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Biomedical subjects

G Federici

Publications and source records attributed to G Federici.

At least 109 records · Page 6Linked to original sources
PubMed1990

Differential activity of human, rat, mouse and bacteria glutathione transferase isoenzymes towards 4-nitroquinoline 1-oxide.

The conjugation capacity of 4-nitroquinoline 1-oxide (4-NQO) with GSH by a number of human, rat, mouse and bacteria glutathione transferases (GSTs) was investigated. Pi and mu classes GSTs exhibited maximum conjugation capacity. Alpha class glutathione transferases as well as bacteria glutathione transferases were found to be unable to conjugate GSH to 4-NQO. The Km values as well as the catalytic efficiency (Kcat/Km) for most of the GSTs investigated were also determined. Mouse liver GST MIII (class mu) was the most efficient of the various isoenzymes tested. Its Kcat/Km value was 162 times higher than that of mouse liver GST MI (class alpha). The relatively high catalytic efficiency exhibited by GST-alpha (class pi) is prevalently due to its low affinity for 4-NQO.

4-Nitroquinoline-1-oxide↗
PubMed1990

Effect of ischaemia-reperfusion on glutathione peroxidase, glutathione reductase and glutathione transferase activities in human heart protected by hypothermic cardioplegia.

The activities of glutathione peroxidase (GSH-px), glutathione reductase (GSSG-rx) and glutathione transferase (GST) were measured in myocardial specimens obtained from right atria of patients subjected to different period of ischaemic arrest (aortic clamping ranging from 10 min to 90 min) followed by 60 min. of reperfusion, during open heart surgery 41-90 min. period of aortic clamping induced a significant increase of GSH-px activity with both H2O2 (p less than 0.05) and cumene hydroperoxide (p less than 0.025) as substrates when compared with baseline levels. Aortic clamping and reperfusion, however did not significantly change the myocardial activities of glutathione transferase and glutathione reductase. It is suggested that the increase of GSH-px in ischaemic-reperfused human hearts may render the myocardium less susceptible to oxidative attack particularly during the reoxygenation period when the level of active oxygen species is greatly elevated.

Female↗
PubMed1990

Effect of propionyl-L-carnitine on rat spinal cord ischaemia and post-ischaemic reperfusion injury.

In this study we have examined the effect of propionyl-L-carnitine (PC) on rat spinal cord ischaemia and post-ischaemic reperfusion injury by evaluating two lipid peroxidation indices, thiobarbituric acid reactive substances (TBARS) and diene conjugation, before and after the addition of an ADP-Fe+2 complex to spinal cord homogenates. Aerobic, ischaemic, and post ischaemic reperfusion rat spinal cord homogenates from PC treated and untreated animals did not show any statistically significant difference in their TBARS and conjugated diene content. The addition of the ADP-Fe+2 complex to these homogenates resulted in an increased production of both the lipid peroxidation indices, though the magnitude of such formation was related to the type of experimental intervention. The post-ischaemic reperfusion samples of untreated rats showed the highest TBARS and conjugated diene content, while ischaemic samples in either treated and untreated rats did not show any statistically significant difference with respect to the aerobic samples. The post-ischaemic reperfusion samples of treated rats showed a statistically significant decrease of TBARS and conjugated diene production in comparison to the untreated samples. In addition, PC was also able to partially inhibit TBARS and conjugated diene formation in linoleic acid micelles exposed to hemoglobin, though it did not protect albumin fragmentation from the irradiation of water with an X-ray source.

Animals↗
PubMed1989

Tryptophan environments in glutathione transferase of human placenta from temperature-dependent phosphorescence studies.

An investigation of the tryptophan emission properties of glutathione transferase from human placenta was conducted in order to characterize the environments of the two aromatic residues. The low-temperature phosphorescence spectra and temperature dependence of the phosphorescence quantum yield of the tryptophan residues revealed a difference in the chemical nature and dynamical structure of the surrounding protein matrix. Thus, one tryptophan residue seems to be deeply embedded within the polypeptide in a rigid weakly polar environment, characteristic of a beta-type secondary structure. The other is located in a more polar site, probably near the surface, in a rather flexible region of the macromolecule. At high temperature, the heterogeneity in the triplet lifetime of the internal residue attests to the presence of multiple conformers which are not in rapid equilibrium in the phosphorescence time scale. The anisotropy of the phosphorescence emission of glutathione transferase indicates that no energy transfer occurs between the two residues, and measurement of the rotational correlation time yields an hydrodynamic volume which is in good agreement with the molecular weight reported in the literature for the dimer.

Female↗
PubMed1989

Glutathione transferase isoenzymes from human testis.

