A protective function of superoxide dismutase during respiratory chain activity.

(1) Aerobic incubation of heart muscle submitochondrial particles in phosphate buffer after treatment with NADH causes a progressive and substantial inhibition of the NADH oxidation system. Succinate oxidation remains almost unaffected by NADH treatment. (2) The loss of NADH oxidase activity is due to an inhibition of the respiratory chain-linked NADH dehydrogenase. This inhibition of the enzyme is very similar to that caused by combination of the organic mercurial mersalyl with NADH dehydrogenase. (3) The inhibition of NADH oxidation is largely prevented by compounds that are known to react with superoxide ions (02-.), including superoxide dismutase, cytochrome c, tiron and Mn2+. EDTA also has a protective effect, but a number of other metal chelating agents, and several proteins, including catalase, are without effect. (4) It is concluded that the inhibition of NADH oxidation of NADH oxidation by superoxide ions or by mersalyl is reversible and is therefore not due to the loss of oxidoreduction components from the respiratory chain or to an irreversible change in protein conformation. (6) The function of mitochondrial superxide dismutase is discussed in relation to the key role of NADH dehydrogenase in energy-conserving reactions and the formation of hydrogen peroxide during mitochondrial oxidations.     

The kinetics of flavine oxidation-reduction. II. Metal ion interactions.

The oxidation-reduction reactions of tetraacetylriboflavine in the presence of various metal ions in dimethylformamide have been investigated using the stopped-flow technique under anaerobic conditions. Dismutation kinetics in the presence of redox-inactive dissociated divalent metal ions such as Cd2+, Zn2+, and Fe2+ are typically triphasic. Metal ions act primarily upon an intermediate flavine dimer formed by fast association of flavoquinone and flavohydroquinone, resulting in a parallel formation and neutral and chelated radicals. A competition between metal ions and proton donors, e.g. the neutral flavohydroquinone (FredH3), is observed at the level of this intermediate complex. Small spectral changes occur secondarily as an ill-defined intermediate phase which could correspond to the reorganization of the solvation of radical chelate. The neutral radical is finally chelated at a much slower rate, the yield of total radical formation remaining almost unchanged during this kinetic phase. The oxidation of flavohydroquinone by ferric ions, either dissociated or strongly coordinated within a porphyrin, is complete and proceeds through biphasic kinetics. The first phase (Fred leads to F) is much faster than the second one (F leads to Fox). Dismutation resulting from the transient accumulation of neutral flavosemiquinone competes with the direct oxidation with ferric ions for the completion of the second oxidation step. The relative rate of dismutation is essentially limited by acidic-basic reactions in the absence of an excess of ferrous ion. The kinetic analysis of the direct oxidation reactions favors an outer-sphere mechanism for the electron transfer to the ferric ion, either free or strongly coordinated. The formation of a ferrous radical chelate can result from the dismutation reactions only when the amount of ferric ion initially present is not sufficient for complete oxidation.     

Inhibition of chemical oxidation and reduction of cytochromes ƒ and b-559 by carbonylcyanide p-trifluoromethoxy phenylhydrazone

The presence of low (1–4 μM) concentrations of carbonylcyanide p-trifluoromethoxyphenylhydrazone during actinic illumination of chloroplasts generally inhibits the rate of subsequent dark chemical oxidation-reduction reactions of cytochrome ƒ and b-559. Ferricyanide oxidation and ascorbate reduction of cytochromes ƒ and b-559 are inhibited, as is hydroquinone reduction of cytochrome b-559. Inhibition by carbonylcyanide p-trifluoromethoxyphenylhydrazone of hydroquinone reduction of cytochrome ƒ, the most rapid of these chemical oxidation-reduction reactions, cannot be detected. The rate of the chemical redox reactions of the cytochromes in the presence of carbonylcyanide p-trifluoromethoxyphenylhydrazone are all markedly dependent upon the concentration of oxidant or reductant except the hydroquinone reduction of cytochrome b-559 photooxidized in the presence of carbonylcyanide p-trifluoromethoxyphenylhydrazone.

