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Biomedical subjects

J Chaudhuri

Publications and source records attributed to J Chaudhuri.

At least 37 records · Page 2Linked to original sources
PubMed1994

Purification and characterization of bacterially expressed mammalian translation initiation factor 5 (eIF-5): demonstration that eIF-5 forms a specific complex with eIF-2.

Eukaryotic translation initiation factor 5, eIF-5, has been purified to apparent electrophoretic homogeneity from overproducing Escherichia coli cells expressing the cDNA of the initiation factor under the control of the T7 promoter-T7 RNA polymerase system. Purified recombinant eIF-5 mimics natural eIF-5 isolated from mammalian cells in size, in specific activity, in its ability to catalyze the hydrolysis of GTP bound to the 40S initiation complex, and in the subsequent joining with 60S ribosomal subunits to form the 80S initiation complex. Further characterization of eIF-5 demonstrates that eIF-5 specifically associates with eIF-2, forming an eIF-2.eIF-5 complex. The protein complex sediments in glycerol gradients with an apparent M(r) of 160,000, suggesting that the two proteins associate in a 1:1 stoichiometry. The association between the two initiation factors is highly specific. Addition of 32P-labeled eIF-5 to a partially purified rabbit reticulocyte initiation factor preparation that contained, in addition to eIF-2 and eIF-5, other initiation factors and many other proteins resulted in the specific binding of labeled eIF-5 only to eIF-2, forming a 160-kDa protein complex. In agreement with these observations, we found that in crude initiation factor preparations derived from rabbit reticulocyte lysates, eIF-5 was present as an eIF-2.eIF-5 complex. The significance of eIF-2.eIF-5 complex formation in the overall mechanism of GTP hydrolysis in protein synthesis initiation is discussed.

Animals↗
PubMed1993

Characterization of mammalian translation initiation factor 5 (eIF-5). Demonstration that eIF-5 is a phosphoprotein and is present in cells as a single molecular form of apparent M(r) 58,000.

Eukaryotic translation initiation factor 5 (eIF-5) promotes the hydrolysis of GTP bound to the 40 S initiation complex (40 S.mRNA.Met-tRNA(f).eIF-2.GTP). Using assays that measure (a) the release of 32Pi from [gamma-32P]GTP bound to the 40 S initiation complex and (b) the eIF-5-dependent formation of an 80 S initiation complex from a preformed 40 S initiation complex containing bound [35S]Met-tRNA(f), we devised a novel and rapid procedure for purifying eIF-5 from rabbit reticulocyte lysates in high yield. Highly purified eIF-5 is a monomeric protein with a M(r) of about 58,000. However, in partially purified preparations, 58-kDa eIF-5 is associated with other cellular protein(s) and sediments in glycerol gradient centrifugation as a protein of M(r) approximately 160,000. Chicken antibodies to native eIF-5 were isolated from egg yolks of laying hens immunized with rabbit reticulocyte eIF-5. 35S-Labeled eIF-5, isolated from rat anterior pituitary GH3 cells using affinity-purified anti-eIF-5 antibodies, also has an apparent M(r) of 58,000. eIF-5 immunoprecipitated from extracts of 32P-labeled GH3 cells was phosphorylated on serine residues. Phosphopeptide mapping revealed two major sites of phosphorylation, which are distinct from rabbit reticulocyte eIF-5 sites phosphorylated in vitro by casein kinase II. These results demonstrate that eIF-5 is a phosphoprotein that is present in cells as a single molecular form of apparent molecular weight 58,000.

Animals↗
PubMed1993

Differential modulation of organophosphate-sensitive muscarinic receptors in rat brain by parathion and chlorpyrifos.