By using affinity chromatography and isoelectric focusing techniques, several forms of glutathione transferase (GSTs) were resolved from human testis obtained from patients operated on for malignant diseases. Large interindividual variations in the expression of different isoenzymes resulted in the samples investigated. Five out of six samples analysed expressed GST-4.4 that resulted in being structurally and immunologically identical to GST-pi (class Pi). All the cationic GSTs of human testis, except for GST-8.36, GST-9.1 and GST-10.1, are homodimers of 24,500 Mr subunit and cross reacted with antisera raised against class Alpha GST. Some of the forms isolated (GST-3.8, GST-8.36, GST-9.1 and GST-10.1) can not apparently be related to any of GSTs so far characterized in other human tissues. Upon SDS/polyacrylamide gel electrophoresis, GST-8.36 and GST-9.1 appeared to be heterodimers of 24,500 and 26,500 Mr subunits and were found only in the testis seminoma suggesting that they might be tumour specific isoenzymes. GST-3.8 appeared to be formed by heterodimers of 23,000 and 26,500 Mr subunits whereas, GST-10.1 was found to be dimers of 22,000 and 24,500 Mr subunits. In addition, the results of immunohistochemical studies with antisera raised against both class Pi and Alpha GSTs are reported.

Aged↗
PubMed1989

Effect of oxidative stress on membrane phospholipid and protein organization in human erythrocytes.

Membrane phospholipid and protein organization was studied in intact human erythrocytes exposed to phenylhydrazine, an oxidative agent inducer. The evaluation of the membrane phospholipid and protein organization was carried out in terms of asymmetric distribution across the membrane bilayer for the phospholipids, and in terms of accessibility of cleavable sites present on the outer membrane surface for the proteins. Treatment of phenylhydrazine-exposed erythrocytes either with bee venom phospholipase A2 or with trinitrobenzenesulfonic acid indicated that phosphatidylserine (PS), which is the only phospholipid not formally present on the outer leaflet of the membrane, was translocated to the outer surface of the cell membrane. The extent of this phenomenon was directly proportional to the concentration of the oxidant having a peak value at 0.1 mM. Phosphatidylcholine and phosphatidylethanolamine conserved their original distribution across the erythrocyte membrane throughout the study. The oxidant, at a dose which did not induce any modification of the sodium dodecyl sulfate-polyacrylamide gel electrophoresis cytoskeleton membrane protein pattern, did not provoke any alteration of the membrane protein surface architecture, although the translocation of PS to the membrane outer leaflet in intact erythrocytes was present.

Analysis of Variance↗
PubMed1989

N-terminal region of Proteus mirabilis glutathione transferase is not homologous to mammalian and plant glutathione transferases.

The N-terminal amino acid sequence of glutathione transferase, Pm-GST-6.0, purified from Proteus mirabilis [(1988) Biochem. J. 255, 971-975] up to residue 38 and a comparative peptide fingerprint are reported. No obvious homology with the sequences of alpha, pi and mu classes of mammalian glutathione transferases as well as with those of plant glutathione transferases has been noted. These results suggest that the classification so far adopted for glutathione transferases cannot be extended to the bacterial enzyme.

Amino Acid Sequence↗
PubMed1989

Nonequivalence of the two subunits of horse erythrocyte glutathione transferase in their reaction with sulfhydryl reagents.

Glutathione transferase (EC 2.5.1.18) from horse erythrocytes has been purified and some molecular and kinetic properties have been investigated. It appears to be a dimeric protein composed of subunits of about 23 kDa, indistinguishable either in sodium dodecyl sulfate or in urea electrophoresis. Amino acid composition, substrate specificities, sensitivity to inhibitors, CD spectra, and immunological studies provide evidence that the horse enzyme is related to the pi class transferases. This enzyme has only two reactive thiol groups/dimer whose integrity appears to be essential for the activity. A peculiar feature of these protein thiol groups is that they react nonidentically with a number of thiol blocking reagents, i.e. iodacetamide, bromopyruvate, N-ethylmaleimide, and 1-chloro-2,4-dinitrobenzene. Also many disulfides react with one thiol group 5- to 10-fold more rapidly than with the other. The two mixed disulfides so formed also have different rates of reactivation by dithiothreitol. All the structural and kinetic data reported in this paper indicate a nonsymmetrical association of two identical subunits, or alternatively heterodimeric structure with subunits of very similar charge and size.

Amino Acids↗
PubMed1989

Electrophoretic and immunological analysis of glutathione transferase isoenzymes of human kidney carcinoma.

