Effects of 2,4-substituents of deuteroheme upon peroxidase functions. II. Reactions of peroxidases and metmyoglobin with azide, cyanide and fluoride

Artificial horseradish peroxidases and metmyoglobins were reconstituted from their apoproteins and the following unnatural hemes: mesoheme, deuteroheme, hematoheme, chlorocruoroheme, and diacetyldeuteroheme. The electron-withdrawing effects of the 2,4-substituents upon affinities of these hemoproteins for azide, cyanide, and fluoride were investigated. Peroxidase preparations used were acidic (A₁ + A₂) and neutral (B + C) enzymes.The electron withdrawal of the substituents increased the strength of ligaud binding. Plotting the logaritms of dissociation constants for the hemoprotein-ligand complexes against pK₃, a measure of relative basicities for metal-free porphyrins, the Hammett relationship was found to be applicable to this case except for a few. The affinities of deuterohemoproteins for ligands were definitely higher than those expected from the above relationship.The constant ρ, slope of the Hammett equation, varied with the ligand. For any particular ligand, the two peroxidase preparations gave the same ρ value, but it was different from the value for metmyoglobin.     

Theoretical study of X- · 1 · YF (1 = triazine,X = Cl,Br and I,Y = H,Cl, Br,I, PH2 and AsH2): noncovalently electron-withdrawing effects on anion-arene interactions

The ternary complexes X^- · 1 · YF (1 = triazine, X = Cl, Br and I, Y = H, Cl, Br, I, PH₂ and AsH₂) have been investigated by MP2 calculations to understand the noncovalently electron-withdrawing effects on anion-arene interactions. The results indicate that in binary complexes (1 · X^-), both weak σ-type and anion-π complexes can be formed for Cl^- and Br^-, but only anion-π complex can be formed for I^-. Moreover, the hydrogen-bonding complex is the global minimum for all three halides in binary complexes. However, in ternary complexes, anion-π complex become unstable and only σ complex can retain in many cases for Cl^- and Br^-. Anion-π complex keeps stable only when YF = HF. In contrast with binary complexes, σ complex become the global minimum for Cl^- and Br^- in ternary complexes. These changes in binding mode and strength are consistent with the results of covalently electron-withdrawing effects. However, in contrast with the covalently electron-withdrawing substituents, Cl^- and Br^- can attack the aromatic carbon atom to form a strong σ complex when the noncovalently electron-withdrawing effect is induced by halogen bonding. The binding behavior for I^- is different from that for Cl^- and Br^- in two aspects. First, the anion-π complex for I^- can also keep stable when the noncovalent interaction is halogen bonding. Second, the anion-π complex for I^- is the global minimum when it can retain as a stable structure.     

Decrease in oxygen affinity of myoglobin by formylation of vinyl groups of heme.

Three kinds of green synthetic myoglobin were prepared by recombination of horse heart apomyoglobin with spirographis (2-formyl-4-vinyl-), isospirographis (2-vinyl-4-formyl-), and 2,4-diformyldeuterohemins. The optical and oxygen binding properties of the reconstituted myoglobins containing two isomeric monoformyl-monovinylhemins were found to be different. The oxygen affinities (P50) of spirographis and 2,4-diformylmyoglobins are 2.7 and 2.8 mm Hg, respectively, at 25 degrees, and about 2.5 times lower than that of native protomyglobin, while that of isospirographis myoglobin is 1.0 mm Hg and is similar to native myoglobin. Spirographis oxymyoglobin has absorption maxima (alpha, beta, and Soret bands) at 601, 556.5, and 435 nm, isospirographis oxymyoglobin at 595, 550, 429 nm, and 2,4-diformyl oxymyoglobin at 603, 563.5, and 447 nm. The optical red shifts as well as the decrease in the oxygen affinities of these myoglobins are attributed mainly to the presence of strongly electron-attractive formyl side chains. Since the free isomers of monoformyl-monovinyl heme have similar properties, the differences observed after recombination with apoprotein must be caused by interactions with apomyoglobins. The degree of such a protein effect may be estimated by comparing the absorption spectra of heme before and after recombination and was found to differ among the various myoglobins. Comparison of the oxygen affinities of the myoglobins taking account of this protein factor showed that the increase in the P50 values are inversely related to that in the pK3 values of the free porphyrins. These results suggest the involvement of pi bonding in determining the oxygen-iron bond strength.     