The data is interpreted in terms of an effect of carbonylcyanide p-trifluoromethoxyphenylhydrazone on thylakoid membrane structure which generally inhibits accessibility to the hydrophobic interior of the membrane, possibly through an increase in membrane microviscosity. The question of whether such an effect on membrane structure could be involved in uncoupling or inhibition effects of the carbonylcyanidephenylhydrazone compounds is discussed, as is the special effect of these compounds on the cytochrome b-559 photoreactions at room temperature.     

The fate of cytoplasmic vanadium. Implications on (NA,K)-ATPase inhibition.

The fate of vanadate (+5 oxidation state of vanadium) taken up by the red cell was studied using EPR spectroscopy. The appearance of an EPR signal indicated that most of the cytoplasmic vanadate is reduced to the +4 oxidation state with axial symmetry characteristic of vanadyl ions. The signal at 23 degrees C was characteristic of an immobilized system indicating that the vanadyl ions in the cytoplasm are associated with a large molecule. [48V]Vanadium eluted with hemoglobin when the lysate from Na3[48V[O4-treated red cells was passed through a Sephadex G-100 column and rabbit anti-human hemoglobin serum caused a hemoglobin-specific precipitation of 48V when added to the red cell lysate. Both results indicate that hemoglobin is the protein which binds cytoplasmic vanadyl ions. However, neither sodium vanadate nor vanadyl sulfate bound to purified hemoglobin in vitro. Finally, transient kinetics of vanadyl sulfate interaction with the sodium-and potassium-stimulated adenosine triphosphatase showed that the +4 oxidation state of vanadium is less effective than the +5 oxidation state in inhibiting this enzyme. These results indicate that oxidation-reduction reactions in the cytoplasm are capable of relieving vanadate inhibition of cation transport.     

Developmental changes in the form of the Arrhenius plots of rat liver plasma membrane adenylate cyclase and 5'-nucleotidase activities

Human α-1-proteinase inhibitor is inactivated by human myeloperoxidase in the presence of hydrogen peroxide and chloride ion. Several antiarthritic drugs and related compounds, including many containing gold, were tested as inhibitors of the myeloperoxidase system. Of the twenty-six compounds used, twenty-two inhibited. The most important feature of these was the presence of a sulfhydryl group. The most effective compounds also were the most hydrophobic. The presence of gold, on the other hand, made little difference to the amount of inhibition. These drugs appear to have many effects, and their inhibition of the myeloperoxidase system suggests that this could be one of them.     

Inhibition of copper-catalyzed cysteine oxidation by nanomolar concentrations of iron salts

Problems caused by the presence of adventitious metals in buffers and reagents are well recognized in studies of metal-catalyzed oxidation reactions. In most cases, metal contamination leads to an increase in rate, and chelating agents are inhibitory. In the present study, however, the rate of copper-catalyzed oxidation of cysteine was found to be increased by buffer purification with Chelex resin or by addition of micromolar concentrations of the specific iron chelator desferrioxamine (DFO). These effects are attributable to inhibition of copper-catalyzed oxidation by adventitious iron. In purified buffer at pH 7.25, containing 0.4 microM copper, cysteine was oxidized at a rate of 32 microM/min. Addition of iron salts to this buffer caused a dose-related decrease in this rate, up to a maximum of 85%. A 50% decrease in rate was recorded at an iron concentration of only 11 nM. Other transition metals were without effect. Similar effects of purification or addition of DFO on the rate of cysteine oxidation were seen in Tris, glycylglycine, Mops, and Pipes buffers. Catalase decreased the rate of cysteine oxidation, but the sensitivity to iron was similar in the presence and absence of catalase. Copper-catalyzed oxidation of cysteamine and reduced glutathione was much less sensitive to inhibition by iron. Our results offer an explanation for the conflicting literature reports of the effects of chelating agents and catalase on cysteine oxidation, and emphasize the need for buffer purification or addition of DFO in studies concerned with the oxidation or cytotoxicity of this thiol. The exceptional sensitivity of copper-catalyzed cysteine oxidation to iron makes this an attractive system for monitoring the iron content of buffers, and may also have application for determining the free iron content of physiological fluids.     

