Chemiluminescence of cobalt(II)-catalysed auto-oxidation of sulphite and its application to cobalt analysis.

The oxidation of sulphite by dissolved oxygen in aqueous solution catalysed by cobalt(II) was investigated. A weak chemiluminescence (CL) emission was observed when the reaction took place in a strong alkaline solution without any special CL reagent. Further studies showed that in the presence of fluorescein sodium the CL signal was enhanced significantly. The CL emission is linear with Co(II) concentration in the range 0.6-80 nmol/L and the detection limit is 0.3 nmol/L. In addition to Co(II), other transition metal ions were also tested, and the results showed that the proposed system was highly selective for Co(II). The method was successfully applied to the determination of Co(II) in pharmaceutical preparations. The possible CL mechanism was also discussed.     

Multiple forms of the cobalt(II)-carnosine complex.

Cobalt(II) ion and L-carnosine produce two different complexes when mixed in aqueous solution at pH 7.2. One complex has coordination of N-3 of the imidazole ring to the cobalt(II) and is produced when the concentration of peptide exceeds that of cobalt(II). The second complex has chelation of three nitrogen atoms of a single carnosine. This second complex produces a reversible oxygen carrier by making stable mixed chelates with additional carnosine, histidine or cysteine. These results indicate that cobalt complexes with mixed ligands should be of more importance $\text{in}\text{vivo}$ than those with carnosine as the only ligand. They provide an explanation for the high activity and substrate specificity of carnosinase in kidney.     

pH-dependent properties of cobalt(II) carboxypeptidase A-inhibitor complexes.

1H NMR spectroscopy of the isotropically shifted signals in cobalt carboxypeptidase, CoCPD, permits a direct and selective detection of protons belonging to the residues liganded to the metal. The chemical shift of these protons in the free enzyme and enzyme-inhibitor complexes with changing pH monitors the state of ionization of the ligands directly and of other residues in the active center indirectly. The 1H NMR spectrum of CoCPD at pH 6 shows three well-resolved isotropically shifted signals in the downfield region at 62 (a), 52 (c), and 45 (d) ppm which have been assigned to the NH proton of His-69 and to the C-4 H's of His-69 and His-196, respectively. Titration of signal a with pH is characterized by a pKa of 8.8 which is identical to that seen in prior electronic absorption and kinetic studies. The fact that the signal reflecting the NH of His-69 is still observed at pH 10 and no major shifts occur for the signals reflecting the C-4 H's indicates the alkaline pKa in carboxypeptidase A catalysis, pKEH, cannot be ascribed to ionization of the histidyl NH of either His-69 or His-196. Binding of L-Phe shifts this pKa to 7.7 while not greatly perturbing the downfield 1H NMR signals that reflect the ligation shell of the cobalt coordination sphere. These results indicate the pKa of 8.8 in CoCPD and the pKa of 7.7 in the CoCPD.L-Phe adduct reflect ionization of the same group.(ABSTRACT TRUNCATED AT 250 WORDS)     

Identification of a key structural feature of cobalt(III)-bleomycins: an exogenous ligand (e.g. hydroperoxide) bound to cobalt

A series of cobalt(III) complexes of the anticancer antibiotic bleomycin has been prepared. Mass spectrometry and enzymatic analysis show that the green cobalt-bleomycin complex contains a hydroperoxide (-OOH) group bound to cobalt with unusual stability. Under appropriate conditions, cobalt-bleomycins containing other monodentate ligands to cobalt can be formed; fast-atom bombardment mass spectra of such complexes show peaks for cobalt-bleomycin at the expected mass, and also peaks for the intact complexes at the required higher mass.     

Magnetic circular dichroic spectra of cobalt(II) substituted metalloenzymes.

