RESUMEN
The Bacillus subtilis redox regulator Fnr controls genes of the anaerobic metabolism in response to low oxygen tension. An unusual structure for the oxygen-sensing [4Fe-4S](2+) cluster was detected by a combination of genetic experiments with UV-visible and Mössbauer spectroscopy. Asp-141 was identified as the fourth iron-sulfur cluster ligand besides three Cys residues. Exchange of Asp-141 with Ala abolished functional in vivo complementation of an fnr knock-out strain by the mutagenized fnr gene and in vitro DNA binding of the recombinant regulator FnrD141A. In contrast, substitution of Asp-141 with Cys preserved [4Fe-4S](2+) structure and regulator function.
Asunto(s)
Bacillus subtilis/metabolismo , Proteínas Bacterianas/metabolismo , Proteínas Hierro-Azufre/metabolismo , Factores de Transcripción/metabolismo , Sustitución de Aminoácidos , Ácido Aspártico/química , Ácido Aspártico/genética , Ácido Aspártico/metabolismo , Bacillus subtilis/química , Bacillus subtilis/genética , Proteínas Bacterianas/química , Proteínas Bacterianas/genética , Proteínas Hierro-Azufre/química , Proteínas Hierro-Azufre/genética , Ligandos , Mutación Missense , Estructura Terciaria de Proteína , Factores de Transcripción/química , Factores de Transcripción/genéticaRESUMEN
Isothermal calorimetric studies of the binding of iron(III) citrate to ferric ion binding protein from Neisseria gonorrhoeae suggested the complexation of a tetranuclear iron(III) cluster as a single step binding event (apparent binding constant K(app) (ITC) = 6.0(5) × 10(5) M(-1)). High-resolution Fourier transform ion cyclotron resonance mass spectrometric data supported the binding of a tetranuclear oxo(hydroxo) iron(III) cluster of formula [Fe(4)O(2)(OH)(4)(H(2)O)(cit)](+) in the interdomain binding cleft of FbpA. The mutant H9Y-nFbpA showed a twofold increase in the apparent binding constant [K(app) (ITC) = 1.1(7) × 10(6) M(-1)] for the tetranuclear iron(III) cluster compared to the wild-type protein. Mössbauer spectra of Escherichia coli cells overexpressing FbpA and cultured in the presence of added (57)Fe citrate were indicative of the presence of dinuclear and polynuclear clusters. FbpA therefore appears to have a strong affinity for iron clusters in iron-rich environments, a property which might endow the protein with new biological functions.
Asunto(s)
Proteínas Bacterianas/química , Compuestos Férricos/química , Proteínas de Unión a Hierro/química , Proteínas Bacterianas/genética , Sitios de Unión , Calorimetría , Clonación Molecular , Proteínas de Unión a Hierro/genética , Espectrometría de Masas , Modelos Moleculares , Estructura Molecular , Neisseria gonorrhoeae , Espectroscopía de MossbauerRESUMEN
Nuclear resonant vibrational spectra of the reduced and oxidized form of a mutant of rubredoxin from Pyrococcus abyssii were measured and are compared with simulated spectra that were calculated by a combined quantum mechanics (QM) and molecular mechanics (MM) method. Density functional theory was used for the QM level. Calculations were performed for different models of rubredoxin. Realistic spectra were simulated with reduced models that include at least the iron center, the four cysteins coordinating it, and the residues connected to the cysteins together with a QM layer that comprises the first two coordination shells of the iron center. Larger QM layers did not lead to significant changes of the simulated spectra.
Asunto(s)
Simulación de Dinámica Molecular , Teoría Cuántica , Rubredoxinas/química , Hierro/química , Pyrococcus abyssi/metabolismo , Azufre/química , VibraciónRESUMEN
We describe here an extraordinary purple-colored DNA ligase, LigFa, from the acidophilic ferrous iron-oxidizing archaeon Ferroplasma acidiphilum, a di-ferric enzyme with an extremely low pH activity optimum. Unlike any other DNA ligase studied to date, LigFa contains two Fe(3+)-tyrosinate centers and lacks any requirement for either Mg(2+) or K(+) for activity. DNA ligases from closest phylogenetic and ecophysiological relatives have normal pH optima (6.0-7.5), lack iron, and require Mg(2+)/K(+) for activity. Ferric iron retention is pH-dependent, with release resulting in partial protein unfolding and loss of activity. Reduction of the Fe(3+) to Fe(2+) results in an 80% decrease in DNA substrate binding and an increase in the pH activity optimum to 5.0. DNA binding induces significant conformational change around the iron site(s), suggesting that the ferric irons of LigFa act both as structure organizing and stabilizing elements and as Lewis acids facilitating DNA binding at low pH.