We previously reported similar levels of brain cholinesterase inhibition but marked differences in toxicity following acute maximum tolerated doses of the organophosphate pesticides parathion and chlorpyrifos. Because extensive acetylcholinesterase inhibition often induces compensatory changes in cholinergic receptor populations, we compared the effects of parathion and chlorpyrifos on brain muscarinic receptors. Adult male rats were treated with vehicle or the maximum tolerated dose of parathion (18 mg/kg, sc) or chlorpyrifos (279 mg/kg, sc) and observed for signs of acute toxicity. Similarly treated animals were sacrificed at 2, 7, or 14 days after treatment for measurement of cholinesterase activity and binding to the nonselective muscarinic antagonist [3H]quinuclidinyl benzilate, the M2-preferential antagonist [3H]AFDX-384, and the high-affinity agonist [3H]cis-methyldioxolane. More acute toxicity was noted after parathion treatment. Both insecticides caused similar levels (> 85%) of maximal cholinesterase inhibition and reductions (up to 55%) in atropine-sensitive quinuclidinyl benzilate binding (i.e., total muscarinic receptors) and [3H]AFDX-384 binding in cortex and striatum. Parathion also reduced, whereas chlorpyrifos increased, total muscarinic receptor binding and [3H]AFDX-384 binding in the cerebellum. When tissues were preincubated with paraoxon (10 microM), radiolabeling of a subset of quinuclidinyl benzilate binding sites was blocked and the apparent densities of these organophosphate-sensitive receptors in all three tissues were decreased (16% maximal) by parathion but increased (up to 37%) by chlorpyrifos. Similarly, parathion decreased whereas chlorpyrifos increased [3H]cis-methyldioxolane binding sites in all three brain regions. We propose that differential modulation of these organophosphate-sensitive muscarinic receptors contributes to differences in acute toxicity following exposure to these pesticides.

Animals↗
PubMed1993

Purification of two azoreductases from Escherichia coli K12.

Two azoreductases (I and II) were purified to homogeneity from extracts of E. coli K12. Azoreductase I was a dimer of two identical subunits of molecular weight 28000 whereas azoreductase II was a monomer of 12,000 molecular weight. Both NADH and NADPH functioned as electron donors for the azoreductases. Azoreductase I and II used Ponceau SX, Tartrazine, Amaranth and Orange II as substrates. Ponceau SX was the best substrate for both the enzymes. However, azoreductase II utilized tartrazine, amaranth and orange II less efficiently than azoreductase I.

Azo Compounds↗
PubMed1992

Purification and partial characterization of two azoreductases from Shigella dysenteriae type 1.

Two azoreductases (I and II) were purified to homogeneity from extracts of Shigella dysenteriae (type 1). Azoreductase I was a dimer of identical subunits of M(r) 28,000, whereas azoreductase II was a monomer of 11,000 M(r). Both were flavoproteins, each containing 1 mol of FMN per mol enzyme. Both NADH and NADPH functioned as electron donors for the azoreductases. Azoreductase I used Ponceau SX, Tartrazine, Amaranth and Orange II as substrates. Azoreductase II utilized all the dyes except Amaranth.

Azo Compounds↗
PubMed1991

A PMA degrading constitutive organomercurial lyase in a broad-spectrum mercury resistant Bacillus pasteurii strain DR2.

A broad-spectrum Hg-resistant strain of B. pasteurii DR2 utilized phenylmercuric acetate (PMA) as sole source of carbon. This bacterial strain contained a constitutive organomercurial lyase which specifically degraded PMA but not other organo-mercurials. This PMA-lyase activity was also stimulated to different extents when this bacterial strain was grown in presence of different organic compounds as sole source of carbon.

Bacillus↗
PubMed1990

Hydrogen peroxide: a potent activator of dioxygenase activity of soybean lipoxygenase.

Hydrogen peroxide, an ubiquitous biologically occurring peroxide, was found to stimulate the dioxygenase activity of soybean lipoxygenase at the physiologically attainable concentration. The increase in enzyme specific activity was directly proportional to hydrogen peroxide concentration up to 0.5 nM. A decrease in the stimulation of dioxygenase activity was observed at higher concentrations. At low enzyme concentration up to 28-fold stimulation was noted when the formation of lipid hydroperoxide was monitored spectrophotometrically. The stimulation was further confirmed by increased oxygen uptake. It is proposed that the mechanism for in vivo activation involves hydrogen peroxide.