The magnetic circular dichroic (MCD) spectra of cobalt(II) sugstituted metalloenzymes have been studied and compared to a series of four-, five-, and six-coordinate cobalt(II) model complexes previously examined (T. A. Kaden et al. (1974), Inorg. Chem. 13, 2582). The MCD spectra of cobalt substituted carboxypeptidase A, procarboxypeptidase ta, and thermolysin are consistent with earlier deductions of tetrahedral coordination from absorption spectra and also with X-ray structure analysis. Inhibitors fail to alter their MCD spectra significantly. The MCD spectra of cobalt alkaline phosphatase and carbonic anhydrase are more complex and their pH dependence and alteration by inhibitors are discussed in terms of known cobalt(II) models.     

Temperature dependence of the magnetic susceptibility of cobalt (II) stellacyanin.

Magnetic susceptibility measurements on cobalt(II) stellacyanin ($\text{Rhus vernicifera}$) have been performed between 2.2 and 50 K. The effective magnetic moment of Co(II) in the protein is 3.91 ± 0.12 (μ_B). Nonlinear behavior below 3 K evidences the presence of zero-field splitting attributable to a low-symmetry component of the ligand field. The results are consistent with a structural model based on a distorted tetrahedral Co(II) site involving one or more extremely covalent metal-ligand bonds.     

Novel cobalt(II) complexes with pyridyl/ether or pyridyl/thioether ligands. The conversion of pyridyl/thioether cobalt(II) complex to pyridyl/sulfinato cobalt(III) compound

Two new cobalt(II) complexes of symmetric hexadentate mixed-ligand N,O [1,12-bis(2-pyridyl)-5,8-dioxa-2,11-diazadodecane (pydado)] and N,S [1,12-bis(2-pyridyl)-5,8-dithia-2,11-diazadodecane (pydadt)] donor atoms have been synthesized as perchlorate salts. The crystal structures show that [Co(pydado)](ClO₄)₂ · H₂O (1) crystallizes in the triclinic space group and [Co(pydadt)](ClO₄)₂ (2) crystallizes in the monoclinic space group P2_1/c. The cation [Co(pydado)]²⁺ is pseudo-octahedral with the two pyridyl groups in trans position. However, in [Co(pydadt)]²⁺ complex, the size of thioether sulfur atoms imposes a distorted octahedral geometry; the pyridyl groups and the sulfur atoms are in trans position. The reaction of the complex 2 and hydrogen peroxide resulted to the oxidation of Co^II into Co^III and the thioether groups of the ligand to sulfinate groups with elimination of the central ethylenic group of pydadt. Thus, complex 2 was converted to bis[3-(2-pyridylmethylamino)ethanesulfinate] cobalt(III) complex (3) {[Co(pynso)₂](ClO₄) · 0.5H₂O}. The X-ray crystal structure reveals that the compound 3 crystallizes in the triclinic space group with the same donor atoms (N_pyridyl, N_amine and S) belonging to the two ligands in cis-position. In aqueous solution, the stability constants of the Co(II) chelates with these two ligands, determined by potentiometry, show the formation of [Co(LH)]³⁺ and [CoL]²⁺ species in all cases. The chelating power of pydadt ligand is slightly greater than that of pydado.     

Interactions between alpha-amino acids and cobalt(II) bovine-carbonic anhydrase.

The results of a study on the interaction between cobalt(II) bovine carbonic anhydrase and the alpha-amino acids L(+) and D(-)alanine, glycine and betaine are reported. L(+)alanine and glycine have been found to have larger affinity for the enzyme than D(-)alanine whereas no sizable affinity is shown by betaine. The electronic spectra indicate that these systems are similar to that containing the acetate ion. Utilizing the inhibition properties of L(+)alanine at various pH an analysis of the species involved in the inhibition reaction is presented.     

Investigation of the system cobalt(II) bovine carbonic anhydrase B-trichloroacetaldehyde.

The interactions between hydrated trichloroacetaldehyde and cobalt(II)bovine carbonic anhydrase B have been investigated as a function of pH by means of electronic spectroscopy of FT nmr spectroscopy. The hydrated aldehyde is bound to the metal ion and its apparent affinity constant is pH dependent with a bell-shaped profile. The kinetic parameters of the dissociation process have also been determined.     

The 1H nuclear-magnetic-resonance spectroscopy of cobalt(II)-beta-lactamase II.

