Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Several studies have suggested that pulmonary toxicity to asbestos and silica may be mediated through oxidant-induced cell injury. We have reported recently that surface radicals associated with freshly fractured silica may be an important factor in cell injury and induction of pulmonary disease. Although the generation of oxygenated radicals in dust-cell interactions has been demonstrated, there are no data correlating the toxicity of a dust with the level of oxygen radical generation by the dust during its interaction with phagocytic cells. In the present study, we have investigated the in vitro generation of oxygen free radicals from human neutrophils and rat alveolar macrophages stimulated with freshly fractured silica, aged silica, amosite, crocidolite, chrysotile, and nontoxic dust, barite. Electron spin resonance (ESR) with the aid of a spin trap phenyl-N-tert-butyl nitrone (PBN) was used to measure the oxygen radicals generated during phagocytosis of the dusts. The relative toxicity index and ESR peak heights, on an equal surface area basis and normalized to barite as one, showed a direct relationship. The normalized toxicity indices and peak heights were: silica, 3.5 versus 2; chrysotile, 4 versus 2; crocidolite, 11 versus 8; and amosite, 26 versus 13. Addition of hydroxyl radical scavengers such as catalase, dimethyl sulfoxide, 1,3 dimethyl-2-thiourea (DMTU), sodium benzoate, and mannitol prevented the radical generation. Carmustine, a glutathione reductase-glutathione peroxidase inhibitor, caused a 5-fold increase in the radical generation. These results indicate that a nontoxic dust such as barite generates toxic oxygen radicals at a minimal level that can be quenched by the normal cellular defense system. For toxic dusts such as silica, amosite, chrysotile, and crocidolite, the potential for oxygen radical generation is enhanced by their surface properties, physical dimensions, and the surface-based radical-generating redox sites. The enhanced radical generation may impair the cellular defense system, resulting in cell injury. Use of scavengers, chelators, and potentiating agents suggests the membrane-based oxidase system as the probable primary source of the radical-generating system. The data presented herein suggest the generation of oxygen free radicals as an important primary event in silica- as well as asbestos-induced cell injury.
Am J Respir Cell Mol Biol 1992 Apr
PMID:Enhanced generation of free radicals from phagocytes induced by mineral dusts. 131 51

The activity of pure calf-liver and Escherichia coli thioredoxin reductases decreased drastically in the presence of NADPH or NADH, while NADP+, NAD+ and oxidized E. coli thioredoxin activated both enzymes significantly, particularly the bacterial one. The loss of activity under reducing conditions was time-dependent, thus suggesting an inactivation process: in the presence of 0.24 mM NADPH the half-lives for the E. coli and calf-liver enzymes were 13.5 and 2 min, respectively. Oxidized E. coli thioredoxin fully protected both enzymes from inactivation, and also promoted their complete reactivation after only 30 min incubation at 30 degrees C. Lower but significant protection and reactivation was also observed with NADP+ and NAD+. EDTA protected thioredoxin reductase from NADPH inactivation to a great degree, thus indicating the participation of metals in the process; EGTA did not protect the enzyme from redox inactivation. Thioredoxin reductase was extensively inactivated by NADPH under aerobic and anaerobic conditions, thus excluding the participation of O2 or oxygen active species in redox inactivation. The loss of thioredoxin reductase activity promoted by NADPH was much faster and complete in the presence of NAD+ glycohydrolase, thus suggesting that inactivation was related to full reduction of the redox-active disulfide. Those results indicate that thioredoxin reductase activity can be modulated in bacteria and mammals by the redox status of NADP(H) and thioredoxin pools, in a similar way to glutathione reductase. This would considerably expand the regulatory potential of the thioredoxin-thioredoxin reductase system with the enzyme being self-regulated by its own substrate, a regulatory protein.
Mol Cell Biochem 1992 Jan 15
PMID:NADPH and oxidized thioredoxin mediate redox interconversion of calf-liver and Escherichia coli thioredoxin reductase. 131 49