Inhibition of cellulase-catalyzed lignocellulosic hydrolysis by iron and oxidative metal ions and complexes

Enzymatic lignocellulose hydrolysis plays a key role in microbially driven carbon cycling and energy conversion and holds promise for bio-based energy and chemical industries. Cellulases (key lignocellulose-active enzymes) are prone to interference from various noncellulosic substances (e.g., metal ions). During natural cellulolysis, these substances may arise from other microbial activities or abiotic events, and during industrial cellulolysis, they may be derived from biomass feedstocks or upstream treatments. Knowledge about cellulolysis-inhibiting reactions is of importance for the microbiology of natural biomass degradation and the development of biomass conversion technology. Different metal ions, including those native to microbial activity or employed for biomass pretreatments, are often tested for enzymatic cellulolysis. Only a few metal ions act as inhibitors of cellulases, which include ferrous and ferric ions as well as cupric ion. In this study, we showed inhibition by ferrous/ferric ions as part of a more general effect from oxidative (or redox-active) metal ions and their complexes. The correlation between inhibition and oxidation potential indicated the oxidative nature of the inhibition, and the dependence on air established the catalytic role that iron ions played in mediating the dioxygen inhibition of cellulolysis. Individual cellulases showed different susceptibilities to inhibition. It is likely that the inhibition exerted its effect more on cellulose than on cellulase. Strong iron ion chelators and polyethylene glycols could mitigate the inhibition. Potential microbiological and industrial implications of the observed effect of redox-active metal ions on enzymatic cellulolysis, as well as the prevention and mitigation of this effect in industrial biomass conversion, are discussed.     

Evaluation of Factors Affecting the Survival of Escherichia coli in Sea Water: VI. Cysteine

The relationship between death of cells of Escherichia coli in artificial sea water and time was established as linear, and statistical tests demonstrated that the most suitable measure of survival was log per cent after 24 hr. Survival of E. coli in water supplemented with cysteine at levels of 0.284 × 10^-6 to 284 × 10^-6m was increased greatly over that in untreated water. To provide an insight into the mode of action of cysteine, the effect of concentration of various sulfhydryl and disulfide compounds was measured, and the influence of several compounds that lack a functional sulfur group but which are capable of affecting oxidation-reduction potential was determined. Moreover, a number of substances related structurally to cysteine were tested to ascertain their influence on the survival of cells of E. coli in artificial sea water. It appeared that the beneficial effect of cysteine was not due to the sulfhydryl group of the amino acid or to the ability of the compound to influence oxidation-reduction potential. Some sulfhydryl compounds had no favorable effect and, in general, disulfides were more active than the corresponding sulfhydryl compounds. Substances that lack a functional sulfur group but influence oxidation-reduction potential had no significant activity. The beneficial effect of a number of compounds related structurally to cysteine indicates that both an amino and carboxyl group are required for activity. It is suggested that cysteine and other amino acids act to increase survival of cells of E. coli in sea water by a chelation mechanism.     

Transition metal ions and selenite modulate the methylation of arsenite by the recombinant human arsenic (+3 oxidation state) methyltransferase (hAS3MT)

