Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.5.1.47 (cysteine synthase)
625 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The cytotoxic effects of alloxan are not understood in any great detail, although they are considered to involve reactions mediated by oxygen-derived free radicals. These reactive species may form extra-or intracellularly following alloxan reduction, and result in cell damage through a number of complex interactions with a variety of macromolecules. The purpose of the present study was to elucidate further the early intracellular effects of alloxan on a model system of macrophage-like cells in culture. Addition of alloxan (15 mM), without reducing agents, to the medium surrounding the cells (phosphate-buffered saline, PBS, 37 degrees C, pH 7.4) resulted in rapid lysosomal damage (disappearance of the proton gradient over the membrane) followed by severe cellular degeneration (swelling and blebbing) and 50% cell death (trypan blue dye exclusion test) within fifty min. Cells pretreated with the gamma-glutamyl cysteine synthetase-inhibiting agent BSO, to decrease levels of intracellular glutathione, showed enhanced sensitivity to alloxan. The results are interpreted as indicating the cytotoxicity to result from intracellular formation of superoxide radicals, hydrogen peroxide and hydroxyl radicals, the latter within secondary lysosomes containing trace amounts of reactive iron (inducing Fenton reactions). The ensuing lysosomal membrane damage may result in leakage of lysosomal hydrolases and further cellular degeneration.
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PMID:Alloxan cytotoxicity involves lysosomal damage. 158 Oct 39

Glutathione (GSH) is one of several intracellular hydrogen donating species thought to compete with O2, a damaging species, to repair radiation induced free radical damage. The O2 K factor was determined for normal Chinese hamster V79 cells, V79 cells made acutely thiol deficient (no detectable GSH or NPSH) using the gamma-glutamyl-cysteine synthetase enzyme inhibitor D,L-Buthionine-S,R-sulfoximine (D,L-BSO), or a human skin fibroblast cell line GM3877 which, because of the nature of its genetic defect, has chronically low levels of GSH (7% of normal skin fibroblasts). The K factors for normal V79 cells, treated with BSO or GSH deficient human fibroblasts, were 0.54, 0.15, and 0.1% O2, respectively. While thiol depletion affects the O2 K factor, V79 cells without any detectable GSH were still not as sensitive as the genetically deficient line GM3877 with 7% of normal GSH values. Other factors which may influence the results are whether the GSH levels remain low or regenerate following irradiation and the intracellular distribution of GSH.
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PMID:Oxygen concentration and the OER for acutely or chronically thiol deficient cells. 374 29

Extracts of Desulfovibrio vulgaris were found to contain serine transacetylase and cysteine synthase activities. When extracts were incubated with bisulfite and o-acetylserine, or acetyl coenzyme A plus L-serine, under a hydrogen atmosphere, cysteine was formed. Pyruvate served as a reductant for bisulfite reduction to sulfide and concomitantly provided the acetyl moiety for acetyl coenzyme A formation. Consequently, when extracts were incubated with pyruvate, bisulfite, and L-serine, cysteine synthesis resulted.
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PMID:Cysteine synthesis by Desulfovibrio vulgaris extracts. 736 31

The O-acetylserine sulfhydrylase (OASS) reaction has been studied using a number of spectral probes including UV--visible, fluorescence, circular dichroism, and 31P NMR spectroscopy. The addition of L-cysteine, L-alanine, and glycine to OASS results in a shift in lambda max of 412 nm for the internal Schiff base to 418 nm resulting from the formation of the external Schiff base. The addition of L-serine or O-methyl-D,L-serine gives decreases of the absorbance of unliganded enzyme at 412 nm of about 50% and 20%, respectively, concomitant with an increase in the absorbance at 320 nm and a shift in the lambda max of the remaining visible absorbance to 418 nm. The spectral shifts observed in the presence of L-serine are suggestive of establishing an equilibrium between different forms of external Schiff base. The concentration dependence of the changes at 440 (L-cysteine) and 320 nm (L-serine) provides an estimate of the dissociation constant for the external aldimine. The pH dependence of the dissociation constant suggests the alpha-amine of the amino acid must be unprotonated for nucleophilic attack at C4' of PLP, and an enzyme side chain must be unprotonated to hydrogen-bond the thiol or hydroxyl side chain of the amino acid. When L-cysteine is the amino acid, the thiol side chain must be protonated to hydrogen-bond to the unprotonated enzyme side chain. The 31P NMR chemical shift is increased from 5.2 ppm for unliganded enzyme to 5.3 ppm in the presence of L-cysteine, signaling a tighter interaction at the 5'-phosphate upon formation of the external Schiff base.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Identification and spectral characterization of the external aldimine of the O-acetylserine sulfhydrylase reaction. 754 55

