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Biomedical subjects

A Shrift

Publications and source records attributed to A Shrift.

At least 19 recordsLinked to original sources
PubMed1985

Single transporter for sulfate, selenate, and selenite in Escherichia coli K-12.

A Michaelis-Menten kinetic analysis of the transport of sulfate, selenate, and selenite into Escherichia coli K-12 showed that the three dianions were transported by the same carrier. Km values, used as a measure of the affinity of each ligand for the carrier, showed that sulfate was bound 5 times more tightly than selenate and 37 times more tightly than selenite. The specificity ratio, Vmax/Km, also indicated that sulfate was the preferred ligand. There was little difference in the ratios for selenate and selenite.

Biological Transport↗
PubMed1982

Aerobic, Selenium-Utilizing Bacillus Isolated from Seeds of Astragalus crotalariae.

Bacillus sp. strain SS, an aerobic, gram-positive sporeformer, was isolated from seeds of Astragalus crotalariae, a selenium-accumulating plant. This bacillus grew in a nutrient broth (containing beef extract and peptone) if the medium was supplemented with high concentrations of selenium. Concentrations of Na(2)SeO(3) that supported growth ranged from 3 to 100 mM. After 24 h of growth, the culture developed a deep red color characteristic of elemental selenium. When selenium was provided in the form of selenate, the pattern of growth showed a prolonged lag period, from 24 to 48 h. Final growth remained below that of cells cultured in the presence of selenite, and only a light red color developed. Concentrations of selenate below 40 mM failed to support growth. Tellurate, though not tellurite, could replace selenite, but only over a narrow concentration range, 5 to 10 mM. By 24 h, the typical black color of elemental tellurium developed. Bacillus sp. strain SS grew also in brain heart infusion broth and Trypticase soy broth (BBL Microbiology Systems, Cockeysville, Md.) without the addition of selenium or tellurium compounds. When added to these media, 50 mM selenite was tolerated and metabolized by the organism. The crucial distinction between this bacillus and other selenium-tolerant organisms (e.g., Salmonella) remains: under certain conditions, growth requirements of Bacillus sp. strain SS are fulfilled by selenium (and tellurium) compounds.

Journal Article↗
PubMed1982

Selective assimilation of selenite by Escherichia coli.

The assimilation of selenite by Escherichia coli involves a transport process specific for this anion. Cystine, a repressor of sulfate and selenite uptake, when added to the growth medium, had no effect on selenite uptake; nor did sulfite inhibit assimilation of selenite. Cells grown aerobically in a basic salts medium transported selenite at an initial rate of 0.14 mumol . g-1 min-1 and a Vmax of 393.0 mumol. g-1 . min-1. In contrast, cells grown in a medium that contained the trace elements necessary for synthesis of the selenoenzyme formate dehydrogenase took up selenite at a significantly faster rate (initial rate = 0.27 mumol . g-1 . min-1, Vmax = 658.2 mumol . g-1 . min-1). Km values for the transport process in the two media, however, were found to be similar. The results suggest the existence of a metabolic pathway, specific for selenite, responsible for the incorporation of selenium into formate dehydrogenase.U

Biological Transport↗
PubMed1981

Exclusion of selenium from proteins of selenium-tolerant astragalus species.

Protein fractions from three selenium-tolerant and three selenium-sensitive Astragalus species, grown in the presence of [(75)Se]selenate, were analyzed for their selenium content. Though tolerant species are known to accumulate considerably more selenium than do sensitive plants, protein fractions from the three selenium accumulators were found to contain significantly less selenium (0.46 to 0.57 picomoles selenium per milligram protein) than did protein fractions from the three nonaccumulators (4.17 to 5.02 picomoles selenium per milligram protein). Under similar conditions, seedlings of Vigna radiata (L.) Wilczek had taken up selenium (6.31 picomoles selenium per milligram protein) at levels comparable to those observed in the proteins of the nonaccumulator Astragali. These results establish that the ability to tolerate and to circumvent the toxic effects of selenium, characteristic of the accumulator species of Astragalus, is associated with a reduced incorporation of this element into protein.

Journal Article↗
PubMed1981

Selenium toxicity: aminoacylation and Peptide bond formation with selenomethionine.

Selenomethionine and methionine were compared as substrates for in vitro aminoacylation, ribosome binding, and peptide bond formation with preparations from wheat germ. Selenomethionine paralleled methionine in all steps of the translation process except peptide bond formation. Peptide bond formation with the initiating species of tRNA(Met) demonstrated that selenomethionyl-tRNA(Met) was less effective as a substrate than was methionyl-tRNA(f) (Met). Participation of selenomethionine in the initiation process of translation could be expected to reduce the overall rate of protein synthesis and might aid in explaining selenium toxicity in selenium-sensitive plants.

