SEARCH · PubMed Health

Results for “Ferrous Compounds”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 199 records · Page 11Linked to original sources

PubMed2001

The effects of pH on the mechanism of hydrogen peroxide and lipid hydroperoxide consumption by myoglobin: a role for the protonated ferryl species.

Myoglobin catalyses the breakdown of lipid hydroperoxides (e.g., HPODE) during which the absorption band of the lipid conjugated diene (234 nm) is partially bleached. The constant for this process is strongly pH-dependent (k = 9.5 x 10(-3)s(-1), pH 7: k = 2.3 x 10(-1)s(-1), pH 5). This rate enhancement is not due to acid-induced changes in protein conformation or the involvement of protein-based radical species, as demonstrated by an almost identical pH dependence of the same reaction catalyzed by ferric haemin. The rate constants for ferryl formation and auto-reduction show different pH dependencies, with a pK of 8.3 for ferryl formation and a projected pK of 3.5 for ferryl auto-reduction. The pH dependence for the auto-reduction of the ferryl species is the same as that of the myoblobin catalyzed breakdown of HPODE. We propose that the protonated form of ferryl myoglobin (Fe(4+) - OH(-)) is the reactive species regulating the peroxidatic activity of myoglobin. The protonated ferryl species abstracts an electron from either the protein or porphyrin, allowing fast regeneration of the ferric species. Alkaline conditions stabilize the ferryl species, making myoglobin considerably less reactive towards lipids and lipid hydroperoxides. These findings are significant for understanding myoglobin-induced oxidative stress in vivo and the development of therapies.

Animals↗

PubMed2001

Does the redox status of cytochrome C act as a fail-safe mechanism in the regulation of programmed cell death?

It has now become recognized that one of the key events in the induction of apoptosis, or programmed cell death, in both plants and animals is the release of cytochrome c from mitochondria. It is also known that oxidative stress imposed on cells can have a profound effect on the onset or progression of apoptosis. Here, we discuss how the redox status of cytochrome c, and thus its structure, can be altered by the presence of reactive oxygen species (ROS) and reduced glutathione (GSH). We suggest that cytochrome c will only induce programmed cell death if present in the cytoplasm in the oxidized state, and that the presence of high levels of cytoplasmic GSH maintain cytochrome c in an inactive (reduced) state, thus behaving as a fail-safe mechanism if cytochrome c is released by mitochondria when programmed cell death is not the required outcome. If the redox status of the cell is disturbed however, perhaps in the presence of hydrogen peroxide, GSH concentrations will drop, the cellular E(h) will rise, and cytochrome c will tend towards the oxidized state, allowing programmed cell death to proceed. Therefore, we propose that the redox state of cytoplasmic cytochrome c may be a key regulator of programmed cell death.

Animals↗

PubMed1998

Structure-activity relationships for antioxidant activities of a series of flavonoids in a liposomal system.

Structurally diverse plant phenolics were examined for their abilities to inhibit lipid peroxidation induced either by Fe(II) and Fe(III) metal ions or by azo-derived peroxyl radicals in a liposomal membrane system. The antioxidant abilities of flavonoids were compared with those of coumarin and tert-butylhydroquinone (TBHQ). The antioxidant efficacies of these compounds were evaluated on the basis of their abilities to inhibit the fluorescence intensity decay of an extrinsic probe, 3-(p-(6-phenyl)-I,3,5-hexatrienyl)phenylpropionic acid (DPH-PA), caused by the free radicals generated during lipid peroxidation. All the flavonoids tested exhibited higher antioxidant efficacies against metal-ion-induced peroxidations than peroxyl-radical-induced peroxidation, suggesting that metal chelation may play a larger role in determining the antioxidant activities of these compounds than has previously been believed. Distinct structure-activity relationships were also revealed for the antioxidant abilities of the flavonoids. Presence of hydroxyl substituents on the flavonoid nucleus enhanced activity, whereas substitution by methoxy groups diminished antioxidant activity. Substitution patterns on the B-ring especially affected antioxidant potencies of the flavonoids. In cases where the B-ring could not contribute to the antioxidant activities of flavonoids, hydroxyl substituents in an catechol structure on the A-ring were able to compensate and become a larger determinant of flavonoid antioxidant activity.

Amidines↗

PubMed1999

Dihydrofluorescein diacetate is superior for detecting intracellular oxidants: comparison with 2',7'-dichlorodihydrofluorescein diacetate, 5(and 6)-carboxy-2',7'-dichlorodihydrofluorescein diacetate, and dihydrorhodamine 123.

