Akitsugu Nakahara

Some properties of a blue copper protein ‘plantacyanin’ from cucumber peel

Absorption, circular dichroism, electron spin resonance and resonance Raman spectra of a blue copper protein, plantacyanin from cucumber peel have been measured and these spectral properties compared with those of other blue copper proteins. From the spectral properties, amino acid analysis and redox potential, we discuss the active site and redox properties of this protein.

Copper(II) complexes with tripodal imidazole-containing ligands. Structure-electrochemistry relationship

Mono- and dimeric copper(II) complexes of tris- (benzimidazol-2-ylmethyl)amine (L1) and of its derivatives with the imidazole NH protons substituted by a methyl (L2), ethyl (L3), benzyl (L4) or o-methylbenzyl (L5) group were prepared and characterized based on magnetic susceptibility, conductivity, cyclic voltammetric half-wave potential (E12), infrared, electronic and ESR spectra. The complexes containing H2O, NO3−, Cl−, Br−, N3−, and N-methylimidazole with L1–L5 were mononuclear copper(II) complexes, while the complexes with NCS− or NCO− and L1–L3 [except [Cu(L1)NCS](NO3)] were dinuclear ones, which dissociate to monomeric complexes in DMF solution. The E12 values of copper(II) in a series of complexes containing H2O or NO3− decrease in the order L5 > L1 > L2 > L3 > L4, whereas those in complexes with Br− and N-methylimidazole decrease in the orders L4 ≳ L5 > L1 > L2 > L3 and L5 > L1 > L4 ≳ L2 > L3, respectively. The relationship between the structure of the complexes and the E12 values are discussed in some detail.

Copper(III) and nickel(III) complexes of tripeptides, dipeptide schiff bases and related ligands

Abstract Trivalent copper and nickel complexes of tripeptides, dipeptide Schiff bases and related ligands were comparatively studied by cyclic voltammetry. Depending upon the nature of the coordinated groups and the structure of fused ring systems the electrode potentials of Cu(III)Cu(II) and Ni(III)Ni(II) couples vary in the range 0.7–1.4 V. The stronger donating groups such as amino and deprotonated amide nitrogens assist the stabilizing of metal ions in the higher oxidation state, although aromatic nitrogen and oxygen do not contribute effectively for this. The trivalent copper and nickel with smaller ionic radii are stabilized by the 5-5-5 but not by the 5-6-5 or 6-5-6 fused ring structure as in bivalent copper and nickel. These redox characters were related with ESR and the absorption spectral feature of parent and trivalent metal complexes.

X-Ray crystal structures and magnetic properties of azide-bridged binuclear copper(II) complexes containing the Schiff-base ligand derived from 2-pyridinecarbaldehyde and histamine. Structure–magnetism relationship

The structures of the complexes [Cu2(L1)2(N3)2][ClO4]2(1){ L1= 1-(imidazol-4-yl)-2-[(2-pyridylmethylene)amino]ethane} and [Cu2(L1)2(N3)3]Cl·2H2O (2) have been determined by X-ray methods. Complex (1) contains discrete [Cu2(L1)2(N3)2]2+ cations which are located at centres of inversion. The geometry around each copper(II) ion is square pyramidal. The two basal planes in the complex cation are parallel and separated by 2.64 A. The two azide ions which are parallel to each other bridge in an end-on fashion between a basal position of one square pyramid and an apical position of the other. Magnetic susceptibility data for (1) show a ferromagnetic interaction (Weiss constant θ=+23 K) above 200 K. On the other hand, below 200 K, it is considered that there is a weaker exchange interaction than that above 200 K. In differential scanning calorimetry curves for (1), small thermal anomalies have been observed over the temperature range 150–250 K. Complex (2) contains an azide-bridged, centrosymmetric copper(II) dimeric cation. The five-coordinate geometry around each copper(II) ion is square pyramidal. An azide ion bridges in an endto-end fashion between the two apical positions of two square pyramids. A very weak antiferromagnetic exchange interaction is found for (2)(0 J–0.5 cm–1, where J denotes the exchange integral defined by the Hamiltonian ℋ=–2JS1S2).