Journal Article↗
PubMed1981

In vitro incorporation of selenomethionine into protein by astragalus polysomes.

Selenium-accumulator plants synthesize selenium compounds that differ from those produced by nonaccumulators. To determine if there are any subcellular differences between accumulators and nonaccumulators in the use of selenomethionine in vitro, polysomes from Astragalus crotalariae (accumulator) and Astragalus lentiginosis (nonaccumulator) were translated in the presence of selenomethionine. Polysomes from both species efficiently used selenomethionine in vitro during the translation process. Inasmuch as no differences in the incorporation of selenomethionine into protein were observed between polysomes from the two types of Astragalus, it can be inferred that in accumulators there exists a mechanism that either prevents synthesis of selenomethionine or modifies this selenocompound to a derivative that cannot be incorporated into protein.

Journal Article↗
PubMed1980

Reduction of DL-selenocystine and isolation of L-seleoncysteine.

Cystine, selenocytsine, and several analogs were reduced by dithiothreitol (DTT), beta-mercaptoethanol (ME) and sodium borohydride (NaBH4). DTT was the most effective; DTT to cystine ratios from 10 to 80 were equally effective. With selenocysteine, however, absorption was considerably reduced at all ratios. Selenocysteine was identified as the reduction product by reaction with Gaitonde's reagent, comparison of absorption spectra, paper chromatograhy, utilization by cysteinyl-tRNA synthetase fro Paracoccus denitrificans and Vigna radiata, changes in solubility after DTT treatment, and comparison of infrared spectra. During the ATP-PPi exchange assay, DTT and ME convert cysteine and selenocysteine derivatives to cysteine and selenocysteine which serve as substrates for cysteinyl-tRNA synthetase.

Amino Acyl-tRNA Synthetases↗
PubMed1980

In Vitro Incorporation of Selenomethionine into Protein by Vigna radiata Polysomes.

Vigna radiata polysomes efficiently incorporated [(75)Se]selenomethionine, [(14)C]methionine, and [(14)C]leucine in vitro. The optimal conditions for translation were determined to be 4.8 millimolar Mg(2+), 182 millimolar K(+), and pH 7.4. The rates of incorporation of [(75)Se]selenomethionine and [(14)C]methionine were similar when measured separately, but [(75)Se]selenomethionine incorporation was 35% less than [(14)C]methionine incorporation when both amino acids were present in equal molar concentrations. Polyacrylamide gel electrophoresis of the hot trichloroacetic acid precipitable translation products demonstrated synthesis of high molecular weight labeled proteins in the presence of [(75)Se]selenomethionine or [(35)S]methionine. No major differences in molecular weights could be detected in the electrophoretic profiles. Utilization of selenomethionine during translation by Vigna radiata polysomes establishes a route for the assimilation of selenomethionine by plants susceptible to selenium toxicity.

Journal Article↗
PubMed1980

Identification of Selenocysteine in the Proteins of Selenate-grown Vigna radiata.

Selenocysteine, the selenium analog of cysteine, was identified in proteins of Vigna radiata (L.) Wilczak grown with selenate. To stabilize selenocysteine and prevent its breakdown, the carboxymethyl derivative was synthesized by the addition of iodoacetic acid to the protein extract from [(75)Se]selenate-grown plants. A (75)Se-labeled component of the carboxymethylated protein hydrolysate possessed chromatographic properties identical to those of a (14)C-labeled carboxymethylselenocysteine standard during paper and thin layer chromatography and during gel-exclusion, anion-exchange, and cation-exchange column chromatography. Detection of selenocysteine in proteins of a selenium-sensitive plant, and the possibility that the presence of this compound alters normal functions, provides an explanation for the toxic effects of selenium.

Journal Article↗
PubMed1980

Assimilation of selenate and selenite by Salmonella typhimurium.

A comparative study of selenate and selenite assimilation by Salmonella typhimurium revealed that selenite was not transported by the sulphate permease. Selenite uptake could be detected both in wild-type cells repressed for sulphate transport and in mutants that lacked a functional sulphate permease. In contrast, selenate was assimilated by the same process as was sulphate; selenate transport was repressed under the same conditions which repressed sulphate uptake and was absent in permeaseless mutants. Selenite transport was absent if cells were glucose starved or treated with either azide or p-chloromercuribenzoate. The pH optimum was between pH 6 and pH 7; transport was most rapid at 36 degrees C. The double reciprocal plot for selenite transport at different substrate concentrations was biphasic: between 10 and 50 microM SeO32(-) the apparent Km was 37.8 microM, and at higher concentrations, 2.87 mM. The transport rate for 0.1 mM SeO32(-) was significantly stimulated by sulphite concentrations up to 5.0 mM, with a maximum at 3.0 mM SO32-. The results establish a selenite transport process, in S. typhimurium, as the initial step of an assimilatory pathway selective for selenium.