Ethylisocyanide equilibria of hemoglobins M Iwate, M Boston, M Hyde Park, M Saskatoon, and M Milwaukee-I in half-ferric and fully reduced states.

The ethylisocyanide equilibria of all the five known hemoglobins M, namely Hb M Iwate (alpha287 Tyrbeta2), Hb M Boston (alpha258 Tyrbeta2), Hb M Hyde Park (alpha2beta292 Tyr), Hb M Saskatoon (alpha2beta263 tyr), and Hb M Milwaukee-I (alpha2beta267 Glu), were studied both in the half-ferric and fully reduced heme states. In the half-ferric state, no heme-heme interaction was observed for Hb M Iwate, Hb M Boston, and Hb M Hyde Park, but Hb M Saskatoon and Hb M Milwaukee-I show small but definite heme-heme interaction with Hill's n of 1.3. The beta chain mutants, Hb M Hyde Park and Hb M Saskatoon, have almost normal affinity for ethylisocyanide and a normal Bohr effect, whereas the alpha chain mutants, Hb M Iwate and Hb M Boston, have abnormally low affinity and almost no Bohr effect. Hb M Milwaukee-I showed a large Bohr effect and low affinity. These results are consistent qualitatively with those on oxygen equilibria reported previously. In the fully reduced state, in which all four hemes were in the ferrous state and capable of binding ethylisocyanide distinct differences were found in the extent of heme-heme interaction. Namely, the n values for proximal histidine mutants, Hb M Iwate and Hb M Hyde Park, were 1.1 and 1.0, respectively, whereas the distal histidine mutants, Hb M Boston and Hb M Saskatoon, showed high n values of 2.4 and 1.6, respectively. Hb M Milwaukee-I also exhibited a high n value of 2.0 The ethylisocyanide affinity of the four histidine mutants was high compared with that of Hb A, while that for Hb M Milwaukee-I was almost normal. All five Hbs M had approximately normal magnitudes of Bohr effect. In the half-ferric state, the proximal and distal histidine mutants of the same chain showed similar affinity for ethylisocyanide and Bohr effect, rather different from those of the mutants of the opposite chain. These differences seem to be derived from the difference of abnormal bonding of ferric iron to tyrosine or glutamic acid. On the other hand, the reduction of iron, which abolished the abnormal bonding and made all of the chains capable of binding ligand, extinguished the differences of alpha and beta chains, and the effect of amino acid side chains close to iron on ligand binding properties became clear. Proximal histidine, which is considered to trigger the transition between the T and R states, seems to be essential to the heme-heme interaction.     

Kinetic and equilibrium studies of the reactions of heme-substituted horse heart myoglobins with oxygen and carbon monoxide.