This report demonstrates that transition metal ions and selenite affect the arsenite methylation by the recombinant human arsenic (+3 oxidation state) methyltransferase (hAS3MT) in vitro. Co²⁺, Mn²⁺, and Zn²⁺ inhibited the arsenite methylation by hAS3MT in a concentration-dependent manner and the kinetics indicated Co²⁺ and Mn²⁺ to be mixed (competitive and non-competitive) inhibitors while Zn²⁺ to be a competitive inhibitor. However, only a high concentration of Fe²⁺ could restrain the methylation. UV-visible, CD and fluorescence spectroscopy were used to study the interactions between the metal ions above and hAS3MT. Further studies showed that neither superoxide anion nor hydrogen peroxide was involved in the transition metal ion or selenite inhibition of hAS3MT activity. The inhibition of arsenite methylating activity of hAS3MT by selenite was reversed by 2 mM DTT (dithiothreitol) but neither by cysteine nor by β-mercaptoethanol. Whereas, besides DTT, cysteine can also prevent the inhibition of hAS3MT activity by Co²⁺, Mn²⁺, and Zn²⁺. Free Cys residues were involved in the interactions of transition metal ions or selenite with hAS3MT. It is proposed that the inhibitory effect of the ions (Co²⁺, Mn²⁺, and Zn²⁺) or selenite on hAS3MT activity might be via the interactions of them with free Cys residues in hAS3MT to form inactive protein adducts.     

Studies on the mechanism of induction of haem oxygenase by cobalt and other metal ions.

Cobalt ions (Co2+) are potent inducers of haem oxygenase in liver and inhibit microsomal drug oxidation probably by depleting microsomal haem and cytochrome P-450. Complexing of Co2+ ions with cysteine or glutathione (GSH) blocked ability of the former to induce haem oxygenase. When hepatic GSH content was depleted by treatment of animals with diethyl maleate, the inducing effect of Co2+ on haem oxygenase was significantly augmented. Other metal ions such as Cr2+, Mn2+, Fe2+, Fe3+, Ni2+, Cu2+, Zn2+, Cd2+, Hg2+ and Pb2+ were also capable of inducing haem oxygenase and depleting microsomal haem and cytochrome P-450. None of these metal ions had a stimulatory effect on hepatic haem oxidation activity in vitro. It is suggested that the inducing action of Co2+ and other metal ions on microsomal haem oxygenase involves either the covalent binding of the metal ions to some cellular component concerned directly with regulating haem oxygenase or non-specific complex-formation by the metal ions, which depletes some regulatory system in liver cells of an essential component involved in controlling synthesis or activity of the enzyme.     

Interactions of selenium with metal ions at the cellular level

In a supplementation study in which organic selenium asl-selenomethionine was administered in low doses during 1 yr, alterations in the concentrations of metal ions in the erythrocytes and the neutrophil granulocytes were observed. In the erythrocytes, altered concentrations of zinc were parallel with selenium. The concentrations of magnesium, calcium, manganese, copper, and sulfur were not significantly altered. However, altered concentrations of iron and zinc were observed in the neutrophils. The concentrations of magnesium, calcium, manganese, copper, and sulfur were not significantly altered. The accumulation of selenium in individual blood cells was different from that obtained with supplementation of inorganic selenium. When organic selenium was supplemented, the thrombocytes accumulated more selenium than the erythrocytes and the neutrophil granulocytes. The observations indicate that selenium interacts with metal ions at the cellular level when supplemented in low doses. The chemical form of selenium might be important in nutrition and therapy in view of the interaction and distribution pattern at the cellular level.     

Full replacement of 2-mercaptoethanol by cysteine plus selenium compounds in augmenting DNA synthesis of mitogen-stimulated mouse spleen lymphocytes

Mouse spleen lymphocytes require 2-mercaptoethanol for maximal mitogenic activation in vitro. Previous studies indicate that the lymphocytes are defective in the cystine transport activity and that they require 2-mercaptoethanol to utilize cystine. 2-Mercaptoethanol catalytically carries cysteine moiety into the cells in a mixed disulfide form. Because cysteine is easily oxidized to cysteine in the culture medium, it has been not easy to precisely examine the effect of near-physiological concentrations of cysteine on the activation of lymphocytes. By controlling the cysteine content in the medium, we have reviewed the effect of cysteine to see if cysteine replaces 2-mercaptoethanol in enhancing the DNA synthesis of lipopolysaccharide-stimulated lymphocytes. It was found that cysteine was less effective than 2-mercaptoethanol, and that cysteine fully replaced 2-mercaptoethanol when a selenium compound was supplemented. The effects of cysteine and selenium compounds were apparently independent and additive. Among the selenium compounds examined, sodium selenite and L-selenocystine were much more effective in stimulating DNA synthesis than sodium selenate and L-selenomethionine.     

