A possible model of hemoprotein-peroxide complexes formed in an iron-tetraphenylporphyrin system

A ferric low-spin species with an anomalously small g3-g1 separation generated by the reaction of Fe(III)tetraphenylporphyrin with t-butylhydroperoxide in the presence of tatramethylammonium hydroxide was characterized by ESR spectroscopy. The reaction kinetics, investigated by monitoring ESR intensity, indicated that this low-spin complex is highly reactive and easily changed to non-heme type iron complexes. The rhombic parameters of this complex are very similar to those of heme-peroxide adducts such as [Fe(II)-hemoglobin-O2]- and [Fe(II)-horseradish peroxidase-O2]-.     

Spectroscopic studies on bleomycin-iron complexes with carbon monoxide, nitric oxide, isocyanide, azide, and cyanide and comparison with iron-porphyrin complexes

The bleomycin-iron complexes with CO, NO, C2H5NC, OH-, N-3, CN-, and CH3NH2 were characterized by electronic, ESR, 1H-NMR, and M?ssbauer spectroscopies and the findings were compared with the corresponding hemoprotein complexes. The 1H-NMR and M?ssbauer features for the CO and C2H5NC adducts of the bleomycin-Fe(II) complex are consistent with an S = 0 ferrous assignment. The OH-, CH3NH2, and N-3 adducts of the bleomycin-Fe(III) complex show the ESR, 1H-NMR, and M?ssbauer spectra typical of a low-spin Fe(III). The unique M?ssbauer parameters of the bleomycin-Fe(II)-NO complex demonstrate mixing between the NO pi- and the Fe 3d-orbitals. The magnitude of the proton chemical shifts over +/- 50 ppm indicates a high-spin ferric type for the bleomycin-Fe(III)-CN complex. The M?ssbauer parameters (delta EQ = 0.89 and delta = 0.48 mm/s) of the CN- adduct differ substantially from those of typical low-spin hemoprotein-cyanide complexes. Except for the CN- adduct, the M?ssbauer and crystal field parameters of these bleomycin-iron complexes are similar to those of the corresponding hemoprotein complexes.     

Models for coordination site of cytochrome P-450, characterization of hemin-thiolato complexes with S, O, and N donor ligands by electronic absorption and electron spin resonance spectra

Bisthiolato-hemin complexes exhibiting "two split Soret bands" at 370 and 460 nm, classified into "hyperporphyrin spectrum" was prepared with naturally occurring porphyrins (Fe(III)protoporphyrin IX and its dimethyl ester), thioglycolate esters, and tetramethylammonium hydroxide in organic solvents. The structure of the complexes was characterized by electronic absorption and electron spin resonance (ESR) spectrometries. These complexes were stable under air at room temperature, their apparent half-lives being about 30 min monitored by the intensities of the two Soret bands. Thus the bisthiolato-hemin complex containing thioglycolate ester was shown to be a model for the cytochrome P450(P450)-thiolato binding complex. Ligand exchange reactions of the bisthiolato-hemin complex with imidazole or methanol indicated that the intermediate species are stabilized as thiolato-hemin-imidazole or -methanol complexes. The latter intermediate complex was suggested to be a good model for low-spin ferric P450 as characterized by distinct beta- and alpha-bands at 530 and 560 nm, respectively, as well as a single Soret peak at approximately 410 nm. The result of the analysis on ESR g values and crystal field parameters for the bisthiolato-hemin, thiolato-hemin-imidazole, and thiolato-hemin-oxygen ligand complexes comparing with those for P450 itself and the ligand binding complexes revealed that the sixth ligand trans to the fifth thiolato ligand of the low-spin ferric P450 can be an oxygen atom of water molecule.     

Low-spin heme b(3) in the catalytic center of nitric oxide reductase from Pseudomonas nautica