In order to study the effects of chemical modifications of the vinyl groups of heme on oxygen and carbon monoxide binding to myoglobin, apomyoglobins from horse heart were reconstituted with six different hemins with various side chains. Laser flash photolysis experiments of these reconstituted myoglobins showed that the combination rate constants for oxygen (k') and carbon monoxide (l') were closely related to the electron-attractive properties of the side chains. The k' values obtained in 0.1 M potassium phosphate buffer, pH 7.0, at 20 degrees were 0.83 (meso-), 2.4 (deutero-), 1.1 (reconstituted proto-), 1.2 (native proto-), 1.5 (2-formyl-4-vinyl-), 1.9 (2-vinyl-4-formyl-), and 2.7 X 10(7) M-1 S-1 (2,4-diformylmyoglobins), and the corresponding l' values were 2.8, 18, 4.8, 5.1, 7.1, 15, and 35 X 10(5) M-1 S-1, respectively. These rate constants tend to increase as the electron-withdrawing power of the side chains increases, indicating that reduced electron density of the iron atom of heme in myoglobin favors the combination reaction for both oxygen and carbon monoxide. Equilibrium constants (L) between carbon monoxide and various myoglobins were also determined by measuring the partition coefficients (M) between oxygen and carbon monoxide for the myoglobins, and were also found to be closely related to the electronic properties (pK3 of porphyrin) of the heme side chains. The equilibrium association constants for carbon monoxide thus obtained increased with a decrease in pK3 value of the porphyrin. This order was completely opposite to the case of the oxygen binding reaction. The dissociation rate constants for oxygen (k) and carbon monoxide (l) were calculated from the equilibrium and the combination rate constants. The dissociation rate constants showed a similar characteristic to the combination rate constants and increased with the increase in electron attractivity of heme side chains. The concomitant increase in both the combination and dissociation rate constants with increase in electronegativity of the iron atom suggests that these reactions have different rate determining steps, although such a reaction process is contradictory to the generally accepted concept that in a reversible reaction, both on and off reactions proceed through the same transition state. In the on reaction sigma bond formation appears to be dominant, while in the off reaction eta bond break-up is more important.     

Nuclear-magnetic-resonance investigation of the cooperative homodimeric hemoglobin from the mollusc Scapharca inaequivalvis. Molecular and electronic structure of the cyano-met derivative

The proton nuclear-magnetic-resonance spectra of the cyano-met complexes of the cooperative dimeric and tetrameric hemoglobins from the mollusk Scapharca inaequivalvis have been investigated and compared to those of other structurally characterized oxygen binding hemoproteins. For these proteins, cooperativity is displayed even in the homodimer and preliminary X-ray structural data reveal an unusual back-to-front assembly with intersubunit contacts involving the EF helices [Royer, W. E., Love, W. E. + Fenderson, F. F. (1985) Nature (Lond.) 316, 277-280]. The pattern of hyperfine shifts is very similar for the dimer and tetramer chains, but distinctly different from those of previously characterized low-spin, ferric heme proteins. Individual heme resonances are identified by reconstituting the protein with specifically deuterated hemes. While the axial interactions involving the proximal and distal histidines are very similar to that in myoglobins and other hemoglobins, both the heme contact shift pattern and the amino acid dipolar shift pattern reflect a significantly reduced asymmetry. The decreased spread of the non-cordinated amino acid signals is interpreted in terms of a rotation of the magnetic axes relative to those in myoglobin or other hemoglobins, rather than a change in the magnetic anisotropy. The decreased spread of the heme methyl contact shifts supports this conclusion and is consistent with an orientation of the proximal histidine with the imidazole ring rotated by about 30-40 degrees relative to that in other structurally characterized proteins. Although resonances associated with a complex pattern of alternate heme orientations can be detected immediately after reconstitution of the protein, the isolated protein was found to exhibit insignificant equilibrium heme rotational disorder.     

Crystal structures of modified myoglobins. II. Relation between oxygen affinity properties and structural changes around heme in myoglobins reconstituted with 2,4-diisopropyldeuteroheme, 2-isopropyl-4-vinyldeuteroheme, and 2-vinyl-4-isopropyldeuteroheme

The crystal structures of sperm whale metmyoglobins reconstituted with three kinds of modified hemes, 2,4-diisopropyldeuteroheme, 2-isopropyl-4-vinyldeuteroheme, and 2-vinyl-4-isopropyldeuteroheme, have been determined and refined at 2.2 A resolution to R = 0.216, 0.219, and 0.195, respectively. All the crystals of these myoglobins are isomorphous with that of native metmyoglobin. The 2-vinyl-4-isopropyldeuteroheme was found to be in a reverse orientation, in which the heme plane is rotated by 180 degrees about an axis through the alpha-gamma-meso carbons, whereas the orientations of the other two hemes were the same as that of protoheme in native myoglobin. In the myoglobins with 2,4-diisopropyldeuteroheme and 2-vinyl-4-isopropyldeuteroheme, both of which have lower oxygen affinities than native myoglobin, the bulky isopropyl side chain pushes Phe 43 0.7 A toward His 64 (the distal histidine) in the former, and the whole E helix at most 1.5 A, including a 0.7 A shift of the His 64 imidazole ring, in the latter. The changes of the structures prevent His 64 from forming a hydrogen bond with the liganded oxygen molecule, so that these two modified myoglobins show low oxygen affinities. On the other hand, there is no such drastic displacement in myoglobin with 2-isopropyl-4-vinyldeuteroheme, which has a slightly higher oxygen affinity than native myoglobin.     

