Hideo Kon

Study on the Quinquevalent Chromium Complex

Accurate values of the parallel and perpendicular components of g‐factor and hyperfine coupling tensor were determined for [CrOCln]—(n−3) where n=4 and 5, by the ESR line shape analysis. It was shown by a molecular orbital treatment that these magnetic parameters can be explained consistently by taking into account of an excited electronic configuration in which an electron in a filled orbital is excited to the half‐filled level. An assignment of a hitherto unexplained visible absorption band was proposed on the same ground.

An interpretation of the three line EPR spectrum of nitric oxide hemeproteins and related model systems:: The effect of the heme environment

The EPR spectra of the nitric oxide (NO) derivatives of structurally perturbed Fe (II) hemeproteins show various patterns, all of which are characterized by the conspicuous three-line hyperfine splitting due to 14NO, in contrast to that of the native proteins. For the purpose of obtaining structural information from these three line spectra, the model systems were studied, which consist of NO, heme (or TPP-Fe(II), where TTP means alpha, beta, gamma, delta-tetraphenylporphine) and the nitrogenous base, pyridine or quinoline, which, respectively, give the native type or the three line (perturbed type) EPR spectrum. The ring proton paramagnetic shift of quinoline in this system shows that it is not coordinated to NO-TPP-Fe(II) as a normal axial ligand, in contrast to pyridine which gives the shift pattern of the ordinary axial ligation. This observation suggests that in the NO-hemeproteins some perturbations of the protein structure cause the rupture or distortion of the bond between the imidazole nitrogen on the fifth coordination site and the heme iron, resulting in the three line spectrum. The EPR study of the model systems, the pentacoordinated complex, NO-heme and NO-TPP-Fe(II), in various media revealed that the pentacoordinated species indeed exhibits, depending upon its environment, a variety of spectra, which closely reproduce the three line patterns observed in the perturbed proteins and some related model systems. Such spectral variation can be attributed to the difference in the degree of quenching the internal motion and/or the structural heterogeneity caused by molecular environment.

An investigation of the copper site(s) of dopamine-β-hydroxylase by electron paramagnetic resonance

Abstract 1. 1. Dopamine-β-hydroxylase (3,4-dihydroxyphenylethylamine, ascorbate: oxygen oxidoreductase (β-hydroxylating), EC 1.14.17.1) contains approximately four copper atoms per tetramer. These can be fully reduced by ascorbate and fully oxidized by potassium ferricyanide or by oxygen in the presence of fumarate, tyramine or octopamine. 2. 2. Upon the addition of tyramine (substrate) or octopamine (product) to dopamine-β-hydroxylase, several superhyperfine lines with a splitting of about 15 G are detected at the g 1 region of the EPR spectrum of the enzyme. The involvement of nitrogen ligands in the binding of the enzyme copper is suggested. The absence of superhyperfine lines upon the addition of the substrate, phenylthylamine, implicates the involvement of the 4-hydroxyl group on tyramine and octopamine in the appearance of these lines. 3. 3. A decrease in the amplitude of the EPR signal of dopamine-β-hydroxylase upon the deoxygenation of the enzyme was consistently observed and could suggest that oxygen is reversibly bound in a protein-copper oxygen intermediate complex. 4. 4. By the use of EPR as a probe for the extent of copper oxidation, the midpoint potential of dopamine-β-hydroxylase is estimated to be +310 mV.

Effect of shear stress and free radicals induced by ultrasound on erythrocytes

The present study was undertaken to elucidate the mechanism of hemolysis induced by ultrasound. Ar or N2O gas was used to distinguish between cavitation with or without free radical formation (hydroxyl radicals and hydrogen atoms). Free radical formation was examined by the method of spin trapping combined with ESR. After sonication of erythrocyte suspensions, several structural and functional parameters of the erythrocyte membrane--hemolysis, membrane fluidity, membrane permeability, and membrane deformability--were examined. Although free radical formation was observed in the erythrocyte suspensions sonicated in the presence of Ar, no free radical formation was observed in the presence of N2O. However, the hemolysis behavior induced by ultrasound was similar in the presence of Ar or N2O. The membrane fluidity, permeability, and deformability of the remaining unlysed erythrocytes after sonication in the presence of Ar or N2O were unchanged and identical to those of the control cells. On the other hand, after gamma irradiation (700 Gy), the hemolysis behavior was quite different from that after sonication, and the membrane properties were significantly changed. These results suggest that hemolysis induced by sonication was due to mechanical shearing stress arising from cavitation, and that the membrane integrity of the remaining erythrocytes after sonication was the same as that of control cells without sonication. The triatomic gas, N2O, may be useful for ultrasonically disrupting cells without accompanying free radical formation.

Electron Paramagnetic Resonance and Polarized Absorption Spectra of Cu(II)‐Doped Single Crystal of L‐Histidine Hydrochloride Monohydrate

The electron paramagnetic resonance of copper(II) doped in a single crystal of L‐histidine· HCl monohydrate indicates that the Cu2+ ion is incorporated in a different environment from that in the ordinary bis(L‐histidinyl)copper(II) complex. The coefficients of the ground state Kramers doublet were determined from the observed g tensor (gz = 2.013, gx = 2.262, gv = 2.153) in such a way that the hyperfine parameters calculated from the coefficients closely reproduce the observed values. The results show that the half‐filled d orbital is of the dz2 type, suggesting an axially compressed structure. The polarized single crystal absorption spectra taken in the three perpendicular crystal faces exhibit three d‐d transitions at 14 000, 12 800 and 11 100 cm−1, which are polarized mostly along the c, b, and a crystal axes, respectively. By combining the optical and the paramagnetic resonance results, the one‐electron orbital sequence was found to be dz2>dx2‐y2>dyz>dxy>dxz. For a possible site of copper incorporat...