Respiratory nitric oxide reductase (NOR) was purified from membrane extract of Pseudomonas (Ps.) nautica cells to homogeneity as judged by polyacrylamide gel electrophoresis. The purified protein is a heterodimer with subunits of molecular masses of 54 and 18 kDa. The gene encoding both subunits was cloned and sequenced. The amino acid sequence shows strong homology with enzymes of the cNOR class. Iron/heme determinations show that one heme c is present in the small subunit (NORC) and that approximately two heme b and one non-heme iron are associated with the large subunit (NORB), in agreement with the available data for enzymes of the cNOR class. Mo?ssbauer characterization of the as-purified, ascorbate-reduced, and dithionite-reduced enzyme confirms the presence of three heme groups (the catalytic heme b(3) and the electron transfer heme b and heme c) and one redox-active non-heme Fe (Fe(B)). Consistent with results obtained for other cNORs, heme c and heme b in Ps. nautica cNOR were found to be low-spin while Fe(B) was found to be high-spin. Unexpectedly, as opposed to the presumed high-spin state for heme b(3), the Mo?ssbauer data demonstrate unambiguously that heme b(3) is, in fact, low-spin in both ferric and ferrous states, suggesting that heme b(3) is six-coordinated regardless of its oxidation state. EPR spectroscopic measurements of the as-purified enzyme show resonances at the g ～ 6 and g ～ 2-3 regions very similar to those reported previously for other cNORs. The signals at g = 3.60, 2.99, 2.26, and 1.43 are attributed to the two charge-transfer low-spin ferric heme c and heme b. Previously, resonances at the g ～ 6 region were assigned to a small quantity of uncoupled high-spin Fe(III) heme b(3). This assignment is now questionable because heme b(3) is low-spin. On the basis of our spectroscopic data, we argue that the g = 6.34 signal is likely arising from a spin-spin coupled binuclear center comprising the low-spin Fe(III) heme b(3) and the high-spin Fe(B)(III). Activity assays performed under various reducing conditions indicate that heme b(3) has to be reduced for the enzyme to be active. But, from an energetic point of view, the formation of a ferrous heme-NO as an initial reaction intermediate for NO reduction is disfavored because heme [FeNO](7) is a stable product. We suspect that the presence of a sixth ligand in the Fe(II)-heme b(3) may weaken its affinity for NO and thus promotes, in the first catalytic step, binding of NO at the Fe(B)(II) site. The function of heme b(3) would then be to orient the Fe(B)-bound NO molecules for the formation of the N-N bond and to provide reducing equivalents for NO reduction.     

M?ssbauer and EPR studies of the photoactivation of nitrile hydratase.

The alphabeta dimer of active nitrile hydratase from Rhodococcus sp. R312 contains one low-spin ferric ion that is coordinated by three Cys residues, two N-amide groups from the protein backbone, and one OH(-). The enzyme isolated from bacteria grown in the dark is inactive and contains the iron site as a six-coordinate diamagnetic Fe-nitrosyl complex, called NH(dark). The active state can be obtained from the dark state by photolysis of the Fe-NO bond at room temperature. Activation is accompanied by the conversion of NH(dark) to a low-spin ferric complex, NH(light), exhibiting an S = (1)/(2) EPR signal with g values of 2.27, 2.13, and 1.97. We have characterized both NH(dark) and NH(light) with M?ssbauer spectroscopy. The z-axis of the 57Fe magnetic hyperfine tensor, A, of NH(light) was found to be rotated by approximately 45 degrees relative to the z-axis of the g tensor (g(z) = 1.97). Comparison of the A tensor of NH(light) with the A tensors of low-spin ferric hemes indicates a substantially larger degree of covalency for nitrile hydratase. We have also performed photolysis experiments between 2 and 20 K and characterized the photolyzed products by EPR and M?ssbauer spectroscopy. Photolysis at 4.2 K in the M?ssbauer spectrometer yielded a five-coordinate low-spin ferric species, NH(A), which converted back into NH(dark) when the sample was briefly warmed to 77 K. We also describe preliminary EPR photolysis studies that have yielded new intermediates.     

Hexaheme nitrite reductase from Desulfovibrio desulfuricans. M?ssbauer and EPR characterization of the heme groups

M?ssbauer and EPR spectroscopy were used to characterize the heme prosthetic groups of the nitrite reductase isolated from Desulfovibrio desulfuricans (ATCC 27774), which is a membrane-bound multiheme cytochrome capable of catalyzing the 6-electron reduction of nitrite to ammonia. At pH 7.6, the as-isolated enzyme exhibited a complex EPR spectrum consisting of a low-spin ferric heme signal at g = 2.96, 2.28, and 1.50 plus several broad resonances indicative of spin-spin interactions among the heme groups. EPR redox titration studies revealed yet another low-spin ferric heme signal at g = 3.2 and 2.14 (the third g value was undetected) and the presence of a high-spin ferric heme. M?ssbauer measurements demonstrated further that this enzyme contained six distinct heme groups: one high-spin (S = 5/2) and five low-spin (S = 1/2) ferric hemes. Characteristic hyperfine parameters for all six hemes were obtained through a detailed analysis of the M?ssbauer spectra. D. desulfuricans nitrite reductase can be reduced by chemical reductants, such as dithionite or reduced methyl viologen, or by hydrogenase under hydrogen atmosphere. Addition of nitrite to the fully reduced enzyme reoxidized all five low-spin hemes to their ferric states. The high-spin heme, however, was found to complex NO, suggesting that the high-spin heme could be the substrate binding site and that NO could be an intermediate present in an enzyme-bound form.     