Mechanism and biological role of nitric oxide binding to cytochrome c'

The binding of nitric oxide to ferric and ferrous Chromatium vinosum cytochrome c' was studied. The extinction coefficients for the ferric and ferrous nitric oxide complexes were measured. A binding model that included both a conformational change and dissociation of the dimer into subunits provided the best fit for the ferric cytochrome c' data. The NO (nitric oxide) binding affinity of the WT ferric form was found to be comparable to the affinities displayed by the ferric myoglobins and hemoglobins. Using an improved fitting model, positive cooperativity was found for the binding of NO to the WT ferric and ferrous forms, while anticooperativity was the case for the Y16F mutant. Structural explanations accounting for the binding are proposed. The NO affinity of ferrous cytochrome c' was found to be much lower than the affinities of myoglobins, hemoglobins, and pentacoordinate heme models. Structural factors accounting for the difference in affinities were analyzed. The NO affinity of ferrous cytochrome c' was found to be in the range typical of receptors and carriers. In addition, cytochrome c' was found to react with cytosolic light-irradiated membranes in the presence of succinate and carbon monoxide. With these results, a biochemical model of cytochrome c' functioning as a nitric oxide carrier was proposed.     

Effects of formylation of vinyl side chains of heme on optical and ligand binding properties of horse heart ferric myoglobin.

Effects of substitution of vinyl groups of hemin with formyl groups on the optical and ligand binding properties of horse heart ferric myoglobin were investigated. The peak positions as well as the line shapes of the absorption spectra of the ferric derivatives of three kinds of formylmyoglobin, 2-vinyl-4-formyl-, 2-formyl-4-vinyl-, and 2,4-diformylmyoglobins depend on the number and the position of the formyl groups. Absorption maxima in the Soret region of the acid forms of these ferric formylmyoglobins in 0.1 M potassium phosphate buffer, pH 6.0, at 20 degrees were 415.2, 422, and 429 nm, respectively. The acid forms of these formylmyoglobins exhibit absorption spectra of the mixture of high- and low spin states at ambient temperature. Since proto-, deutero- and mesomyoglobins have a high spin state under the same condition, the increase of the low spin iron in these formylmyoglobins may be due to the strong electron withdrawal by the formyl groups toward the periphery of the porphyrin ring. The affinities of these ferric formylmyoglobins and protomyoglobin for N3-, F-, OCN-, and SCN- increased in the order of proto-, monoformyl-monovinyl-, 2,4-diformyl-myoglobin, which corresponds to the increasing order of electron-withdrawing power of the porphyrin side chains. The pKa values of the acid-alkaline transition decreased in the same order. Although the ferric forms of the two isomeric monoformyl-monovinylmyoglobins exhibited different optical spectra, the dissociation constants of the complexes of these isomers for various ligands were similar to each other. The pKa values of the acid-alkaline transition were also similar. These results indicate that affinities of ferric myoglobin for ligands, in contrast to those of the ferrous form for oxygen and carbon monoxide (Sono, M., and Asakura, T. (1975) J. Biol. Chem. 250, 5527-5232 and Sono, M., Smith, P.D., McCray, J.A., and Asakura, T. (1976) J. Biol. Chem 251, 1418-1426), are not affected by the position of modifications at the two vinyl groups, but are determinedby the number of the formyl groups and that two vinyl groups at position 2 and 4 are equivalent in the binding of various ligands by ferric myoglobin. The electron density of the ferric iron appears to be similar for the two isomeric monoformyl-monovinylmyoglobins.     