To detect intracellular oxidant formation during reoxygenation of anoxic endothelium, the oxidant-sensing fluorescent probes, 2',7'-dichlorodihydrofluorescein diacetate, dihydrorhodamine 123, or 5(and 6)-carboxy-2',7'-dichlorodihydrofluorescein diacetate were added to human umbilical vein endothelial cells during reoxygenation. None of these fluorescent probes were able to differentiate the controls from the reoxygenated cells in the confocal microscope. However, dihydrofluorescein diacetate demonstrated fluorescence of linear structures, consistent with mitochondria, in reoxygenated endothelium. This work tests the hypothesis that dihydrofluorescein diacetate is a better fluorescent probe for detecting intracellular oxidants because it is more reactive toward specific oxidizing species. To investigate this, dihydrofluorescein diacetate was exposed to various oxidizing species (hydrogen peroxide, superoxide [KO2], peroxynitrite, nitric oxide, horseradish peroxidase, ferric iron, xanthine oxidase, cytochrome c, and lipoxygenase) and compared with the three other popular probes. Though oxidized dihydrofluorescein has higher molar fluorescence, comparison of the reactions of dihydrofluorescein with these other three probes in a cell-free system indicates that dihydrofluorescein is sometimes less fluorescent than the other probes. In addition, we find that the reactivity of all of the probes is very complex. Based on the results reported here, it is no longer appropriate to think of these probes as detecting a specific oxidizing species in cells, such as H2O2, but rather as detectors of a broad range of oxidizing reactions that may be increased during intracellular oxidant stress. Cell-loading studies indicate that dihydrofluorescein achieves higher intracellular concentrations than the second brightest intracellular probe, 2',7'-dichlorodihydrofluorescein. This fact and its higher molar fluorescence may account for the superior brightness of dihydrofluorescein diacetate. Dihydrofluorescein diacetate may be a superior fluorescent probe for many cell-based studies.

Arachidonate 5-Lipoxygenase↗

PubMed1999

Nitric oxide-forming reactions of the water-soluble nitric oxide spin-trapping agent, MGD.

The objective of this study was to elucidate the nitric oxide-forming reactions of the iron-N-methyl-D-glucamine dithiocarbamate (Fe-MGD) complex from the nitrogen-containing compound hydroxyurea. The Fe2+(MGD)2 complex is commonly used in electron paramagnetic resonance (EPR) spectroscopic detection of NO both in vivo and in vitro. The reaction of Fe2+(MGD)2 with NO yields the resultant NO-Fe2+(DETC)2 complex, which has a characteristic triplet EPR signal. It is widely believed that only NO reacts with Fe2+(MGD)2 to form the NO-Fe2+(MGD)2 complex. In this report, the mechanism leading to the formation of NO-Fe2+(MGD)2 was investigated using oxygen-uptake studies in conjunction with the EPR spin-trapping technique. We found that the air oxidation of Fe2+(MGD)2 complex results in the formation of the Fe3+(MGD)3 complex, presumably concomitantly with superoxide (O3*-). Dismutation of superoxide forms hydrogen peroxide, which can subsequently reduce Fe3+(MGD)3 back to Fe2+(MGD)2. The addition of NO to the Fe3+(MGD)3 complex resulted in the formation of the NO-Fe2+(MGD)2 complex. Hydroxyurea is not considered to be a spontaneous NO donor, but has to be oxidized in order to form NO. We present data showing that in the presence of oxygen, Fe2+(MGD)2 can oxidize hydroxyurea to yield the stable NO-Fe2+(MGD)2 complex. These results imply that hydroxyurea can be oxidized by reactive oxygen species that are formed from the air oxidation of the Fe2+(MGD)2 complex. Formation of the NO-Fe2+(MGD)2 complex in this case could erroneously be interpreted as spontaneous formation of NO from hydroxyurea. The chemistry of the Fe2+(MGD)2 complexes in aerobic conditions must be taken into account in order to avoid erroneous conclusions. In addition, the use of these complexes may contribute to the overall oxidative stress of the system under investigation.

Chelating Agents↗

PubMed1997

Effects of 1,10-phenanthroline and hydrogen peroxide in Escherichia coli: lethal interaction.