Asunto(s)
ADN Ligasas/metabolismo , Hierro/metabolismo , Thermoplasmales/enzimología , Secuencia de Aminoácidos , ADN/metabolismo , ADN Ligasa (ATP) , ADN Ligasas/química , Concentración de Iones de Hidrógeno , Datos de Secuencia Molecular , Mutación , Conformación Proteica , Proteínas Recombinantes/metabolismo , Espectroscopía de Mossbauer , Especificidad por SustratoRESUMEN
The MEP pathway for the biosynthesis of isoprene units is present in most pathogenic bacteria, in the parasite responsible for malaria, and in plant plastids. This pathway is absent in animals and is accordingly a target for the development of antimicrobial drugs. LytB, also called IspH, the last enzyme of this pathway catalyzes the conversion of (E)-4-hydroxy-3-methylbut-2-enyl diphosphate (HMBPP) into a mixture of isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP) using an oxygen sensitive iron sulfur cluster. The exact nature of this iron sulfur cluster is still a matter of debate. We have used (57)Fe Mössbauer spectroscopy to investigate the LytB cluster in whole E. coli cells and in the anaerobically purified enzyme: In LytB an unusual [4Fe-4S](2+) cluster is attached to the protein by three conserved cysteines and contains a hexacoordinated iron linked to three sulfurs of the cluster and three additional oxygen or nitrogen ligands.
Asunto(s)
Eritritol/análogos & derivados , Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/metabolismo , Hierro , Oxidorreductasas/química , Oxidorreductasas/metabolismo , Fosfatos de Azúcar/metabolismo , Azufre , Terpenos/metabolismo , Anaerobiosis , Eritritol/metabolismo , Escherichia coli/enzimología , Escherichia coli/genética , Espectroscopía de MossbauerRESUMEN
Deoxyhypusine hydroxylase (DOHH) is a non-heme diiron enzyme involved in the posttranslational modification of a critical lysine residue of eukaryotic translation initiation factor 5A (eIF-5A) to yield the unusual amino acid residue hypusine. This modification is essential for the role of eIF-5A in translation and in nuclear export of a group of specific mRNAs. The diiron center of human DOHH (hDOHH) forms a peroxo-diiron(III) intermediate (hDOHHperoxo) when its reduced form reacts with O2. hDOHHperoxo has a lifetime exceeding that of the peroxo intermediates of other diiron enzymes by several orders of magnitude. Here we report the 1.7-Å crystal structures of hDOHHperoxo and a complex with glycerol. The structure of hDOHHperoxo reveals the presence of a µ-1,2-peroxo-diiron(III) species at the active site. Augmented by UV/Vis and Mössbauer spectroscopic studies, the crystal structures offer explanations for the extreme longevity of hDOHHperoxo and illustrate how the enzyme specifically recognizes its only substrate, deoxyhypusine-eIF-5A.
Asunto(s)
Lisina/análogos & derivados , Oxigenasas de Función Mixta/química , Oxigenasas de Función Mixta/metabolismo , Factores de Iniciación de Péptidos/metabolismo , Proteínas de Unión al ARN/metabolismo , Dominio Catalítico , Cristalografía por Rayos X , Estabilidad de Enzimas , Glicerol/metabolismo , Humanos , Lisina/metabolismo , Oxigenasas de Función Mixta/genética , Modelos Moleculares , Simulación del Acoplamiento Molecular , Factores de Iniciación de Péptidos/química , Proteínas de Unión al ARN/química , Especificidad por Sustrato , Factor 5A Eucariótico de Iniciación de TraducciónRESUMEN
The vulnerability of the dopaminergic neurons of the substantia nigra (SN) in Parkinson's disease has been related to the presence of the pigment neuromelanin (NM) in these neurons. It is hypothesised that NM may act as an endogenous storage molecule for iron, an interaction suggested to influence free radical production. The current study quantified and characterised the interaction between NM and iron. Iron-binding studies demonstrated that both NM and synthetically-produced dopamine melanin contain equivalent numbers of high and low-affinity binding sites for iron but that the affinity of NM for iron is higher than that of synthetic melanin. Quantification of the total iron content in iron-loaded NM and synthetic melanin demonstrated that the iron-binding capacity of NM is 10-fold greater than that of the model melanin. This data was in agreement with the larger iron cluster size demonstrated by Mössbauer spectroscopy in the native pigment compared with the synthetic melanin. These findings are consistent with the hypothesis that NM may act as an endogenous iron-binding molecule in dopaminergic neurons of the SN in the human brain. The interaction between NM and iron has implications for disorders such as Parkinson's disease where an increase in iron in the SN is associated with increased indices of oxidative stress.