Characterization of cucumber ascorbate oxidase and its reaction with hexacyanoferrate (II).

The spectroscopic features of cucumber ascorbate oxidase (AOase) and its type-2 copper-depleted (T2D) derivative, and the electron pathway among the copper sites in the enzyme have been investigated. The electronic and CD spectra of native and T2D AOase in the visible region bear a striking resemblance to those of plastocyanin or azurin, which contain type-1 copper alone. The electronic absorption shoulder of the native enzyme at around 330 nm for the native enzyme which has been assigned to type-3 copper disappears with the depletion of the type-2 copper. The reduction of AOase with a large excess of hexacyanoferrate(II) results in a selective reduction of the type-2 Cu, giving rise to an additional EPR-detectable species which is considered to be originated from partly reduced type-3 copper. The type-1 copper is, however, not reduced even in the presence of excess hexacyanoferrate(II). The redox potential of type-1 Cu was determined to be +350 mV, which is distinctly lower than that of hexacyanoferrate(II-III). Type-2 copper was supposed to be a mediator of the electron transfer between type-1 and type-3 coppers in consideration of the extremely low activity of the T2D enzyme under the same condition. A comparison of the electron pathway in AOase with that in laccase is also argued.

The monomeric and dimeric copper(II) complexes containing imidazole and dipeptides

Abstract The mixed ligand copper(II) complexes with imidazole and glycylglycine or glycyl-β-alanine were prepared in neutral and basic solutions. The copper(II) complexes obtained in neutral solutions were mononuclear complexes, while those in basic solutions were binuclear complexes bridged by imidazolate. In the cases of the binuclear complexes, it was found that the two copper(II) ions are antiferromagnetically coupled with -J values of 19–17 cm −1 . It has also been disclosed on the basis of visible and ESR spectral measurements that these dimeric structures are maintained in the concentration of 1.3 × 10 −2 to 1.2 × 10 −3 M /Cu.

Monomeric and dimeric copper(II) complexes composed of some terdentate schiff bases and oxalate, oxamate, azide, thiocyanate or cyanate ions

Abstract Monomeric and dimeric copper(II) complexes containing the Schiff bases derived from 2-pyridinecarbaldehyde and histamine (2-PcaHist), 2-(2-aminoethyl)pyridine (2-Pca2-Aep), or β-alanine (2-Pcaβ-Ala), and oxalate (ox), oxamate (om), azide, thiocyanate, or cyanate ions have been prepared and characterized based on their electronic, infrared and electron spin resonance spectra, and magnetic susceptibilities. The structure of the monomeric copper(II) complex, [Cu(2-PcaHist)(N3)2] has been determined by the X-ray diffraction method. The crystal of [Cu(2-PcaHist)(N3)2] is triclinic, with a = 10.262(8), b = 9.177(6), c = 7.688(5) A, α = 104.42(4), β = 94.09(4), γ = 92.64(4)°, Z = 2, and space group P 1 . The five-coordinate geometry around copper(II) ion is intermediate between trigonal-bipyramid and square-pyramid. The half-field absorption in the ΔMs = 2 region of powdered X-band ESR spectra was scarcely observed for [Cu(2-Pca2-Aep)(N3)2], [Cu(2-Pca2-Aep)(NCS)2], [Cu(2-Pca2-Aep)(NCO)2] and [Cu(2-PcaHist(N3)2]. The compounds [Cu2(2-PcaHist)2(N3)3]Cl·2H2O and [Cu2(2-PcaHist)2(N3)2](ClO4)2 which are considered to have a dimeric structure exhibited the half-field absorption in the same region. However, an exchange interaction was hardly observed down to 4.2 °K in the magnetic susceptibility measurement for [Cu2(2-PcaHist)2(N3)3](Cl·2H2O. The susceptibility of an oxalate-bridged copper(II) complex, [Cu2(2-PcaHist)2(ox)](ClO4)2, showed an antiferromagnetic interaction (J = −21.5 cm−1) and the X-band ESR spectrum for the powdered sample showed a very weak absorption for the triplet state of such a dimer in the ΔMs = 2 region.