It has been observed that when Escherichia coli cells are treated simultaneously with phenanthroline and H2O2, there is a lethal interaction. In order to analyze the mechanism of this lethal interaction, wild-type and xthA mutant cells of E. coli were treated with 2.5 mM H2O2 and 1 mM phenanthroline. This treatment was preceded by treatments with different metal chelators (dipyridyl for Fe2+, desferal for Fe3+ and neocuproine for Cu2+) or conducted simultaneously to other treatments with chelators and radical scavengers (thiourea, ethanol and sodium benzoate). The lethal interaction was observed in both the E. coli wild-type strain and xthA mutant strain, which is deficient in the exonuclease III repair enzyme. Nevertheless, the mutant strain was much more sensitive than the wild-type one. Dipyridyl pretreatment protected the cells against the lethal interaction, while desferal pretreament was unable to do so. This suggests that the lethal interaction requires Fe2+ and not Fe3+ ions. Ethanol and sodium benzoate were incapable of protecting bacterial cells against the lethal interaction. Even a 20-min pretreatment with benzoate did not confer protection. On the other hand, thiourea protected the cells completely. Based on our results, we propose that the lethal interaction may be caused not only by the reaction kinetics of phenanthroline and Fe, but also by the ability of phenanthroline to intercalate in DNA. After forming the mono and bis complexes, phenanthroline would serve as a shuttle and take the Fe2+ ions to the DNA. So, the Fenton reaction would take its course with the consequent generation of OH. radicals near DNA. This proximity to the DNA would protect the OH. radicals against the scavengers' action, thus optimizing the Fenton reaction.

2,2'-Dipyridyl↗

PubMed2000

Mössbauer studies on laser evaporated iron atoms and their reactions with oxygen in argon matrices.

Laser-evaporated iron atoms were isolated in low-temperature Ar matrices and their chemical reactions with oxygen were investigated by means of Mössbauer spectroscopy. Reactions of iron atoms with oxygen produce FeO, Fe(O2), FeO3, (O2)FeO2 and OFeO isolated in the Ar matrices and their yields vary depending on the concentration of oxygen. Similarly, FeO and Fe(O2) were obtained by the reaction of iron atoms with N2O. Infrared spectroscopy and molecular orbital calculations were applied to support their assignments.

Calorimetry↗

PubMed1998

Stereochemistry of the chloroperoxidase active site: crystallographic and molecular-modeling studies.

BACKGROUND: Chloroperoxidase (CPO) is the most versatile of the known heme enzymes. It catalyzes chlorination of activated C-H bonds, as well as peroxidase, catalase and cytochrome P450 reactions, including enantioselective epoxidation. CPO contains a proximal heme-thiolate ligand, like P450, and polar distal pocket, like peroxidase. The substrate-binding site is formed by an opening above the heme that enables organic substrates to approach the activated oxoferryl oxygen atom. CPO, unlike other peroxidases, utilizes a glutamate acid-base catalyst, rather than a histidine residue. RESULTS: The crystal structures of CPO complexed with exogenous ligands, carbon monoxide, nitric oxide, cyanide and thiocyanate, have been determined. The distal pocket discriminates ligands on the basis of size and pKa. The refined CPO-ligand structures indicate a rigid active-site architecture with an immobile glutamate acid-base catalyst. Molecular modeling and dynamics simulations of CPO with the substrate cis-beta methylstyrene and the corresponding epoxide products provide a structural and energetic basis for understanding the enantioselectivity of CPO-catalyzed epoxidation reactions. CONCLUSIONS: The various CPO-ligand structures provide the basis for a detailed stereochemical mechanism of the formation of the intermediate compound I, in which Glu183 acts as an acid-base catalyst. The observed rigidity in the active site also explains the relative instability of CPO compound I and the formation of the HOCI chlorinating species. Energetics of CPO-substrate/ product molecular modeling provides a theoretical basis for the P450-type enantioselective epoxidation activities of CPO.

Binding Sites↗

PubMed2002

Active transport of iron and siderophore antibiotics.

Bacteria solubilize iron (Fe(3+)) with secreted siderophores, which are then taken up as Fe(3+)-siderophore complexes. Some bacteria also use iron in heme, hemoglobin, hemopexin, transferrin and lactoferrin of eukaryotic hosts. Crystal structures of two outer membrane transport proteins, FhuA and FepA, and biochemical data reveal strong long-range conformational changes of the proteins upon binding of Fe(3+)-siderophore complexes and in response to energy transfer from the cytoplasmic membrane into the outer membrane via the TonB-ExbB-ExbD protein complex. The crystal structure of the periplasmic binding protein FhuD strongly deviates from the uniform overall structure of binding proteins hitherto determined. Sideromycins, antibiotics that contain Fe(3+)-siderophore complexes as carriers, are highly effective, as they enter cells via Fe(3+)-siderophore transport systems. In this review, recently published data is discussed to demonstrate the state of understanding of iron transport across the outer membrane and the cytoplasmic membrane.