Asunto(s)
Hierro/metabolismo , Melaninas/metabolismo , Enfermedad de Parkinson/metabolismo , Sustancia Negra/química , Humanos , Melaninas/síntesis química , Melaninas/farmacología , Espectroscopía de MossbauerRESUMEN
The binuclear iron complex [{Fe("EtN(2)S(2)")}(2)] (1b, "EtN(2)S(2)" = N,N'-diethyl-3,7-diazanonane-1,9-dithiolate) was prepared from the free ligand and ferrous bis[bis(trimethylsilyl)amide] in toluene. In dichloromethane 1b reacts with the [Fe(4)S(4)I(4)](2-) cubane cluster to displace two iodo ligands and to form the neutral hexanuclear cluster [{Fe("EtN(2)S(2)")}(2)Fe(4)S(4)I(2)] (2), which is isolated as black crystals in 87% yield. As elucidated by an X-ray structure analysis, 2 contains the novel hexanuclear stair-like [Fe(6)(&mgr;(3)-S)(4)(&mgr;(2)-SR)(4)] core, which exhibits crystallographic inversion symmetry. The compound crystallizes as a solvate with two molecules of CH(2)Cl(2) per formula unit in the monoclinic space group P2(1)/n with a = 1570.5(2) pm, b = 1060.2(1) pm, c = 1604.0(2) pm, beta = 114.93(1) degrees, and Z = 2. In the aprotic polar solvents DMF, 1,2-propylenecarbonate, and DMSO, 2 dissolves with decomposition and formation of the cluster [{Fe("EtN(2)S(2)")}(2)Fe(4)S(5)] (3), which is isolated as black needles from DMF. 3.2DMF crystallizes in the triclinic space group P&onemacr; with a = 950.9(1) pm, b = 1086.0(1) pm, c = 2381.5(2) pm, alpha = 101.81(1) degrees, beta = 91.94(1) degrees, gamma = 97.01(1) degrees, and Z = 2. The neutral compound contains a nest-like [Fe(6)(&mgr;(4)-S)(&mgr;(3)-S)(2)(&mgr;(2)-S)(2)(&mgr;(2)-SR)(4)] core of idealized C(2)(v)() symmetry that is closely related to that of other well-known clusters, e.g., the cluster anion [Fe(6)S(9)(SR)(2)](4-). The zero-field (57)Fe Mössbauer spectrum of 3 is in accordance with four Fe(2.5+)S(4) centers (delta = 0.46 mm/s; DeltaE(Q) = 1.14 mm/s) and two N(2)S(3)-bound high-spin Fe(2+) sites (delta = 0.83 mm/s; DeltaE(Q) = 3.64 mm/s). A total cluster spin of 0 is deduced from the Mössbauer spectrum at 4.2 K and 5.3 T, which yields magnetic splitting from the applied field only. For 2, three subspectra are observed in the Mössbauer spectrum (a, delta = 0.45 mm/s, DeltaE(Q) = 1.05 mm/s; b, delta = 0.55 mm/s, DeltaE(Q) = 1.61 mm/s, c, delta = 0.80 mm/s; DeltaE(Q) = 3.83 mm/s) reflecting different coordination environments of the iron atoms rather than different oxidation states. The electrochemical properties of 1b, 2, and 3 were determined by cyclic voltammetry. 1b can be quasi-reversibly oxidized in dichloromethane solution at -75 mV (vs SCE). Whereas 2 shows only an irreversible redox behavior in N,N'-dimethylimidazolidin-2-one solution, 3 in the same solvent can be quasi-reversibly reduced in two consecutive steps at -830 and -1630 mV (vs SCE) to the dianion, which consists entirely of Fe(II).
