High-spin Pentacoordinate Research Articles

A Quantum Chemistry View on Two Archetypical Paramagnetic Pentacoordinate Nickel(II) Complexes Offers a Fresh Look on Their NMR Spectra.

Quantum chemical methods for calculating paramagnetic NMR observables are becoming increasingly accessible and are being included in the inorganic chemistry practice. Here, we test the performance of these methods in the prediction of proton hyperfine shifts of two archetypical high-spin pentacoordinate nickel(II) complexes (NiSAL-MeDPT and NiSAL-HDPT), which, for a variety of reasons, turned out to be perfectly suited to challenge the predictions to the finest level of detail. For NiSAL-MeDPT, new NMR experiments yield an assignment that perfectly matches the calculations. The slightly different hyperfine shifts from the two “halves” of the molecules related by a pseudo-C2 axis, which are experimentally divided into two well-defined spin systems, are also straightforwardly distinguished by the calculations. In the case of NiSAL-HDPT, for which no X-ray structure is available, the quality of the calculations allowed us to refine its structure using as a starting template the structure of NiSAL-MeDPT.

Slow magnetic relaxation in a high-spin pentacoordinate Fe(iii) complex.

A mononuclear pentacoordinate iron(iii) complex shows slow magnetic relaxation with three relaxation channels. The high-frequency relaxation time of the order of microseconds is prolonged on cooling in accordance with the direct and Raman processes. The low frequency relaxation time is little dependent on temperature and varies in the range τ(LF) = 0.52-0.79 s.

Anomalous Stoichiometry, 3-D Bridged Triangular/Pentagonal Layered Structured Artificial Antiferromagnet for the Prussian Blue Analogue A3MnII5(CN)13 (A = NMe4, NEtMe3). A Cation Adaptive Structure.

The size of the organic cation dictates both the composition and the extended 3-D structure for hybrid organic/inorganic Prussian blue analogues (PBAs) of A aMnII b(CN) a+2 b (A = cation) stoichiometry. Alkali PBAs are typically cubic with both MC6 and M'N6 octahedral coordination sites and the alkali cation content depends on the M and M' oxidation states. The reaction of MnII(O2CCH3)2 and A+CN- (A = NMe4, NEtMe3) forms a hydrated material of A3MnII5(CN)13 composition. A3MnII5(CN)13 forms a complex, 3-D extended structural motif with octahedral and rarely observed square pyramidal and trigonal bipyramidal MnII sites with a single layer motif of three pentagonal and one triangular fused rings. A complex pattern of MnIICN chains bridge the layers. (NMe4)3MnII5(CN)13 possesses one low-spin octahedral and four high-spin pentacoordinate MnII sites and orders as an antiferromagnet at 11 K due to the layers being bridged and antiferromagnetically coupled by the nonmagnetic cyanides. These are rare examples of intrinsic, chemically prepared and controlled artificial antiferromagnets and have the advantage of having controlled uniform spacing between the layers as they are not physically prepared via deposition methods. A3Mn5(CN)13 (A = NMe4, NEtMe3) along with [NEt4]2MnII3(CN)8, [NEt4]MnII3(CN)7, and Mn(CN)2 form stoichiometrically related A aMnII b(CN) a+2 b ( a = 0, b = 1; a = 2, b = 3; a = 1, b = 3; and a = 3, b = 5) series possessing unprecedented stoichiometries and lattice motifs. These unusual structures and stoichiometries are attributed to the very ionic nature of the high-spin N-bonded MnII ion that enables the maximization of the attractive van der Waals interactions via minimization of void space via a reduced ∠MnNC. This A aMnII b(CN) a+2 b family of compounds are referred to as being cation adaptive in which size and shape dictate both the stoichiometry and structure.

Scorpionate-supported models of nickel-dependent superoxide dismutase

Ligation of nickel(II) by Trofimenko’s hydrotris(3,5-dimethyl-1-pyrazolyl)borate anion (Tp Me,Me) and zwitterionic organoxanthate or dithiocarbamate co-ligands affords neutral high-spin pentacoordinate complexes with formally trianionic N 3S 2 ligand fields, similar to that of the nickel-dependent superoxide dismutase active site. Given this analogy to NiSOD, the structure, dynamics, and redox properties of the product complexes were examined. X-ray structures revealed rotation of the dithioacid chelates against the scorpionate face, giving coordination geometries between square pyramidal and trigonal bipyramidal limits. The complexes accordingly adopt paramagnetic ( S = 1) d 8 electron configurations, but magnetic susceptibilities suggest partial isomerization to a diamagnetic state in solution. The complexes also exhibit quasi-reversible one-electron redox couples at potentials suitable for SOD activity.

