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)

Colominic acid (CA), an alpha-(2-->8) N-acetylneuraminic acid (sialic acid) polymer (average molecular weight of 10 kDa) was activated by periodate oxidation of carbon 7 at the non-reducing end of the saccharide. The oxidized CA was then coupled to catalase by reductive amination in the presence of sodium cyanoborohydride. The extent of sialylation of catalase, estimated by ammonium sulfate precipitation as 3.8+/-0.4 (mean+/-S.D.) moles of CA per mole of catalase, did not improve significantly when depolymerized CA was used in the coupling reaction. At the end of the coupling reaction, sialylated catalase exhibited a two-fold (70%) retention of initial activity compared to enzyme controls (29-35%) subjected to the same conditions. Formation of sialylated catalase was confirmed by ammonium sulfate or trichloroacetic acid precipitation, molecular sieve chromatography and SDS-PAGE electrophoresis. Enzyme kinetics studies revealed an increase in the apparent Km of the enzyme from 70.0 (native) to 122.9 mmol l-1 H2O2 (sialylated catalase) indicating a reduction of enzyme affinity for the substrate (hydrogen peroxide) on sialylation. Compared to native enzyme, sialylated catalase was much more stable in the presence of specific proteinases, completely resisting degradation by chymotrypsin and losing only some of its activity in the presence of trypsin. The increased stability conferred to catalase by sialylation agrees with similar observations on enzymes modified by other hydrophilic molecules (e.g., monomethoxypoly(ethyleneglycol)) and suggests that steric stabilization with the biodegradable polysialic acid may prove an alternative means to render therapeutic proteins more effective in vivo.
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PMID:Synthesis, characterization and properties of sialylated catalase. 865 33

Cynomolgus (Macaca fascicularis) monkeys from three families, which showed symptoms of early onset macular degeneration was studied. Two proteins, albumin and glyceraldehyde 3-phosphate dehydrogenase, were found to have markedly altered concentrations in whole retina of the monkeys with early onset macular degeneration, compared with normal controls. SDS-polyacrylamide gel patterns detected a 40-70% increase in the concentration of albumin and about 65% decrease in the concentration of glyceraldehyde 3-phosphate dehydrogenase in these affected retinas. There was however no significant difference in the relative concentrations of albumin in the plasma samples of affected and normal monkeys belonging to the three families studied and to an unrelated family. These initial findings suggest that degradative as well as antioxidant enzymes might be involved in the mechanisms leading to macular degeneration. In addition, the results also correlate with a possible role of these two proteins in H2O2 toxicity and appear to indicate that oxidative stress is significant in the etiology of early onset macular degeneration.
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PMID:Studies on the mechanism of early onset macular degeneration in cynomolgus (Macaca fascicularis) monkeys. I. Abnormal concentrations of two proteins in the retina. 869 30

The 100000Xg supernatant parasite platyhelminth Schistosoma mansoni exhibits a glutathione peroxidase activity with the substrate phosphatidylcholine hydroperoxide. Purification yielded a protein of 20 kDa molecular mass both on gel filtration column chromatography and SDS/PAGE, thus suggesting that S. mansoni expresses a protein similar to the mammalian selenoenzynic phospholipid-hydroperoxide glutathione peroxidase. Kinetic analysis and substrate specificity corroborated this assumption, the second-order rate constants for the oxidation of the ground-state enzyme (k+1) being higher with phosphatidylcholine hydroperoxide than with other peroxide substrates, such as cumene liydroperoxide or H2O2, and quantitatively similar to those of mammalian phospholipid-hydroperoxide glutathione peroxidase. Partial sequencing of the protein and selenium measurement by neutron activation analysis established that the purified peroxidase corresponded to the product of the S. mansoni gene previously reported and supposed to encode a selenium-containing glutathione peroxidase [Roche, C., Williams, D. L., Khalife, J., LePresle, T., Capron, A. & Pierce, R. J. (1994) Cloning and characterization of gene encoding Schistosoma mansoni glutathione peroxidase, Gene 138, 149 - 152]. S. mansoni thus contains a scienoperoxidase sharing molecular mass, catalytic efficiency and substrate specificity with phospholipid-hydroperoxide glutathione peroxidase, dismantling the concept that those enzymes are unique to vertebrate organisms.
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PMID:A selenium-containing phospholipid-hydroperoxide glutathione peroxidase in Schistosoma mansoni. 870 88

