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Biomedical subjects

M Chevion

Publications and source records attributed to M Chevion.

At least 91 records · Page 5Linked to original sources
PubMed1984

Roles of copper and O(2) in the radiation-induced inactivation of T7 bacteriophage.

The effect of copper on the radiation damage induced in T7 bacteriophage has been investigated. The phages were gamma-irradiated and the effects of copper(II) ions in the presence of various additives and radical scavengers were examined in an attempt to better understand the effect of transition metal ions on the role of free radicals, particularly superoxide, in biological damage. The present work extends a study previously done on isolated enzyme to a whole biological entity. Copper(II) ions even at very low concentrations enhanced the lethal effect of radiation. This sensitization was observed in both the presence and the absence of oxygen. The effect of copper could be reverted by chelating agents such as EDTA or 1,10-phenanthroline. Hydrogen peroxide enhanced the sensitizing effect of copper, though little if any protection was provided by catalase or SOD. High molecular weight scavengers of free radicals in the presence of both copper(II) and hydrogen peroxide had no protective effect. (This is in contrast to metal-free systems where, although such scavengers are incapable of penetrating the phages, they protect them against inactivation.) These scavengers, without added H2O2, afforded only slight protection to the irradiated phages in the presence of Cu. Low molecular weight scavengers of free radicals reduced but did not eliminate the sensitizing effect of copper. The sensitizing effect of copper was also observed with other T-odd phages, but not with the T-even series. Copper(II) ions under similar experimental conditions did not sensitize T4 or T2 phages but rather had a protective effect. The results are interpreted in terms of a site-specific Fenton mechanism according to which the binding of the metal ion to the phages is a prerequisite for the occurrence of the biological damage. The results also indicate that most of the copper effect is endogenous. This is in accord with the failure of copper to sensitize the T-even phages, which differ by the rigidity and permeability of their outer coat structures.

Cesium Radioisotopes↗
PubMed1984

The effects of isouramil on erythrocyte mechanics: implications for favism.

Isouramil is one of the two aglycones derived from broad beans (Vicia faba) with the capacity to autoxidize and generate hydrogen peroxide. During its incubation with erythrocytes in vitro, it causes a marked decrease in cellular deformability at low shear forces, as measured by cone plate viscometry. At the same time the membrane shear modulus of elasticity, as determined by the dimensions of pressure induced pseudopods (PIPs), is increased. These experiments suggest that alterations in the mechanical properties of erythrocyte membranes may be the basis for changes in cellular deformability and hence, erythrocyte sequestration in G-6PD deficient cells exposed to peroxide-generating agents, such as isouramil.

Barbiturates↗
PubMed1983

The analogous mechanisms of enzymatic inactivation induced by ascorbate and superoxide in the presence of copper.

The mechanism of enzymatic inactivation of purified and membrane-bound acetylcholine esterase by ascorbate and copper was investigated. While the exposure of the enzyme to either ascorbate or copper did not cause enzymatic inactivation, the incubation of the enzyme with a combination of both ascorbate and copper resulted in a loss in acetylcholine esterase activity, which was time dependent. The enzymatic inactivation required either molecular oxygen or hydrogen peroxide under anaerobic conditions. Scavengers of hydroxyl radicals at concentrations of up to 100 mM did not provide protection to acetylcholine esterase. Only mannitol at very high concentrations (above 1 M) efficiently prevented the inactivation of the enzyme. The kinetics of the aerobic oxidation of reduced ascorbate in the presence of acetylcholine esterase and copper closely followed the rate of enzyme inactivation. Addition of the chelating agents EDTA and diethylenetriaminepentaacetic acid prevented both the oxidation of ascorbate and the inactivation of the enzyme. In the presence of low concentrations of histidine (0.5-2.0 mM), which forms high affinity complexes with copper, the rate of ascorbate oxidation was similar to that recorded in its absence. On the other hand, no enzyme inactivation was indicated in the presence of histidine. Low temperature EPR measurements have demonstrated the binding of copper to the enzyme, and have shown the reduction of the cupric enzyme to the corresponding cuprous complex. In view of these results, a general "site-specific" mechanism for biological damage can be offered, in which copper(II) ions are bound to enzymes or other biological macromolecules. Ascorbate plays a dual role: it reduces the cupric complex to the corresponding cuprous state and serves as a source for H2O2, which, in turn, reacts with the reduced copper complex, in a Fenton reaction. In this reaction, secondary hydroxyl radicals are site specifically formed, and react preferentially with the protein, at the site of their formation, causing its inactivation. This mechanism is analogous to that previously proposed (Samuni, A., Chevion, M., and Czapski, G. (1981) J. Biol. Chem. 256, 12632-12635) for the enhancement of the biological damage caused by superoxide in the presence of copper.