The pH dependence of kinetic parameters using natural and alternative reactants was determined in order to obtain information on the chemical mechanisms of the A and B isozymes of O-acetylserine sulfhydrylase (OASS) from Salmonella typhimurium. A general mechanism is proposed for OASS in which OAS binds with its alpha-amine unprotonated to carry out a nucleophilic attack on C4' of the protonated Schiff base and with the acetyl carbonyl hydrogen-bonded to a protonated enzyme group (or a water molecule), which aids in the beta-elimination of acetate. The enzyme lysine that was in Schiff base linkage with the active site pyridoxal 5'-phosphate deprotonates the alpha-carbon in the beta-elimination reaction, and a proton is likely released with the acetate product. Sulfide likely binds as HS- to undergo nucleophilic attack on the alpha-aminoacrylate intermediate, followed by protonation of the alpha-carbon by the enzyme lysine. In OASS-A, HS- is hydrogen-bonded to the enzyme group that assists in the beta-elimination of acetate, but this is not the case for OASS-B. The pH independent equilibrium constant for the first half-reaction of OASS-A is 1.6 x 10(-3), while the second half-reaction is practically irreversible.
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PMID:Acid-base chemical mechanism of O-acetylserine sulfhydrylases-A and -B from pH studies. 754 74

Exposure to low level noise prior to a high level exposure reduces noise-induced hearing loss in mammals. This phenomenon is known as sound conditioning or 'toughening'. Reactive oxygen intermediates have been implicated in noise-induced cochlear damage. To evaluate if in situ antioxidant processes may play a role in the toughening phenomenon initiated by low level noise exposure we analyzed glutathione reductase, gamma-glutamyl cysteine synthetase, and catalase in stria vascularis and organ of Corti fractions from cochleae of chinchillas exposed to a sound conditioning paradigm. Chinchillas were either (A) kept in quiet cages (control), (B) exposed to conditioning noise of a 0.5 kHz octave band (90 dB for 6 h/day for 10 days), (C) exposed to high level noise (105 dB for 4 h) or (D) exposed to conditioning noise (B) followed by exposure to the higher level noise (C). Each of the noise exposure conditions (B, C, D) induced changes in the levels of these three antioxidant enzymes. The enzyme-specific activity data for the four subject groups support the following two hypotheses. (1) Changes in glutathione reductase, gamma-glutamyl cysteine synthetase, and catalase play a role in attenuating hearing loss associated with sound conditioning followed by high level noise. (2) Hair cells in the organ of Corti are protected from noise-induced damage by increasing stria vascularis levels of catalase, a hydrogen peroxide scavenging enzyme, and of enzymes involved in maintaining glutathione in the reduced state. The model formulated by these hypotheses suggests that agents that protect or augment the glutathione system in the cochlea may be protective against noise-induced hearing loss.
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PMID:Changes in cochlear antioxidant enzyme activity after sound conditioning and noise exposure in the chinchilla. 955 76

A wide spectrum of human lung diseases is characterized by the presence of granulomas. Although understanding of the pathways leading to their development remains incomplete, data from in vitro studies suggest that neutrophils, monocytes, and their secreted products (eg, hydrogen peroxide, H2O2) influence the pathogenesis of pulmonary granulomatous disease through the regulation of local chemokine and cytokine production. Using a well-characterized rat model of glucan-induced pulmonary granulomatous vasculitis, we sought to determine the role of intracellular glutathione (GSH) redox status in the expression of monocyte chemoattractant protein-1 (MCP-1). Previous studies have revealed that vascular wall MCP-1 expression is obligatory for granuloma development and that both neutrophils and hydrogen peroxide are required for MCP-1 induction. Because in vitro expression of MCP-1 is in part mediated by the redox-sensitive transcription factors nuclear factor-kappa B (NF-kappaB) and activator protein-1 (AP-1), we studied their activation as a function of varying intracellular GSH redox status in the pathogenesis of glucan-induced pulmonary granulomatosis. Infusion of particulate yeast cell wall glucan into rats resulted in a rapid decrease in intracellular GSH concentrations which was accompanied by the activation of NF-kappaB and AP-1. The pattern of AP-1 and NF-kappaB activation in turn correlated temporally with the expression of MCP-1. Administration of L-buthionine-S, R-sulfoximine, a specific inhibitor of gamma-glutamyl cysteine synthetase, resulted in a significant reduction in intracellular GSH pools. GSH depletion resulted in a more than 100% increase in pulmonary MCP-1 concentrations and increased cytosolic to nuclear translocation of NF-kappaB while having no effect on AP-1 levels. These observations suggest that in the pathogenesis of pulmonary granulomatous disease, intracellular glutathione redox status modulates the expression of MCP-1 through redox-sensitive transcription factors.
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PMID:Intracellular glutathione redox status modulates MCP-1 expression in pulmonary granulomatous vasculitis. 1041 24

