Biomedical subjects

R K Banerjee

Publications and source records attributed to R K Banerjee.

At least 37 records · Page 2Linked to original sources

PubMed1996

Lactoperoxidase-catalysed oxidation of indomethacin, a nonsteroidal antiinflammatory drug, through the formation of a free radical.

Lactoperoxidase (LPO, EC 1.11.1.7; donor-H2O2 oxidoreductase) catalyses the oxidation of indomethacin, a nonsteroidal antiinflammatory drug by H2O2 as measured by time-dependent decay of indo-methacin extinction at 280 nm and concurrent appearance of stable oxidation product(s) at 412 nm. From a plot of log Vmax against varying pH of indomethacin oxidation, involvement of an ionizable group of the enzyme having pka = 5.7 could be ascertained for controlling the oxidation process. Spectral studies revealed that LPO-compound II oxidises indomethacin through one-electron transfer and is reduced to the native ferric state as shown by its spectral shift from 430 nm to 412 nm through an isosbestic point at 421 nm. The one-electron oxidation product is a nitrogen-centered free radical detected as a 5,5-dimethyl-l-pyrroline N-oxide (DMPO) adduct (alpha N = 15 G, alpha H beta = 16 G) in electron spin resonance spectroscopy. The free radical is scavenged by reaction with O2 as shown by O2 consumption sensitive to the free-radical trap, DMPO. Binding studies by optical difference spectroscopy indicate that indomethacin binds to LPO with an apparent KD value of 24.5 microM. The free energy change, delta G', for the binding is -26.3 KJ mol-1, suggesting that the interaction is favourable for oxidation. Indomethacin binding remains unaltered by a change of pH from 5.25 to 7.5, presumably because of hydrophobic interaction. The binding is competitive with resorcinol, an aromatic electron donor, showing the KD value to be as high as 100 microM. We suggest that indomethacin interacts at the aromatic donor binding site and is oxidised by one-electron transfer by LPO catalytic intermediates to stable oxidation product(s) through the formation of a free radical.

Animals↗

PubMed1996

EDTA inhibits lactoperoxidase-catalyzed iodide oxidation by acting as an electron-donor and interacting near the iodide binding site.

Ethylenediamine tetraacetate (EDTA) inhibits lactoperoxidase (LPO)-catalyzed rate of iodide oxidation in concentration and pH-dependent manner. A plot of log Kiapp values against various pH yields a sigmoidal curve from which an ionisable group of pKa value 6.0 could be ascertained for controlling the inhibition of catalytically active LPO by EDTA. Kinetic studies indicate that EDTA competitively inhibits iodide oxidation by acting as an electron donor. EDTA al so reduces LPO-compound-11 to the native ferric state by one-electron transfer as evidenced by the spectral shift from 428 to 412 nm. Optical difference spectroscopic studies indicate that EDTA binds to LPO with the apparent equilibrium dissociation constant (KD) of 12 +/- 2 mM at pH 6.5. A plot of log KD values against various pH produces a sigmoidal curve from which an ionisable group of LPO having pKa = 5.47 could be calculated, deprotonation of which favours EDTA binding. EDTA also binds to LPO-CN-complex indicating its binding site away from heme iron centre. The KD of LPO-EDTA complex is significantly increased (62 +/- 5 mM) by iodide suggesting that EDTA binds close to the iodide binding site. EDTA also increases the KD value of LPO-hydroquinone complex from 62 +/- 5 mM to 200 +/- 21 mM indicating that EDTA and aromatic donor binding sites are also close. We suggest that EDTA inhibits iodide oxidation competitively as an electron donor by interacting at or near the iodide binding site and these sites are close to the aromatic donor binding site.

Binding Sites↗

PubMed1996

Probing the active site residues in aromatic donor oxidation in horseradish peroxidase: involvement of an arginine and a tyrosine residue in aromatic donor binding.