Acetylcholinesterase↗
PubMed1983

On the cytotoxicity of vitamin C and metal ions. A site-specific Fenton mechanism.

The toxicity of ascorbate towards phage lambda and the phages T2-T7 has been investigated. At room temperature the T-odd and lambda bacteriophages are highly susceptible to ascorbate-induced damage, whereas the T-even phages are practically resistant. The toxicity of ascorbate is dependent on the presence of copper (or iron) and oxygen, although oxygen is not required in the presence of H2O2. Hydrogen peroxide is essential for the ascorbate-induced phage inactivation and the damage is prevented by catalase. At the concentrations used, most of the copper ions are bound to the phage particles. Chelating agents such as EDTA or histidine fully protect the phages, whereas salicylate only reduces the rate of phage inactivation. OH scavengers such as sucrose, formate, mannitol, tert-butyl alcohol or poly(ethylene glycol) have no protective effect. Experiments with DNA labeled phages indicate that both phage adsorption and DNA injection are impaired as a result of the exposure to ascorbate and copper. The failure to express the viral genetic information as a result of single and double-strand breaks in the DNA, probably also contribute to the loss of the plaque-forming ability of the phages. The results are interpreted in terms of a 'site-specific' Fenton mechanism according to which the binding of the transition metal ions to the biological target is a prerequisite for the production of damage. The bound metal ion is reduced either by O(2), ascorbate or other reductants and is subsequently reoxidized by H2O2 yielding OH. radicals. This cyclic redox reaction of the metal generates OH. radicals which react with vital macromolecules with a high probability of causing 'multi-hit' damage. This 'site-specific' formation of OH. radicals, which takes place near the target molecules, accounts both for the high damaging efficiency and for the failure of OH. scavengers to protect against it.

Ascorbic Acid↗
PubMed1983

Copper and zinc levels in normal and malignant tissues.

The copper and zinc levels in 53 malignant and 47 normal human tissue samples were measured. In the malignant tissues, the mean copper concentration was 46% higher (P less than 0.001) than in the normal ones. Analysis of the individual organs showed this increment to be statistically significant in malignancies of the large bowel, stomach, urinary bladder and female reproductive organs, while in cancer of the breast, kidney and testis, the increase in copper level was not significant. The mean zinc concentration in the malignant tissues was not significantly different from that in the corresponding normal tissue specimens (-11%; P less than 0.2). In breast cancer, however, tissue zinc levels were increased by 72% (P less than 0.01), and decreased markedly in carcinoma of the kidney (-73%). A hypothesis for the possible mechanism involving elevated tissue copper levels in biological damage (previously caused by free radicals) which may be responsible for the malignant process, is presented and discussed.

Breast Neoplasms↗
PubMed1983

Reaction of nitric oxide with heme proteins: studies on metmyoglobin, opossum methemoglobin, and microperoxidase.

Kinetic and EPR studies show that the first step in the reaction of NO with ferric myoglobin, opossum hemoglobin, and microperoxidase is the reversible formation of the H-NO complex: H + NO in equilibrium H-NO (where H = Mb+, or Hb+ OP, or MP+). The NO-combination rates are markedly affected by the presence or absence of the distal histidine. The distal histidine significantly reduces the NO-combination rates, perhaps by interaction between the distal histidine and the ferric iron. Thus the beta-chains of Hb+ OP and metmyoglobin show similar combination rates. In the absence of a distal histidine, the NO-combination rates in the alpha-chains of Hb+ OP are much faster and similar to those observed for the five-coordinate heme in microperoxidase. The loss of a water molecule from the six-coordination site is assumed to be the rate-limiting step.

Animals↗
PubMed1983

A novel method for quantitation of favism-inducing agents in legumes.