By sequential use of GSH-affinity chromatography and chromatofocusing, the isoenzymes of glutathione transferase from tumor and non-tumor kidney tissues have been purified and their properties compared. On the basis of electrophoretic mobilities on SDS/polyacrylamide gel, substrate specificities toward the diagnostic substrates cumene hydroperoxide and ethacrynic acid and immunoreactivity with antisera raised against alpha, mu and pi class glutathione transferases, it was found that most of the isoenzymes purified from both tumor and non-tumor kidney can be identified as members of either alpha or pi classes. All the samples investigated lacked mu class glutathione transferase. In addition, we could identify in tumor samples two transferases GST-7.6 and GST-5.8/5.9 which on the basis of immunological properties cannot be related to any of the members of the three major classes of glutathione transferases. The latter do not appear to have corresponding forms in non-tumor tissues. It was suggested that specific transferases can be selectively expressed by tumor kidney carcinoma.

Carcinoma, Renal Cell↗
PubMed1989

Mechanism of spectrin degradation induced by phenylhydrazine in intact human erythrocytes.

The exposure of human erythrocytes to phenylhydrazine results in the degradation of both monomers of spectrin, a major cytoskeleton membrane protein. The degradative process, characterized by a loss of spectrin without the appearance of high-molecular-weight products, either under reducing conditions or not, is almost complete in 10 min when a 5% erythrocyte suspension is treated with 1 mM phenylhydrazine. Under these conditions, we found a loss of 62.3 and 48.5% for the alpha and beta monomer, respectively. A similar degradative extent was obtained when the membrane ghost plus cellular free extracts, were dialyzed, and the membrane ghost plus hemoglobin was exposed to 1 mM phenylhydrazine for 10 min. The presence of different proteinase inhibitors and effectors, such as EDTA, diethylenetriaminepentaacetic acid, EGTA, leupeptin, aprotinin, phenylmethylsulfonyl fluoride, pepstatin, Ca2+ and ATP plus Mg2+, in the membrane ghost plus cellular free extract system (undialyzed) did not affect the degree of the spectrin-degradative process induced by phenylhydrazine. In addition, a purified spectrin tetramer preparation exposed to 1 mM phenylhydrazine in the presence of hemoglobin was degraded to an extent comparable to that with intact cells. Our data suggest that the initial degradative step of spectrin induced by phenylhydrazine in intact erythrocytes may be ascribed more to a direct oxidative breakdown, probably involving main-chain cleavage and side-chain cleavage processes, than to an eventual proteolytic system.

Erythrocyte Membrane↗
PubMed1989

Glutathione transferases in human nasal mucosa.

Glutathione transferase (GST) was investigated with 1-chloro-2,4-dinitrobenzene as substrate in tissues specimens of human nasal mucosa. The average +/- (SD) of GST activity in the cytosol was 76.8 +/- 21 nmol/min/mg with a range of 47-113. Using affinity chromatography and isoelectric focusing, the isozymes of GST from human nasal mucosa have been purified and characterized. On the criteria of isoelectric point, substrate specificities, apparent subunit molecular weight, sensitivity to characteristic inhibitors and immunological properties the major GST purified (about 85% of total activity) can be identified as class pi GST. Although a limited amount of class alpha GST was expressed by human nasal mucosa, no class mu isoenzymes was noted. In addition, we have also identified a GST subunit that cannot be related to any of three major classes of human GST.

Adolescent↗
PubMed1989

NPGB-induced inhibition of superoxide anion production by normal Lewis rat macrophages.

The treatment of Lewis rat peritoneal macrophages with p1-nitrophenyl p-guanidinobenzoate (NPGB) inhibited the superoxide anion production stimulated with phorbol myristate acetate (PMA). The addition of NPGB at the time of maximum superoxide generation was still able to block the superoxide release. It appears from these findings that NPGB may block either the activation process of the membrane bound NAD(P)H oxidase or directly on the active enzyme. Other protease inhibitors such as, epsilon-amino caproic acid (EACA), pepstatin, trans aminomethyl cyclohexane carboxylic acid (AMCA), aprotinin, and leupeptin did not inhibit the superoxide release. The superoxide anion release by the xanthine-xanthine oxidase system was not inhibited by NPGB. This finding indicates that NPGB does not itself react with superoxide. It has been also demonstrated that NPGB is a good reactant toward sulfhydryl group. The relevance of these finding to experimental allergic encephalomyelitis (EAE) is discussed.

Animals↗
PubMed1989

Induction of rat liver glutathione transferase subunit 7 by lead nitrate.

The effect of a single dose of lead nitrate (10 microM/100 g body wt), a hepatic mitogen, on rat liver glutathione transferase (GST) subunit expression was investigated. Using SDS-polyacrylamide gel electrophoresis and Western blot technique evidence for the induction of GST 7-7 is shown. This occurrence is identical to that observed in preneoplastic nodules generated in rat liver by different models of chemical carcinogenesis, suggesting that lead nitrate may be a very simple model for investigation of the mechanism of glutathione transferase 7-7 gene expression in chemical hepatocarcinogenesis.

Animals↗
PubMed1989

Glutathione transferase in bacteria: subunit composition and antigenic characterization.