Amino Acid Sequence↗

PubMed2001

Metal complexes of antibiotic drugs. Studies on dicluxacillin complexes of FeII, FeIII, CoII NiII and CuII.

The synthesis and characterization of dicluxacillin (DC) complexes with di- and tri-valent metal ions are described. The nature of bonding of the chelated DC and its metal complexes structures have been elucidated on the basis of their spectroscopic (infrared, solid reflectance, magnetic spectra, mass and thermal analysis) properties. In all the complexes studied, the DC acts as a chelate monoanionic ligand with coordination involving the carboxylate O atom and the endocyclic N of the thiazole ring. The DC ligand forms mono-ligand complexes of the general formula [M(DC)(H2O)x(A)]. yH2O where DC is the uninegatively charged bidentate ligand and A = OAc in case of CuII and Cl in case of FeIII, FeII, CoII and NiII ions. IR, solid reflectance spectra and magnetic moment measurement are used to infer the structure and to illustrate the coordinating capacity of the ligand. From the thermal decomposition curves, the water molecules of crystallization are removed in a single stage while the decomposition of the ligand and coordinated water molecules occur in the second and subsequent stages. Different thermodynamic kinetic parameters namely activation energy (E*), enthalpy of activation (AH*), entropy of activation (AS*) and free energy change of activation (AG*) are calculated using Coats and Redfern equation.

Anti-Bacterial Agents↗

PubMed2004

Vibrational study of the Fe(phen)2(NCS)2 spin-crossover complex by density-functional calculations.

The geometries and vibrational frequencies of an iron(II) spin-crossover complex Fe(phen)2(NCS)2 in the low- and high-spin states have been calculated using DFT (BP86 functional with 6-311G* + Wachters + f, 6-31G* and LANL2DZ basis sets). A redundancy-free set of internal coordinates has been defined and used to obtain a valence force field. The use of the symmetry coordinates of a perfect octahedron as internal coordinates for the FeN6 framework of the complex has enabled the description of the stretching-bending couplings. A comparison of the force constants of free ligands with the corresponding constants of the two spin states has provided a basis for the discussion of the changes in electronic charge distribution in the complex. By use of the generalized harmonic mode scrambling, it is shown that in the low-lying normal vibrations of the complex, the ligands participate as almost rigid units. Energies of these modes are close to the oscillators describing the interaction of the complex with the surrounding molecules in the solid phase.

Electrons↗

PubMed2000

Hyperfine interaction in K2Ba[Fe(NO2)6].

Magnetic hyperfine splitting observed in the low temperature Mössbauer spectrum of potassium barium hexanitro ferrate(II), in the absence of any external field, is attributed to the 5T2g state of the central metal atom further split into a ground 5Eg state and a first excited 5B2g state under a distorted octahedral symmetry in contrast to the earlier prediction of 1A1g ground state on the basis of room temperature Mössbauer spectral and other properties. The central iron atom is coordinated to six nitrito groups (NO2-), having an oxidation state of +2. The temperature dependence of Mössbauer spectra is explained on the basis of electronic relaxation among the spin-orbit coupled levels of the 5Eg ground state. Various kinds of electronic relaxation mechanisms have been compared to explain the proposed mechanism. The observed temperature dependent spectra with varying internal magnetic field and line width can be explained by simple spin lattice relaxation.

Barium Compounds↗

PubMed2000

Altering the specificity of CooA, the carbon monoxide-sensing transcriptional activator: characterization of CooA variants that bind cyanide in the Fe(II) form with high affinity.

CooA is a carbon monoxide- (CO-) sensing homodimeric heme protein that activates the transcription of genes required for the anaerobic oxidation of CO to CO(2) in the phototrophic bacterium Rhodospirillum rubrum. In this study, we demonstrate that mutational alteration of the histidine residue (His(77)) that serves as a heme ligand in the Fe(II) form of CooA allows high-affinity binding of cyanide (K(d) approximately 0.4 mM) to the heme. In contrast, neither these same variants in the Fe(III) form nor wild-type CooA in either oxidation state was able to bind cyanide even at high concentrations (50 mM). Examination of the pH dependence of spectral changes upon addition of cyanide suggested that the cyanide anion coordinated the heme iron. In addition, the UV-visible absorption spectrum of H77Y Fe(II) CooA without added effectors is also pH-dependent, suggesting that an ionizable amino acid has become solvent-accessible in the absence of His(77). Finally, we demonstrate that the transcriptional activity of H77Y CooA shows a small (1.4-fold) increase in the presence of cyanide, suggesting that the binding of cyanide to this variant promotes the active conformation of H77Y CooA.