Pentacoordinate and hexacoordinate ferric hemes in acid medium: EPR, UV–Vis and CD studies of the giant extracellular hemoglobin of Glossoscolex paulistus

The equilibrium complexity involving different axially coordinated hemes is peculiar to hemoglobins. The pH dependence of the spontaneous exchange of ligands in the extracellular hemoglobin from Glossoscolex paulistus was studied using UV–Vis, EPR, and CD spectroscopies. This protein has a complex oligomeric assembly with molecular weight of 3.1 MDa that presents an important cooperative effect. A complex coexistence of different species was observed in almost all pH values, except pH 7.0, where just aquomet species is present. Four new species were formed and coexist with the aquomethemoglobin upon acidification: (i) a “pure” low-spin hemichrome (Type II), also called hemichrome B, with an usual spin state ( d xy ) 2( d xz , d yz ) 3; (ii) a strong g max hemichrome (Type I), also showing an usual spin state ( d xy ) 2( d xz , d yz ) 3; (iii) a hemichrome with unusual spin state ( d xz , d yz ) 4( d xy ) 1 (Type III); (iv) and a high-spin pentacoordinate species. CD measurements suggest that the mechanism of species formation could be related with an initial process of acid denaturation. However, it is worth mentioning that based on EPR the aquomet species remains even at acidic pH, indicating that the transitions are not complete. The “pure” low-spin hemichrome presents a parallel orientation of the imidazole ring planes but the strong g max hemichrome is a HALS (highly anisotropic low-spin) species indicating a reciprocally perpendicular orientation of the imidazole ring planes. The hemichromes and pentacoordinate formation mechanisms are discussed in detail.

The role of the length and sequence of the linker domain of cytochrome b5 in stimulating cytochrome P450 2B4 catalysis.

Cytochrome b(5) (cyt b(5)) is a 15-kDa amphipathic protein with a cytosolic amino-terminal catalytic heme domain, which is anchored to the microsomal membrane by a hydrophobic transmembrane alpha-helix at its carboxyl terminus. These two domains are connected by an approximately 15-amino acid linker domain, Ser(90)-Asp(104), which has been modified by site-directed mutagenesis to investigate whether the length or sequence of the linker influences the ability of cyt b(5) to bind ferric cytochrome P450 2B4 and donate an electron to oxyferrous (cyt P450 2B4), thereby stimulating catalysis. Because shortening the linker by 8 or more amino acids markedly inhibited the ability of cyt b(5) to bind cyt P450 2B4 and stimulate catalysis by this isozyme, it is postulated 7 amino acids are sufficient to allow a productive interaction. All mutant cyts b(5) except the protein lacking the entire 15-amino acid linker inserted normally into the microsomal membrane. Alternatively, lengthening the linker by 16 amino acids, reversing the sequence of the amino acids in the linker, and mutating conserved linker residues did not significantly alter the ability of cyt b(5) to interact with cyt P450 2B4. A model for the membrane-bound cyt b(5)-cyt P450 complex is presented.

Formation of a bis(histidyl) heme iron complex in manganese peroxidase at high pH and restoration of the native enzyme structure by calcium.

Manganese peroxidase (MnP) from Phanerochaete chrysosporium undergoes a pH-dependent conformational change evidenced by changes in the electronic absorption spectrum. This high- to low-spin alkaline transition occurs at approximately 2 pH units lower in an F190I mutant MnP when compared to the wild-type enzyme. Herein, we provide evidence that these spectral changes are attributable to the formation of a bis(histidyl) heme iron complex in both proteins at high pH. The resonance Raman (RR) spectra of both ferric proteins at high pH are similar, indicating similar heme environments in both proteins, and resemble that of ferric cytochrome b(558), a protein that contains a bis-His iron complex. Upon reduction with dithionite at high pH, the visible spectra of both the wild-type and F190I MnP exhibit absorption maxima at 429, 529, and 558 nm, resembling the absorption spectrum of ferrous cytochrome b(558). RR spectra of the reduced wild-type and F190I mutant proteins at high pH are also similar to the RR spectrum of ferrous cytochrome b(558), further suggesting that the alkaline low-spin species is a bis(histidyl) heme derivative. No shift in the low-frequency RR bands was observed in 75% (18)O-labeled water, indicating that the low-spin species is most likely not a hydroxo-heme derivative. Electronic and RR spectra also indicate that addition of Ca(2+) to either the ferric or ferrous enzymes at high pH completely restores the high-spin pentacoordinate species. Other divalent metals, such as Mn(2+), Mg(2+), Zn(2+), or Cd(2+), do not restore the enzyme under the conditions studied.