Structure of the active site of hemocyanin. Cobalt(II)-substituted squid hemocyanin

Abstract Cobalt(II)-substituted squid ( Sepioteuthis lessoniana ) hemocyanin was prepared by dialyzing the apohemocyanin against pH 8.0 Tris-HCl buffer containing 5 m M Co(II) ion. The electronic absorption, circular dichroism and magnetic circular dichroism spectra of Co(II)-substituted hemocyanins containing one mol of Co(II) and 2.6 mol of Co(II) per mol of protein were studied comparatively to characterize the coordination geometry and the ligating groups around the binuclear copper sites in native squid hemocyanin. The two copper sites at one active site in squid deoxyhemocyanin may differ from each other in coordination geometry, and may consist of a tetrahedral-like copper site and a tetragonal-like copper site. Three histidine imidazoles are expected to be coordinated around each copper(II). The structure of the active site of squid and horseshoe crab hemocyanins was comparatively discussed on the basis of spectral data of the respective cobalt(II)-substituted hemocyanins of the two phyla.

Spectroscopic studies on the active site of Sepioteuthis lessoniana hemocyanin

Abstract The absorption, circular dichroism (CD), and magnetic circular dichroism (MCD) spectra in the visible region have been measured for Sepioteuthis lessoniana hemocyanin at 77, 198, and 293K. From the temperature dependence of the CD spectra of oxyhemocyanin, the bands observed at 450, 565, and 700 nm were resolved into those centered at 430, 490, 565, 600, and 700 nm. Since these five peaks are most probably due to the d-d transitions, the two copper ions at the oxygenated active center are inferred to be Cu(II) ions each in a non-equivalent coordination geometry of very low symmetry. The MCD spectral data confirm the view and reasonably explain the diamagnetism of oxyhemocyanin.

Properties of the type II copper-depleted cucumber ascorbate oxidase and its reaction with azide

Abstract On the selective removal of type II copper, the absorption band at around 330 nm of native cucumber ascorbate oxidase disappeared, while the band at 610 nm was unaffected. The spectral feature of 330 nm was recovered by treatment of the type II copper-depleted ascorbate oxidase with hydrogen peroxide. However, the modification of the circular dichroic spectral features in the near-ultraviolet region resulting from depletion of the type II copper was not restored by treatment of type II copper-depleted ascorbate oxidase with H 2 O 2 but was restored after the incorporation of Cu 2+ into the type II copper site. Type II copper-depleted ascorbate oxidase was fully reduced only with 3 e − -equivalents of l -ascorbate, indicating that the type III coppers had been reduced during the procedure to remove the type II copper. On the other hand, nearly 7 e − -equivalents of l -ascorbate were required to reduce H 2 O 2 -treated type II copper-depleted ascorbate oxidase, since type III coppers were reoxidized by H 2 O 2 . The action of excess azide on native, type II copper-depleted and H 2 O 2 -treated type II copper-dpleted ascorbate oxidase indicated that at least two azide anions bind to both the type II and III copper ions. The present results demonstrate that type II copper is present relatively near the type III copper site, affecting the oxidation state of the coupled coppers. It was also revealed that the negative CD band at around 320 nm, where the coupled type III coppers exhibit a prominent shoulder in the absorption spectrum, is at least partly contributed by type II copper. Furthermore, reoxidation of the reduced type II copper-depleted ascorbate oxidase by O 2 suggested that the reoxidation process of the type I coppers is not hampered owing to the absence of the type II copper.