Covalent binding of L-methionine as an external aldimine to the pyridoxal 5'-phosphate-cofactor in the K41A mutant of O-acetylserine sulfhydrylase from Salmonella typhimurium induces a large conformational change in the protein. Methionine mimics the action of the substrate O-acetyl-L-serine during catalysis. The alpha-carboxylate moiety of L-methionine in external aldimine linkage with the active site pyridoxal 5'-phosphate forms a hydrogen bonding network to the "asparagine-loop" P67-T68-N69-G70 which adopts a different conformation than in the native protein. The side-chain nitrogen of Asn69 moves more than 7 A to make a hydrogen bond to the alpha-carboxylate group of the inhibitor. As the external aldimine is formed, the PLP tilts by 13 degrees along its longitudinal axis such that C4' moves toward the entrance to the active site and the side-chain of the methionine is directed toward the active site entrance. The local rearrangement acts as a trigger to induce a large global conformational change in the protein. A subdomain comprised of beta-strand 4, alpha-helix 3, beta-strand 5 and alpha-helix 4 moves towards the active site by a rotation of 7 degrees. This subdomain movement results in a reduction of the severe twist of its central beta-sheet and reduces the active site entrance to a small hole, giving access only to small molecules like sulfide, the second substrate, or acetate, the first product.
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PMID:Ligand binding induces a large conformational change in O-acetylserine sulfhydrylase from Salmonella typhimurium. 1045 98

The enteric protist parasites Entamoeba histolytica and Entamoeba dispar possess a cysteine biosynthetic pathway, unlike their mammalian host, and are capable of de novo production of L-cysteine. We cloned and characterized cDNAs that encode the regulated enzyme serine acetyltransferase (SAT) in this pathway from these amoebae by genetic complementation of a cysteine-auxotrophic Escherichia coli strain with the amoebic cDNA libraries. The deduced amino acid sequences of the amoebic SATs exhibited, within the most conserved region, 36-52% identities with the bacterial and plant SATs. The amoebic SATs contain a unique insertion of eight amino acids, also found in the corresponding region of a plasmid-encoded SAT from Synechococcus sp., which showed the highest overall identities to the amoebic SATs. Phylogenetic reconstruction also revealed a close kinship of the amoebic SATs with cyanobacterial SATs. Biochemical characterization of the recombinant E. histolytica SAT revealed several enzymatic features that distinguished the amoebic enzyme from the bacterial and plant enzymes: 1) inhibition by L-cysteine in a competitive manner with L-serine; 2) inhibition by L-cystine; and 3) no association with cysteine synthase. Genetically engineered amoeba strains that overproduced cysteine synthase and SAT were created. The cysteine synthase-overproducing amoebae had a higher level of cysteine synthase activity and total thiol content and revealed increased resistance to hydrogen peroxide. These results indicate that the cysteine biosynthetic pathway plays an important role in antioxidative defense of these enteric parasites.
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PMID:Characterization of the gene encoding serine acetyltransferase, a regulated enzyme of cysteine biosynthesis from the protist parasites Entamoeba histolytica and Entamoeba dispar. Regulation and possible function of the cysteine biosynthetic pathway in Entamoeba. 1054 89

We investigated immunohistochemical expression of manganese superoxide dismutase (MnSOD) and three hydrogen peroxide (H(2)O(2)) scavenging pathways, i.e. catalase (CAT), gamma-glutamyl cysteine synthetase (gammaGCS) and thioredoxin (Trx) system in normal bronchial epithelium, bronchial metaplasia and dysplasia and correlated their expression with NF-kappaB activation (p50) and proliferation (Ki67). Normal bronchial epithelium was positive for MnSOD, heavy and light subunits of gammaGCS, CAT and Trx and TrxR. Metaplastic epithelium showed strongest expression of gammaGCSh and Trx, whereas dysplastic epithelium expressed most prominently MnSOD and CAT. There was a significant correlation between expression of gammaGCSh and gammaGCSl (P=0.034) and Trx and TrxR (P=0.037). Trx expression also correlated with gammaGCSh (P<0.001) and gammaGCSl (P=0.012) and TrxR with gammaGCSh (P<0.001) but not with gammaGCSl immunoreactivity (P=0.744). Expression of p50 was highest in metaplastic epithelium while Ki67 was highest in dysplastic lesions. Expression of Trx and gammaGCSh correlated inversely with age of the patients (R=-0.6038, P<0.001 for Trx and R=-0.6162, P<0.001 for gammaGCSh). Changes in the expression of these enzymes in bronchial lesions might be due to alterations of antioxidative mechanisms due to irritation via exogenous toxins and activation of reactive oxygen species (ROS) known to be associated with induction of metaplasia and dysplasia in the bronchial tree.
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PMID:Expression of antioxidant enzymes in bronchial metaplastic and dysplastic epithelium. 1249 89


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