Azides↗
PubMed1979

Phylogenetic distribution of glutathione peroxidase.

1. The enzyme glutathione peroxidase (E.C.1.11.1.9), known to be a selenoprotein from mammalian sources, was detected in the following vertebrates: fish, frog, salamander, and turtle. 2. Among invertebrates, the enzyme was detected in crayfish and snail but not in insects or earthworm. 3. No plant tissues or microorganisms showed any evidence of the enzyme activity. 4. The presence of the enzyme activity in so many animal groups implies the widespread occurrence of genetic information for the specific assimilation of the selenium atom.

Animals↗
PubMed1979

Uptake of selenium-75 by human lymphocytes in vitro.

Selenite uptake by human lymphocytes was studied both in whole blood and in isolated cells. When 75Se-selenite (75Se-SeO3-2--) is supplied to whole blood, it is converted by the erythrocytes to a form which rapidly becomes bound to plasma proteins. Studies with a variety of inhibitors indicated that the process is not energy dependent but that sulfhydryl groups are required. The 75Se bound to plasma proteins is absorbed by lymphocytes in preference to 75SeO3-2-. By the use of selective inhibitors (respiratory, sulfhydryl, protein biosynthetic) it was demonstrated that the uptake of either form of 75Se requires neither energy nor protein synthesis; however, all the sulfhydryl inhibitors cause a decrease of absorption. A scheme which summarizes the pathway of selenite conversion in human blood and uptake by lymphocytes is presented; the data indicate that plasma proteins function as carriers of selenium to lymphocytes.

Blood Proteins↗
PubMed1976

Utilization of Selenocysteine by a Cysteinyl-tRNA Synthetase from Phaseolus aureus.

An l-cysteinyl-tRNA synthetase (EC 6.1.1.16) from Phaseolus aureus has been purified approximately 200-fold. The enzyme uses selenocysteine as substrate in the ATP-PPi exchange assay; other cysteine analogs were inactive. The molecular weight as determined by Sephadex G-200 column chromatography is about 61,000; sodium dodecyl sulfate and 8 m urea acrylamide gel electrophoresis indicate that the enzyme is a dimer consisting of two identical monomers of molecular weight 30,000. A method for the preparation of selenocysteine from selenocystine is described.

Journal Article↗
PubMed1974

Form of selenium in selenite enrichment media for isolation of salmonellae.

Selenite-F and selenite-cystine media, commercially available for the routine isolation of salmonellae, were treated by anion exchange chromatography to separate the selenium from other components of the media. A chemical assay, based on an ascorbic acid reduction, showed that the selenium was all in the form of selenite.

Ascorbic Acid↗
PubMed1971

Response to selenium by callus cultures derived from astragalus species.

Callus cultures were obtained from five selenium accumulator and three nonaccumulator species of Astragalus. Their morphological characteristics and their growth responses to light, sucrose, kinetin, and 2,4-dichlorophenoxyacetic acid are described. Calluses derived from accumulator species characteristically retained their tolerance to high concentrations of selenate and selenite, whereas calluses derived from nonaccumulator species were markedly inhibited by these two forms of selenium. Competition between sulfate and selenate was demonstrated. The two types of calluses could not be distinguished on the basis of (75)Se-labeled selenate or selenite uptake. Neutron activation analysis failed to show differences in selenium content between the two types of calluses grown on media to which no selenium had been added.

Journal Article↗
PubMed1969

Transport of selenate and selenite into astragalus roots.

After incubation for 1 hr with (75)Se-selenate, excised roots of Astragalus crotalariae, a selenium-accumulating species, and A. lentiginosus, a nonaccumulator, had absorbed radioactivity to levels well over the external concentration. About 98% of the radioactivity was ethanol-soluble, and when analyzed by column and paper chromatography and by electrophoresis proved to be selenate. This and previous evidence shows an active transport for selenate. Considerably less radioactivity was absorbed when (75)Se-selenite was supplied to the excised roots, and levels of the ethanol-soluble radioactivity did not exceed the external concentration. A good deal of the radioactivity was ethanol-insoluble. Analysis of the soluble radioactivity from both species showed appreciable conversion of selenite to other forms.

Biological Transport, Active↗