Mechanism of the inhibition of milk xanthine oxidase activity by metal ions: a transient kinetic study

The nature and mechanism of the inhibition of the oxidoreductase activity of milk xanthine oxidase (XO) by Cu(2+), Hg(2+) and Ag(+) ions has been studied by steady state and stopped flow transient kinetic measurements. The results show that the nature of the inhibition is noncompetitive. The inhibition constants for Cu(2+) and Hg(2+) are in the micromolar and that for Ag(+) is in the nanomolar range. This suggests that the metal ions have strong affinity towards XO. pH dependence studies of the inhibition indicate that at least two ionisable groups of XO are involved in the binding of these metal ions. The effect of the interaction of the metal ions on the reductive and oxidative half reactions of XO has been investigated, and it is observed that the kinetic parameters of the reductive half reaction are not affected by these metal ions. However, the interaction of these metal ions with XO significantly affects the kinetic parameters of the oxidative half reaction. It is suggested that this may be the main cause for the inhibition of XO activity by the metal ions.     

Gold complexes inhibit mitochondrial thioredoxin reductase: consequences on mitochondrial functions

The effects of gold(I) complexes (auranofin, triethylphosphine gold and aurothiomalate), gold(III) complexes ([Au(2,2'-diethylendiamine)Cl]Cl(2), [(Au(2-(1,1-dimethylbenzyl)-pyridine) (CH(3)COO)(2)], [Au(6-(1,1-dimethylbenzyl)-2,2'-bipyridine)(OH)](PF(6)), [Au(bipy(dmb)-H)(2,6-xylidine)](PF(6))), metal ions (zinc and cadmium acetate) and metal complexes (cisplatin, zinc pyrithione and tributyltin) on mitochondrial thioredoxin reductase and mitochondrial functions have been examined. Both gold(I) and gold(III) complexes are extremely efficient inhibitors of thioredoxin reductase showing IC(50) ranging from 0.020 to 1.42 microM while metal ions and complexes not containing gold are less effective, exhibiting IC(50) going from 11.8 to 76.0 microM. At variance with thioredoxin reductase, auranofin is completely ineffective in inhibiting glutathione peroxidase and glutathione reductase, while gold(III) compounds show some effect on glutathione peroxidase. The mitochondrial respiratory chain is scarcely affected by gold compounds while the other metal complexes and metal ions, in particular zinc ion and zinc pyrithione, show a more marked inhibitory effect that is reflected on a rapid induction of membrane potential decrease that precedes swelling. Therefore, differently from gold compounds, the various metal ions and metal complexes exert their effect on different targets indicating a lower specificity. It is concluded that gold compounds are highly specific inhibitors of mitochondrial thioredoxin reductase and this action influences other functions such as membrane permeability properties. Metal ions and metal complexes markedly inhibit the activity of thioredoxin reductase although to an extent lower than that of gold compounds. They also inhibit mitochondrial respiration, decrease membrane potential and, finally, induce swelling.     

