Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0272170 (SDS)
 50,377 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Hydroxylamine is a direct-acting hematotoxic agent leading to hemolytic anemia in animals and man. The effect of hydroxylamine on the morphology, sulfhydryl status and membrane skeletal proteins of human erythrocytes were studied. Loss of reduced glutathione (GSH) from the red blood cells was directly proportional to the hydroxylamine concentration used. This loss of GSH was larger than the sum of the increase in the amounts of extracellular glutathione and intracellular oxidized glutathione (GSSG). The extracellular glutathione is mainly present as GSSG, which is in agreement with the fact that only GSSG is exported from the erythrocytes by membrane bound ATPases. Lack of GSSG export was not limited by decreased ATP levels in the erythrocytes and we concluded that the GSH that disappeared did not become available as intracellular GSSG. After reduction of the erythrocyte incubates the lost GSH was almost completely recovered indicating that the lost GSH is present in the cell as protein-glutathione mixed disulfides. Glutathione thus stored within the cell can be quickly recovered by combined thioltransferase and glutathione reductase activity when conditions become more favorable again. SDS-polyacrylamide gel electrophoresis of membrane ghosts from human red cells revealed changes in skeletal proteins with a smearing of bands 1, 2 and 3 to the higher molecular weight end of the gel and the appearance of new monomeric and dimeric hemoglobin bands at about 16 and 30 kD. The observed alterations are probably a consequence of disulfide bridge formation between cellular proteins (mainly hemoglobin) and skeletal proteins as well as between hemoglobin monomers. Exposure of hydroxylamine to erythrocytes caused severe Heinz body formation but the outside morphology of the cells was only marginally altered. The described changes in sulfhydryl status of the red blood cells are likely to play a major role in the premature splenic sequestration of hydroxylamine-damaged erythrocytes.
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PMID:Hydroxylamine treatment increases glutathione-protein and protein-protein binding in human erythrocytes. 939 34

microPx-11, a ferriheme undecapeptide proteolytic degradation product of cytochrome C is shown to be a peroxidase with broad specificity degrading H2O2 and tertiary butyl hydroperoxide. It is also capable of effectively eliminating superoxide and hydroxyl radical. The peroxidase loses activity in the presence of peroxide unless it is stabilized by ascorbate (Asc) or solutions such as aqueous humor or medium 199. While thiol but not disulfides inactivates the microPx-11, it is not inhibited in the presence of the rat lens which has a high GSH content. microPx-11 at concentrations 10 to 50 fold greater than are required to achieve good protective activity exhibits no toxicity based on cell viability, ATP levels and lens transparency after long-term incubations of alpha TN4-1 cells or cultured rat lens. The peroxidase is capable of protecting cultured rat lenses from photochemical stress where H2O2, O2.- and OH. are generated based on transparency, choline transport, epithelial cell viability and protein integrity as indicated by SDS-PAGE of the rat lens protein. In the absence of the peroxidase, extensive epithelial cell death and other degradative changes are observed. The DNA of alpha TN4-1 cells can also be protected from H2O2 induced single strand breaks by the microPx-11. The overall results suggest that a number of cytochrome C proteolytic degradation products are peroxidases which may be effective anti-cataract agents protecting the lens from oxidative stress.
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PMID:Microperoxidases catalytically degrade reactive oxygen species and may be anti-cataract agents. 946 80

The purpose of this communication is to elucidate if selenium plays a role in the function of granulocytes and lymphocytes. Thus, the incorporation of selenium in proteins from granulocytes and lymphocytes cultured with 1 microCi/mL radioactive Na2(75)SeO3 was studied. The protein peaks containing 75Se from two columns of Heparin Sepharose CL-6B and Sephacryl S-200 HR were separated further by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis. The results showed that the incorporation of 75Se into granulocytes was about six times higher than that of lymphocytes during a 96-h cultivation, however, the GSH-Px activity in granulocytes did not change significantly. On the other hand, the GSH-Px activity of lymphocytes rose significantly after three days cultivation. These data indicated that the main chemical form of selenium in granulocytes was not GSH-Px. Results from SDS-PAGE revealed a strongly 75Se-labeled protein band with subunit molecular weight of 15 kDa in the supernatant of granulocyte homogenate. However, the main chemical forms of selenium in the culture media of granulocytes and lymphocytes were found to be selenoprotein P. The different forms of selenium-containing proteins in the intracellular and extracellular media of granulocytes indicated the different functions of these proteins.
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PMID:Different selenium-containing proteins in the extracellular and intracellular media of leucocytes cultivated in vitro. 953 63