A spin-label study of the correlation between stomatocyte formation and membrane fluidization of erythrocytes.

Change in the membrane fluidity of human erythrocytes on transformation to stomatocytes was observed by ESR spectroscopy using 12-doxyl stearic acid or its methyl ester as a probe. When the transformation to stomatocytes was induced by four qualitatively different methods, i.e. (a) addition of cationic amphiphilic agents such as chloropromazine, tetracaine, chloroquine or primaquine, (b) addition of Triton X-100, a non-ionic detergent, (c) lowering the pH, and (d) depleting membrane cholesterol, membrane fluidization was always observed. This indicates the existence of a close correlation between stomatocyte formation and increase in the membrane fluidity. Furthermore, since the stomatocytes fixed by diamide treatment exhibited membrane fluidization only in the presence of the reagent, the enhanced membrane fluidity was a direct consequence of the reagent interacting with, and changing the state of, the lipid bilayer itself, and not through the influence of some structural alteration of spectrin. These results provide experimental support for the theoretical prediction made by Brailsford et al. [J. theoret. Biol. 86, 531 (1980)]. Plausible mechanisms for the discocyte-stomatocyte transformation are discussed.

Free-Radical Distributions in Several Gamma-Irradiated Dry Proteins as Determined by the Free-Radical Interceptor Technique

Free-radical distributions in several dry, γ-irradiated proteins have been compared by means of the free-radical interceptor technique, which involves reaction of carbon free radicals with tritiated hydrogen sulfide to form carbon-tritium bonds within the protein. Subsequent hydrolysis, amino acid analysis, and counting then reveal the tritium distribution among the amino acids which is taken as an indicator of the free-radical distribution. A comparison of the tritium distributions among the proteins studied indicates many similarities, among which are high specific activities for methionine and proline, with relatively low activities for many other residues (for example, glycine, alanine, valine, isoleucine, and leucine). Despite this general pattern, many differences in labeling were found. Lysine, for example, was the most heavily labeled amino acid for ribonuclease. For lysozyme, lysine and methionine were the most radioactive, with nearly equal specific activities. Methionine was the most active ami...

Nature of the Unpaired Electron Orbital in Some Low Spin Oxygen-Co Complexes

Abstract The CoII complexes, Co(II)-bissalicylal-3,3′-diimino-di-n-propylamine peroxide I and decaammine-μ-peroxodicobalt cation II, were investigated by the electron spin resonance (esr). On the basis of the observed esr parameters, it was concluded that the unpaired electron is largely localized on the O2 group and, therefore, these complexes are better described as super-oxides. The equilibrium in solution of I between the paramagnetic and dia-magnetic forms implied by the previous studies was also demonstrated.

Anomalous magnetic properties of tetraphenylporphinato Co(II) complex in the solid state

Abstract The α,β,γ,δ-tetraphenylporphinatocobalt(II) complex is found to exist in two distinct, but interconvertible, polycrystalline forms. The one with a tetragonal crystal symmetry (species B) gives the EPR spectrum which has been attributed to the low-spin electronic configuration of Co(II) ion in an axial crystal field. The other form (species A) having a triclinic crystal symmetry shows no easily detectable EPR signal even at liquid helium temperature. Magnetic susceptibility and magnetization meaurements demonstrated that the complex is paramagnetic in both forms, but the species (A) is characterized by ferromagnetic exchange coupling, while the species (B) behaves as a normal paramagnet. The experimental susceptibility versus 1/ T curve can be reproduced quite well by using the Ising method. The g values thus obtained ( g | = 5.2, g ⊥ = 0) can not be explained by a low-spin electronic configuration, but are consistent with a high-spin ground state. Assigning a high-spin state to the species (A), the first such case in Co(II) porphine complexes, can not only explain the absence of EPR signal, but is also supported by the results of X-ray structural analyses.

EPR studies on the high spin FeIII tetraphenylporphine with rhombic character, determination of zero-field splitting parameters from the middle kramers transition

Abstract Electron paramagnetic resonance transitions (EPR) in the middle Kramers doublet is observed for the first time in the 6A1 ground state of Fe(III) complex of tetraphenylporphine (TPP) diluted in the polycrystalline free base tetraphenylporphine (THHP2). The system shows the EPR spectra and resulting from several distinct species of high spin (Fe(III) TPP with various degress of rhombicity. The middle Kramers as absorptions are in general of much weaker intensity than the corresponding lowest Kramers absorptions, and their g values show marked systematic deviations from the calculated first order prediction. The deviation can be explained by carrying the Zeeman perturbation to the third order. The analysis also rendered it possible to determine the zero-field parameters D and E separately. The results indicate the similarity in the crystal field of the present system with that of cyctochrome P450.