The 1H n.m.r. spectra of beta-lactamase II in the presence of Co(II) were studied. Analysis of the spectra suggests that Co(II) binds at the same two metal-binding sites as does Zn(II). The binding of Co(II) at the first site is much weaker than the binding of Zn(II) at this site, whereas the binding of Co(II) at the second site is tighter than the binding of Zn(II). The binding of Co(II) to the mono-zinc(II)-enzyme caused only one marked change in the spectrum, namely a decrease in the intensity of the resonances assigned to the C-2 and C-4 protons of one histidine residue (residue E). However, when the spectra of the apoenzyme and the Co(II)-enzyme were compared, there were many differences. A significant fraction of the protons in the whole molecule are affected by the binding of Co(II) at the first metal-ion-binding site (where the ligands are the enzyme's sole thiol group and three histidine residues). This may be because the first site is internal, or because of a difference in conformation between the apoenzyme and the mono-Co(II)-enzyme. The second site may be located on the surface of the molecule.     

A crystallographic study of deoxy cobalt (II) mesoporphyrin IX myoglobin.

The crystal structures of acid metmyoglobin and deoxy cobalt(II)mesoporphyrin IX myoglobin were compared by a difference Fourier analysis at 2.5 A resolution. No large differences in protein conformation were observed. The greatest density of structural differences was found in the heme region. There was a loss of the histidine-bound sulfate ion and of the metal-bound water molecule, as well as a shift in the position of the prosthetic group with associated changes in the adjacent globin. The structural changes resulting from the substitution of ethyl for the vinyl side chains of the porphyrin were clearly observed. There was also a suggestion of a conformational change of the porphyrin ring. It was not clear whether there was any change of the metal position relative to the porphyrin plane or proximal histidine.     

Decrease in hepatic cytochrome P-450 by cobalt. Evidence for a role of cobalt protoporphyrin.

Exposure of cultured chick-embryo hepatocytes to increasing concentrations of CoCl2 in the presence of allylisopropylacetamide results in formation of cobalt protoporphyrin, with a reciprocal decrease in haem and cytochrome P-450. Treatment of rats with CoCl2 (84 mumol/kg) and 5-aminolaevulinate (0.2 mmol/kg) also results in formation of cobalt protoporphyrin and a decrease in cytochrome P-450 in the liver. Hepatic microsomal fractions from rats treated with phenobarbital, CoCl2 and 5-aminolaevulinate were analysed by polyacrylamide gel electrophoresis. Cobalt protoporphyrin was associated mainly with proteins of 50000-53000 mol.wt. The results suggest that the formation of cobalt protoporphyrin occurred at the expense of the synthesis of haem, leading to a decrease in cytochrome P-450. Furthermore, the cobalt protoporphyrin that was formed may itself have been incorporated into apocytochrome P-450.     

The distribution of cobalt inT. subterraneum

Summary The distribution of cobalt in plants ofT. subterraneum grown on different levels of supply of cobalt and nitrate nitrogen has been measured, and the nodules and to a lesser extent the roots have been found to accumulate much more than the leaves.The distribution of vitamin-B₁₂-type compounds and of Co⁶⁰ in the nodule have also been determined following fractionation of the nodule components by hgh speed centrifugation. Most of the cobalt whether present as vitamin B₁₂, or as indicated by the measurement of Co⁶⁰, is present in the supernatant and in the bacteroids. The distribution of Co⁶⁰ is much the same in the nodules from both effective and non-effective strains. The quantities of vitamin B₁₂ present in both bacteroids and the supernatant are greatly increased, in relation to the fresh weight of the nodule tissue, when the supply of cobalt in the nutrient is raised.When Co⁶⁰ is incorporated into the plant a proportion of the activity is found to be present as vitamin B₁₂, free from bacteroids, within four days of addition. After this time the supernatant still contained about 90 per cent of the activity that entered. Of this some 40 per cent was present as ionic cobalt, 19 per cent as vitamin B₁₂ like compounds, and the remainder as a compound, presently termed Factor N, which is so far unidentified.Most of the data on which this paper is based are from the thesis submitted by S.B.W. for the degree of Ph. D. of the University of Nottingham.     