1-Aminocyclopropane-1-carboxylate (ACC) synthase is a key enzyme regulating the biosynthesis of the plant hormone ethylene. A wound-inducible zucchini ACC synthase cDNA was isolated by reverse-transcription polymerase chain reaction (RT-PCR) and expressed in a heterologous Escherichia coli BL21(DE3)pLysS:pET30a protein expression system. A method was developed and optimized for the renaturation of the ACC synthase expressed in the form of inclusion bodies. The optimum conditions were found to be unfolding in a buffer containing 100 mM Mops, pH 9.5, 6 M urea, and 50 mM DTT, for 3 h at 4 degrees C and refolding by a combined process of dialysis and dilution in 100 mM Mops, pH 8, 30 mM Chaps, and 5 mM GSH at a protein concentration of 45 microg/ml. The purified enzyme has a specific activity of 90,000 U mg-1 and exhibits an apparent homogeneity on SDS-PAGE fractionation. Biochemical characterization of the refolded enzyme revealed a high degree of similarity to the enzyme purified from the soluble source. The refolded enzyme was found to be a dimer with a native size of 110 kDa, a Km of 23 microM, and a Vmax of 112,000 U mg-1.
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PMID:Renaturation of 1-aminocyclopropane-1-carboxylate synthase expressed in Escherichia coli in the form of inclusion bodies into a dimeric and catalytically active enzyme. 953 97

A 2-hydroxychromene-2-carboxylate isomerase was purified from a cell-free extract of naphthalenesulfonate-assimilating Pseudomonas sp. TA-2 to an electrophoretically homogeneous state by successive column chromatography on DEAE-cellulose, DEAE-Toyopearl 650M, Sephadex G-75, Hydroxyapatite, and Mono Q. The enzyme had a molecular mass of 25 and 27 kDa as estimated by SDS-PAGE and Superdex 200, respectively. Its N-terminal 30 amino acid sequence had high homology with the deduced amino acid sequences of the 2HC2CA isomerase of nahD (a gene of naphthalene metabolism), pahD (a gene of naphthalene and phenanthrene metabolism), and doxJ (a gene of dibenzothiophene metabolism). The enzymatic product was a trans isomer. The isomerase activity was inhibited in the presence of monoiodoacetate or Hg2+, but not by preincubation with monoiodoacetate or N-ethylmaleimide. GSH functioned as a cofactor and activated the enzyme at above 0.15 mM.
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PMID:Purification and some properties of 2-hydroxychromene-2-carboxylate isomerase from naphthalenesulfonate-assimilating Pseudomonas sp. TA-2. 972 70

Thioltransferase is a general GSH-disulfide reductase of importance for redox regulation. The protein thioltransferase has been purified to apparent homogeneity on SDS-PAGE from the Arabidopsis thaliana seed. The purification procedures included DEAE-cellulose ion exchange chromatography, Sephadex G-75 gel filtration, Q-Sepharose ion exchange chromatography, and DEAE-Sephadex A-25 ion exchange chromatography. The enzyme has a molecular mass of 22 kDa and a pI of 4.8, and it is heatstable. The protein had broad specificities for substrates ranging from low-molecular disulfides (S-sulfocysteine and cystine) to protein disulfides (trypsin and insulin). However, it could not reduce the disulfide linkages of ribonuclease A and bovine serum albumin. It could utilize non-disulfide substrates such as dehydroascorbic acid and alloxan. The protein can reduce the disulfide bond in 2-hydroxyethyl disulfide with an optimum pH of 8.5. Its activity was greatly activated by monothiol compounds such as reduced glutathione and L-cysteine.
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PMID:Thioltransferase from Arabidopsis thaliana seed: purification to homogeneity and characterization. 985 42