Discovering protein−ligand chalcogen bonding in the protein data bank using endocyclic sulfur-containing heterocycles as ligand search subsets

The chalcogen bond, the noncovalent, electrostatic attraction between covalently bonded atoms in group 16 and Lewis bases, is present in protein?ligand interactions based on X-ray structures deposited in the Protein Data Bank (PDB). Discovering protein?ligand chalcogen bonding in the PDB employed a strategy that focused on searching the database for protein complexes of five-membered, heterocyclic ligands containing endocyclic sulfur with endo electron-withdrawing groups (isothiazoles; thiazoles; 1,2,3-, 1,2.4-, 1,2,5-, 1,3,4-thiadiazoles) and thiophenes with exo electron-withdrawing groups, e.g., 2-chloro, 2-bromo, 2-amino, 2-alkylthio. Out of 930 ligands investigated, 33 or 3.5% have protein?ligand S---O interactions of which 31 are chalcogen bonds and two appear to be S---HO hydrogen bonds. The bond angles for some of the chalcogen bonds found in the PDB are less than 90°, and an electrostatic model is proposed to explain this phenomenon.     

Comparative analysis of spatial organization of myoglobins. II. Secondary structure]

An analysis of probability of distribution curves of alpha-helical sites and bends of polypeptide chains of myoglobins in half-water mammals (beaver, nutria, muskrat, otter) carried out in comparison with those of myoglobins of the horse and Sperm whale (X-ray diffraction analysis has revealed their tertiary structure) has revealed a coincidence of the secondary structure sites end bends of the chain in the studied respiratory hemoproteins of muscles. Despite a considerable number of amino acid substitutions the profiles of alpha-helicity and B-bends of the compared proteins are practically identical. This indicates to the "resistance" of the probability curves to amino acid substitutions and to retention of the tertiary structure of myoglobins in evolutionary remote species of the animals.     

The oscillator strengths of hemoproteins. Their relation to the coordination structure and magnetic susceptibility

The oscillator strengths of hemoproteins in the light frequency range of 1.11 X 10(4) to 3.23 X 10(4) cm-1 (wavelength range of 900 to 310 nm) were measured by means of computer-assisted spectrophotometry. The obtained values of oscillator strength per molar heme ranged from about 1.4 to 2.2. By comparing the oscillator strength values of the ferric and ferric cyanide-bound forms of hemoproteins and also the values of low molecular weight ferric heme complexes, it was found that the oscillator strength was lower for those hemoproteins whose heme was coordinated with strong field ligands. It was also found that the hemoproteins showing a smaller pH-dependent change in the carbon monoxide-difference spectrum had lower oscillator strengths. The following linear relation was observed, with various ligand complexes of bovine methemoglobin, horse metmyoglobin, and ferric horseradish peroxidase, between the oscillator strength (f) determined in the present study and the respective magnetic susceptibility (10(6) X chi 20 degrees M) values in the literature: f = A (10(6) X chi 20 degrees M) + B. The values of constants A and B in the equation were estimated for horseradish peroxidase, methemoglobin, and metmyoglobin. On varying the temperature in the range of 0 to 40 degrees C, the oscillator strength of the metmyoglobin-azide complex changed in parallel with the change in the spin state. Taking advantage of the fact that fluoride complexes of many hemoproteins show 10(6) X chi 20 degrees M values close to 14,500 and also that the values of intersection B are around 86.4% of the respective values of the fluoride complexes of ferric horseradish peroxidase, methemoglobin, and metmyoglobin, an empirical equation was evolved for the calculation of an approximate 10(6) X chi 20 degrees M value from the f value of a given complex (fobs) and that of the fluoride complex (fF) of a hemoprotein. The approximate magnetic susceptibilities of various ligand complexes of bovine lactoperoxidase could be thus calculated with the equation. The oscillator strengths of ferrous hemoproteins were also investigated and ligand-dependent regular changes were found.     