The presence of glutathione transferase (GST; EC 2.5.1.18) in Escherichia coli ATCC 25922, E. coli ATCC 25422, Proteus vulgaris ATCC 8427, Pseudomonas aeruginosa ATCC 27853, Klebsiella oxytoca CIP 666, K. oxytoca AF 101, Enterobacter cloacae CIP 6085, Serratia marcescens CIP 6755, and Proteus mirabilis AF 2924 was investigated. Using 1-chloro-2,4-dinitrobenzene as substrate, GST activity was found in the glutathione-(GSH-)affinity-purified fraction of all strains tested. SDS-PAGE analysis of GSH-affinity-purified enzyme indicated that the GSTs of all these bacteria are dimers of two identical subunits of Mr about 22,500. Rabbit antiserum directed against the major isoenzyme present in Proteus mirabilis AF 2924, Pm-GST-6.0, was used to investigate the antigenic properties of bacterial GSTs. Western blot analysis indicated that a GST antigenically identical to Pm-GST-6.0 is present in Enterobacter cloacae CIP 6085, Escherichia coli ATCC 25422 and Proteus vulgaris ATCC 8427, but absent in Escherichia coli ATCC 25922, Klebsiella oxytoca CIP 666, K. oxytoca AF 101 and Serratia marcescens CIP 6755. The presence of Pm-GST-6.0, but not mammalian GST, increased the MIC values of amikacin, ampicillin, cefotaxime, cephalothin and nalidixic acid for E. coli ATCC 25922. It is suggested that bacterial GST may represent a defense against the effects of antibiotics.

Anti-Bacterial Agents↗
PubMed1988

Purification and characterization of three forms of glutathione transferase from Proteus mirabilis.

Three forms of glutathione transferase (GST) with pI values of 6.0, 6.4 and 7.3 were isolated from Proteus mirabilis AF 2924 by glutathione-affinity chromatography followed by isoelectric focusing, and their structural, kinetic and immunological properties were investigated. Upon SDS/polyacrylamide-slab-gel electrophoresis, all forms proved to be composed of two subunits of identical (22,500) Mr. GST-6.0 and GST-6.4 together account for about 95% of the total activity, whereas GST-7.3 is present only in trace amounts. Extensive similarities have been found between GST-6.0 and GST-6.4. These include subunit molecular mass, amino acid composition, substrate specificities and immunological characteristics. GST-7.3 also cross-reacted (non-identity) with antisera raised against bacterial GST-6.0. None of the antisera raised against a number of human, rat and mouse GSTs cross-reacted with the bacterial enzymes, indicating major structural differences between them and the mammalian GSTs. This conclusion is further supported by c.d. spectra.

Amino Acids↗
PubMed1988

Purification and characterization of five forms of glutathione transferase from human uterus.

Five glutathione transferase (GST) forms were purified from human uterus by glutathione-affinity chromatography followed by chromatofocusing, and their structural, kinetic and immunological properties were investigated. Upon SDS/polyacrylamide slab gel electrophoresis all forms resulted composed of two subunits of identical molecular size. GST V (pI 4.5) is a dimer of 23-kDa subunits. GST I (pI 6.8) and GST IV (pI 4.9) are dimers of 24-kDa subunits whereas GST II (pI 6.1) and GST III (pI 5.5) are dimers of 26.5-kDa subunits. GST V accounts for about 85-90% of the activity whereas the other isoenzymes are present in trace quantities. On the basis of the molecular mass of the subunits, amino acid composition, substrate specificities, sensitivities to inhibitors, CD spectra and immunological studies, GST V appeared very similar to transferase pi. Structural and immunological studies provide evidence that GST IV is closely related to the less 'basic' transferase (GST pI 8.5) of human skin. Extensive similarities have been found between GST II and GST III. The comparison includes amino acid compositions, subunits molecular size and immunological properties. The two enzymes, however, are kinetically distinguishable. The data presented also indicate that GST II and GST III are related to transferase mu and to transferase psi of human liver. Even though GST I has a subunit molecular mass identical to GST IV, several lines of evidence, including catalytic and immunological properties, indicate that they are different from each other. GST I seems not to be related to any of known human transferases, suggesting that it may be specific for the uterus.

Affinity Labels↗
PubMed1988

Glutathione and glutathione metabolizing enzymes in yeasts.

Total glutathione content, glutathione peroxidase, glutathione transferase and glutathione reductase activities have been measured in 12 species of yeasts. All the strains tested contained glutathione, though in different amounts, as well as the above mentioned enzymes. To discriminate between the selenium-dependent and the selenium-independent form, glutathione peroxidase activity has been measured with both H2O2 and cumene hydroperoxide. Rhodotorula glutinis appeared to be the only strain in which the selenium-dependent form was not found, but this yeast exhibited the highest level of selenium-independent glutathione peroxide activity as compared to the other strains.

Glutathione↗