Neurogranin (Ng) is a prominent protein kinase C (PKC) substrate which binds calmodulin (CaM) in the absence of Ca2+. Rat brain Ng contains four cysteine residues that were readily oxidized by nitric oxide (NO) donors, 1,1-diethyl-2-hydroxy-2-nitrosohydrazine (DEANO) and sodium nitroprusside, and by oxidants, H2O2 and o-iodosobenzoic acid. NO oxidation of Ng resulted in a conformational change detectable by increased electrophoretic mobility upon SDS-polyacrylamide gel electrophoresis. The NO-mediated mobility shift was reversed by treatment with dithiothreitol and was blocked by modification of Ng sulfhydryl groups with 4-vinylpyridine. Both the nonphosphorylated and PKC-phosphorylated Ng were susceptible to NO oxidation. Modification of Ng by DEANO was blocked by CaM in the absence of Ca2+; while in the presence of Ca2+, CaM did not protect Ng from oxidation by DEANO. CaM also failed to protect DEANO-mediated oxidation of PKC-phosphorylated Ng with or without Ca2+. Oxidation of Ng by the various oxidants apparently resulted in the formation of intramolecular disulfide bond(s) as judged by a reduction of apparent Mr on SDS-polyacrylamide gel electrophoresis; this oxidized form, unlike the reduced form, did not bind to CaM-affinity column. The oxidized Ng was also a poorer substrate for PKC; both the reduced and oxidized forms had similar Km values, but the Vmax of the oxidized form was about one-fourth of the reduced one. When comparing the rate of DEANO-mediated nitrosation of Ng with other sulfhydryl-containing compounds, it became evident that Ng ranked as one of the best NO acceptors among those tested, including serum albumin, glutathione, and dithiothreitol. Ng present in the rat brain synaptosomal preparations was also oxidized by DEANO in a dose-dependent manner when analyzed by immunoblot with a polyclonal antibody against this protein. These results suggest that Ng is a likely target of NO and other oxidants and that oxidation/reduction may serve as a mechanism for controlling both the PKC phosphorylation and the CaM-binding affinity of this protein.
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PMID:Nitric oxide modification of rat brain neurogranin affects its phosphorylation by protein kinase C and affinity for calmodulin. 879 3

Reactive oxygen species are autocrine and paracrine modulators of cell behavior. Hydrogen peroxide, a cellular oxidant, has been shown to stimulate mesangial cell proliferation. In the present study we analyzed the H2O2-induced early signaling events. Immunofluorescence analysis revealed a H2O2 induced dose-dependent increase in tyrosine phosphorylation. Short treatment (2 or 5 min) with 5 mM H2O2 induced a mitogenic response and a significant (P < 0.01) increase in the number of cells compared to non-treated controls. Proteins extracted from H2O2 (0.1 to 10 mM) treated cells were separated on SDS-PAGE and subjected to immunoblot analysis with anti-phosphotyrosine. A dose-dependent induction of tyrosine phosphorylation of 180 kDa, 120 kDa and 60 kDa proteins was observed within 1 to 10 minutes. By sequentially using immunoprecipitation and immunoblotting the 180 kDa tyrosine phosphorylated band was shown to represent both PDGF alpha- and beta-receptors. The tyrosine phosphorylated 60 kDa protein was identified as the cytoplasmic protein tyrosine kinase pp60c-src. The c-src phosphorylation was associated with an inhibition of c-src kinase activity, suggesting phosphorylation of tyrosine 527 in the c-src regulatory domain. Pretreatment with catalase completely abrogated the H2O2-induced PDGF receptor and c-src tyrosine phosphorylation. These data support the notion that the activation of a signaling pathway involving the PDGF receptors and c-src contributes to the mitogenic effects of reactive oxygen species.
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PMID:Oxidative stress induces tyrosine phosphorylation of PDGF alpha-and beta-receptors and pp60c-src in mesangial cells. 880 85

