Search PubMed⌕ Search

Biomedical subjects

A Virion

Publications and source records attributed to A Virion.

At least 37 records · Page 2Linked to original sources
PubMed1986

Solubilization and characteristics of the thyroid NADPH-dependent H2O2 generating system.

Solubilization of the thyroid particulate-associated NADPH-dependent H2O2 generating system has been tested with different detergents; (3-(3-cholamidopropyl)-dimethylammonio)1-propane sulfonate (CHAPS) was found to be the best of the six detergents tested. The ratio of H2O2 generation to NADPH oxidation was similar for CHAPS extract and native particulate material. CHAPS was also the only detergent able to preserve the Ca++-sensitivity of the NADPH oxidase. Solubilization of this enzyme allowed the determination of some of its characteristics: specificity for divalent cations, apparent Km for NADPH, optimum pH and sensitivity to SH- reagents.

Animals↗
PubMed1985

NADPH-dependent generation of H2O2 in a thyroid particulate fraction requires Ca2+.

The action of Ca2+ on H2O2 formation catalyzed by an NADPH-dependent H2O2-generating system was studied in a thyroid particulate fraction. It was shown that H2O2 formation was inhibited by EGTA and restored in the presence of micromolar concentrations of free Ca2+, and that Mg2+ was unable to restore the activity of the EGTA-inhibited particulate fraction. Consequently, NADPH-dependent H2O2 formation requires Ca2+, and the Ca2+ stimulation of this process is specific and reversible.

Animals↗
PubMed1985

NADPH oxidation catalyzed by the peroxidase/H2O2 system. Guaiacol-mediated and scopoletin-mediated oxidation of NADPH to NADPH+.

We have examined the respective roles played by guaiacol and scopoletin in NADPH oxidation catalyzed by the peroxidase/H2O2 system. It was shown that NADPH was not oxidized by either the horseradish or lactoperoxidase/H2O2 systems alone; oxidation occurred immediately after the addition of guaiacol or scopoletin. In both cases, the oxidation product was enzymatically active NADP+. Differences were observed in the NADPH oxidation mechanism depending on whether guaiacol or scopoletin was the mediator molecule. In guaiacol-mediated NADPH oxidation, the stoichiometry between H2O2 and oxidized NADPH was about 1; superoxide dismutase did not affect the oxidation rate. In scopoletin-mediated oxidation, the stoichiometry was much higher (1:14 in the present experiments); superoxide dismutase considerably increased the oxidation rate. It is concluded that catalysis of NADPH oxidation by the horse radish peroxidase/H2O2 system requires the presence of a mediator molecule. The NADPH oxidation mechanism depends on the intermediary oxidation state of this molecule.

Catalysis↗
PubMed1985

NADPH oxidation catalyzed by the peroxidase/H2O2 system. Iodide-mediated oxidation of NADPH to iodinated NADP.

Oxidation of NADPH catalyzed by the peroxidase/H2O2 system is known to require the presence of mediating molecules. Using either lactoperoxidase or horseradish peroxidase, we demonstrated that in the peroxidase/H2O2 system, NADPH oxidation was mediated by iodide. The oxidation product was the iodinated NADP. This product was shown to possess spectral characteristics different from those of NADP+ and NADPH, since for iodinated NADP, increased absorbance was observed in the 280-nm region and was directly proportional to the rate of iodination. It is suggested that oxidation and iodination of NADPH proceed via a single reaction between the intermediary iodide oxidation species and NADPH. Experiments with different molecules of NADPH analogues indicated that iodination occurred in the nicotinamide part of the NADPH molecule.

Catalysis↗
PubMed1985

Thyroid hormone synthesis and thyroglobulin iodination related to the peroxidase localization of oxidizing equivalents: studies with cytochrome c peroxidase and horseradish peroxidase.

Cytochrome c peroxidase (CcP) and horseradish peroxidase (HRP), when combined with a stoichiometric amount of H2O2, form stable compounds I which are known as FeIV Ro and FeIV o pi + structures, respectively. These compounds were assayed in the catalysis of thyroid hormone synthesis and the iodination reaction. As previously shown for the lactoperoxidase FeIV Ro compound, the CcP FeIV Ro compound was involved in the coupling and not in the iodination reactions. In contrast, the HRP FeIV o pi + compound catalyzed both iodination and hormone formation. The possible role of the different peroxidase-H2O2 compounds in the two sequential reactions, thyroglobulin iodination and thyroid hormone formation, is discussed.