Effect of ionizing radiation on haemoglobin: the oxy-derivative of haemoglobin Iwate

It is well established that exposure of oxyhaemoglobin to ionizing radiation results in remarkably selective electron addition to the (FeO2) unit, giving a novel species, (FeO2)-, in which the extra electron is largely localized on iron and dioxygen. This work has now been extended to haemoglobin (Hb.) Iwate. The haemoglobin M. Iwate used is a mutant haemoglobin having only Fe(III) alpha-chains by oxy beta-chains (alpha 2 Met beta 2 oxy). The haem iron atoms in the alpha-chains are coordinated in the fifth site by a proximal tyrosine in place of histidine. This unit is a high-spin Fe(III) with axial symmetry and prominent electron spin resonance (ESR) features in the g = 6 and g = 2 regions. On exposure to 60Co gamma-rays at 77 K, efficient electron addition occurred at both types of iron centre, giving Fe(II) and (FeO2)- units. The former was monitored by the decrease of the g = 6 feature for Fe(III) and the latter by the growth of g-features at 2.254 (gx), 2.149 (gy) and 1.967 (gz). These values are close to those for the FeO2- centre formed in the beta-chains of normal oxyhaemoglobin. On annealing above 77 K, two changes occurred: first there was a small but clear increase in gx and gy, followed by a marked reduction in gx and gy giving g-values close to those for the centre formed directly in the alpha-chains of the normal protein. Finally, this intermediate species gave a centre having gx = 2.310, gy = 2.180 and gz = 1.935. These values are typical of low-spin Fe(III) haemoglobin and are assigned to the protonated complex, Fe(III)O2H. Ultimately at ca. room temperature, this was converted into the high-spin, met-form, with a gain in the g = 6 feature. These results established that the beta-chain centre in Hb. Iwate behave in the same way as isolated beta-chains. They also confirm that electron addition to the oxy-units is facile, even in the presence of Fe(III) units in each tetramer. The results also confirm that electron capture to give (FeO2)- units is not followed by internal electron-transfer to give Fe(II) from the Fe(III) centres in the alpha-chains.     

Azide- and cyanide-binding to the Escherichia coli bd-type ubiquinol oxidase studied by visible absorption, EPR and FTIR spectroscopies.

Cytochrome bd-type ubiquinol oxidase contains two hemes b (b(558) and b(595)) and one heme d as the redox metal centers. To clarify the structure of the reaction center, we analyzed Escherichia coli cytochrome bd by visible absorption, EPR and FTIR spectroscopies using azide and cyanide as monitoring probes for the exogenous ligand binding site. Azide-binding caused the appearance of a new EPR low-spin signal characteristic of ferric iron-chlorin-azide species and a new visible absorption band at 647 nm. However, the bound azide ((14)N(3)) anti-symmetric stretching infrared band (2, 010.5 cm(-1)) showed anomalies upon (15)N-substitutions, indicating interactions with surrounding protein residues or heme b(595) in close proximity. The spectral changes upon cyanide-binding in the visible region were typical of those observed for ferric iron-chlorin species with diol substituents in macrocycles. However, we found no indication of a low-spin EPR signal corresponding to the ferric iron-chlorin-cyanide complexes. Instead, derivative-shaped signals at g = 3.19 and g = 7.15, which could arise from the heme d(Fe(3+))-CN-heme b(595)(Fe(3+)) moiety, were observed. Further, after the addition of cyanide, a part of ferric heme d showed the rhombic high-spin signal that coexisted with the g(z) = 2.85 signal ascribed to the minor heme b(595)-CN species. This indicates strong steric hindrance of cyanide-binding to ferric heme d with the bound cyanide at ferric heme b(595).     

