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 effects of the synthetic dibromo-pyrethroid insecticide deltamethrin on some hepatic phase I and II enzyme activities were studied in rat liver. The animals were treated with daily doses of 5 and 10 mg/kg of both pure insecticide or its commercial formulation (Decis), administered i.p. in corn oil for 7 days. The following enzyme activities were studied: NADPH-cytochrome-P450 reductase, aryl-hydrocarbon hydroxylase, aminopyrine N-demethylase, glutamyl cysteine synthetase, glutathione S-transferase, glutathione peroxidase, peroxisomal acyl-CoA oxidase, catalase, and urate oxidase. Both deltamethrin and its commercial formulation were effective in modifying the activities of several of these hepatic xenobiotic-metabolizing enzymes. However, some differences in enzyme modifications were found between treatment with pure or commercial deltamethrin, the latter being more active. This effect could be ascribed to additives, solvents, and chemical intermediates present in the Decis formulation. These results suggest that exposure to this deltamethrin commercial formulation could be more dangerous than exposure to deltamethrin alone, both in terms of its hepatotoxicity and/or alterations in the hepatic biotransformation of other occupational/environmental xenobiotics.
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PMID:Studies on hepatic xenobiotic-metabolizing enzymes in rats treated with insecticide deltamethrin. 747 74

A procedure has been developed to prepare the apoenzyme of O-acetylserine sulfhydrylase (apoOASS) by first converting the native enzyme to the alpha-aminoacrylate intermediate and dialyzing against 5 M guanidinium chloride. Aposulfhydrylase is stable for at least a month in buffers containing phosphate or phosphate analogues. Reconstitution of aposulfhydrylase with pyridoxal 5'-phosphate (PLP), 2'-methyl PLP (2'-MePLP), and pyridoxal 5'-deoxymethylenephosphonate (PDMP) results in enzymatically competent proteins. Pyridoxal in the absence and presence of phosphate and pyridoxal 5'-phosphate monomethyl ester are unable to form a Schiff base with apoOASS. The reconstitution of apoOASS with PLP is highly cooperative judged by the initial rate of activity regained and shows no evidence of saturation with PLP. The reconstituted enzymes have been studied using 31P NMR spectroscopy. The 31P NMR of the aposulhydrylase reconstituted with PLP exhibits a chemical shift of 5.2 ppm, identical to that of native enzyme. The latter has been interpreted in terms of a strong ionic interaction between enzyme and the 5'-phosphate of PLP (P. F. Cook, S. Hara, S. Nalabolu, and K. D. Schnackerz, 1992, Biochemistry 31, 2298-2303). Reconstitution with 2'-MePLP gives a lower chemical shift of 4.95 ppm, suggesting a weaker ionic interaction at the 5'-phosphate when compared to native enzyme. The PDMP-reconstituted enzyme gives a chemical shift of 23.7 ppm, consistent with the monoanionic form of the bound phosphonate. All of the chemical shifts are pH independent. The apoenzyme has also been reconstituted with pyridoxal 5'-sulfate. Although the resulting enzyme is not active in the overall reaction, it forms the external Schiff base. The PDMP- and 2'-MePLP-reconstituted enzymes have also been studied in the presence of amino acid reactants and analogues, and results are discussed in terms of the mechanism of OASS.
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PMID:Resolution of pyridoxal 5'-phosphate from O-acetylserine sulfhydrylase from Salmonella typhimurium and reconstitution of apoenzyme with cofactor and cofactor analogues as a probe of the cofactor binding site. 750 62

Nitric oxide (NO) has been demonstrated to play a protective role in cell injury. In this study, we have explored the effect of NO and two NO donors (sodium nitroprusside [SNP] and isosorbide dinitrate [ISDN]) on cellular glutathione (GSH) levels in a rat lung fibroblast cell line (RFL6 cells). SNP and ISDN significantly increased cellular GSH in RFL6 cells (5 x 10(-4) M SNP: 21.9 +/- 3.6 nmol/10(6) cells and 5 x 10(-3) M ISDN: 27.6 +/- 1.7 nmol/10(6) cells versus control: 13.2 +/- 0.4 nmol/10(6) cells; P < 0.05). The stimulatory effect of SNP and ISDN on GSH was first seen at 6 h and peaked at 12 to 24 h. A similar increase in GSH was observed in RFL6 cells exposed to 400 ppm NO for 7.5 h (NO: 20.5 +/- 3.4 nmol/10(6) cells versus control: 11.9 +/- 2.4; P < 0.05). SNP and ISDN also increased cellular GSH in bovine pulmonary artery smooth muscle cells (BPSMC) and bovine pulmonary artery endothelial cells (BPAEC). Buthionine sulfoximine (BSO) (0.01 mM), an inhibitor of the GSH synthetic enzyme gamma-glutamyl cysteine synthetase, blocked the increase in GSH in RFL6 cells seen with both SNP and ISDN. In BPAEC, exposure to NO donors for 24 h stimulated glutamate uptake (SNP: 441 +/- 19 pmol/10 min/10(6) cells and ISDN: 677 +/- 48 pmol/10 min/10(6) min/10(6) cells versus control: 222 +/- 9 pmol/10 min/10(6); P < 0.05). This effect paralleled the increase in GSH. In RFL6 cells, only SNP increased glutamate uptake after 24 h of incubation. In summary, NO and NO donors increase cellular GSH in RFL6 cells, BPAEC, and BPSMC. The mechanism of this effect is unclear but may involve upregulation of the normal GSH synthetic pathways. This observation may explain in part the protective effect of NO seen in some cell culture systems and may contribute to a protective effect against oxidant injury in vivo.
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PMID:Nitric oxide increases cellular glutathione levels in rat lung fibroblasts. 754 74

