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1.
The selenophosphate synthetases from several organisms contain a selenocysteine residue in their active site where the Escherichia coli enzyme contains a cysteine. The synthesis of these enzymes, therefore, depends on their own reaction product. To analyse how this self-dependence is correlated with the selenium status, e.g. after recovery from severe selenium starvation, we expressed the gene for the selenocysteine-containing selenophosphate synthetase from Haemophilus influenzae (selD HI) in an E. coliΔselD strain. Gene selD HI gave rise to a selenium-containing gene product and also supported – via its activity – the formation of E. coli selenoproteins. The results provide evidence either for the suppression of the UGASec codon with the insertion of an amino acid allowing the formation of a functional product or for a bypass of the selenophosphate requirement. We also show that the selenocysteine synthesis and the insertion systems of the two organisms are fully compatible despite conspicuous differences in the mRNA recognition motif. Received: 8 July 1997 / Accepted: 3 September 1997  相似文献   

2.
In vivo and in vitro studies were conducted to investigate the chemical forms by ion-exchange chromatography of selenium (Se) present in rat and ovine spermatozoa. After injection with 75Se-selenite, the form of 75Se in rat sperm was selenocysteine, but selenocysteine and selenomethionine (SeMet) were present in ovine sperm. Presumably, synthesis of SeMet by rumen microbes are responsible for its presence in ovine sperm. In vitro incubation of ram sperm with selenocysteine or SeMet produced no changes, but incubation with selenite produced a compound that eluted one fraction before SeMet from the ion-exchange column. After treatment of this fraction with mercaptoethanol, it eluted in a later fraction upon rechromatography, suggesting it to be selenodicysteine. This compound is apparently formed because of high levels of cysteine in semen. Cysteine, reduced glutathione, and oxidized glutathione were also found in semen. The significance of the results is discussed.  相似文献   

3.
4.
The path of unspecific incorporation of selenium in Escherichia coli   总被引:2,自引:0,他引:2  
The path of unspecific selenium incorporation into proteins was studied in Escherichia coli mutants blocked in the biosynthesis of cysteine and methionine or altered in its regulation. Selenium incorporation required all enzymatic steps of cysteine biosynthesis except sulfite reduction, indicating that intracellular reduction of selenite occurs nonenzymatically. Cysteine (but not methionine) supplementation prevented unspecific incorporation of selenium by repressing cysteine biosynthesis. On the other hand, when the biosynthesis of cysteine was derepressed in regulatory mutants, selenium was incorporated to high levels. These findings and the fact that methionine auxotrophic strains still displayed unspecific incorporation show that selenium incorporation into proteins in E. coli occurs mainly as selenocysteine. These findings also provide information on the labeling conditions for incorporating 75Se only and specifically into selenoproteins. Received: 2 May 1997 / Accepted: 23 June 1997  相似文献   

5.
Nitrogen from atmospheric deposition serves as the dominant source of new nitrogen to forested ecosystems in the northeastern U.S. By combining isotopic data obtained using the denitrifier method, with chemical and hydrologic measurements we determined the relative importance of sources and control mechanisms on nitrate (NO3 ) export from five forested watersheds in the Connecticut River watershed. Microbially produced NO3 was the dominant source (82–100%) of NO3 to the sampled streams as indicated by the δ15N and δ18O of NO3 . Seasonal variations in the δ18O–NO3 in streamwater are controlled by shifting hydrologic and temperature affects on biotic processing, resulting in a relative increase in unprocessed NO3 export during winter months. Mass balance estimates find that the unprocessed atmospherically derived NO3 stream flux represents less than 3% of the atmospherically delivered wet NO3 flux to the region. This suggests that despite chronically elevated nitrogen deposition these forests are not nitrogen saturated and are retaining, removing, and reprocessing the vast majority of NO3 delivered to them throughout the year. These results confirm previous work within Northeastern U.S. forests and extend observations to watersheds not dominated by a snow-melt driven hydrology. In contrast to previous work, unprocessed atmospherically derived NO3 export is associated with the period of high recharge and low biotic activity as opposed to spring snowmelt and other large runoff events.  相似文献   

6.
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 [75Se]selenate-grown plants. A 75Se-labeled component of the carboxymethylated protein hydrolysate possessed chromatographic properties identical to those of a 14C-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.  相似文献   