Significant interaction between low-spin iron(III) site and pyrroloquinoline quinone in active center of nitrile hydratase

Nitrile hydratase, a unique non-heme iron enzyme, contains low-spin Fe(III) site and pyrroloquinoline quinone in the active center. The enzymatic activity of nitrile hydratase is strongly inhibited by typical carbonyl reagents such as phenylhydrazine and semicarbazide. Of special interest is the fact that these carbonyl reagents induced significant alteration of the ESR features of the native iron(III) enzyme at pH 7.2. In addition, the isobutyronitrile(inhibitor)-bound enzyme was also remarkably affected by phenylhydrazine and then the transformed ESR characteristics were different from those of phenylhydrazine-treated enzyme. The present results reveal the first evidence for an important interaction between the low-spin Fe(III) site and pyrroloquinoline quinone which is essential for the activity of nitrile hydratase.     

Heme spin states and peroxide-induced radical species in prostaglandin H synthase

We have examined the optical, magnetic circular dichroism, and electron paramagnetic resonance (EPR) spectra of pure ovine prostaglandin H synthase in its resting (ferric) and ferrous states and after addition of hydrogen peroxide or 15-hydroperoxyeicosatetraenoic acid. In resting synthase, the distribution of heme between high- and low-spin forms was temperature-dependent: 20% of the heme was low-spin at room temperature whereas 50% was low-spin at 12 K. Two histidine residues were coordinated to the heme iron in the low-spin species. Anaerobic reduction of the synthase with dithionite produced a high-spin ferrous species that had no EPR signals. Upon reaction with the resting synthase, both hydroperoxides quickly generated intense (20-40% of the synthase heme) and complex EPR signals around g = 2 that were accompanied by corresponding decreases in the intensity of the signals from ferric heme at g = 3 and g = 6. The signal generated by HOOH had a doublet at g = 2.003, split by 22 G, superimposed on a broad component with a peak at g = 2.085 and a trough at g = 1.95. The lipid hydroperoxide generated a singlet at g = 2.003, with a linewidth of 25 G, superimposed on a broad background with a peak at g = 2.095 and a trough around g = 1.9. These EPR signals induced by hydroperoxide may reflect synthase heme in the ferryl state complexed with a free radical derived from hydroperoxide or fragments of hydroperoxide.     

Study of the respiratory chain in Micrococcus luteus (lysodeikticus) by electron-spin-resonance spectroscopy

Low-temperature electron spin resonance spectroscopy was used to investigate the redox centres of Micrococcus luteus membranes. Three different types of iron-sulphur centres were distinguished. Two of these, a [4Fe-4S]3+-type cluster giving rise to a signal at g = 2.01 in the oxidized state and a [2Fe-2S] cluster with a spectrum at g = 2.03 and 1.93 in the reduced state, were attributable to succinate dehydrogenase. Another, generating signals in the reduced state at g = 2.027, 1.90 and 1.78 was identified as a 'Rieske' iron-sulphur centre. This latter cluster had a mid-point potential (pH 7.0) of +130 mV. In addition, signals characteristic of high-spin ferric haem (g = 6.20), low-spin ferric haem (g = 3.67, 3.36 and 3.01) and Cu2+ (g = 2.18 and 2.02) were also detected. The ferric-haem features, together with the Cu2+ and 'Rieske' centres, were enriched in membrane residues insoluble in Triton X-100, which are known from difference spectroscopy to contain cytochromes b-560, c-550 and a-601 (aa3 oxidase). The signals demonstrated by electron spin resonance for M. luteus membranes showed marked similarities to those documented for the complexes II, III, and IV of mitochondria. However, signals analogous to complex I (NADH-ubiquinone reductase) could not be demonstrated for M. luteus membranes.     

Synthetic analogs of the active site of cytochrome P-450cam characterization of thiolpeptide fragment-hemin complexes by optical and ESR spectrometries

Thiol-containing penta (Leu-Ala-Cys-Ser-Leu) and nona (Leu-Ala-Cys-Ser-Leu-Ile-Glu-Ser-Leu) peptides corresponding to residues 132-136 and 132-140, respectively, of apo-P450 from Psuedomonas putida were synthesized to examine heme-binding by the enzymes in the oxidized (ferric) form. The peptide-hemin complexes prepared in solution were characterized by their optical and ESR spectra. In these complexes without nitrogenous ligands, no ferric complexes in the low-spin state were observed, suggesting that simultaneous axial coordination of Cys-134 (thiolate) and Ser-139 (hydroxyl) of apo-P450cam to the heme is unlikely to occur. In the presence of nitrogenous ligands, such as py, Im and MeIm, the resulting complexes were good models of apo-P450cam-nitrogenous ligand adducts in the low-spin ferric state retaining a thiolate-Fe(III)-nitrogen axial coordination mode, as judged by their spectral pattern as well as by crystal field analysis of ESR g-values.     