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

Serine acetyltransferase (SATase; EC 2.3.1.30), which catalyzes the reaction connecting serine and cysteine/methionine metabolism, plays a regulatory role in cysteine biosynthesis in plants. We have isolated a cDNA clone encoding SATase by direct genetic complementation of a Cys- mutation in Escherichia coli using an expression library of Citrullus vulgaris (watermelon) cDNA. The cDNA encodes a polypeptide of 294 amino acids (31,536 Da) exhibiting 51% homology with that of E. coli SATase. DNA-blot analysis indicated the presence of a single copy of the SATase gene (sat) in watermelon. RNA hybridization analysis suggested the relatively ubiquitous and preferential expression in the hypocotyls of etiolated seedlings. Immunoblot analysis indicated the accumulation of SATase predominantly in etiolated plants. L-Cysteine, an end product of the cysteine biosynthetic pathway, inhibited the SATase in an allosteric manner, indicating the regulatory function of SATase in this metabolic pathway, whereas beta-(pyrazole-1-yl)-L-alanine, a secondary metabolite formed partly through the cysteine biosynthetic pathway, showed no inhibitory effect. A multi-enzyme complex was formed from recombinant proteins of SATase and cysteine synthase (O-acetylserine(thiol)-lyase) from watermelon, suggesting efficient metabolic channeling from serine to cysteine, preventing the diffusion of intermediary O-acetyl-L-serine.
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PMID:Molecular cloning and characterization of a plant serine acetyltransferase playing a regulatory role in cysteine biosynthesis from watermelon. 760

Three types of cysteine synthase (CSase, EC 4.2.99.8) isozymes were purified from spinach leaves. Each isozyme was isolated to homogeneity by preparative PAGE. These isozymes were revealed to have different primary structures by amino-acid and proteinase digestion analyses, respectively. The enzymes designated as CSase 1, CSase 2 and CSase 3 with reference to the mobility on native PAGE were characterized with respect to physicochemical and enzymatic properties, and it was found that those enzymes had similar properties. It was also found that CSase 1 could be attributed to chloroplasts.
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PMID:Comparative studies on cysteine synthase isozymes from spinach leaves. 766 16

The coding sequences of the cysE and cysK genes from Escherichia coli, which encode the enzymes of the cysteine biosynthetic pathway, namely, serine acetyltransferase (EC 2.3.1.30) and O-acetylserine sulfhydrylase (or cysteine synthase [EC 4.2.99.8]), were modified for expression in eukaryotic cells and introduced into murine L cells. A number of fusion genes comprising the cysE or cysK coding sequences joined to the promoter of the ovine metallothionein-Ia (MT-Ia) gene and various portions of the ovine growth hormone (GH) gene were prepared. Significant differences in the level of transcription were observed, depending on the amount and arrangement of the GH gene sequences used, the highest levels being obtained with the constructs MTCE10 and MTCK7, which contained only the GH 3' untranslated gene sequences. These two constructs were fused to produce the gene MTCEK1. In this single DNA sequence, each bacterial gene is under independent MT-Ia promoter control. Expression of the cysK sequence in this construct (MT-Ia promoter-cysE-3' GH sequence-MT-Ia promoter-cysK-3' GH sequence) was elevated compared with expression of the cysK gene in MTCK7. However, expression of the cysE sequence in MTCEK1 was only 40% of that of the cysE gene cloned into MTCE10. The double-promoter configuration, which enhances the expression of the second gene in MTCEK1, is proposed as a model for the modification of bacterial genes in general.
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PMID:Introduction and expression of the bacterial genes cysE and cysK in eukaryotic cells. 768 85

Liver homogenate glutathione (GSH) content, lipid peroxide levels and the activities of GSH metabolizing enzymes were studied in rats after 24 hours of galactosamine (GalN) treatment. Lipid peroxide levels increased whereas hepatic GSH content was decreased significantly. On the other hand, hepatic gamma-glutamyl cysteine synthetase activity was unaffected by GalN administration but gamma-glutamyl transpeptidase activity increased.
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PMID:Hepatic gamma-glutamyl cysteine synthetase and gamma-glutamyl transpeptidase activities in galactosamine-treated rats. 774 60

The main enzymes of the gamma-glutamyl cycle in the testis were studied during the onset of spermatogenesis. The activities of gamma-glutamyl transpeptidase, 5-oxoprolinase, and gamma-glutamyl cysteine synthetase, and levels of glutathione were measured in testis homogenates and Sertoli cell preparations obtained from 10-, 18-, and 26-day-old rats. A significant increase of all enzyme activities with the animal age was observed. Level of glutathione also increased in an age-dependent manner. Since the gamma-glutamyl cycle is involved in the cellular incorporation of amino acids, the present findings suggest that this uptake mechanism may be relevant during spermatogenic onset in which synthesis and secretion of specific proteins are essential for germ cell development.
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PMID:Glutathione and gamma-glutamyl cycle enzymes in rat testis during sexual maturation. 785 69


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