Catalysis↗
PubMed1985

Spectral characteristics and catalytic properties of thyroid peroxidase-H2O2 compounds in the iodination and coupling reactions.

Hog thyroid peroxidase (TPO) was highly purified in order to study the spectral properties and catalytic specificities of its H2O2 compounds in iodothyronine biosynthesis. Purified TPO exhibited a Soret spectrum with an absorption maximum at 410 nm and had an A410/A280 value of 0.55. Protein iodination was only catalyzed under conditions which allowed formation of the transient TPO compound I (Fe(IV)-pi o+). On addition of an equimolar amount of H2O2, TPO formed a stable compound with an absorption maximum at 417 nm. This compound efficiently catalyzed the coupling reaction, but was unable to iodinate proteins. It catalyzed the formation of 1 mol iodothyronines/mol TPO, and therefore retained two oxidizing equivalents per molecule. It is proposed that this compound constitutes a second form of compound I whose structure might be Fe(IV)-Ro, analogous to that of cytochrome c peroxidase compound I. In the presence of an excess of H2O2, it formed TPO-compound III with an absorption maximum at 420 nm. TPO-compound III catalyzed neither the iodination nor the coupling reaction.

Animals↗
PubMed1985

Relation between thyroid peroxidase, H2O2 generating system and NADPH-dependent reductase activities in thyroid particulate fractions.

In thyroid gland, iodination takes place on the apical plasma membrane and requires the presence of the thyroid peroxidase and H2O2 generating system. H2O2 generation and NBT (nitro blue tetrazolium) reductase activity (both of which are NADPH-dependent) as well as peroxidase activity were compared for their respective orientations in membrane vesicles. The possible role of NADPH-NBT reductase activity in H2O2 generation was also examined. Results favor the conclusion that thyroid peroxidase is oriented towards the luminal side of the vesicles, whereas the NADPH site of NADPH oxidase-dependent H2O2 generation is located on the external side of the same or of different vesicles. Furthermore, it is shown that different NADPH-NBT reductase activities are present on both the outer and inner surfaces of the membrane vesicles, and that none of these activities is able to produce either H2O2 or O-2. The idea that a multi-component complex is involved in H2O2 generation is discussed, and a model is proposed which takes into account the possible spatial separation of the thyroid peroxidase site from the NADPH site of this H2O2 generation system on the apical membrane of the thyrocyte.

Animals↗
PubMed1984

Reduction of lactoperoxidase-H2O2 compounds by ferrocyanide: indirect evidence of an apoprotein site for one of the two oxidizing equivalents.

The titration by ferrocyanide and the localization of the oxidizing equivalents of lactoperoxidase "compound II" were studied as a function of pH. It was demonstrated that 1) whatever the pH, the structure of lactoperoxidase "compound II" was compatible with a Fe IV R degree state, 2) at acidic pH, ferrocyanide preferentially reduced the oxidizing equivalent localized on the heme iron to give an Fe III R degree compound, 3) at pH 4.2 only the Fe III R degree form was obtained after reduction of lactoperoxidase "compound II" with one mole of ferrocyanide and whereas at pH greater than 4.2, a mixture of both Fe III R degree and Fe IV R forms was present, 4) lowering the pH from 7.2 to 4.0 induced a transition of Fe IV R state to Fe III R degree state, but increasing the pH from 4.0 to 7.2 did not permit the formation of Fe IV R compound from Fe III R degree compound.

Apoenzymes↗
PubMed1984

NADPH-dependent H2O2 generation and peroxidase activity in thyroid particular fraction.