Resonance Raman spectral properties and stability of manganese protoporphyrin IX cytochrome b5.

The structure and stability of cytochrome b5 reconstituted with manganese protoporphyrin IX instead of iron protoporphyrin IX has been investigated by resonance Raman spectroscopy and stopped-flow visible spectroscopy. The resonance Raman spectrum of MnIII cytochrome b5 was consistent with a high-spin hexacoordinate MnIII protoporphyrin IX structure that converted to a high-spin pentacoordinate structure at higher laser power. The resonance Raman spectrum of MnII cytochrome b5 indicated a high-spin pentacoordinate structure which was independent of laser power. Studies of the binding of MnIII protoporphyrin IX to apocytochrome b5 indicated that the MnIII-containing porphyrin bound much less tightly to the protein than did heme. Although the second-order rate constant at 20 degrees C for the association of heme with apocytochrome b5 (4.5 x 10(7) M(-1) s(-1)) was estimated to be only 1 order of magnitude higher than that with Mn protoporphyrin IX (3.3 x 10(6) M(-1) s(-1)), the dissociation of manganese substituted cytochrome b5 into the apoprotein and free Mn protoporphyrin IX occurs with a first-order rate constant of 1.2 x 10(-2) s(-1) at 20 degrees C while the dissociation of heme from cytochrome b5 at room temperature occurs 3 orders of magnitude more slowly with a first-order rate constant of 1.67 x 10(-5) s(-1) [Vergeres, G., Chen, D. Y., Wu, F.F., & Waskell, L. (1993) Arch. Biochem. Biophys. 305, 231-241]. The equilibrium dissociation constant for manganese-substituted cytochrome b5 increased with temperature from 4 nM at 20 degrees C to 14 nM at 37 degrees C. These results suggest that, in the reconstituted cytochrome P450 metabolizing system, especially in studies done with low protein concentrations (0.1 microM), and at elevated temperatures (37 degrees C), as much as 30% of the manganese-substituted cytochrome b5 may dissociate to free Mn-protoporphyrin IX and apocytochrome b5.

Structural characterization of oxidized dimeric Scapharca inaequivalvis hemoglobin by resonance Raman spectroscopy.

Resonance Raman spectra of the ferric homodimeric hemoglobin from Scapharca inaequivalvis have been measured over the pH range 5.8-8.3 in buffers of ionic strengths 0.01 and 0.1 M to determine the spin and coordination state of the iron atom. Three species contribute to the spectra: a low spin hexacoordinate, a high spin pentacoordinate, and a high spin hexacoordinate component. Optical absorption and EPR spectra measured under the same conditions allowed the identification of the ligands in the sixth coordination position, namely the distal histidine in the low spin derivative and a water molecule in the high spin one. The relative concentrations of these three species depend on pH in an unusual way. Thus, the aquomet derivative is present over the whole pH range, albeit in small amounts as most of the hemoglobin converts to the low spin hemichrome at acid pH values and to the pentacoordinate derivative at neutral and slightly alkaline ones. The formation of a pentacoordinate heme as the pH is increased has not been reported previously for other myoglobins and hemoglobins. Low ionic strength and high protein concentration favor the formation of the high spin pentacoordinate species, while at high ionic strength and low protein concentration the low spin hexacoordinate species prevails. Ionization of the iron-bound water molecule occurs at pH > or = 9.3; accordingly, signals from the hydroxyl derivative were not observed in the Raman spectra over the pH range studied.