Bacterial Proteins↗

PubMed1991

Enzymatic redox chemistry: a proposed reaction pathway for the six-electron reduction of SO3(2-) to S2- by the assimilatory-type sulfite reductase from Desulfovibrio vulgaris (Hildenborough).

A detailed reaction pathway for the six-electron reduction of SO3(2-) to S2- by the assimilatory-type sulfite reductase (SiR) from Desulfovibrio vulgaris (Hildenborough) has been deduced from experiments with 35S-labeled enzyme and the relative reaction rates of nitrogenous substrates. The ligand bridging the prosthetic [Fe4S4]-siroheme center is apparently exchanged by 35S2- in both oxidized and reduced enzyme. This 35S2- label was retained in the course of SO3(2-) reduction, implicating substrate binding to the nonbridging axial site of the siroheme. A reaction mechanism is proposed in which SO3(2-) binds to Fe2+ through the sulfur atom, followed by a series of two-electron reductive cleavages of S-O bonds. Protonation of oxygen facilitates bond cleavage, giving hydroxide as leaving group. The bridge remains intact throughout the course of the reaction, providing an efficient coupling pathway for electron transfer between the cluster and siroheme.

Binding Sites↗

PubMed1991

Conformational change and histidine control of heme chemistry in cytochrome c peroxidase: resonance Raman evidence from Leu-52 and Gly-181 mutants of cytochrome c peroxidase.

Resonance Raman (RR) spectra are reported for Fe(III), Fe(II), and Fe(II)CO forms of site-directed mutants of the cytochrome c peroxidase variant CCP(MI), cloned in Escherichia coli. The Fe(II) form is five-coordinate (5-c) and high-spin at low pH, but it is six-coordinate (6-c) and low-spin at high pH except when the distal His-52 residue is replaced with Leu, showing the sixth ligand to be the His-52 imidazole. Although the Leu-52 mutant stays 5-c, it does undergo an alkaline transition, as revealed by upshifts and broadening of bands assigned to vinyl C = C stretching (1620 cm-1) and C beta-vinyl bending (402 cm-1). Similar changes are seen for CCP(MI) and other mutants. Thus the alkaline transition induces a conformational change that affects the vinyl groups, probably through changes in their orientation, and that permits the His-52 imidazole to bind the Fe. The RR band arising from the stretching of the proximal Fe(II)-imidazole bond contains components at ca. 235 and 245 cm-1 for CCP(MI), which are believed to reflect a double well potential for the H-bond between the proximal His-175 imidazole and the Asp-235 carboxylate group. Loss of this H-bond by mutation of Asp-235 to Asn results in the loss of these two bands and their replacement by a single band at 205 cm-1. Although the Fe(II)-imidazole stretching mode cannot be observed in the 6-c alkaline form of the enzyme, the sixth ligand in the alkaline form of CCP(MI) is photolabile, and the status of the Fe(II)-imidazole bond can be determined in the resulting 5-c-photoproduct. For CCP(MI) at alkaline pH, the conformation change induces an increase in the 235/245-cm-1 ratio, reflecting a perturbation of the H-bond potential. In the His-52----Leu mutant, a 205-cm-1 band appears along with the 235/245-cm-1 doublet at alkaline pH, indicating partial loss of the proximal H-bond due to the distal alteration. The effect of mutations that perturb the H-bonding network that extends from the distal to the proximal side of the heme is more dramatic: at alkaline pH, His-181----Gly, Arg-48----Leu, and Trp-51----Phe mutants show an Fe(II)-imidazole stretching mode at 205 cm-1 exclusively, indicating complete loss of the proximal Asp-235-His-175 H-bond.(ABSTRACT TRUNCATED AT 400 WORDS)

Alkenes↗

PubMed1994

The dioxygenation rate in lipoxygenase catalysis is determined by the amount of iron (III) lipoxygenase in solution.