7.
Cysteine S-conjugate β-lyases are pyridoxal 5′-phosphate-containing enzymes that catalyze β-elimination reactions with cysteine S-conjugates that possess a good leaving group in the β-position. The end products are aminoacrylate and a sulfur-containing fragment. The aminoacrylate tautomerizes and hydrolyzes to pyruvate and ammonia. The mammalian cysteine S-conjugate β-lyases thus far identified are enzymes involved in amino acid metabolism that catalyze β-lyase reactions as non-physiological side reactions. Most are aminotransferases. In some cases the lyase is inactivated by reaction products. The cysteine S-conjugate β-lyases are of much interest to toxicologists because they play an important key role in the bioactivation (toxication) of halogenated alkenes, some of which are produced on an industrial scale and are environmental contaminants. The cysteine S-conjugate β-lyases have been reviewed in this journal previously (Cooper and Pinto in Amino Acids 30:1–15, 2006). Here, we focus on more recent findings regarding: (1) the identification of enzymes associated with high-M r cysteine S-conjugate β-lyases in the cytosolic and mitochondrial fractions of rat liver and kidney; (2) the mechanism of syncatalytic inactivation of rat liver mitochondrial aspartate aminotransferase by the nephrotoxic β-lyase substrate S-(1,1,2,2-tetrafluoroethyl)-l-cysteine (the cysteine S-conjugate of tetrafluoroethylene); (3) toxicant channeling of reactive fragments from the active site of mitochondrial aspartate aminotransferase to susceptible proteins in the mitochondria; (4) the involvement of cysteine S-conjugate β-lyases in the metabolism/bioactivation of drugs and natural products; and (5) the role of cysteine S-conjugate β-lyases in the metabolism of selenocysteine Se-conjugates. This review emphasizes the fact that the cysteine S-conjugate β-lyases are biologically more important than hitherto appreciated.  相似文献   

8.
To determine the chemical and physicochemical characteristics of dissolved organic carbon in the Ado River and the Yasu River, the main rivers flowing into Lake Biwa, the adsorption behavior onto hydrous iron oxide (HIO) and the reactivity to KMnO4 oxidant were investigated in parallel with measurement of the distribution profiles of dissolved organic carbon (DOC) along the rivers. In one year of observation at the mouths of the two rivers, DOC concentrations were found to vary in the Ado over the range 0.28–1.21 mg C l−1 and in the Yasu over the range 1.01–2.68 mg C l−1. Act-DOC, one of the fractions separated from the total DOC by its adsorption-active character onto HIO at pH 4, was thought primarily to control the variation of total DOC, as in Lake Biwa. The int-DOC, another fraction separated by its adsorption-inert or -inactive character onto HIO, remained at almost a steady value around 0.18 ± 0.07 mg C l−1 in the Ado, which was lower than that (0.35 ± 0.05 mg C l−1) in Lake Biwa. The act-DOC in river waters was reactive to KMnO4 oxidant, showing a linear relation with the amount of permanganate consumed for the reaction (chemical oxygen demand: COD). In river waters, the relation can be approximated by a straight line expressed as COD (mg O2 l−1) = 0.64 × act-DOC (mg C l−1) − 0.02. In contrast, in the lake water the relation was COD (mg O2 l−1) = 0.97 × act-DOC (mg C l−1) − 0.50. Received: March 3, 1999 / Accepted: December 2, 1999  相似文献   

9.
In eukaryotes, the synthesis of selenoproteins depends on an exogenous supply of selenium, required for synthesis of the novel amino acid, selenocysteine, and on the presence of a “selenium translation element” in the 3′ untranslated region of mRNA. The selenium translation element is required to re-interpret the stop codon, UGA, as coding for selenocysteine incorporation and chain elongation. Messenger RNA lacking the selenium translation element and/or an inadequate selenium supply lead to chain termination at the UGA codon. We exploited these properties to provide direct translational control of protein(s) encoded by transfected cDNAs. Selenium-dependent translation of mRNA transcribed from target cDNA was conferred by mutation of an in-frame UGU, coding for cysteine, to UGA, coding for either selenocysteine or termination, then fusing the mutated coding region to a 3′ untranslated region containing the selenium translation element of the human cellular glutathione peroxidase gene. In this study, the biological consequences of placing this novel amino acid in the polypeptide chain was examined with two proteins of known function: the rat growth hormone receptor and human thyroid hormone receptor β1. UGA (opal) mutant-STE fusion constructs of the cDNAs encoding these two polypeptides showed selenium-dependent expression and their selenoprotein products maintained normal ligand binding and signal transduction. Thus, integration of selenocysteine had little or no consequence on the functional activity of the opal mutants; however, opal mutants were expressed at lower levels than their wild-type counterparts in transient expression assays. The ability to integrate this novel amino acid at predetermined positions in a polypeptide chain provides selenium-dependent translational control to the expression of a wide variety of target genes, allows facile 75Se radioisotopic labeling of the heterologous proteins, and permits site-specific heavy atom substitution. © 1996 Wiley-Liss, Inc.  相似文献   