RESUMEN
NAD is an important cofactor and essential molecule in all living organisms. In many eubacteria, including several pathogens, the first two steps in the de novo synthesis of NAD are catalyzed by l-aspartate oxidase (NadB) and quinolinate synthase (NadA). Despite the important role played by these two enzymes in NAD metabolism, many of their biochemical and structural properties are still largely unknown. In the present study, we cloned, overexpressed and characterized NadA and NadB from Bacillus subtilis, one of the best studied bacteria and a model organism for low-GC Gram-positive bacteria. Our data demonstrated that NadA from B. subtilis possesses a [4Fe-4S]2+ cluster, and we also identified the cysteine residues involved in the cluster binding. The [4Fe-4S]2+ cluster is coordinated by three cysteine residues (Cys110, Cys230, and Cys320) that are conserved in all the NadA sequences reported so far, suggesting a new noncanonical binding motif that, on the basis of sequence alignment studies, may be common to other quinolinate synthases from different organisms. Moreover, for the first time, it was shown that the interaction between NadA and NadB is not species-specific between B. subtilis and Escherichia coli.
Asunto(s)
Aminoácido Oxidorreductasas/química , Bacillus subtilis/enzimología , Complejos Multienzimáticos/química , Aminoácido Oxidorreductasas/genética , Proteínas Bacterianas/química , Proteínas Bacterianas/genética , Sitios de Unión , Clonación Molecular , Secuencia Conservada , Cisteína , Proteínas de Escherichia coli , Proteínas Hierro-Azufre , Complejos Multienzimáticos/genética , Especificidad de la EspecieRESUMEN
The vibrational contribution to DeltaS of the low-spin ((3)T(1)) to high-spin ((5)E) spin transition in two 3d(4) octahedral systems [Mn(III)(pyrol)(3)tren] and [Cr(depe)(2)I(2)] have been estimated by means of DFT calculations (B3LYP/CEP-31G) of the vibrational normal-modes frequencies. The obtained value at the transition temperature for the Mn(iii) complex is DeltaS(vib)(44 K) = 6.3 J K(-1) mol(-1), which is comparable with the proposed Jahn-Teller contribution of R ln3 = 9.1 J K(-1) mol(-1) and which is approximately half of the experimentally determined 13.8 J K(-1) mol(-1). The corresponding value for the Cr(ii) complex is DeltaS(vib)(171.45 K) = 46.5 J K(-1) mol(-1), as compared to the experimental value of 39.45 J K(-1) mol(-1). The analysis of the vibrational normal modes reveals that for the d(4) systems under study, contrary to Fe(ii) d(6) systems, not all metal-ligand stretching vibrations make a contribution. For the Mn(iii) complex, the only vibration that contributes to DeltaS(vib) involve the nitrogens occupying the Jahn-Teller axis, while in the case of Cr(ii) the contributing vibrations involve the Cr-I bonds. Low-frequency modes due to ring vibrations, metal-ligand bending and movement of the molecule as a whole also contribute to the vibrational entropy associated with the spin transition.
RESUMEN
High resolution crystal structures of myoglobin in the pH range 5.2-8.7 have been used as models for the peroxide-derived compound II intermediates in heme peroxidases and oxygenases. The observed Fe-O bond length (1.86-1.90 A) is consistent with that of a single bond. The compound II state of myoglobin in crystals was controlled by single-crystal microspectrophotometry before and after synchrotron data collection. We observe some radiation-induced changes in both compound II (resulting in intermediate H) and in the resting ferric state of myoglobin. These radiation-induced states are quite unstable, and compound II and ferric myoglobin are immediately regenerated through a short heating above the glass transition temperature (<1 s) of the crystals. It is unclear how this influences our compound II structures compared with the unaffected compound II, but some crystallographic data suggest that the influence on the Fe-O bond distance is minimal. Based on our crystallographic and spectroscopic data we suggest that for myoglobin the compound II intermediate consists of an Fe(IV)-O species with a single bond. The presence of Fe(IV) is indicated by a small isomer shift of delta = 0.07 mm/s from Mössbauer spectroscopy. Earlier quantum refinements (crystallographic refinement where the molecular-mechanics potential is replaced by a quantum chemical calculation) and density functional theory calculations suggest that this intermediate H species is protonated.