Stimulatory effect of phosphate, ATP and ADP on iron transfer from Fe(III)-bleomycin to apotransferrin

1. The rate of ferric ion transfer from Fe(III)-bleomycin to apotransferrin was increased in the presence of orthophosphate, ATP and ADP, while AMP was without effect. 2. Ortho phosphate activation probably involves formation of a Fe(III)-bleomycin-phosphate complex. The optical absorption of Fe(III)-bleomycin at 450 nm is enhanced in the presence of phosphate. 3. ATP and ADP remove the ferric ion from the iron-drug complex; thus making the ferric ion readily available for uptake by apotransferrin. 4. Low concentrations of ATP, ADP and AMP, also enhance the 450 nm absorption of the iron-drug complex. Higher ATP and ADP concentrations reduce both the 450 and 384 nm absorption of Fe(III)-bleomycin.     

A magnetic study of acidic ferric hemoglobin

In this article, we have extensively studied and discussed the magnetic properties of acidic ferric hemoglobin and its isolated chains. The magnetic susceptibility, EPR and optical spectra of those samples were measured in the temperature region below 77 degrees K. By the magnetic susceptibility measurements, it could be made clear that at an acidic pH value, both ferric hemoglobin and its isolated chains were constituted of a mixture of two spin states (high-spin state S = 5/2 and low-spin state S = 1/2) and the ratio of this mixture varied in each protein sample, but was independent of the temperature change below 77 degrees K. The co-existence of these two components could be ascertained by the observation of EPR spectra at liquid hydrogen temperature. Acidic ferric hemoglobin and its isolated chains exhibited the two components of EPR spectra which corresponded to their magnetic susceptibility, and it was found that the relaxation time of the low-spin state was longer than that of the high-spin state. The low-spin component of EPR spectra was almost undetectable at liquid nitrogen temperature. The three principal g values of this low-spin were gz = 2.80, gy = 2.20, and gx = 1.70. At alkaline pH values these low-spin components and the high-spin component of EPR spectra were displaced by the different low-spin spectra which corresponded to the ferric hemoglobin-hydroxide complex. It seems that the magnetic properties of the high-spin component are the same as the acidic ferric myoglobin, and the fine structure of the iron ion also seems to be same. Optical spectroscopy also gave similar magnetic properties which corresponded to the magnetic measurements.     

Low-temperature magnetic circular dichroism studies of the photoreaction of horseradish peroxidase compound I

Horseradish peroxidase (HRP) compound I is photolabile at all temperatures between room temperature and 4 K. The photoredox reaction has been studied in frozen glassy solutions by using optical absorption and magnetic circular dichroism spectra following photolysis of HRP compound I with visible-wavelength light at 4.2 and 77 K. The photochemical process is characterized as a concerted two-electron transfer reaction which results in the conversion of the Fe(IV) heme pi-cation radical species of HRP compound I into a low-spin Fe(III) heme species. This reaction occurs even when photolysis is carried out at 4.2 K. Spectra recorded between 4.2 and 80 K for the low-spin ferric hydroxide complex of HRP closely resemble the data measured for the photochemical product. The proposed mechanism for the photoreaction is (formula; see text) No evidence is found for the formation of an Fe(II) heme at these temperatures.     

Generation of Oxygen Radicals from Iron Complex of Orellanine, A Mushroom Nephrotoxin; Preliminary ESR and Spin-Trapping Stukes

Orellanine, [2,2'-bipyridine]-3,3',4,4'-tetrol-l,I'-dioxide, is the toxin responsible for the lethal nephrotoxicity of some Cortinarius mushrooms. Our present ESR and spin-trapping studies of the redox properties of the system of non-illuminated orellanine, ferrous iron and dioxygen contribute to understanding the molecular mechanism of its toxicity. UV-visible spectrophoto-metry, cyclic voltammetry and ESR in frozen medium showed the formation of a wine-red tris complex, Fe(III)Or₃. This ferric complex is easily reducible (E_P =-565 mV vs Ag/AgCl/3M KCl at pH7), involving a one-electron reversible process. Spin-trapping using DMPO is employed to detect the generation of super-oxide anion and hydroxyl radicals. The instantaneous one-electron oxidation of ferrous ions in the presence of the toxin under air is concomitant with dioxygen consumption as supported by dioxygen consumption. GSH involves the toxin and ferrous ions under air in a redox cycling process resulting in the production of glutathionyl and oxygen free radicals, observed for the first time with an iron complex of a mushroom toxin. In most cases, EDTA is not able to prevent the Fe(III)Or₃ and radical formation. The ortho-dihydroxylated groups borne by the di-N-oxidized bipyridine structure and not the bipyridine structure itself, are responsible for the formation of a stable ferric complex at pH 7, as they are for the generation of an apparently stable ortho-semiquinone anion radical. These one-electron mechanisms may play a major role in some of the known toxic effects of orellanine.     