The plausible role of arginine and tyrosine residues at the active side of horseradish peroxidase (HRP) in aromatic donor (guaiacol) oxidation was probed by chemical modification followed by characterization of the modified enzyme. The arginine-specific reagents phenylglyoxal (PGO), 2,3-butanedione and 1,2-cyclohexanedione all inactivated the enzyme, following pseudo-first-order kinetics with second-order rate contents of 24M(-1.)min(-1), 0.8M(-1.)min(-1) and 0.54M(-1.)min(-1) respectively. Modification with tetranitromethane, a tyrosine-specific reagent, also resulted in 50% loss of activity following pseudo-first-order kinetics with a second-order rate constant of 2.0M(-1.)min(-1). The substrate, H2O2, and electron donors such as I- and SCN- offered no protection against inactivation by both types of modifier, whereas the enzyme was completely protected by guaiacol or o-dianisidine, an aromatic electron donor (second substrate) oxidized by the enzyme. These studies indicate the involvement or arginine and tyrosine residues at the aromatic donor site of HRP. The guaiacol-protected phenylglyoxal-modified enzyme showed almost the same binding parameter (Kd) as the native enzyme, and a similar free energy change (deltaG')for the binding of the donor. Stoicheiometric studies with [7-14C]phenylglyoxal showed incorporation of 2 mol of phenylglyoxal per mol of enzyme, indicating modification of one arginine residue for complete activation. The difference absorption spectrum of the tetranitromethane-modified against the native enzyme showed a peak at 428 nm, characteristic of the nitrotyrosyl residue, that was abolished by treatment with sodium dithionite, indicating specific modification of a tyrosine residue. Inactivation stoicheiometry showed that modification of one tyrosine residue per enzyme caused 50% inactivation. Binding studies by optical difference spectroscopy indicated that the arginine-modified enzyme could not bind guaiacol at all, whereas the tyrosine-modified enzyme bound it with reduced affinity (Kd 35mM compared with 10mM for the native enzyme). Both the modified enzymes, however, retained the property of the formation of compound II (one-electron oxidation state higher than native ferriperoxidase) with H2O2, but reduction of compound II to native enzyme by guaiacol did not occur in the PGO-modified enzyme, owing to lack of binding. No non-specific change in protein structure due to modification was evident from circular dichromism studies. We therefore suggest that the active site of HRP for aromatic donor oxidation is composed of an arginine and an adjacent tyrosine residue, of which the former plays an obligatory role in aromatic donor binding whereas the latter residue plays a facilitatory role, presumably by hydrophobic interaction or hydrogen bonding.

Arginine↗

PubMed1996

Characterization of sheep lacrimal-gland peroxidase and its major physiological electron donor.

A soluble sheep lacrimal-gland peroxidase was purified to apparent homogeneity. It had a native molecular mass of 75 kDa with a subunit molecular mass of 82 kDa and an isoelectric point of 6.5. Western blotting showed that it shares some of the enzyme antigenic determinants in common with other soluble peroxidases. The enzyme exhibits a Soret peak at 410 nm which is shifted to 431 nm by 5 equiv. of H2O2 due to the formation of compound II. The latter is, however, unstable and gradually returns to the native state. The enzyme forms complexes with CN- and N3- and is reduced by dithionite showing a characteristic reduced peroxidase spectrum. Although the enzyme oxidizes I-, SCN- and Br- optimally at pH 5.5., 5.25 and 5.0 respectively, at physiological pH, it oxidizes I- and SCN- only. Since extracellular SCN- concentration is much higher than I-, SCN- may act as the major electron donor to the enzyme. The second-order rate constants for the reaction of the enzyme with H2O2 (k+1) and of compound I with SCN- (k+2) were 4 X 10(7) M-1 X s-1 and 8.1 X 10(5) M-1 X s-1 respectively. A plot of log Vmax against pH yields a sigmoidal curve consistent with a single ionizable group on the enzyme with a pK(a) value of 5.75, controlling thiocyanate oxidation. In a coupled system with the peroxidase, H2O2, SCN-, GSH, NADPH and glutathione reductase, peroxidase-catalysed SCN- oxidation by H2O2 could be coupled to NADPH consumption. The system is proposed to operate in vivo for the efficient elimination of endogenous H2O2.

Animals↗

PubMed1996

Hemodynamic changes in recurrent intracranial terminal aneurysm after endovascular treatment.