Lung surfactant protein A (SP-A) is the most abundant surfactant-associated protein present in the lung and respiratory tract. SP-A binds to several pathogens via its C-type lectin domains, and may act as an opsonin, mediating adhesion to cells via the collectin receptor. Binding studies using SP-A are made difficult by its apparent instability following radioiodination. This study investigated the effect of oxidation (via radioiodination and exposure to H2O2) on the structural and functional characteristics of SP-A. Radioiodinated SP-A, stored at 4 degrees C, retained carbohydrate binding activity after labeling. After 10 days storage, the radioiodinated SP-A was indistinguishable on SDS-PAGE from freshly radioiodinated SP-A, but sedimentation coefficient and Stokes radius values changed dramatically, indicating SP-A depolymerization. Such a quaternary structural breakdown, with a concomitant reduction in carbohydrate binding activity, is likely to be due to oxidative cleavage of disulfide bonds. Comparable results were observed upon radioiodination of the structurally similar molecule C1q. Consequently, the effect of prolonged incubation with H2O2 upon SP-A was investigated, with similar results. Thus, exposure to oxidizing agents leads to breakdown of the hexameric quaternary structure of SP-A, often to native polypeptides, with an attendant loss of binding activity. Such an effect may have consequences for the physiological role of SP-A in the lung.
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PMID:Characterization of radioiodinated lung surfactant protein A (SP-A) and the effects of oxidation on SP-A quaternary structure and activity. 887 89

Peroxidase activity is detectable in Aedes aegypti ovaries, containing developing eggs, at 24 h following blood feeding, and peak peroxidase activity is reached at 36-48 h after the blood-meal. Peroxidase is associated with the chorion layer in mature eggs and the majority of the enzyme is released from the chorion layer by treating the isolated chorion fraction with SDS/urea. Analysis of the SDS/urea solubilized chorion proteins using SDS-PAGE with tropolone/H2O2 or dopa staining verified the presence of both peroxidase and phenol oxidase in the released chorion proteins. The molecular weight of chorion peroxidase is about 61,000 Da as determined by SDS-PAGE analysis. Incubation of the solubilized chorion proteins with tyrosine and H2O2 produces dityrosine, and hyrolysis of hardened egg chorion results in the detection of dityrosine and trityrosine in the chorion hydrolysate. Data suggest that chorion peroxidase is involved in the hardening of the mosquito egg chorion by catalyzing the formation of ditryrosine through tyrosine residues on structural proteins. The overall hardening of the A. aegypti egg chorion includes both peroxidase-mediated chorion protein crosslinking through dityrosine formation and phenol oxidase-catalyzed chorion melanization.
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PMID:Involvement of peroxidase in chorion hardening in Aedes aegypti. 890 May 99

Oxidants play a significant role in endothelial cell dysfunction through modulation of diverse biochemical reactions and signal transduction pathways. Towards understanding the role of oxidants in vascular injury, we studied the effect of hydrogen peroxide (H2O2), vanadate, and pervanadate (V(4+)-OOH) on [32Pi] uptake and protein phosphorylation in bovine pulmonary artery endothelial cells (BPAEC). The incorporation of labelled [32Pi] into BPAEC was dependent on the concentration of the oxidant employed and time of incubation. Of the oxidants tested, pervanadate (10 microM) induced maximum incorporation of [32Pi] into cells (two- to threefold over control) followed by H2O2 (1 mM) and vanadate (100 microM) and clear differences in labeled protein profiles were noticed between control and oxidant treated cells. The proteins, analyzed by SDS-PAGE, showed distinct increases in labeling patterns ranging from 21-205 kDa, as evidenced by autoradiography. While the majority of the incorporated [32Pi] was in serine/threonine residues, immunoprecipitation and immunoblotting of cell lysates, using an antiphosphotyrosine antibody, revealed that oxidant treatment resulted in significant increases in total protein tyrosine phosphorylation. Most significantly, immunoprecipitation of cell lysates, from pervanadate treatment showed distinct tyrosine phosphorylation of 22 kDa protein, which was identified as caveolin, a marker of caveolae. Pervanadate-mediated phosphorylation was effectively inhibited by staurosporine (5 microM), while genistein showed only partial attenuation. Furthermore, H2O2 treatment resulted in enhanced phosphorylation of 24 kDa protein, which was attenuated by genistein. In addition, oxidant-treated cells exhibited increased tyrosine kinase activity and decreased phosphatase activity. These data show differences in labeling profiles of proteins in response to different oxidants, suggesting differential modulation of distinct protein kinases/phosphatases.
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PMID:Activation of protein phosphorylation by oxidants in vascular endothelial cells: identification of tyrosine phosphorylation of caveolin. 895 27