Crystal structures, antioxidation and DNA binding properties of Yb(III) complexes with Schiff-base ligands derived from 8-hydroxyquinoline-2-carbaldehyde and four aroylhydrazines

X-ray crystal and other structural analyses indicate that Yb(III) and all four newly synthesized ligands can form a binuclear Yb(III) complex with a 1:1 metal to ligand stoichiometry by octacoordination at the Yb(III) center. Investigations of DNA binding properties show that all the ligands and Yb(III) complexes can bind to Calf thymus DNA through intercalations with the binding constants at the order of magnitude 10⁵–10⁷ M⁻¹, but Yb(III) complexes present stronger affinities to DNA than ligands. All the ligands and Yb(III) complexes may be used as potential anticancer drugs. Investigations of antioxidation properties show that all the ligands and Yb(III) complexes have strong scavenging effects for hydroxyl radicals and superoxide radicals but Yb(III) complexes show stronger scavenging effects for hydroxyl radicals than ligands. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Interaction between alkaline earth cations and oxo ligands: a DFT study of the affinity of Mg2+ for carbonyl ligands

The affinities of Mg(2+) for various substituted carbonyl ligands were determined at the DFT (B3LYP/6-31+G(d)) and semi-empirical (PM6) levels of theory. Two sets of carbonyl ligands were studied: monosubstituted [aldehydes R-CHO and RPh-CHO] and homodisubstituted [ketones R(2)C=O and (RPh)(2)C=O], where R = NH(2), OCH(3), OH, CH(3), H, F, Cl, Br, CN, or NO(2)). In the (RPh)(2)CO case, the R group was bonded to the para position of a phenyl ring. The enthalpies of interaction between the ligands and a pentaaquomagnesium(II) complex were calculated to determine the affinity of each ligand for the Mg(2+) cation and to correlate with geometrical and electronic parameters. These parameters exhibited the same trends for all of the ligands studied, showing that the affinity of Mg(2+) for electron-donating ligands is higher than its affinity for electron-withdrawing ligands. In the complexes, electron-donating groups increase both the electrostatic and the covalent components of the Mg-ligand interaction. This behavior correlates with the Mg-O(carbonyl) distance and the ligand electron-donor strength.     

Oxygen binding of myoglobins of diving animals]

When studying oxygen binding by myoglobins of diving animals it is shown that half-saturation of myoglobins obtained from the muscles of animals who can stop the external respiration for a long period of time occurs at oxygen strength of 0.62-0.67 mm Hg. Such indices of oxygenation of respiratory hemoprotein of muscles are characteristic of most mammals. Temperature being decreased from 37 to 15 degrees C, myoglobin affinity to oxygen considerably increases.     

Crystal structures of modified myoglobins. I. Heme orientation and structural changes around heme in myoglobins reconstituted with isopemptoheme, pemptoheme, 2-ethyldeuteroheme, and 4-ethyldeuteroheme

The crystal structures of sperm whale metmyoglobins reconstituted with four modified hemes, isopemptoheme, pemptoheme, 2-ethyldeuteroheme, and 4-ethyldeuteroheme, have been determined and refined at 2.2 A resolution to R = 0.217, 0.218, 0.213, and 0.222, respectively. All the crystals of these myoglobins are isomorphous with that of native metmyoglobin. The structural changes of the modified myoglobin from the native myoglobin were examined on difference Fourier maps; the orientation of 4-ethyldeuteroheme in the heme pocket is such that the heme is rotated by 180 degrees about an axis through the alpha-gamma-meso carbons, whereas the orientations of the other three hemes are the same as that of the protoheme in the native myoglobin. The changes of the structures around the heme become greater in the order of isopemptoheme, 2-ethyldeuteroheme less than pemptoheme less than 4-ethyldeuteroheme. The magnitudes of the changes seem to be related to the oxygen affinities of these four reconstituted myoglobins.     