Activated phagocyte cells generate hypochlorite (HOCl) via the release of H2O2 and the enzyme myeloperoxidase. Plasma proteins are major targets for HOCl, although little information is available about the mechanism(s) of oxidation. In this study the reaction of HOCl (at least 50 microM) with diluted fresh human plasma has been shown to generate material that oxidizes 5-thio-2-nitrobenzoic acid; these oxidants are believed to be chloramines formed from the reaction of HOCl with protein amine groups. Chloramines have also been detected with isolated plasma proteins treated with HOCl. In both cases chloramine formation accounts for approx. 20-30% of the added HOCl. These chloramines decompose in a time-dependent manner when incubated at 20 or 37 degrees C but not at 4 degrees C. Ascorbate and urate remove these chloramines in a time- and concentration-dependent manner, with the former being more efficient. The reaction of fresh diluted plasma with HOCl also gives rise to protein-derived nitrogen-centred radicals in a time- and HOCl-concentration-dependent manner; these have been detected by EPR spin trapping. Identical radicals have been detected with isolated HOCl-treated plasma proteins. Radical formation was inhibited by excess methionine, implicating protein-derived chloramines (probably from lysine side chains) as the radical source. Plasma protein fragmentation occurs in a time- and HOCl-concentration-dependent manner, as evidenced by the increased mobility of the EPR spin adducts, the detection of further radical species believed to be intermediates in protein degradation and the loss of the parent protein bands on SDS/PAGE. Fragmentation can be inhibited by methionine and other agents (ascorbate, urate, Trolox C or GSH) capable of removing chloramines and reactive radicals. These results are consistent with protein-derived chloramines, and the radicals derived from them, as contributing agents in HOCl-induced plasma protein oxidation.
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PMID:Hypochlorite-induced oxidation of proteins in plasma: formation of chloramines and nitrogen-centred radicals and their role in protein fragmentation. 1033

Diacylglycerol lipase (DGL) was solubilized from human platelet microsomes with heptyl-beta-D-thioglucoside, and purified to homogeneity on SDS-PAGE using a combination of chromatographic and electrophoretic methods. The molecular mass of the purified DGL was estimated to be 33 kDa. Its apparent pI was pH 6.0, as determined by Immobiline isoelectro-focusing. The enzymatic activity of the partially purified DGL was investigated in the presence of a variety of inhibitors and reagents, as well as its pH and calcium dependence. Thiol reagents such as p-chloromercurubenzoic acid (pCMB), N-ethylmaleimide (NEM), and HgCl2 inhibited the activity, while dithiothreitol (DTT) and reduced glutathione (GSH) enhanced it. In addition, the enzymatic activity was inhibited by two serine blockers, phenylmethylsulfonyl fluoride (PMSF) and diisopropyl fluorophosphate (DFP), and by a histidine modifying reagent, p-bromophenacyl bromide (pBPB). These results suggest that cysteine, serine and histidine residues are required for the enzymatic activity of DGL. DGL was optimally active in the pH range of 7-8 and its activity did not change significantly in the presence of various calcium concentrations, even in the presence of 2 mM EGTA. This indicates that DGL can hydrolyze substrates with a basal cytosolic free Ca2+ level in the physiological pH range. A DGL inhibitor, RHC-80267, inhibited DGL activity in a dose-dependent manner with an IC50 (the concentration required for 50% inhibition) of about 5 microM. Unexpectedly, several phospholipase A2 (PLA2) inhibitors were potent inhibitors of DGL activity (IC50<5 microM), suggesting that the catalytic mechanisms of DGL and PLA2 may be similar. Finally, we show that DGL activity was inhibited by 2-monoacylglycerols (2-MGs), the reaction products of this enzyme. Among the three 2-MGs tested (2-arachidonoyl glycerol, 2-stearoyl glycerol, and 2-oleoyl glycerol), 2-arachidonoyl glycerol was the most potent inhibitor.
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PMID:Purification and characterization of diacylglycerol lipase from human platelets. 1034 10