The role of sulfhydryl compounds in mammalian melanogenesis: the effect of cysteine and glutathione upon tyrosinase and the intermediates of the pathway

The effect of cysteine and glutathione on mammalian melanogenesis has been studied. It has been shown that their action is mediated by two different mechanisms. (a) The reaction of the thiol groups with dopaquinone after the tyrosinase-catalyzed oxidation of tyrosine and dopa. This mechanism leads to the formation of sulfhydryl-dopa conjugates and finally sulfur-containing pigments, phaeomelanins instead of eumelanins. This fact might produce an inhibition of melanogenesis due to the slower rate of chemical reactions involved in the polymerization of such thiol-conjugates when compared to that of indoles. (b) The direct interaction between the sulfhydryl compounds and the tyrosinase active site. This interaction may regulate the activity of the enzyme. It is shown that Harding-Passey mouse melanoma tyrosinase is more sensitive to sulfhydryl compounds than mushroom tyrosinase. Cysteine always produces an inhibition of the tyrosinase hydroxylase and dopa oxidase activities of melanoma tyrosinase, this inhibition becoming greater as the cysteine concentration increases. On the other hand, glutathione produces an activation of the tyrosine hydroxylase activity below 3 mM and an inhibition at higher concentrations. The limit between the enzymatic activation and inhibition appears at glutathione concentrations similar to the physiological levels of this compound found in melanocytes. Although the switch from eumelanogenesis to phaeomelanogenesis occurs at much lower concentrations of glutathione, taking into account these data it is discussed that this sulfhydryl compound may regulate not only the type but also the amount of melanin formed inside melanocytes.     

Oxygen Enhancement of bactericidal activity of rifamycin SV on Escherichia coli and aerobic oxidation of rifamycin SV to rifamycin S catalyzed by manganous ions: the role of superoxide

Oxygen enhanced the bactericidal activity of rifamycin SV to Escherichia coli K12. Anaerobically grown cells, which had a low level of superoxide dismutase, were more susceptible to the bactericidal activity than aerobically grown cells, which contained a high level of superoxide dismutase. Oxygen also enhanced the inhibition of RNA polymerase activity of rifamycin SV, when Mn2+ was used as a cofactor. Rifamycin S was reduced to rifamycin SV by NADPH catalyzed by cell-free extracts of Escherichia coli K12. These results indicate that the inhibition of bacterial growth by rifamycin SV is due to the production of active species of oxygen resulting from the oxidation-reduction cycle of rifamycin SV in the cells. The aerobic oxidation of rifamycin SV to rifamycin S was induced by metal ions, such as Mn2+, Cu2+, and Co2+. The most effective metal ion was Mn2+. In the presence of Mn2+, accompanying the consumption of 1 mol of oxygen and the oxidation of 1 mol of rifamycin SV, 1 mol of hydrogen peroxide and 1 mol of rifamycin S were formed. Superoxide was generated during the autoxidation of rifamycin SV. Superoxide dismutase inhibited the formation of rifamycin S, but scavengers for hydrogen peroxide and the hydroxyl radical did not affect the oxidation. A mechanism of Mn2+-catalyzed oxidation of rifamycin SV is proposed and its relation to bactericidal activity is discussed.     

Inhibition of glutathione-insulin transhydrogenase by metal ions and activation by histidine and other chelating agents

The catalytic activity of purified glutathione-insulin transhydrogenase (thiol:protein-disulfide oxidoreductase/isomerase, EC 1.8.4.2) from bovine pancreas is markedly stimulated by histidine and other chelating agents. The activation produced was highest with EDTA, followed by EGTA, 8-hydroxyquinoline and 1,10-phenanthroline. Of the many amino acids tested, histidine was the only one that activated the enzyme; the structurally related compounds, 3-methylhistidine and imidazole also stimulated the enzyme, but 1-methylhistidine and histamine were without effect. The activation of EDTA was negated by metal ions, most effectively by Se2+, Hg2+, Cu2+ and Zn2+, and less effectively by Ca2+ and Ni2+. Likewise, activation by histidine was negated by Zn2+ but not by Ca2+ or Mg2+. Thus, activation of glutathione-insulin transhydrogenase is apparently achieved in part by the chelation of inhibitory metal ion(s). These findings are consistent with a regulatory scheme for glutathione-insulin transhydrogenase in which (a) the enzyme is inhibited by selenium and heavy metal ions normally present in tissues and (b) this inhibition can be relieved by the addition of histidine or chelating agents.     