RATIONALE AND OBJECTIVES: Multiple cases of recurrence of aneurysms after endovascular treatment have been reported. The purpose of the current hemodynamic study was to identify changes in shear stress and pressure associated with the recurrence of terminal intracranial aneurysms after endovascular occlusion. METHODS: Using a finite element method, a pulsed flow with a non-Newtonian viscosity of blood was simulated within the aneurysm cavity. A recurrent terminal intracranial aneurysm of a patient originally treated with balloon occlusion was then studied. This was based on a physiologic pulsatile flow, which was observed in the middle cerebral artery. Before and after the balloon occlusion, local maximum wall shear stress and pressure drop at the neck of the aneurysm were calculated and compared with the normal shear stress. RESULTS: Although the maximum shear stress at the right neck of the aneurysm was significantly reduced after balloon insertion, it was still 2.5 times greater than the normal maximum shear stress. This was attributable to the presence of a portion of the aneurysmal neck (residual neck), which was not obliterated by the balloon. The balloon also helped to reduce the maximum pressure inside the aneurysm by approximately 15%. Hemodynamic changes in the residual aneurysm neck, where the shear stress and the pressure are high, and other factors may be responsible for the recurrence of aneurysms after balloon or coil occlusion. CONCLUSION: Residual necks after balloon occlusion, coil occlusion, or both are attributable to the geometric orientation of the aneurysm with respect to parent and daughter vessels and the variety of configurations of the balloon and coils used to occlude the aneurysms. Inadequate reduction in local shear stress found in these residual necks is an important factor in the recurrence and rupture of the aneurysm after endovascular occlusion.

Blood Flow Velocity↗

PubMed1996

Hamadryas baboons Papio hamadryas as maintenance hosts of Schistosoma mansoni in Saudi Arabia.

Free ranging hamadryas baboons (Papio hamadryas) in four localities in the west and north of Saudi Arabia were examined for natural infection with Schistosoma mansoni. Faecal examination revealed infection with S. mansoni on four occasions within one year (at a prevalence rate of 2.5-4.0%) in only one locality, the Al-Baha area. The eggs were viable, as shown by miracidial hatching tests, and were recorded at a density of 140-280 eggs/g of faeces (7000-14,000 eggs/day). Post-mortem examination of 13-24 baboons from each locality revealed infection with S. mansoni (adult worms and eggs in tissue) in only one locality, the Al-Baha area, at a prevalence rate of 4.16%. Viable eggs were found in the faeces and tissue of the infected baboons. The low prevalence rate of S. mansoni in hamadryas baboons in Saudi Arabia is in accordance with the low prevalence rate of S. mansoni in humans in the area. This natural baboon isolate was highly infective to snail intermediate hosts and mammalian hosts under experimental conditions. The epidemiological significance of the role of P. hamadryas (considering their large overall population of 250,000) as maintenance hosts of S. mansoni in Saudi Arabia is discussed.

Animals↗

PubMed1995

Concurrent reduction of iodine and oxidation of EDTA at the active site of horseradish peroxidase: probing the iodine binding site by optical difference spectroscopy and steady state kinetic analysis for the formation of active enzyme-I(+)-EDTA ternary complex for iodine reductase activity.

Horseradish peroxidase (HRP) catalyzes the reduction of iodine to iodide by EDTA with pseudocatalatic degradation of H2O2 to O2 (Banerjee et al., (1986) J. Biol. Chem. 261, 10592-10597; and Banerjee (1989) J. Biol. Chem. 264, 9188-9194). The reduction of iodine (I+) is dependent on EDTA concentration and is blocked by spin trap, DMPO, indicating the involvement of free radical species in the reduction process. Incubation of EDTA with both HRP and H2O2 results in the appearance of triplet ESR signal of spin-trapped EDTA radical (aN = 15 G), indicating its one-electron oxidation to a nitrogen-centered monocation radical (N-N+). The latter oxidizes H2O2 to evolve O2 and regenerate EDTA. In the presence of I+, a ternary complex of compound I-I(+)-EDTA is formed, which generates compound II-I. complex and both nitrogen-centered dication radical (N(+)-N+) through intermolecular electron transfer from EDTA nitrogens. Compound II-I. complex is further reduced similarly by another molecule of EDTA to form ferric enzyme, I-, and (N(+)-N+).(N(+)-N+) the oxidation product of EDTA, which may be released from the active site and, being more reactive, oxidizes H2O2 to O2 at a faster rate to regenerate EDTA. The existence of (N(+)-N+) is suggested from the similarity of its ESR signal with that of single nitrogen-centered monocation radical (N-N+). EDTA degradation by oxidative decarboxylation due to two-electron oxidation from the same or both nitrogen, atoms is not evident, and EDTA concentration remains the same throughout the reactions.(ABSTRACT TRUNCATED AT 250 WORDS)

Binding Sites↗

PubMed1995

Irreversible inactivation of lactoperoxidase by mercaptomethylimidazole through generation of a thiyl radical: its use as a probe to study the active site.