The gene encoding the streptococcal flavoprotein NADH oxidase (NOXase), which catalyzes the four-electron reduction of O2-->2H2O, has been cloned and sequenced from the genome of Streptococcus (Enterococcus) faecalis 10C1 (ATCC 11700). The deduced NOXase protein sequence corresponds to a molecular mass of 48.9 kDa and contains three previously sequenced cysteinyl peptides obtained with the purified enzyme. In Escherichia coli, the expressed nox gene produced a catalytically active product, which retained its immunoreactivity to affinity-purified NOXase antisera. Alignment of the NOXase protein sequence with that of streptococcal NADH peroxidase (NPXase) revealed that the proteins are 44% identical. Among the most highly conserved segments is a sequence containing Cys42; this residue is known to exist as a stabilized cysteine-sulfenic acid (Cys-SOH) in NPXase and serves as the non-flavin redox center. In addition, three previously identified NPXase segments, known to be involved in FAD and NAD(P)-binding in other pyridine nucleotide-linked flavoprotein oxidoreductases, are strongly conserved in NOXase. Overall, the extensive homology observed between NOXase and NPXase suggests that the monomer chain fold of the oxidase closely resembles that of the peroxidase. Both sequences share limited but significant homology to those of glutathione reductase and other members of the flavoprotein disulfide reductase family. These and other considerations suggest that these two unusual streptococcal flavoproteins constitute a distinct class of FAD-dependent oxidoreductases, the flavoprotein peroxide reductases, easily contrasted with enzymes such as glutathione reductase and thioredoxin reductase.
J Mol Biol 1992 Oct 05
PMID:Molecular cloning and analysis of the gene encoding the NADH oxidase from Streptococcus faecalis 10C1. Comparison with NADH peroxidase and the flavoprotein disulfide reductases. 140 82

Previous studies demonstrated that preconditioning of a heart by repeated stunning can reduce the cellular injury to the heart from subsequent acute ischemic insult. To examine the possible biochemical mechanism for such myocardial preservation afforded by preconditioning, swine heart was subjected to four episodes of 5 min. stunning by occluding the left anterior descending coronary artery (LAD), followed by 10 min. of reperfusion after each stunning. Heart was then made regionally ischemic for 60 min. by LAD occlusion, followed by 6 hrs. reperfusion. Control heart was perfused for 60 min., followed by 60 min. ischemia and 6 hrs. reperfusion. The results of our studies indicated the stimulation of a number of antioxidative enzymes, including Mn-superoxide dismutase (Mn-SOD), catalase, glutathione peroxidase, and glutathione reductase, after repeated stunning and reperfusion. In addition, a number of new proteins were expressed after preconditioning the heart, including some oxidative-stress related proteins and 72 kDa heat-shock protein. These results suggest that preconditioning of a heart by repeated stunning may lead to strengthening of the oxidative defense system of the heart, which is likely to play a role in myocardial preservation during subsequent ischemic and reperfusion injury.
Cell Mol Biol (Noisy-le-grand) 1992 Nov
PMID:Preconditioning of heart by repeated stunning. Adaptive modification of antioxidative defense system. 147 1

The X-ray crystal structure of the enzyme trypanothione reductase, isolated from the trypanosomatid organism Crithidia fasciculata, has been solved by molecular replacement. The search model was the crystal structure of human glutathione reductase that shares approximately 40% sequence identity. The trypanosomal enzyme crystallizes in the tetragonal space group P4(1) with unit cell lengths of a = 128.9 A and c = 92.3 A. The asymmetric unit consists of a homodimer of approximate molecular mass 108 kDa. We present the structural detail of the active site as derived from the crystallographic model obtained at an intermediate stage of the analysis using diffraction data to 2.8 A resolution with an R-factor of 23.2%. This model has root-mean-square deviations from ideal geometry of 0.026 A for bond lengths and 4.7 degrees for bond angles. The trypanosomid enzyme assumes a similar biological function to glutathione reductase and, although similar in topology to human glutathione reductase, has an enlarged active site and a number of amino acid differences, steric and electrostatic, which allows it to process only the unique substrate trypanothione and not glutathione. This protein represents a prime target for chemotherapy of several debilitating tropical diseases caused by protozoan parasites belonging to the genera Trypanosoma and Leishmania. The structural differences between the parasite and host enzymes and their substrates thus provides a rational basis for the design of new drugs active against trypanosomes. In addition, our model explains the results of site-directed mutagenesis experiments, carried out on recombinant trypanothione reductase and glutathione reductases, designed by consideration of the crystal structure of human glutathione reductase.
J Mol Biol 1992 Sep 05
PMID:Active site of trypanothione reductase. A target for rational drug design. 152 96