Modeling low-pH hemoproteins

A tetracoordinate ferrous heme (iron-porphyrin) has been proposed as an intermediate at low pH (less than 3.0) for respiratory hemoproteins, peroxidases, and model heme complexes. This intermediate is believed to arise via protonation of the N(epsilon) atom of the proximal histidine and consequent cleavage of the Fe-N(epsilon) bond. To establish a spectral signature for the proposed low-pH tetracoordinate species, we have obtained Soret excitation resonance Raman spectra on samples of crystallographically defined, tetracoordinate iron(II)-octaethylporphyrin (Fe.OEP; S = 1). The high-frequency (greater than or equal to 900 cm-1) resonance Raman spectral features of Fe.OEP are clearly distinct from those of high-spin pentacoordinate or low-spin hexacoordinate ferrous hemes. Rather, they are at frequencies more typically observed for low-spin hexacoordinate ferric porphyrins. Comparative spectral analysis of tetracoordinate Fe.OEP and other proposed tetracoordinate ferrous hemes (e.g. iron(II)-protoporphyrin IX) demonstrates little or no macrocycle effect on the resonance Raman frequencies above 900 cm-1. This work thus serves to provide a testable spectral signature by which the existence of the proposed tetracoordinate biological intermediate may be verified and by which its functional significance may be tested.

Comparative Spectral Analysis of Mammalian, Fungal, and Bacterial Catalases: Resonance Raman Evidence for Iron-Tyrosinate Coordination

Resonance Raman spectra are reported for catalases from bovine liver, the ascomycete fungus Aspergillus niger, and the bacterium Micrococcus luteus. The vibrational frequencies of the oxidation-, spin-, and coordination number-sensitive spectral bands are indicative of high spin pentacoordinate hemes in the resting ferric enzymes of each of these organisms. This result is in accord with the crystal structure of bovine catalase (Fita, I., and Rossmann, M.G. (1985) J. Mol. Biol. 185, 21–37). In contrast, the crystallographic study of catalase from the ascomycete Penicillium vitale (Vainshtein, B. K., Melik-Adamyan, W. R., Barynin, V. V., Vagin, A.A., Grebenko, A. I., Borisov, V. V., Bartels, K. S., Fita, I., and Rossmann, M. G. (1986) J. Mol. Biol. 188, 49–61) showed electron density on the distal side of the heme which could imply the presence of a sixth ligand, possibly a water molecule. However, both of these crystallographic studies showed the proximal ligand in catalase to be a tyrosine. The present study confirms tyrosinate coordination in each of the three catalases from the appearance of selected resonance-enhanced tyrosine vibrational modes. The most characteristic band is the tyrosinate ring mode at ∼ 1612 cm−1 which is maximally enhanced with 488.0 nm excitation. The appearance of tyrosinate modes at 1607 and 1245 cm−1 in the resonance Raman spectra of M. luteus cyano catalase serves to identify tyrosine as an axial ligand in bacterial as well as eukaryotic catalases. Unlike non-heme iron tyrosinate proteins, whose resonance Raman spectra are dominated by several intense bands diagnostic of tyrosine ligation, the heme-linked tyrosine modes are not easily distinguished from the large number of porphyrin vibrations.

On the Mechanism of Action of Cytochrome P-450: Spectral intermediates in the reaction with iodosobenzene and its derivatives

Abstract Cytochrome P-450 is known to catalyze the following oxygen transfer reaction: RH + PhIO----ROH + PhI where RH represents a variety of hydroxylatable substrates and PhIO a variety of iodosobenzene derivatives that serve as oxygen donors, and neither molecular oxygen nor an external electron donor is required. To determine whether the cytochrome functions in such reactions by a peroxidase-type mechanism, the kinetics of its interaction with a variety of substituted iodosobenzenes and iodobenzene diacetates have been determined by stopped flow spectrophotometry. The reaction of phenobarbital-induced rabbit liver microsomal cytochrome P-450 form 2 with iodosobenzenes or iodobenzene diacetates leads to the reversible formation of three spectral intermediates, termed E, F, and G. Complex E is characterized by a type I difference spectrum, representing the iodosobenzene-dependent partial shift of the low spin hexacoordinate form of the ferric enzyme to the high spin pentacoordinate form, F represents a transient intermediate whose spectrum cannot be determined for kinetic reasons, and G represents a blue-shifted intermediate with an absorption maximum at about 393 nm in the absolute spectrum. The striking and principal feature of these observations is that the spectrum of Complex G does not vary with structural differences in the iodosobenzene derivatives, in contrast to the transient species observed in previous studies in this laboratory in the reaction between cytochrome P-450 form 2 and aromatic peroxy compounds. Complex G exhibits the spectral properties one might anticipate for an iron-oxo intermediate containing only one oxygen atom derived from the starting iodosobenzene.