A new method for the quantitation of the favism-inducing agents in legumes is described. The procedure involves differential extraction of the glucosides vicine and convicine with acetic acid (25%), followed by an enzymatic hydrolysis by beta-glucosidase under anaerobic conditions. Each of the aglycone moieties, isouramil and divicine, anaerobically reduces two molecules of o-ferriphenanthroline to o-ferrophenanthroline. This reaction is readily followed spectrophotometrically at 515 nm. Using this procedure, it was found that in various strains of Vicia faba, the level of these two glucosides comprises approximately 0.5% of the wet weight of the seeds. In contrast, these glucosides could not be detected in either green peas or chick peas.

Barbiturates↗
PubMed1983

Inhibitory effect of a fava bean component on the in vitro development of Plasmodium falciparum in normal and glucose-6-phosphate dehydrogenase deficient erythrocytes.

We examined the hypothesis that G-6-PD deficiency associated with fava bean ingestion confers resistance to malaria by studying the in vitro interactions between malaria parasites (Plasmodium falciparum), human erythrocytes with varying degrees of G-6-PD deficiency, and isouramil (IU), a fava bean extract that is known to cause oxidant stress and hemolysis of G-6-PD-deficient erythrocytes. Untreated G-6-PD-deficient and normal erythrocytes supported the in vitro growth of P. falciparum equally well. However, after pretreatment with IU, G-6-PD-deficient erythrocytes did not support parasite growth in vitro, whereas growth remained high in normal erythrocytes. Parasite growth was proportional to the G-6-PD activity of the IU-treated erythrocytes. In contrast, when parasitized erythrocytes were exposed to IU, parasites even in normal erythrocytes were destroyed. Ring forms were much less sensitive than late trophozoites and schizonts. The results suggest that there are two modes by which IU affects the development of P. falciparum and demonstrate in vitro that G-6-PD deficiency confers resistance against malaria under conditions of fava-bean-associated oxidant stress.

Barbiturates↗
PubMed1982

The chemistry of favism-inducing compounds. The properties of isouramil and divicine and their reaction with glutathione.

Isouramil and divicine are pyrimidine aglycones of two glucosides found in broad beans. They have been shown to be strong reducing agents. Their reaction with oxygen in a (gas) saturated solution, 26 degrees C, is characterized by tau 1/2 = 1 min and 3 min respectively. Hydrogen peroxide is formed in this reaction stoichiometrically (1:1). The pyrimidines lose two hydrogen and form an intermediate that is structurally analogues to alloxan. This intermediate is not stable, and in the absence of reducing agents it decomposes, possibly by ring-cleavage. In the presence of reduced glutathione the intermediate is reduced and can now react with oxygen once again. Thus, the pyrimidines cycle between the two states and the net reaction is the catalytic oxidation of glutathione by molecular oxygen; in each cycle 4 molecules of glutathione are dissipated. The possible involvement of these pyrimidines in the pathogenesis of favism may be in a similar mechanism. Red blood cells deficient in glucose-6-phosphate dehydrogenase cannot cope with such an oxidative challenge exerted by the pyrimidines. Consequently an irreversible cellular damage can take place leading to the enhanced sequestration of these red blood cells by the reticuloendothelial system.

Animals↗
PubMed1980

Chain inequivalence in bovine methemoglobin.

Using pulse radiolysis, a single heme in the tetramer of bovine methemoglobin was reduced within a few microseconds to the ferro state, producing a valence intermediate. The kinetics of oxygen binding to the valence intermediate as well as the re-oxidation of the ferro-heme to the ferric state were studied as a function of pH. The kinetics of the oxygenation revealed the existence of two species, characterized by high and low affinities for oxygen that are associated with two quaternary structures (R and T, respectively). A sigmoidal curve representing a transition between the two states as a function of pH was derived. Above pH 7.7 only the R state could be observed, while below pH 6.5 the T state was dominant. The reaction between the valence intermediate and ferricyanide at pH 7.75 (R state) consisted of two (about) equal contributions (k1 = 23 x 10(4) M-1 S-1; k2 = 2.1 x 10(4) M-1 S-1) attributed to the beta and alpha subunits within the tetramer, respectively. At pH 6.3 (T state) a similar phenomenon was observed (k1 = 69 x 10(4) M-1 S-1; k2 = 3.7 x 10(4) M-1 S-1), indicating chain inequivalences both in the T and the R states of methemoglobin. In the presence of inositol hexakisphosphate the T leads to R transition, as monitored by oxygenation of the valence intermediate, was shifted up to a higher pH by about 0.35. Yet similar rate constants exhibiting similar chain inequivalences have been measured.

