A. Aceto

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.

Glutathione peroxidase, glutathione reductase and glutathione transferase activities in the human artery, vein and heart

The continuous exposure to blood components, including prooxidants, makes the blood vessel wall susceptible to oxidative stress and free radical mediated reactions (Henning and Chow, 1988; Stamm et al., 1989; Halliwell and Gutteridge, 1984). Free radicals can be produced extracellularly via the respiratory bursts of activated neutrophils, or intracellularly, via oxidation of hypoxanthine by xanthine oxidase (Henning and Chow, 1988; Stamm et al., 1989; Rubanyi, 1988). Microsomal enzymes such as lipoxygenase and cyclooxygenase may also be a source of reactive species of oxygen (Henning and Chow, 1988; Stamm et al., 1989; Rubanyi, 1988; Mason et al., 1980). It has been proposed that free radicals are involved in the initiation and progression of various cardiovascular diseases including arteriosclerosis (Henning and Chow, 1988; Stamm et al., 1989; Yagi, 1988; Jürgens et al., 1987). Thus the adequacy of the defence systems against free radicals is critical for the susceptibility of blood vessel wall to oxidative damage. Among the enzymatic systems capable of protecting the cell against oxidative injury, selenium dependent glutathione peroxidase (Se-GSH-px), glutatione reductase (GSSG-rx) and glutathione transferase (GST) play a crucial role (Flohe et al., 1976; Mannervik and Danielson, 1988). Using glutathione (GSH) as a cofactor, Se-GSH-px reduces H2O2 to water and organic hydroperoxides to the corresponding alcohols (Flohe et al., 1976). This reaction leads to conversion of GSH into its oxidized form (GSSG). In the presence of NADPH, GSSG-rx is able to reduce the oxidized glutathione.(ABSTRACT TRUNCATED AT 250 WORDS)

Glutathione transferases in human nasal mucosa

Glutathione transferase (GST) was investigated with 1-chloro-2,4-dinitrobenzene as substrate in tissues speciments 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 inibitors 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.

Activities of enzymes associated with the metabolism of glutathione in fetal rat liver and placenta.

Glutathione S-transferase, glutathione peroxidase, glutathione reductase and gamma-glutamylcysteine synthetase activities were measured in fetal rat liver and placenta supernatant at the 16th and 20th days of pregnancy. Compared with adult liver, low activities were found in both fetal liver and placenta. Both selenium-dependent and selenium-independent glutathione peroxidase activities were present in fetal liver, but only the selenium-dependent activity augmented as parturition advanced. Selenium-independent glutathione peroxidase was found to be absent in placenta. The progress of gestation is accompanied by a significant increase in conjugating capacity toward 1-chloro-2,4-dinitrobenzene and 1,2-dichloro-4-nitrobenzene and a significant decrease toward 1,2-epoxy-3-(p-nitrophenoxy)propane in fetal liver. Glutathione S-transferase activity in rat placenta diminished from day 15 to day 20 of gestation. The elevation of enzymatic activities involved in the synthesis and recovery of glutathione, which takes place in fetal liver and placenta, was thought to be adaptively responsive to the changes that occurred in glutathione-consuming enzymes.

Characterization of glutathione transferase from Gammarus italicus

1. By using affinity chromatography and chromatofocusing analysis at least two major glutathione transferases, named GST II and GST III can be isolated from Gammarus italicus. 2. GST II has an isoelectric point at pH 5.0 and is composed of two subunits with an apparent molecular mass of 28 KDa. 3. GST III which has an isoelectric point at pH 4.6 was found to be an heterodimer of 27 KDa and 28 KDa. 4. The 28 KDa subunit cross-reacted in immunoblotting analysis with antisera raised against pi class GST, whereas none of the antisera raised against alpha, mu and pi class GSTs cross-reacted with the 27 KDa subunit.

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.

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.

Crystallization and preliminary X-ray analysis of a bacterial glutathione transferase

Crystals of a bacterial glutathione S-transferase from Proteus mirabilis have been grown from polyethylene glycol by the hanging-drop vapour-diffusion method. Successful crystallization required the presence of the substrate glutathione. The crystals belong to the tetragonal space group P4 with cell dimensions a = b = 90.9 and c = 117.3 A. They contain between three and six monomers in the asymmetric unit and diffract to beyond 2.3 A resolution.

Recurrent atrial fibrillation in a patient with ulcerative colitis treated with azathioprine: case report and review of the literature.

We present a clinical case report regarding recurrent atrial fibrillation in a patient with ulcerative colitis treated with azathioprine. Atrial fibrillation represents the most common sustained cardiac arrhythmia, occurring in 1–2% of the general population and characterized by seemingly disorganized atrial depolarizations without effective atrial contraction. Several mechanisms determine this arrhythmia; in particular remodelling (structural, mechanical and electrical alteration related to atrial fibrillation). The pro-arrhythmic effect of azathioprine may be evaluated during immunosuppressive therapy to be aware of this serious but reversible adverse effect.

Changes of Rat Liver Glutathione Peroxidase, Glutathione Reductase and Glutathione Transferase 7–7 by Lead Nitrate Treatment

The importance of glutathione in the detoxication of reactive species of xenobiotics, including those chemical carcinogens generated by cytochrome P-450 mixed function oxidase has been well establishedl. Glutathione conjugates with electrophilic molecules of many chemicals carcinogens directly as well as through the action of glutathione transferases (GST; EC: 2.5.1.18)2,3. This latter are comprised of a family of dimeric proteins that in addition to facilitate the conjugation of glutathione to a large number of electrophilic molecules have the capacity to binds covalently/non-covalently a wide range of lipophilic compounds4,5.