The mechanism of suicidal inactivation of lactoperoxidase (LPO) by mercaptomethylimidazole (MMI) has been studied. Analogue studies indicate a specific requirement for the thiol group of MMI for inactivation of LPO in the presence of H2O2. MMI is oxidized via one-electron transfer by LPO compound II as demonstrated by a spectral shift from 430 to 412 nm through an isosbestic point at 421 nm. A decrease in Soret absorbance at 412 nm and the appearance of visible peaks at 592 and 636 nm are the characteristics of the inactivated enzyme. The one-electron oxidation product of MMI was identified by e.s.r. spectroscopy as the 5,5'-dimethyl-l-pyrroline N-oxide (DMPO) adduct of the sulphur-centred thiyl radical. Both inactivation and spectral change are prevented by the radical trap DMPO, suggesting involvement of the thiyl radical in inactivation. pH-dependent inactivation kinetics indicate the involvement of an ionizable group on LPO (pKa 6.1), deprotonation of which favours inactivation. The enzyme is protected by iodide and not by guaiacol, suggesting that MMI interacts at or near the iodide-binding site which is away from the aromatic-donor-binding site. The inactive enzyme can form compound II and bind aromatic donor, indicating that the MMI oxidation product does not attack haem iron or aromatic-donor-binding site. We suggest that MMI interacts at the iodide-binding site for oxidation and the reactive product, probably the thiyl radical, is incorporated into the adjacent electron-rich site of haem porphyrin to cause inactivation.

Binding Sites↗

PubMed1995

Thiocyanate, a plausible physiological electron donor of gastric peroxidase.

Gastric peroxidase (GPO) was purified to apparent homogeneity to characterize its major physiological electron donor. The enzyme (RZ = 0.7), with a subunit molecular mass of 50 kDa, is a glycoprotein, with a relative abundance of aspartic and glutamic acid over arginine and lysine. It has a Soret maximum at 412 nm, which is shifted to 426 nm by H2O2 due to formation of compound II. Although the physiological electron donors I-, Br- and SCN-, but not Cl-, are oxidized by GPO optimally at acid pH, only I- and SCN- are oxidized appreciably at physiological pH. Considering that the I- concentration in stomach is less than 1 microM, whereas the SCN- concentration is about 250 microM, SCN- may act as a major electron donor for GPO. Moreover, SCN- oxidation remains unaltered in the presence of physiological concentrations of other halides. The second-order rate constant for the reaction of GPO with H2O2 (k1) and compound I with SCN- (k2) at pH 7 was found to be 8 x 10(7) M-1.s-1 and 2 x 10(5) M-1.s-1 respectively. GPO has significant pseudocatalase activity also in the presence of I- or Br-, but it is blocked by SCN-. The SCN- oxidation product OSCN- may be reduced back to SCN- by cellular GSH, and GSSG may be reduced back to GSH by glutathione reductase and NADPH. In a system reconstituted with pure glutathione reductase, NADPH, GSH, SCN- and H2O2. GPO-catalysed SCN- oxidation could be coupled to NADPH oxidation. This system where GPO utilizes SCN- as the major physiological electron donor may operate efficiently to scavenge intracellular H2O2.

Animals↗

PubMed1994

Mechanism of inhibition of horseradish peroxidase-catalysed iodide oxidation by EDTA.