The metal-binding properties of ovotransferrin. An investigation of cobalt(II) derivatives

Cobalt(II) ovotransferrin bicarbonate and oxalate ternary complexes were prepared and investigated in the pH range 7-10.5. Cobalt(II) provides an excellent and unique spectroscopic probe to monitor subtle structural differences in solution between the two sites of ovotransferrin and to investigate the structural dependence on pH. CD spectroscopy on one side and 1H NMR spectroscopy of isotropically shifted signals on the other are extremely sensitive techniques and are particularly suited for high spin cobalt(II)-containing compounds. In the case of the oxalate derivative the metal-binding ability of the protein is different at the two binding sites and is pH dependent; the CD spectra reveal two different sites, one of which is clearly pH dependent with a pKa of 9.5. On the contrary the bicarbonate analogue does not show any spectral difference between the two sites; both of them change with pH, the pKa being again 9.5. 1H NMR spectra of the oxalate derivatives at pH 7-8 reveal the presence of conformers, the distribution of which depends on the H2O/D2O ratio. Such conformers are not revealed in the bicarbonate system; at pH around 10 the NMR spectra of both systems show inequivalence between the two sites and/or the presence of different conformers for each site. Such differences are discussed in terms of the possible implications in mechanism and function. The overall spectral data are consistent with the donor groups being two histidines, two tyrosines, the synergistic anion, and possibly a solvent molecule.     

The interaction of cobalt (II) complexes with bovine apocarbonic anhydrase B.

The second-order rate constants for interaction of a number of cobalt(II) complexes (including the aquated ion) with bovine apocarbonic anhydrase have been measured at pH equal to 7.5, 25 degrees C and I equal 0.1 M. The ease of entry of cobalt decreases, in general, as the number of its coordinated watersdecreases. The highest complexes (bis or tris) do not react. The Mono cobalt(II) -8-quinolinol-5-sulfonate and, particularly, 8-quinolinol complexes react very much more rapidly with apoenzyme than does Co(H20)6-2+ ion. The results are discussed.     

Allosteric transitions in cobalt hemoglobins.

Circular dichroism and difference ultraviolet visible spectra were obtained for cobalt hemoglobin derivatives. At 287 nm the ellipticity difference between the oxy- and deoxycobaltohemoglobin is about one-half as great as that for the native proteins indicating smaller quaternary conformational changes for the former. Deoxygenation increases the Soret rotational strengths of both iron and cobalt hemoglobins to comparable degrees suggesting similar conformational changes for their aromatic residues near the "heme." Deoxygenation causes a much larger decrease of L band ellipticity for iron than cobalt hemoglobin. Circular dichroism spectra of nitrosylcobaltohemoglobin indicate the molecule to have a T quaternary structure. The circular dichroism spectra of cobaltihemoglobin do not seem to fit the patterns of the other cobalt derivatives and its 287 nm ellipticity is pH-dependent. From the shape of the Soret circular dichroism spectra, it is estimated that the transition dipole makes an angle with the line joining the two opposing pyrrole nitrogens of about 60 degrees for oxy- and deoxycobaltohemoglobin, 80 degrees for cobaltihemoglobin, as compared to 70 degrees for the native oxy- and deoxyhemoglobins. Inositol hexaphosphate has little or no effect on the circular dichroism spectra of cobalt hemoglobins in the 287 nm region, but it significantly increases the Soret rotational strength and decreases the L band ellipticity. The results are interpreted to mean that polyphosphates modify primarily the protein structure of hemoglobins at the tertiary level, and that the intersubunit interactions are weak in cobalt hemoglobins.     

Molecular modeling of cobalt(II) hyaluronate

Structural data for complexes of hyaluronic acid and 3d metals(II) of the fourth group of the periodic table are lacking. A combined QM/MM method was used to solve the structure of the first coordination sphere around the cobalt(II) ion. Some available experimental data were compared with the results obtained via computation and were found to be in good agreement. Our results open the way for using molecular modeling to solve the structure of other metal(II) hyaluronates.