A NADPH-dependent H2O2 generating system associated with a thyroid particular fraction is described. H2O2 is measured by two different methods: iodination of NADPH itself when the system is supplemented with lactoperoxidase and [125I]iodide, and by the scopoletin method. It is shown that: H2O2 generation is inhibited by catalase and is dependent on NADPH or particulate protein concentration; radical scavengers of OH and of singlet oxygen have no effect while superoxide dismutase has only a marginal effect; disruption of the particular fraction by phospholipase A2 or digitonin treatment completely abolished H2O2 generation activity while thyroid peroxidase activity appears, suggesting different sites for the two activities in the membrane vesicles.

Animals↗
PubMed1980

Opposite effects of thiocyanate on tyrosine iodination and thyroid hormone synthesis.

The effect of a pseudohalide, SCN-, an anion with the same molecular size as iodide, was studied on two reactions: thyroglobulin iodination and thyroid hormone synthesis (coupling reaction) catalyzed by peroxidases. The coupling reaction was studied separately from the iodination reaction by using labelled thyroglobulin samples previously iodinated but containing little or no hormones. 1. SCN- inhibits iodide oxidation (I- leads to I2) whatever the enzyme, thyroid, lactoperoxidase or horseradish peroxidase. The amount of SCN- required to completely inhibit this reaction varies depending on the enzyme. Similarly tyrosine iodination is inhibited by SCN- with large variations, depending on the peroxidase, in the concentration of this anion required for inhibition. 2. In contrast SCN- stimulates the coupling reaction: (a) this affect is seen with the thyroid and lactoperoxidases but not with horseradish peroxidase; (b) the concentration of SCN- required for half-maximal stimulation of the coupling reaction is much lower (0.5-1 microM) than that required for the inhibition of iodide oxidation (60-80 microM); (c) ClO4(-), an anion with the same molecular size as SCN- and I-, has no effect on the coupling reaction; (d) this stimulatory effect of SCN- does not depend on a modification of the thyroglobulin molecule since it is not seen with horseradish peroxidase or in purely chemical coupling conditions. 3. The stimulatory effect of SCN- is therefore seen as resulting from the binding of this anion to a limited number of high-affinity sites present at the surface of both thyroid and lactoperoxidases. The inhibitory effect depends, in contrast, on the binding of SCN- to the substrate site with lower affinities. Since iodide also behaves both as a substrate for the iodination reaction and as a stimulatory ligand for the coupling reaction, these data provide further support in favour of the existence of an enzyme-iodide (or SCN-) complex with catalytic properties different from those of the native peroxidase.

Dose-Response Relationship, Drug↗
PubMed1977

[Binding sites for 3H-Leu-enkephalin in rat straitum].

The binding of 3H-Leu-Enkephalin to a particulate fraction from Rat striatum has been investigated in the presence of 20 mum bacitracin which prevents its hydrolysis. After deduction of a low-affinity, "non-saturable" component, the saturation kinetics at equilibrium provide evidence for two distinct saturable sites. The first exhibits Michaelis kinetics with a Kd value of 2.7 +/- 0.1 nM; both morphine and naloxone compete with the binding of 3H-Leu-Enkephalin on this site but not on the second one. The marked decrease in binding of 3H-Leu-Enkephalin observed in the presence of high concentration of sodium ions indicates that these two pentapeptides have the same properties as morphinomimetic analgesics. Taken together, these data indicate that the first site is identical to the "opiate receptor", while the nature of the second saturable site remains to be established.

Analgesics↗
PubMed1976

Phosphorylation of microtubule-associated proteins.

1. Tubulin is not an adenosine-3':5'-monophosphate-dependent (cyclic-AMP-dependent) protein kinase. Both entities have been clearly separated by sucrose gradient ultracentrifugation. With a tubulin preparation obtained by the polymerization-depolymerization technique protein kinase had a sedimentation coefficient of 8.7 S whereas tubulin sedimented with 6.4 S. After preincubation with both cyclic AMP and histone the kinase dissociated into its catalytic subunit with a sedimentation coefficient of 3.4 S. 2. Tubulin prepared by the polymerization-depolymerization technique was neither phosphorylated in vivo nor in vitro. On the contrary if this preparation was further purified by the Weisenberg's procedure (DEAE-Sephadex batch absorption) before incubation with [gamma-32 P]ATP, phosphorylation occurred. Thus, phosphorylation depended on the method used to purify tubulin i.e. was likely to an an artefact.

Brain↗