EDTA inhibits horseradish peroxidase (HRP)-catalysed iodide oxidation in a concentration and pH-dependent manner. It is more effective at pH 6 than at lower pH values. A plot of log Kiapp. values as a function of pH yields a sigmoidal curve from which a pKa value of 5.4 can be calculated for an ionizable group on the catalytically active HRP for EDTA inhibition. Among the structural analogues of EDTA, tetramethylethylenediamine (TEMED) is 80% as effective as EDTA, whereas the EDTA-Zn2+ chelate and EGTA are ineffective. Kinetic studies indicate that EDTA competitively inhibits iodide oxidation. Spectral studies show that EDTA can quickly reduce compound I to compound II, but reduction of preformed compound II to the native enzyme is relatively slow, as demonstrated by the time-dependent spectral shift from 417 nm to 402 nm through an isosbestic point at 408 nm. Under steady-state conditions, in a reaction mixture containing HRP, EDTA and H2O2, the enzyme remains in the compound-II form, with absorption maxima at 417, 527 and 556 nm. Direct evidence for one-electron oxidation of EDTA by HRP intermediates is provided by the appearance of an e.s.r. signal of a 5,5-dimethyl-1-pyrroline N-oxide (spin trap)-EDTA radical adduct [aN (hyperfine splitting constant) = 1.5 mT] in e.s.r. studies. The signal intensity, however, decreases in the presence of iodide. The KD of the HRP-EDTA complex obtained from optical difference spectroscopy increases with an increase in iodide concentration, and the double-reciprocal plot for EDTA binding indicates that EDTA and iodide compete for the same binding site for oxidation. We suggest that EDTA inhibits iodide oxidation by acting as an electron donor.

Catalysis↗

PubMed1993

Mechanism-based inactivation of gastric peroxidase by mercaptomethylimidazole.

The mechanism of inhibition of gastric peroxidase (GPO) activity by mercaptomethylimidazole (MMI), an inducer of gastric acid secretion, has been investigated. Incubation of purified GPO with MMI in the presence of H2O2 results in irreversible inactivation of the enzyme. No significant inactivation occurs in the absence of H2O2 or MMI, suggesting the involvement of peroxidase-catalysed oxidized MMI (MMIOX.) in the inactivation process. The inactivation follows pseudo-first-order kinetics consistent with a mechanism-based (suicide) mode. The pseudo-first-order kinetic constants at pH 8 are ki = 111 microM, k(inact.) = 0.55 min-1 and t1/2 = 1.25 min, and the second-order rate constant is 0.53 x 10(4) M-1 x min-1. Propylthiouracil also inactivates GPO activity in the same manner but its efficiency (k(inact./ki = 0.46 mM-1 x min-1) is about 10 times lower than that of MMI (k(inact./ki = 5 mM-1 x min-1). The rate of inactivation with MMI shows pH-dependence with an inflection point at 7.3, indicating the involvement in the inactivation process of an ionizable group on the enzyme with a pKa of 7.3. The enzyme is remarkably protected against inactivation by micromolar concentrations of electron donors such as iodide and bromide but not by chloride. Although GPO oxidizes MMI slowly, iodide stimulates it through enzymic generation of I+ which is reduced back to I- by MMI. Although MMIOX. is formed at a much higher rate in the presence of I-, a constant concentration of I- maintained via the reduction of I+ by MMI, protects the active site of the enzyme against inactivation. We suggest that MMI inactivates catalytically active GPO by acting as a suicidal substrate.

Animals↗

PubMed1993

Chemical and kinetic evidence for an essential histidine residue in the electron transfer from aromatic donor to horseradish peroxidase compound I.

Horseradish peroxidase, when incubated with diethyl pyrocarbonate (DEPC), a histidine-specific reagent, shows time-dependent inactivation to oxidize aromatic electron donor, guaiacol. The inactivation follows pseudo-first order kinetics with a second order rate constant of 0.67 min-1 M-1. The pH dependence of inactivation shows an inflection point at 6.02, indicating histidine derivatization by DEPC. A difference spectrum of modified versus native enzyme shows a peak at 244 nm for N-carbethoxyhistidine that is diminished by hydroxylamine. Stoichiometric studies indicate that out of 2 histidine residues modified, one is responsible for inactivation. A plot of log reciprocal half-time of inactivation against log DEPC concentration suggests that only 1 histidine is essential. From the computer-stimulated structure of horseradish peroxidase, we tentatively suggest that this critical histidine is most likely distal histidine 42. Binding studies show that this histidine is not involved in guaiacol binding. Modified enzyme forms compound I with H2O2 but not compound II, suggesting a block of electron transfer process. Modified compound I cannot oxidize guaiacol as evidenced by the absence of donor-induced spectral shift from 408 nm, suggesting a block of electron transfer from bound donor to compound I. We suggest that this tentatively identified distal histidine controls aromatic donor oxidation by regulating electron transport without affecting donor binding or compound I formation.