Asunto(s)
Cristalografía por Rayos X/métodos , Hierro/química , Mioglobina/química , Oxígeno/química , Espectroscopía de Mossbauer/métodos , Espectrometría Raman/métodos , Animales , Hemo/química , Caballos , Concentración de Iones de Hidrógeno , Conformación Molecular , Miocardio/metabolismo , Peróxidos/química , Protones , Teoría CuánticaRESUMEN
Heptanuclear metal-centered, six-membered, mixed-valent, heterometallic wheels 1-3 of iron, manganese, and indium were prepared in a one-pot reaction from N-benzyldiethanolamine (H2L(1)), cesium carbonate, [PPh4]2[MnCl4], and FeCl3 or InCl3. All three complexes were characterized by the combination of elemental analysis, FAB mass spectroscopy, X-ray diffraction and cyclic voltammetry and in the case of 1 additionally by Mössbauer spectroscopy. In 1, four Mn(II) ions in the periphery are arranged in pairs alternating with one Fe(III) ion each, with an Fe(III) ion located in the center. In 2, three Mn(II) ions alternate with three In(III) ions, whereas in 3, four In(III) ions are arranged in pairs and alternate with one Mn(II) ion each. In 2 and 3 an Mn(II) ion is encapsulated in the center.
Asunto(s)
Bencilaminas/química , Metales/química , Compuestos Organometálicos/síntesis química , Indicadores y Reactivos , Indio/química , Hierro/química , Manganeso/química , Estructura Molecular , Espectroscopía de Mossbauer , Análisis Espectral , Difracción de Rayos XRESUMEN
The oxygen regulator Fnr is part of the regulatory cascade in Bacillus subtilis for the adaptation to anaerobic growth conditions. In vivo complementation experiments revealed the essential role of only three cysteine residues (C227, C230, C235) at the C-terminus of B. subtilis Fnr for the transcriptional activation of the nitrate reductase operon (narGHJI) and nitrite extrusion protein gene (narK) promoters. UV/VIS, electron paramagnetic spin resonance (EPR) and Mössbauer spectroscopy experiments in combination with iron and sulphide content determinations using anaerobically purified recombinant B. subtilis Fnr identified the role of these three cysteine residues in the formation of one [4Fe-4S]2+ cluster per Fnr molecule. The obtained Mössbauer parameters are supportive for a [4Fe-4S]2+ cluster with three cysteine ligated iron sites and one non-cysteine ligated iron site. Gel filtration experiments revealed a stable dimeric structure for B. subtilis Fnr which is independent of the presence of the [4Fe-4S]2+ cluster. Gel mobility shift and in vitro transcription assays demonstrated the essential role of an intact [4Fe-4S]2+ cluster for promoter binding and transcriptional activation. An amino acid exchange introduced in the proposed alphaD-helix of B. subtilis Fnr (G149S) abolished its in vivo and in vitro activities indicating its importance for intramolecular signal transduction. The clear differences in the localization and coordination of the [4Fe-4S] cluster and in the organization of the oligomeric state between Escherichia coli and B. subtilis Fnr indicate differences in their mode of action.