Hydrogen Peroxide↗
PubMed1989

Serotonin receptors in the tissues of adult Ascaris suum.

Serotonin (5-hydroxytryptamine, 5-HT) receptors in the muscle and intestinal tissues of adult Ascaris suum have been investigated. [3H] lysergic acid diethylamide (LSD) exhibited specific and saturable binding to membranes prepared from both intestine and muscle. The intestinal tissue membranes had an equilibrium dissociation constant (Kd) of 2.70 nM for LSD and a Kd of 2.50 microM for 5-HT. As compared to the intestine, the muscle membranes had comparatively higher affinity for both LSD (Kd = 1.80 nM) and 5-HT (Kd = 0.68 microM). The muscle membranes also had a high binding affinity for ketanserin, a 5-HT2 antagonist, (Kd = 16.7 nM) whereas intestinal membranes exhibited no specific binding of ketanserin. Serotonin significantly inhibited the binding of LSD to the intestinal and muscle tissue membranes while adrenergic and cholinergic drugs and histamine did not. This suggested that the binding of LSD, 5-HT and ketanserin to the parasite membranes was specific. Collectively, the data demonstrated the presence of a serotonin receptor in the muscle and intestinal tissues of the adult A. suum. The receptor in the muscle was pharmacologically similar to the mammalian serotonin type 2 receptor.

Animals↗
PubMed1989

Dioxygenase and peroxidase activities of soybean lipoxygenase: synergistic interaction between linoleic acid and hydrogen peroxide.

The interaction of H2O2 with soyabean lipoxygenase was investigated in the presence of linoleic acid. Dioxygenase activity was significantly higher at pH 9.0 than at pH 6.0. H2O2 at concentrations less than 1.0 nM stimulated linoleic acid oxidation synergistically and the magnitude of synergism was higher at pH 9.0. Linoleic acid dependent peroxidase activity towards benzidine, guaiacol, tetramethylbenzidine (TMBD) and tetramethylphenylenediamine (TMPD) was higher at pH 9.0, whereas pyrogallol and ABTS oxidation rates were higher at pH 6.0. H2O2 supported oxidation of benzidine, guaiacol, pyrogallol and ABTS was higher at pH 6.0, whereas TMPD, TMBD exhibited higher oxidation rates at pH 9.0. H2O2 in the presence of linoleic acid produced synergism in xenobotic metabolism and depending upon the substrate in question upto 11-fold increase in oxidation rate was noted.

Drug Synergism↗
PubMed1989

Cadmium resistance in some members of Enterobacteriaceae.

Strains of members of Enterobacteriaceae, namely Escherichia coli (18), Klebsiella aerogenes (16), and Serratia marcescens (16) were screened for Cd resistance or sensitivity. Only one strain each of these was resistant to high levels (25 n moles/0.05 ml) CdCl2. The Minimal inhibitory concentration (MIC) of sensitive strains ranged from 0.8-5 micrograms/ml. All the resistant strains were simultaneously resistant to a number of antibiotics. Treatment with sodium dodecyl sulfate eliminated resistance to Cd and to some antibiotics.

Cadmium↗
PubMed1988

Elimination of mercury, cadmium and antibiotic resistance from Acinetobacter lwoffi and Micrococcus sp. at high temperature.

Resistance determinants for HgCl2 and CdCl2 were eliminated along with a number of antibiotic resistance factors from Acinetobacter lwoffi and Micrococcus sp. at 44 degrees C. These organisms were orginally resistant to HgCl2, merbromin, CdCl2, Pb(NO3)2, benzylpenicillin, erythromycin, carbenicillin, tetracycline and sulfadiazine. Four different types of mutants from A. lwoffi (type I to IV) and one type of mutant from Micrococcus sp. (type V) were obtained, depending on the loss of particular resistance factors for HgCl2, merbromin, CdCl2 and antibiotics. In general, frequency of elimination of all the missing markers was very low (in the range of 10(-3) per bacterium). However, the missing determinants did not revert spontaneously.

Acinetobacter↗