Amino Acid Sequence↗

PubMed1993

Effect of stress on the antioxidant enzymes and gastric ulceration.

The effect of cold-restraint stress on the antioxidant enzymes of the rat gastric mucosa was studied with a view to finding out their role in stress induced gastric ulceration. Histological examination revealed stress induced extensive damage of the surface epithelial cell with lesions extending up to submucosa in some cases. Stress causes time-dependent increase in histamine and pepsin content but decrease in acid content of the gastric fluid with the progress of ulceration (ulcer index) for two hours. The tissue lipid peroxidation was significantly increased as evidenced by accumulation of malondialdehyde. Since lipid peroxidation results from the generation of reactive oxygen species, stress effect was studied on some antioxidant enzymes such as superoxide dismutase, peroxidases and prostaglandin synthetase as a function of time. The time dependent increase in stress ulcer correlates well with the concomitant increase in superoxide dismutase activity and decrease in peroxidase and prostaglandin synthetase activity. This creates a favourable condition for accumulation of endogenous H2O2 and more reactive hydroxyl radical (OH.). Administration of antioxidants such as reduced glutathione or sodium benzoate prior to stress causes significant decrease in ulcer index and lipid peroxidation and protection of gastric peroxidase activity suggesting the involvement of reactive oxygen species in stress induced gastric ulceration. This is supported by the in vitro observation that OH. can also inactivate peroxidase and induce lipid peroxidation. As prostaglandin is known to offer cytoprotection, stress-induced loss of prostaglandin synthetase activity appears to aggravate the oxidative damage caused by reactive oxygen species.

Animals↗

PubMed1993

Effect of dexamethasone on the peptic activity of gastric lumen and mucosa.

Dexamethasone (9 alpha-fluoro-16 alpha methyl-11 beta,17 alpha,21-trihydroxy-1,4-pregnadiene-3,20-dione-21-phosphate), a synthetic glucocorticoid, has a dual role on pepsinogen content of the gastric lumen and mucosa as measured by its peptic activity. Following stimulation the luminal peptic activity gradually decreases after 6 hr, then returns to basal levels at 18 hr and by 24 hr is inhibited by 50%. The luminal peptic activity induced by the secretory compound mercaptomethylimidazole (MMI) is also decreased. Dexamethasone effect on both basal and MMI-induced peptic activity can be reproduced by cycloheximide or puromycin, translational blockers of protein synthesis. This drug also has an independent time and dose-dependent inhibitory effect on gastric mucosal peptic activity which does not correlate with increased peptic activity of the lumen. Dexamethasone appears to be more effective than hydrocortisone and corticosterone in inhibiting the basal peptic activity of both lumen and mucosa. The inhibitory effect of this drug on tissue peptic activity is not mediated through induction of any inhibitory protein as evidenced by the insensitivity of the effect to actinomycin D. Studies on [14C]phenylalanine incorporation into gastric protein indicate that the effect of dexamethasone on tissue pepsinogen content is not due to a generalized block of protein synthesis.

Animals↗

PubMed1993

Inhibition of intestinal peroxidase activity by nonsteroidal antiinflammatory drugs.

The peroxidase activity of the mitochondrial fraction of rat intestine is inhibited in vitro by non-steroidal antiinflammatory drugs (NSAIDs), such as indomethacin (IMN) and acetylsalicylic acid (ASA), the former being more potent than the latter. The peroxidase was solubilised by cetab-NH4Cl extraction and purified to apparent homogeneity by Sephadex G-150 gel filtration and affinity chromatography on Con-A Sepharose. The purified enzyme activity was 80% inhibited by 150 microM IMN and 50% by 2.67 mM ASA. IMN could also inhibit lactoperoxidase activity to the same extent but not the horseradish peroxidase activity. The inhibition of peroxidase-catalysed iodide oxidation by IMN and ASA was optimal at pH 5.5 and 4.5, respectively. Kinetic studies revealed that the inhibition by IMN was competitive with respect to iodide or guaiacol, while the inhibition by ASA was noncompetitive and reversible in nature. Studies of some structural analogues showed that indole-3-acetic acid was as effective as IMN, while salicylic acid was more potent than ASA. Spectral studies showed a small bathochromic shift of the Soret band of the enzyme by IMN, suggesting its possible interaction at or near the heme moiety. The competitive nature of IMN may be explained as due to its oxidation by the peroxidase to a product absorbing at 412 nm, the formation of which is inhibited by iodide. We suggest that IMN inhibits intestinal peroxidase activity by acting as a competitive substrate for the enzyme. As intestinal peroxidase is mainly contributed by the invading eosinophils, NSAIDs may affect the host defence mechanism by inhibiting the activity of the enzyme.