Endogenous hydrogen peroxide (H2O2) release from aortic endothelial cells was studied in the presence of antioxidant enzyme inhibitors, mitochondrial inhibitors, a microsomal cytochrome P-450 inhibitor, and after oxidative stress induced with H2O2 or menadione. Extracellular H2O2 generation was determined spectrofluorometrically using 3-methoxy-4-hydroxy phenylacetic acid, and intracellular H2O2 production (in or near peroxisomes) was measured indirectly using aminotriazole, which inactivates catalase in the presence of H2O2. Extracellular H2O2 release was 0.079 +/- 0.005 nmol/min/mg protein in Hanks' balanced salt solution, was constant during a 120-min incubation period, and was not affected by the cell passage number. The half-life for catalase inactivation with aminotriazole was 23 min. Inhibition of catalase, glutathione reductase, or gamma-glutamylcysteine synthetase did not change the rate of extracellular release of H2O2. Furthermore, inhibition of the mitochondrial respiratory chain (rotenone, antimycin A) or microsomal cytochrome P-450 (8-methoxypsoralen) did not change extracellular H2O2 release or intracellular H2O2 production (at peroxisomes) by endothelial cells or cells in which glutathione reductase was inactivated. When the cells were exposed to exogenous H2O2 (30 microM), extracellular H2O2 was scavenged primarily by the glutathione redox pathway. Exogenously added H2O2 (100 microM) changed intracellular H2O2 production (in or near peroxisomes) only when the glutathione redox cycle was inactivated. Menadione (20 microM), which undergoes intracellular redox cycling, increased extracellular H2O2 release almost 4-fold to 0.3 nmol/min/mg protein. Furthermore, menadione increased peroxisomal H2O2 levels and decreased the half-life for catalase inactivation in the presence of aminotriazole to 13 min. Catalase inhibition increased extracellular H2O2 release during menadione treatment, indicating that H2O2 can diffuse across the plasma membrane during oxidant stress.(ABSTRACT TRUNCATED AT 250 WORDS)
Am J Respir Cell Mol Biol 1992 Feb
PMID:Regulation of hydrogen peroxide generation in cultured endothelial cells. 154 Mar 80

Trypanothione reductase (TR) is a target for drug design since it is unique to trypanosomatids, substituting for the otherwise ubiquitous enzyme, glutathione reductase. We report the cloning and sequencing of several cDNAs and genes encoding Crithidia fasciculata TR, the structure of which has recently been solved by crystallography. Single base polymorphisms are detected in cDNAs (containing 80% of the coding sequence) and two different genomic clones, including a glutamine to glutamate change in the C-terminal region of the TR coding region; other nucleotide changes are silent. Homology (from genomic clones, both of which contained signals appropriate for expression) to the Trypanosoma congolense gene was 63% at the nucleic acid level, with 68% amino acid identity; the significance of homologies to human and Escherichia coli glutathione reductase sequences is discussed. Polymorphic sites in the genomic clones included sites found in the cDNAs, indicating that differences existing in the genomic sequence are real, and propagated to RNA.
Mol Biochem Parasitol 1992 Jan
PMID:Cloning, sequencing, and demonstration of polymorphism in trypanothione reductase from Crithidia fasciculata. 154 16

