(II)-superoxide interaction mechanism could be visualized and the nature of all species that occur along the reaction coordinate could be clarified. The equilibrium between the low-spin and high-spin forms of the starting Fe(II) complex was quantified, which turns out to be the actual activation step toward substitution and subsequent inner-sphere electron transfer reactions. The constructed reaction volume profile demonstrates that the reaction product consists of Fe(III)-peroxo and Fe(II)-superoxo species that exist in equilibrium, which can better account for the versatile reactivity of {Fe(II)-O(2)}(-) adducts toward different substrates.