Animals↗

PubMed1993

Iodide modulation of the EDTA-induced iodine reductase activity of horseradish peroxidase by interaction at or near the EDTA-binding site.

Horseradish peroxidase (HRP) catalyses the reduction of iodinium ion (I+) to iodide by H2O2 in the presence of EDTA. I+ reduction occurs optimally at pH 6 whereas the enzyme catalyses iodide oxidation optimally at pH 3.5. Thus the two activities reside on the same enzyme with two characteristic pH optima. Iodide modulates the expression of the reductase activity by EDTA. Higher concentrations of iodide inhibit the reductase activity by EDTA. Nitrite, an electron donor, acts similarly to iodide. Both EDTA and nitrite competitively inhibit iodide oxidation, indicating that they compete with iodide for the same binding site for electron flow to the haem iron group. However, unlike iodide, EDTA converts compound I, not into the native enzyme, but into a compound absorbing at 416 nm which reduces I+ and then returns to the native form. The apparent equilibrium dissociation constant, KD, for the formation of the EDTA-HRP complex (15 mM) is doubled in the presence of iodide, indicating interference with EDTA binding by iodide. EDTA binds away from the haem iron centre and not through intramolecular Ca2+. The pH-dependence of EDTA binding indicates that an ionizable group of the enzyme with pKa 5.8, presumably a distal histidine, controls the binding. The data suggest that iodide competes with EDTA for compound I and modulates the iodine reductase activity by limiting the formation of the 416 nm-absorbing active compound.

Anions↗

PubMed1993

Numerical studies of three-dimensional arterial flows in reverse curvature geometry: Part I--Peak flow.

A three-dimensional flow simulation at Repeak = 192 and 580 was made in a smooth reverse curvature model that conformed to the gentle "S" shape from a human left femoral artery angiogram. The objective of this numerical investigation was to find the changes in pressure, shear stress, velocity profile, and particle path occurring in the double-curved arterial vessel. Due to the impingement of blood at the outer wall in the first bend region, the wall shear stress approached 40 dyne/cm2--a value over twice as large as in the straight upstream segment. Conversely, at the inner wall in the first bend, a low shear stress region was found where the value of the shear stress was consistently smaller than that in the straight section. The initiation of centrifugal effects caused by the first bend could clearly be seen at Repeak = 580, but due to the close proximity of the reverse curvature segment, the momentum effect due to the second bend overshadowed the centrifugal effect. Hence, only near the end of the second bend did the centrifugal effect due to the second bend result in a double-spiral-secondary motion. In addition, the numerically calculated pressure drop data were in agreement with prior experimental values.

Angiography↗

PubMed1992

Localization of gastric peroxidase and its inhibition by mercaptomethylimidazole, an inducer of gastric acid secretion.

Mercaptomethylimidazole (MMI) is a potent inducer of gastric acid secretion which is associated with significant inhibition of peroxidase activity of rat gastric mucosa in vivo. A time-dependent increase in acid secretion correlates well with time-dependent decrease in the peroxidase activity. In a chamber experiment in vitro using isolated gastric mucosa, MMI stimulates acid secretion, showing an almost linear response up to 600 microM. The time-dependent increase in acid secretion is also correlated with time-dependent inhibition of the peroxidase activity. This effect is not mediated through oxidation of MMI by flavin-containing mono-oxygenase, which is absent from gastric mucosa. The peroxidase has been localized mainly in parietal cells isolated and purified from gastric mucosa by controlled digestion with collagenase followed by Percoll-density-gradient centrifugation. Peroxidase activity was further localized in the outer membrane of the purified mitochondria of the parietal cell by some membrane-impermeant reagents, indicating outward orientation of the enzyme. MMI can inhibit the peroxidase activity of both the parietal cell and its mitochondria in a concentration-dependent manner. The possible involvement of the parietal-cell peroxidase-H2O2 system in MMI-induced acid secretion may be suggested.

4-Chloromercuribenzenesulfonate↗