Spectral characterization of manganese peroxidase, an extracellular heme enzyme from the lignin-degrading basidiomycete, Phanerochaete chrysosporium.

Manganese peroxidase (MnP) is a component of the lignin degradation system of the basidiomycetous fungus, Phanerochaete chrysosporium. This novel MnII-dependent extracellular enzyme (Mr = 46,000) contains a single protoporphyrin IX prosthetic group and oxidizes phenolic lignin model compounds as well as a variety of other substrates. To elucidate the heme environment of this enzyme, we have studied its electron paramagnetic resonance and resonance Raman spectroscopic properties. These studies indicate that the native enzyme is predominantly in the high-spin ferric form and has a histidine as fifth ligand. The reduced enzyme has a high-spin, pentacoordinate ferrous heme. Fluoride and cyanide readily bind to the sixth coordination position of the heme iron in the native form, thereby changing MnP into a typical high-spin, hexacoordinate fluoro adduct or a low-spin, hexacoordinate cyano adduct, respectively. EPR spectra of 14NO- and 15NO-adducts of ferrous MnP were compared with those of horseradish peroxidase (HRP); the presence of a proximal histidine ligand was confirmed from the pattern of superhyperfine splittings of the NO signals centered at g approximately equal to 2.005. The appearance of the FeII-His stretch at approximately 240 cm-1 and its apparent lack of deuterium sensitivity suggest that the N delta proton of the proximal histidine of the enzyme is more strongly hydrogen bonded than that of oxygen carrier globins and that this imidazole ligand may be described as having a comparatively strong anionic character. Although resonance Raman frequencies for the spin- and coordination-state marker bands of native MnP, nu 3 (1487), nu 19 (1565), and nu 10 (1622 cm-1), do not fall into frequency regions expected for typical penta- or hexacoordinate high-spin ferric heme complexes, ligation of fluoride produces frequency shifts of these bands very similar to those observed for cytochrome c peroxidase and HRP. Hence, these data strongly suggest that the iron in native MnP is predominantly high-spin pentacoordinate. Analysis of the Raman frequencies indicates that the dx2-y2 orbital of the native enzyme is at higher energy than that of metmyoglobin. These features of the heme in MnP must be favorable for the peroxidase catalytic mechanism involving oxidation of the heme iron to FeIV. Consequently, it is most likely that the heme environment of MnP resembles those of HRP, cytochrome c peroxidase, and lignin peroxidase.

A high-spin iodo-arsine ruthenium(III) complex

A metathetical reaction conducted between RuCl3(AsPh3)2(MeOH) and a large excess of NH4I afforded a high-spin pentacoordinate complex, RuI3(AsPh3)2 (high-spin /paramagnetic/triphenylarsine/iodide/ruthenium(III) complex).

POLARIZED SPECTRA OF THE TRIGONAL BIPYRAMIDAL CoNP3Br CHROMOPHORE

Abstract The electronic spectra of high-spin pentacoordinate cobalt(II) complexes have been tentatively assigned on the basis of a ligand field approach.1–3 A controversy has arisen about the assignment of the spectrum of a typical chromophore, CoN4Br, having strictly C 3v symmetry in the compound [CoBr(Me6tren)]Br, where Me6tren is N[CH2CH2N(CH3)2]3. This spectrum shows four distinct bands well separated from charge transfer bands at 5.7, 12.3, 16.1 and 19.2–19.8 kK.4 Ciampolini and Bertini2 assigned these bands to transitions from the 4 A 2(F) ground state to the 4 E(F), 4 E E, 4 A 2(P), and 4 E(P) excited states whereas Wood3 assigned the second band to a spin forbidden transition, the third band to a transition to the 4 E(F) level and the last band to the two transitions to the highest two levels. Because of the cubic symmetry of the crystal lattice of this compound,5 polarized spectra were of no aid in deciding the correct assignment.