Vulnerable areas of atherosclerotic plaques often contain lipid-laden macrophages and display matrix metalloproteinase activity. We hypothesized that reactive oxygen species released by macrophage-derived foam cells could trigger activation of latent proforms of metalloproteinases in the vascular interstitium. We showed that in vivo generated macrophage foam cells produce superoxide, nitric oxide, and hydrogen peroxide after isolation from hypercholesterolemic rabbits. Effects of these reactive oxygens and that of peroxynitrite, likely to result from simultaneous production of nitric oxide and superoxide, were tested in vitro using metalloproteinases secreted by cultured human vascular smooth muscle cells. Enzymes in culture media or affinity-purified (pro-MMP-2 and MMP-9) were examined by SDS-PAGE zymography, Western blotting, and enzymatic assays. Under the conditions used, incubation with xanthine/xanthine oxidase increased the amount of active gelatinases, while nitric oxide donors had no noticeable effect. Incubation with peroxynitrite resulted in nitration of MMP-2 and endowed it with collagenolytic activity. Hydrogen peroxide treatment showed a catalase-reversible biphasic effect (gelatinase activation at concentrations of 4 microM, inhibition at > or = 10-50 microM). Thus, reactive oxygen species can modulate matrix degradation in areas of high oxidant stress and could therefore contribute to instability of atherosclerotic plaques.
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PMID:Reactive oxygen species produced by macrophage-derived foam cells regulate the activity of vascular matrix metalloproteinases in vitro. Implications for atherosclerotic plaque stability. 895 20

Oxygen-derived free radicals have been reported to damage the sarcoplasmic reticulum (SR) Ca(2+)-ATPase, potentially contributing to cellular Ca2+ overload and myocardial damage after ischemia and reperfusion. To determine whether the ATP binding site on Ca(2+)-ATPase is involved in oxygen radical injury, SR vesicles containing bound Ca(2+)-ATPase were isolated from rabbit cardiac and skeletal muscle and exposed to a hydroxyl radical (.OH)-generating system consisting of H2O2 and Fe(3+)-nitrilotriacetic acid in amounts that generate a magnitude of .OH similar to that which occurs in the reperfused heart. .OH exposure completely inhibited Ca(2+)-ATPase activity and SR 45Ca uptake for both cardiac and skeletal muscle. In contrast, when the purified vesicles were premixed with 1 mmol/L ATP before exposure to .OH, complete protection was observed: there was no loss of ATPase activity or 45Ca transport. No significant protection occurred with adenosine, sucrose, AMP, or ADP (1 mmol/L each). SDS-gel electrophoresis indicated that .OH did not damage the primary structure of the enzyme. Electron paramagnetic resonance spin-trapping experiments demonstrated that ATP did not scavenge .OH. These results suggest that .OH denatures the SR Ca(2+)-ATPase by directly attacking the ATP binding site, and occupation of the active site by ATP protects against .OH-induced loss of enzymatic activity and SR Ca2+ transport. The depletion of ATP that occurs during ischemia may enhance the toxic effect of .OH at the time of reperfusion.
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PMID:Hydroxyl radical inhibits sarcoplasmic reticulum Ca(2+)-ATPase function by direct attack on the ATP binding site. 897 25


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