The purpose of this study was to determine if in vivo ozone exposure results in elevations in the levels of glutathione and glutathione-dependent enzymes in cells derived from bronchoalveolar lavage fluid (BALF). Our hypothesis was that, as part of a defense mechanism against oxygen toxicity, such cells would have increased levels of glutathione (GSH) in response to an oxidant stress. Female F344/N rats were exposed to 0.8 ppm ozone, 6 hr/day, for 1, 3, or 7 days, after which cells were collected by lung lavage. The GSH and GSH-peroxidase activity per milligram of protein in the cellular fraction, both necessary for reducing cellular peroxides, were elevated after 3 days of ozone exposure. After 7 days of exposure, cellular GSH had returned to control values, but the activity of glutathione reductase, the enzyme that reduces oxidized glutathione to GSH, was increased. Extracellular GSH concentration and glutathione reductase activity in BALF were also increased after 7 days of exposure. The total glutathione equivalents (GSH and GSSG, both cellular and extracellular) in BALF increased throughout the 7-day exposure, with GSH increasing first in the cells, and then in the extracellular fluid. This study demonstrated that the glutathione anti-oxidant system of BALF cells is stimulated by exposure to ozone. This response may serve to protect cells from the toxic effects of oxidant stress.
Exp Mol Pathol 1992 Feb
PMID:Glutathione and GSH-dependent enzymes in bronchoalveolar lavage fluid cells in response to ozone. 154 67

Redox interconversion of glutathione reductase was studied in situ with S. cerevisiae. The enzyme was more sensitive to redox inactivation in 24 hour-starved cells than in freshly-grown ones. While 5 microM NADPH or 100 microM NADH caused 50% inactivation in normal cells in 30 min, 0.75 microM NADPH or 50 microM NADH promoted a similar effect in starved cells. GSSG reactivated the enzyme previously inactivated by NADPH, ascertaining that the enzyme was subjected to redox interconversion. Low EDTA concentrations fully protected the enzyme from NADPH inactivation, thus confirming the participation of metals in such a process. Extensive inactivation was obtained in permeabilized cells incubated with glucose-6-phosphate or 6-phosphogluconate, in agreement with the very high specific activities of the corresponding dehydrogenases. Some inactivation was also observed with malate, L-lactate, gluconate or isocitrate in the presence of low NADP+ concentrations. The inactivation of yeast glutathione reductase has also been studied in vivo. The activity decreased to 75% after 2 hours of growth with glucono-delta-lactone as carbon source, while NADPH rose to 144% and NADPH+ fell to 86% of their initial values. Greater changes were observed in the presence of 1.5 microM rotenone: enzymatic activity descended to 23% of the control value, while the NADH/NAD+ and NADPH/NADP+ ratios rose to 171% and 262% of their initial values, respectively. Such results indicate that the lowered redox potential of the pyridine nucleotide pool existing when glucono-delta-lactone is oxidized promotes in vivo inactivation of glutathione reductase.
Mol Cell Biochem 1992 Mar 25
PMID:Glutathione reductase from Saccharomyces cerevisiae undergoes redox interconversion in situ and in vivo. 158 2

Rats were subjected to bilateral carotid artery occlusion for 30 min, followed by reperfusion for varying time periods. The concentration of reduced and oxidized glutathione, glutathione peroxidase and glutathione reductase were determined in whole brain after varying periods of reperfusion. Lipid peroxidation was also assessed by determining the levels of malondialdehyde (MDA) in the brain. Reperfusion for 1 hr following bilateral carotid artery occlusion resulted in significant decrease in total glutathione (GSH) concentration along with small but significant increase in oxidized glutathione (GSSG) levels. After 4 hr of reperfusion, GSH levels recovered, although GSSG levels remained elevated up to 12 hr of reperfusion. Increase in malondialdehyde levels was also detected in the brain up to 12 hr of reperfusion. Glutathione reductase activity remained significantly low up to 144 hr of reperfusion, while glutathione peroxidase activity remained unaffected. These results demonstrate that oxidative stress is generated in the brain during reperfusion following partial ischemia due to bilateral carotid artery occlusion.
Mol Cell Biochem 1992 Apr
PMID:Glutathione homeostasis in brain during reperfusion following bilateral carotid artery occlusion in the rat. 158 35


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