Prostaglandin (PG) E synthase was solubilized with 6 mM sodium deoxycholate from the microsomal fraction of bovine hearts. The enzyme was purified by about 800-fold to apparent homogeneity. The specific activity of the purified enzyme was about 830 mU/mg of protein, and the K(m) value for PGH(2) was 24 microM. The molecular weight of the enzyme was about 31000 on SDS-polyacrylamide gel electrophoresis and was about 60000 by gel filtration. The enzyme was separated from glutathione (GSH) S-transferase by DEAE-Toyopearl column chromatography, and did not exhibit any GSH S-transferase activity towards four different substrates. The purified enzyme was active in the absence of GSH, but it was activated by various SH-reducing reagents including dithiothreitol, GSH, or beta-mercaptoethanol. This is the first reported purification of membrane-bound PGE synthase to apparent homogeneity.
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PMID:Purification and characterization of membrane-bound prostaglandin E synthase from bovine heart. 1044 27

A glutathione S-transferase (GST) fraction was isolated from cytosol prepared from catfish intestinal mucosa by GSH-agarose affinity chromatography and its molecular weight, isoelectric points, substrate specificities and immunochemical cross-reactivity were examined. Intestinal GSTs were purified 100-fold with respect to cytosolic activity with 1-chloro-2, 4-dinitrobenzene and had high activity with ethacrynic acid, (+/-)benzo(a)pyrene-4,5-oxide, and (+/-)anti-benzo(a)pyrene-7,8-dihydrodiol-9,10-epoxide, but a low activity with 1,2-dichloro-4-nitrobenzene. SDS-polyacrylamide gel electrophoresis revealed the presence of a single band with relative molecular mass of 26700. Gel isoelectric focusing showed a major band with a pI of 8.2. A polyclonal antibody prepared against a GST pi-protein isolated from catfish proximal intestine cross-reacted well with the affinity isolated GST fraction. The catfish antibody also cross-reacted with GST from human placenta which contains predominantly pi-class GST (Mannervik, B., Guthenberg, C., 1981. Glutathione transferase (human placenta). In: Jakoby, W.B. (Ed.), Methods in Enzymology, 77. Academic Press, New York, pp. 231-235; Polidoro, G., Dillio, C., Arduini, A., Frederici, G., 1981. Molecular and catalytic properties of purified glutathione transferase from human placenta. Biochem. Med. 22, 247-259; Dao, D.D., Partridge, C.A., Kurosky, A., Awasthi, Y.C., 1982. Subunit structure of glutathione-S-transferase of human liver and placenta. IRSC Med. Sci, Lib. Compend. 10, 175; Dao, D.D., Partridge, C.A., Kurosky, A., Awasthi, Y.C., 1984. Human glutathione transferase. Characterization of the anionic forms from lung and placenta. Biochem. J. 221, 33-41), but poorly with human liver cytosol. The affinity-isolated protein fraction from whole intestine contained proteins that were immunologically related to all four major classes of human GSTs tested. N-terminal sequence analysis of the predominant band obtained by 2D electrophoresis indicated a marked homology (63-70% identical) to mammalian pi form GST isozymes and very strong similarity (80%) to a salmon hepatic GST that was designated a pi form (Dominey, R.J., Nimmo, I.A., Cronshaw, A.D., Hayes, J.D., 1991. The major glutathione S-transferase in salmonid fish livers is homologous to the mammalian pi class GST. Comp. Biochem. Physiol. (B) 100 (1), 93-98). Other bands contained insufficient protein for N-terminal analysis. Taken together, these results indicate that the predominant intestinal GST isoform is related to the pi-class enzymes, but minor GSTs related to other families are also present.
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PMID:Activities of affinity-isolated glutathione S-transferase (GST) from channel catfish whole intestine. 1081 4


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