Asunto(s)
Bacillus subtilis/genética , Proteínas Bacterianas/química , Regulación Bacteriana de la Expresión Génica , Proteínas Hierro-Azufre/química , Transactivadores/química , Secuencia de Aminoácidos , Proteínas de Transporte de Anión/genética , Bacillus subtilis/metabolismo , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Cisteína/química , Cisteína/genética , ADN Bacteriano/metabolismo , Dimerización , Ensayo de Cambio de Movilidad Electroforética , Proteínas Hierro-Azufre/genética , Proteínas Hierro-Azufre/metabolismo , Datos de Secuencia Molecular , Nitrato-Reductasa/genética , Transportadores de Nitrato , Operón/genética , Oxígeno/metabolismo , Alineación de Secuencia , Transactivadores/genética , Transactivadores/metabolismo , Transcripción GenéticaRESUMEN
The synthesis and characterization of [FeII(trim)2]Cl2 (2), [FeII(trim)2]Br2MeOH (3), and [FeII(trim)2]I2MeOH (4), including the X-ray crystal structure determinations of 2 (50 and 293 K) and 4 (293 K), have been performed and their properties have been examined. In agreement with the magnetic susceptibility results, the Mössbauer data show the presence of high-spin (HS) to low-spin (LS) crossover with a range of T1/2 larger than 300 K (from approximately 20 K for [FeII(trim)2]F2 (1) to approximately 380 K for 4). All complexes in this series include the same [Fe(trim)2]2+ complex cation: the ligand field comprises a constant contribution from the trim ligands and a variable one originating from the out-of-sphere anions, which is transmitted to the metal center by the connecting imidazole rings and hydrogen bonds. The impressive variation in the intrinsic characteristics of the spin-crossover (SCO) phenomenon in this series is then interpreted as an inductive effect of the anions transmitted to the nitrogen donors through the hydrogen bonds. Based on this qualitative analysis, an increased inductive effect of the out-of-sphere anion corresponds to a decreased SCO temperature T1/2, in agreement with the experimental results. Electronic structure calculations with periodic boundary conditions have been performed that show the importance of intermolecular effects in tuning the ligand field, and thus in determining the transition temperature. Starting with the geometries obtained from the X-ray studies, the [FeII(trim)2]X2 complex molecules 1-4 have been investigated both for the single molecules and the crystal lattices with the local density approximation of density functional theory. The bulk geometries of the complex cations deduced from the X-ray studies and those calculated are in fair agreement for both approaches. However, the trend observed for the transition temperatures of 1-4 disagrees with the trend for the spin-state splittings ES (difference EHS-ELS between the energy of the HS and LS isomers) calculated for the isolated molecules, whereas it agrees with the trend for ES calculated with periodic boundary conditions. The latter calculations predict the strongest stabilization of the HS state for the fluoride complex, which actually is essentially HS above T=50 K, while the most pronounced stabilization of the LS state is predicted for 4, in line with the experimental results.
RESUMEN
Star-shaped complex [Fe(III)[Fe(III)(L1)2]3] (3) was synthesized starting from N-methyldiethanolamine H2L1 (1) and ferric chloride in the presence of sodium hydride. For 3, two different high-spin iron(III) ion sites were confirmed by Mössbauer spectroscopy at 77 K. Single-crystal X-ray structure determination revealed that 3 crystallizes with four molecules of chloroform, but, with only three molecules of dichloromethane. The unit cell of 3.4CHCl3 contains the enantiomers (delta)-[(S,S)(R,R)(R,R)] and (lambda)-[(R,R)(S,S)(S,S)], whereas in case of 3.3CH2Cl2 four independent molecules, forming pairs of the enantiomers [lambda-(R,R)(R,R)(R,R)]-3 and [lambda-(S,S)(S,S)(S,S)]-3, were observed in the unit cell. According to SQUID measurements, the antiferromagnetic intramolecular coupling of the iron(III) ions in 3 results in a S = 10/2 ground state multiplet. The anisotropy is of the easy-axis type. EPR measurements enabled an accurate determination of the ligand-field splitting parameters. The ferric star 3 is a single-molecule magnet (SMM) and shows hysteretic magnetization characteristics below a blocking temperature of about 1.2 K. However, weak intermolecular couplings, mediated in a chainlike fashion via solvent molecules, have a strong influence on the magnetic properties. Scanning tunneling microscopy (STM) and scanning tunneling spectroscopy (STS) were used to determine the structural and electronic properties of star-type tetranuclear iron(III) complex 3. The molecules were deposited onto highly ordered pyrolytic graphite (HOPG). Small, regular molecule clusters, two-dimensional monolayers as well as separated single molecules were observed. In our STS measurements we found a rather large contrast at the expected locations of the metal centers of the molecules. This direct addressing of the metal centers was confirmed by DFT calculations.
RESUMEN
C-type cytochromes with histidine-methionine (His-Met) iron coordination play important roles in electron-transfer reactions and in enzymes. Low-temperature electron paramagnetic resonance (EPR) spectra of low-spin ferric cytochromes c can be divided into two groups, depending on the spread of g values: the normal rhombic ones with small g anisotropy and g(max) below 3.2, and those featuring large g anisotropy with g(max) between 3.3 and 3.8, also denoted as highly axial low spin (HALS) species. Herein we present the detailed magnetic properties of cytochrome c(553) from Bacillus pasteurii (g(max) 3.36) and cytochrome c(552) from Nitrosomonas europaea (g(max) 3.34) over the pH range 6.2 to 8.2. Besides being structurally very similar, cytochrome c(553) shows the presence of a minor rhombic species at pH 6.2 (6 %), whereas cytochrome c(552) has about 25 % rhombic species over pH 7.5. The detailed Mössbauer analysis of cytochrome c(552) confirms the presence of these two low-spin ferric species (HALS and rhombic) together with an 8 % ferrous form with parameters comparable to the horse cytochrome c. Both EPR and Mössbauer data of axial cytochromes c with His-Met iron coordination are consistent with an electronic (d(xy))(2) (d(xz))(2) (d(yz))(1) ground state, which is typical for Type I model hemes.