Donor substrate specificity and thiol reduction of glutathione disulfide peroxidase.

By isolation of a mixed disulfide product of glutathione and cysteine, glutathione peroxidase was shown to be highly specific for only one donor substrate. Using the coupled assay of NADPH and yeast glutatione reductase, which is highly specific for flutathione disulfide, it was shown that the apparent inhibition of glutathione peroxidase by mercaptoethanol can be described kinetically and that it is competitive with glutathione. Also, when limiting amounts of hydroperoxide were present in the reaction mixture with mercaptoethanol or cysteine, the total amount of glutathione disulfide produced decreased as compared with that in a reaction mixture without mercaptoethanol or cysteine. This finding is consistent with enzymatic formation of mixed disulfides. Data presented suggest that the selenium in glutathione peroxidase was oxidized to a seleninic acid in the absence of glutathione. These results can be explained by a mechanism for glutathione peroxidase wherein the selenium atom is the only atom in the enzyme that undergoes oxidation reduction.     

The oxidation of Schiff bases of pyridoxal and pyridoxal phosphate with amino acids by manganous ions and peroxidase

1. Oxygen was taken up rapidly when pyridoxal or pyridoxal phosphate was added to mixtures of pea-seedling extracts and Mn(2+) ions. 2. The increases in total oxygen uptake were proportional to the pyridoxal or pyridoxal phosphate added and were accompanied by the disappearance of these compounds. 3. In addition to Mn(2+) ions, the reactions depended on two factors in the extracts, a thermolabile one in the non-diffusible material and a thermostable one in the diffusate; these factors could be replaced in the reactions by horse-radish peroxidase (donor-hydrogen peroxide oxidoreductase, EC 1.11.1.7) and amino acids respectively. 4. When pyridoxal phosphate was added to mixtures of amino acids and Mn(2+) ions oxygen uptake was rapid after a lag period of 30-90min.; the lag period was shortened to a few minutes by peroxidase, particularly in the presence of traces of p-cresol, or by light. 5. When pyridoxal replaced pyridoxal phosphate relatively high concentrations were required and peroxidase had only a small activating effect. 6. Pyridoxal or pyridoxal phosphate disappeared during the reactions and carbon dioxide and ammonia were formed. 7. With phenylalanine as the amino acid present, benzaldehyde was identified as a reaction product. 8. It is suggested that the reactions are oxidations of the Schiff bases formed between pyridoxal or pyridoxal phosphate and amino acids, mediated by a manganese oxidation-reduction cycle, and resulting in oxidative decarboxylation and deamination of the amino acids.     

Inhibition of rat brain prostaglandin D synthase by inorganic selenocompounds

Various inorganic selenocompounds dose-dependently inhibited the rat brain prostaglandin (PG) D synthase, both in the purified enzyme preparation and in the crude brain supernatant. All of the quadrivalent selenium compounds tested had a very limited range of IC50 values in the purified enzyme (11-12 microM) and in the brain supernatant (9-15 microM). A divalent selenium compound was also inhibitory, but a hexavalent selenium compound was ineffective. In contrast, organic selenocompounds such as selenomethionine and selenourea had no effect on the PGD synthase activity. Furthermore, sodium sulfate and sodium sulfite up to 10 mM did not inhibit the activity. The inhibition by selenium required the preincubation of the metal with sulfhydryl compounds such as dithiothreitol (DTT), indicating that the formation of selenotrisulfide or some other adduct(s) is essential for the inhibition. Furthermore, the inhibition was reversed by an excess amount of dithiothreitol, suggesting that the selenotrisulfide derivative of DTT binds to the SH group of the PGD synthase. The kinetic analysis revealed the inhibition by selenite to be noncompetitive with a Ki value of 10.1 microM. On the other hand, glutathione-dependent PGD synthase from rat spleen was much less inhibited, and PGF synthase and PGD2 11-ketoreductase activities were not inhibited by the selenium compound.