Time-resolved absorption and magnetic circular dichroism spectroscopy of cytochrome c3 from Desulfovibrio.

The UV-visible absorption and magnetic circular dichroism (MCD) spectra of the ferric, ferrous, CO-ligated forms and kinetic photolysis intermediates of the tetraheme electron-transfer protein cytochrome c3 (Cc3) are reported. Consistent with bis-histidinyl axial coordination of the hemes in this Class III c-type cytochrome, the Soret and visible region MCD spectra of ferric and ferrous Cc3 are very similar to those of other bis-histidine axially coordinated hemeproteins such as cytochrome b5. The MCD spectra indicate low spin state for both the ferric (S = 1/2) and ferrous (S = 0) oxidation states. CO replaces histidine as the axial sixth ligand at each heme site, forming a low-spin complex with an MCD spectrum similar to that of myoglobin-CO. Photodissociation of Cc3-CO (observed photolysis yield = 30%) produces a transient five-coordinate, high-spin (S = 2) species with an MCD spectrum similar to deoxymyoglobin. The recombination kinetics of CO with heme Fe are complex and appear to involve at least five first-order or pseudo first-order rate processes, corresponding to time constants of 5.7 microseconds, 62 microseconds, 425 microseconds, 2.9 ms, and a time constant greater than 1 s. The observed rate constants were insensitive to variation of the actinic photon flux, suggesting noncooperative heme-CO rebinding. The growing in of an MCD signal characteristic of bis-histidine axial ligation within tens of microseconds after photodissociation shows that, although heme-CO binding is thermodynamically favored at 1 atm CO, binding of histidine to the sixth axial site competes kinetically with CO rebinding.     

Coordination structure and chemical reactivity of hemoprotein-butyl peroxide complex.

ESR and optical spectra ascribed to be the hemoprotein-butylperoxide complex were detected for the frozen aqueous solution containing whale met-Mb and t- or n-butylhydroperoxide at pH 10. The observed ESR and optical parameters of the complex were characteristic to those of six coordinate ferric low-spin complexes, having the butylperoxide anion at the axial position of heme. pH dependent ESR measurements demonstrated the formation of the complex in the biological pH regions (7.0). Time dependent ESR and optical measurements indicated that the complex may be one of the intermediate species in the processes of heme degradation reaction induced by butylperoxide.     

Effective DNA cleavage by bleomycin-vanadium(IV) complex plus hydrogen peroxide

It has been firstly found that the bleomycin-vanadyl(IV) complex is effectively capable of cleaving DNA in the presence of hydrogen peroxide. The 1:1 bleomycin-VO(IV) complex has been characterized by ESR and electronic absorption spectra, and its ESR parameters (go = 1.982 and Ao = 93.5 G) are indicative of VO(N5) coordination type for the metal-binding environment. The mode of nucleotide sequence cleavage induced by the present bleomycin-VO(IV)-H2O2 complex system was appreciably different from the corresponding Fe(III) complex system. Of special interest is the fact that the bleomycin-vanadium complex system more preferentially attacked G-A(5'----3') sequences than the bleomycin-iron complex system.     

Electron exchange in pyridine solutions of iron(III) protoporphyrin IX chloride

Proton magnetic resonance and absorption spectroscopy have been used to examine solutions of mixtures of reduced and oxidised iron protoporphyrin IX chloride in deuterated pyridine. The Fe(II) species are low spin but the Fe(III) complex is an equilibrium mixture of high and low spin forms. The movement to high field of the ring protons of the low-spin Fe(III) signals alone increases regularly with the amount of diamagnetic Fe(II) relative to the paramagnetic Fe(III) haem. The low spin Fe(III) must be in rapid exchange with the low-spin Fe(II) complex but not with the high-spin form. The addition of carbon monoxide to the Fe(II)/Fe(III) mixture effectively blocks electron exchange between the complexes as shown by a return of the proton resonances of the Fe(III) complex to positions seen in the absence of any Fe(II).