Animals↗
PubMed1979

Quaternary structure of methemoglobin. Pulse radiolysis study of the binding of oxygen to the valence hybrid.

The pulse radiolysis of solutions of adult human methemogolbin was used in order to reduce a single heme iron within the protein tetramers. The valence hybrids thus formed were reacted with oxygen. Kinetics of the reactions were studied. The effects of pH and inositol hexaphosphate were examined. The kinetics of the ligation of oxygen to stripped valence hybrids showed a single phase behavior at the pH range 6.5 to 9. As the pH was lowered below 6.5, a second, slower phase became apparent. In the presence of inositol hexaphosphate, above pH 8, the kinetics of oxygen binding was of a single phase. As the pH was lowered, a transition to a second, slower phase was noticed. Below pH 7, the slower phase was the only detectable one. The analysis of the relative contribution of the faster phase to the total reaction as a function of the pH showed a typical transition curve characterized by a pK = 7.5 and a Hill parameter n = 2.9. On this basis, it is concluded that human adult stripped methemoglobin resides in an R quarternary structure, while the presence of IHP stabilizes the T structure at pH below 7.5. This transition between the quaternary structures of methemoglobin cannot be accounted for by the switch between the high spin and the low spin states of the ferric iron. This switch of spin state takes place at pH greater than 8.2.

Heme↗
PubMed1978

Analogous effect of protons and inositol hexaphosphate on the alteration of structure of nitrosyl fetal human hemoglobin.

We have determined the low temperature EPR spectra and room temperature ligand dissociation rate constants of human NO-hemoglobins F and A as a function of pH and inositol hexaphosphate levels in order to assess the contribution of a quaternary structural equilibrium in the two proteins to their spectral and functional properties. Our results are consistent with an increased stability of a ligated low affinity structure in the fetal protein; the functional properties of this structure appear to be essentially the same in the two hemoglobins, even though its stability relative to a high affinity conformation is different. The pH dependence of the NO dissociation constant in both adult and fetal hemoglobin can be assigned primarily to the pH-dependent equilibria of high and low affinity forms as monitored by EPR.

Adult↗
PubMed1978

Quaternary states of methemoglobin and its valence-hybrid. A pulse radiolysis study.

Using the pulse radiolysis technique on solutions of stripped adult human methemoglobin, we found that the heme-iron within a single subunit in the tetramer was reduced to iron(II). The valence-hybrid thus formed was reacted with oxygen and with carbon monoxide. Kinetics of the reactions were studied. The effects of pH, inositol hexaphosphate, and temperature on these reactions were examined. The kinetics of the ligation of O2 and CO were used to characterize the affinity states of the valence-hybrid and its parent methemoglobin. Our results support the description of stripped methemoglobin A as residing in an R state. In the presence of inositol hexaphosphate methemoglobin is stabilized in a T state, but it switches into a high affinity state when the pH is raised a0ove 8.0. This structural transition was not found to coincide with the switch of spin state of the heme-iron that accompanies the ionization of water in aquomethemoglobin A.

Adult↗
PubMed1977

Imidazole, the ligand trans to mercaptide in ferric cytochrome P-450. An EPR study of proteins and model compounds.

A crystal field analysis of EPR data for various low spin ferric cytochromes P-450 suggests that in all of them, regardless of source or method of induction, the heme ligands are a sulfur atom, presumably from cysteine, and an imidazole from histidine. The imidazole can be displaced in the ferric protein by cyanide, guanidine, or by an amine, analogous to its displacement by CO or NO in the ferrous protein. The resulting changes in the EPR parameters for the ferric protein are consistent with similar substitutions in heme thiol model compounds. The analysis of the latter can be understood on the basis of alterations of the electronic structure of the ligands to the heme iron.

Animals↗