Asunto(s)
Grupo Citocromo c/química , Citocromos/química , Espectroscopía de Resonancia por Spin del Electrón/métodos , Histidina/química , Metionina/química , Espectroscopía de Mossbauer/métodos , Anisotropía , Bacillus/metabolismo , Biofisica/métodos , Electrones , Concentración de Iones de Hidrógeno , Ligandos , Espectroscopía de Resonancia Magnética , Nitrosomonas/metabolismoRESUMEN
Crystalline samples of four low-spin Fe(III) octaalkyltetraphenylporphyrinate and two low-spin Fe(III) tetramesitylporphyrinate complexes, all of which are models of the bis-histidine-coordinated cytochromes of mitochondrial complexes II, III, and IV and chloroplast complex b(6)f, and whose molecular structures and EPR spectra have been reported previously, have been investigated in detail by Mössbauer spectroscopy. The six complexes and the dihedral angles between axial ligand planes of each are [(TMP)Fe(1-MeIm)(2)]ClO(4) (0 degree), paral-[(OMTPP)Fe(1-MeIm)(2)]Cl (19.5 degrees), paral-[(TMP)Fe(5-MeHIm)(2)]ClO(4) (26 degrees, 30 degrees for two molecules in the unit cell whose EPR spectra overlap), [(OETPP)Fe(4-Me(2)NPy)(2)]Cl (70 degrees), perp-[(OETPP)Fe(1-MeIm)(2)]Cl (73 degrees), and perp-[(OMTPP)Fe(1-MeIm)(2)]Cl (90 degrees). Of these, the first three have been shown to exhibit normal rhombic EPR spectra, each with three clearly resolved g-values, while the last three have been shown to exhibit "large g(max)" EPR spectra at 4.2 K. It is found that the hyperfine coupling constants of the complexes are consistent with those reported previously for low-spin ferriheme systems, with the largest-magnitude hyperfine coupling constant, A(zz), being considerably smaller for the "parallel" complexes (400-540 kG) than for the strictly perpendicular complex (902 kG), A(xx) being negative for all six complexes, and A(zz) and A(xx) being of similar magnitude for the "parallel" complexes (for example, for [(TMP)Fe(1-MeIm)(2)]Cl, A(zz) = 400 kG, A(xx) = -400 kG). In all cases, A(yy) is small but difficult to estimate with accuracy. With results for six structurally characterized model systems, we find for the first time qualitative correlations of g(zz), A(zz), and DeltaE(Q) with axial ligand plane dihedral angle Deltavarphi.
Asunto(s)
Citocromos/química , Compuestos Férricos/química , Hemo/análogos & derivados , Hemina/química , Espectroscopía de Resonancia por Spin del Electrón , Hemo/química , Membranas/química , Espectroscopía de MossbauerRESUMEN
The vibrational modes of the low-spin and high-spin isomers of the spin crossover complex [Fe(phen)(2)(NCS)(2)] (phen = 1,10-phenanthroline) have been measured by IR and Raman spectroscopy and by nuclear inelastic scattering. The vibrational frequencies and normal modes and the IR and Raman intensities have been calculated by density functional methods. The vibrational entropy difference between the two isomers, DeltaS(vib), which is--together with the electronic entropy difference DeltaS(el)--the driving force for the spin-transition, has been determined from the measured and from the calculated frequencies. The calculated difference (DeltaS(vib) = 57-70 J mol(-1) K(-1), depending on the method) is in qualitative agreement with experimental values (20-36 J mol(-1) K(-1)). Only the low energy vibrational modes (20% of the 147 modes of the free molecule) contribute to the entropy difference and about three quarters of the vibrational entropy difference are due to the 15 modes of the central FeN(6) octahedron.