1H nmr studies of complexes of ferric leghemoglobin with substituted pyridines and nicotinic acids

The ¹H nuclear magnetic resonance (nmr) spectra of complexes of soybean ferric leghemoglobin with 3-substituted pyridines and 5-substituted nicotinic acids have been recorded in order to determine the influence of axial ligands on heme electronic structure. The hyperfine shifted resonances of the heme group were assigned by analogy to previous assignments for the pyridine and nicotinic acid complexes of leghemoglobin. The spectra are characteristic of predominantly low-spin ferric heme complexes. For the pyridine complexes, the rate of ligand exchange was found to increase with decreasing ligand pK_A. For many of the complexes, optical and nmr spectra reveal the presence of an equilibrium mixture of high- and low-spin states of the iron atom. The percentage of high-spin component increases with decreasing ligand pK_A Smaller hyperfine shifts are noted for leghemoglobin complexes with ligands capable of weak ligand → metal π bonding. The pattern of hyperfine shifted resonances is similar for all complexes studied and indicates that the overall heme electronic structure is dominated by the bonding to the proximal histidine.     

The ground-state Na+ affinities of a series of substituted acetophenones: a DFT study

A detailed study of Na⁺ affinities of a series of para-substituted acetophenones and their O–Na⁺ counterparts was performed using density functional theory [Becke, Lee, Yang and Parr (B3LYP)] method using 6-311G(d,p) basis sets with complete geometry optimisation. The gas-phase O–Na⁺ complex formation turns out to be an exothermic case and the local stereochemical disposition of Na⁺ is found to be almost the same in each case. The presence of the para-substituent is seen to cause very little change in the Na⁺ affinity relative to the unsubstituted acetophenones. Electron-releasing p-substituents increase it by 0.0105 hartree and electron-withdrawing p-substituents decrease it by 0.011 hartree. Computed Na⁺ affinities are sought to be correlated with a number of computed system parameters such as the net charge on the Na⁺ and the carbonyl oxygen of the Na⁺ complexes and the net charge on the carbonyl oxygen of the free bases. The energetics, structural and electronic properties of the complexes indicate that the interaction between the Na⁺ ion and a carbonyl base is predominantly an ion–dipole attraction and the ion-induced dipole interaction as well rather than a covalent interaction.     

Effects of 2,4-substituents of deuteroheme upon the stability of the oxy-form and compound I of horseradish peoxidases.

The stability of oxyperoxidases increased in the order meso- < proto < chlorocruoro- < diacetylperoxidases, which was an increasing order of electron-withdrawing capacities of 2,4-substituents of deuteroheme and the ratio of Δlogk₁toΔpK₃ was approximately 0.6 in the two series of isoenzyme preparations, horseradish peroxidases A and (B + C), where k₁andpK₃ represent a rate constant for conversion from an oxyperoxidase to the ferric enzyme and a measure of basicity of pyrrole nitrogen of the substituted deuterohemes, respectively. Deutero-oxyperoxidases A and (B + C) were definitely more stable than expected from the above linear relationship. The stability of peroxidase Compound I also varied with the 2,4-substituents, but it did not necessarily correlate with electron-withdrawing capacities of the substituents. Natural peroxidases formed relatively stable Compound I in both series of the isoenzymes. From these results it was concluded that the stability of oxyperoxidases was affected by the electron density at the iron atom of the enzyme while steric factors might be involved in stabilizing Compound I.     

Synthesis, spectroscopy, electrochemistry and thermal study of vanadyl unsymmetrical Schiff base complexes

The new tetradentate unsymmetrical N₂O₂ Schiff base ligands and VO(IV) complexes were synthesised and characterized by using IR, UV-Vis and elemental analysis. The electrochemical properties of the vanadyl complexes were investigated by means of cyclic voltammetry. The oxidation potentials are increased by increasing the electron-withdrawing properties of functional groups of the Schiff base ligands according to the trend of MeO < H < Br < NO₂. The thermogravimetry (TG) and differential thermoanalysis (DTA) of the VO(IV) complexes were carried out in the range of 20-700 °C. The complexes were decomposed in two stages. Also decomposition of synthesised complexes is related to the Schiff base characteristics. The thermal decomposition of the studied reactions was first order.