The dioxygenation rate in reactions catalyzed by lipoxygenase-1 from soybeans has been measured as a function of the enzyme present in the Fe(III) form with rapid kinetic techniques. The experiments were carried out at pH 10, 25 degree C. The product concentration and the fraction of iron (III) lipoxygenase were monitored by measuring the absorbance at 243 nm and the tryptophan fluorescence at 330 nm (excitation at 287 nm), respectively. In reactions started with 1.3 microM iron (II) lipoxygenase and 9 microM linoleate, the initial rate, r(init) (estimated from the increase in absorbance over the initial 0.02 s of the reaction), is very small (4 s-1). In contrast, when the reactions are started with 1.3 microM (III) lipoxygenase, r(init) is large (150 s-1). In reactions started with mixtures of iron(II) and iron(III) lipoxygenase, r(init) is linearly related to the initial concentration of the Fe (III) enzyme form. Redistributions of the Fe(II) and Fe(III) enzyme forms during the reaction with 12 nM enzyme and 10, 50, or 100 microM linoleate appear to be directly reflected in changes in the dioxygenation rate. The observations provide solid evidence for the hypothesis that only iron (III) lipoxygenase can catalyze the hydrogen abstraction step in the dioxygenation reaction, and thus can be regarded as the active enzyme species. The observed dynamics are accurately predicted by a nonallosteric, two-step model for lipoxygenase catalysis [Schilstra et al. (1992) Biochemistry 31, 7692-7699].

Catalysis↗

PubMed2001

Characterization of the coral allene oxide synthase active site with UV-visible absorption, magnetic circular dichroism, and electron paramagnetic resonance spectroscopy: evidence for tyrosinate ligation to the ferric enzyme heme iron.

Coral allene oxide synthase (AOS), a hemoprotein with weak sequence homology to catalase, is the N-terminal domain of a naturally occurring fusion protein with an 8R-lipoxygenase. AOS converts 8R-hydroperoxyeicosatetraenoic acid to the corresponding allene oxide. The UV--visible absorption and magnetic circular dichroism spectra of ferric AOS and of its cyanide and azide complexes, and the electron paramagnetic resonance spectra of native AOS (high-spin, g = 6.56, 5.22, 2.00) and of its cyanide adduct (low-spin, g = 2.86, 2.24, 1.60) closely resemble the corresponding spectra of bovine liver catalase (BLC). These results provide strong evidence for tyrosinate ligation to the heme iron of AOS as has been established for catalases. On the other hand, the positive circular dichroism bands in the Soret region for all three derivatives of ferric AOS are almost the mirror image of those in catalase. In addition, the cyanide affinity of native AOS (K(d) = 10 mM at pH 7) is about 3 orders of magnitude lower than that of BLC. Thus, while these results conclusively support a common tyrosinate-ligated heme in AOS as in catalase, significant differences exist in the interaction between their respective heme prosthetic groups and protein environments, and in the access of small molecules to the heme iron.

Animals↗

PubMed1994

Spectroscopic study of Ser92 mutants of human myoglobin: hydrogen bonding effect of Ser92 to proximal His93 on structure and property of myoglobin.

Neutron diffraction studies have demonstrated that the hydroxyl group oxygen of Ser92(F7) is hydrogen bonded to the proximal His93(48) N epsilon H proton in myoglobin (Mb) [Cheng, X., & Shoenborn, B. P. (1991) J. Mol. Biol. 220, 381-399]. In order to examine the importance of this hydrogen bond, Ser92 was replaced with Ala and Asp in human Mb. By comparing the optical, 1H-NMR, resonance Raman, and IR spectra of Mb(S92A) in several spin and oxidation states with those of wild-type Mb, it was found that the mutation causes a structural change on the heme proximal side but not on the distal side. Comparison of the NMR spectra of the cyanomet form of Mb(S92A) and Mb(WT) suggests that the imidazole plane of His93 rotates somewhat around the Fe-N delta (His93) bond upon loss of the hydrogen bond between His93 and Ser92. The 2D 1H-NMR measurements of the CO complexes show that mutation of Ser92 to Ala changes the relative position of the His97 imidazole group to the heme plane, but the change is not so drastic as reported in the crystal data of Ser92 mutant of pig Mb [Smerdon et al. (1993) Biochemistry 32, 5132-5138]. On the other hand, ligand (CO, O2) binding is only slightly affected by this mutation. From these results, we conclude that the Ser92-His93 hydrogen bond maintains the protein structure of the proximal heme pocket, but it does not strongly affect the electronic structure of the heme as well as of the His93 imidazole ring.(ABSTRACT TRUNCATED AT 250 WORDS)

Carbon Monoxide↗