10.
The formate dehydrogenases (Fdh) Fdh-O, Fdh-N, and Fdh-H, are the only proteins in Escherichia coli that incorporate selenocysteine at a specific position by decoding a UGA codon. However, an excess of selenium can lead to toxicity through misincorporation of selenocysteine into proteins. To determine whether selenocysteine substitutes for cysteine, we grew Escherichia coli in the presence of excess sodium selenite. The respiratory Fdh-N and Fdh-O enzymes, along with nitrate reductase (Nar) were co-purified from wild type strain MC4100 after anaerobic growth with nitrate and either 2 µM or 100 µM selenite. Mass spectrometric analysis of the catalytic subunits of both Fdhs identified the UGA-specified selenocysteine residue and revealed incorporation of additional, ‘non-specific’ selenocysteinyl residues, which always replaced particular cysteinyl residues. Although variable, their incorporation was not random and was independent of the selenite concentration used. Notably, these cysteines are likely to be non-essential for catalysis and they do not coordinate the iron-sulfur cluster. The remaining cysteinyl residues that could be identified were never substituted by selenocysteine. Selenomethionine was never observed in our analyses. Non-random substitution of particular cysteinyl residues was also noted in the electron-transferring subunit of both Fdhs as well as in the subunits of the Nar enzyme. Nar isolated from an E. coli selC mutant also showed a similar selenocysteine incorporation pattern to the wild-type indicating that non-specific selenocysteine incorporation was independent of the specific selenocysteine pathway. Thus, selenide replaces sulfide in the biosynthesis of cysteine and misacylated selenocysteyl-tRNACys decodes either UGU or UGC codons, which usually specify cysteine. Nevertheless, not every UGU or UGC codon was decoded as selenocysteine. Together, our results suggest that a degree of misincorporation of selenocysteine into enzymes through replacement of particular, non-essential cysteines, is tolerated and this might act as a buffering system to cope with excessive intracellular selenium.  相似文献   

11.
Selenoproteins are a unique family of proteins, characterized by the co-translational incorporation of selenium as selenocysteine, which play key roles in antioxidant defense. Among selenoproteins, selenoprotein P (Sepp1) is particularly distinctive due to the fact that it contains multiple selenocysteine residues and has been postulated to act in selenium transport. Within the brain, Sepp1 delivers selenium to neurons by binding to the ApoER2 receptor. Upon feeding a selenium-deficient diet, mice lacking ApoER2 or Sepp1 develop severe neurological dysfunction and exhibit widespread brainstem neurodegeneration, indicating an important role for ApoER2-mediated Sepp1 uptake in normal brain function. Selenocysteine lyase (Scly) is an enzyme that plays an important role in selenium homeostasis, in that it catalyzes the decomposition of selenocysteine and allows selenium to be recycled for additional selenoprotein synthesis. We previously reported that constitutive deletion of Scly results in neurological deficits only when mice are challenged with a low selenium diet. To gain insight into the relationship between Sepp1 and Scly in selenium metabolism, we created novel transgenic mice constitutively lacking both genes (Scly−/−Sepp1−/−) and characterized the neurobehavioral phenotype. We report that deletion of Scly in conjunction with Sepp1 further aggravates the phenotype of Sepp1−/− mice, as these mice needed supraphysiological selenium supplementation to survive, and surviving mice exhibited impaired motor coordination, audiogenic seizures, and brainstem neurodegeneration. These findings provide the first in vivo evidence that Scly and Sepp1 work cooperatively to maintain selenoprotein function in the mammalian brain.  相似文献   

12.
The cystine-glutamate exchanger, system xc , mediates the Na+-independent exchange of cystine into cells, coupled to the efflux of intracellular glutamate. System xc plays a critical role in glutathione homeostasis. Early studies of brain suggested that system xc was present primarily in astrocytes but not neurons. More recent work indicates that certain brain neurons have an active system xc . In the retina, system xc has been demonstrated in Müller and retinal pigment epithelial cells. We have recently suggested that two protein components of system xc , xCT and 4F2hc, are present in ganglion cells of the intact retina. Here, we have used (1) molecular and immunohistochemical assays to determine whether system xc is present in primary ganglion cells isolated from neonatal mouse retinas and (2) functional assays to determine whether its activity is regulated by oxidative stress in a retinal ganglion cell line (RGC–5). Primary mouse ganglion cells and RGC–5 cells express xCT and 4F2hc. RGC–5 cells take up [3H]glutamate in the absence of Na+, and this uptake is blocked by known substrates of system xc (glutamate, cysteine, cystine, quisqualic acid). Treatment of RGC–5 cells with NO and reactive oxygen species donors leads to increased activity of system xc associated with an increase in the maximal velocity of the transporter with no significant change in the substrate affinity. This is the first report of system xc in primary retinal ganglion cells and RGC–5 cells. Oxidative stress upregulates this transport system in RGC–5 cells, and the process is associated with an increase in xCT mRNA and protein but no change in 4F2hc mRNA or protein. This work was supported by National Institutes of Health grants EY014560 and EY012830.  相似文献   

13.
The role played by a bacterial community composed ofPseudomonas putida, strain 21;Pseudomonas stutzeri, strain 18; andPseudomonas sp., strain 5, and by physical and chemical factors in the degradation of CN and SCN was studied. It was shown that the degradation of CN is determined both by the action of bacteria and by abiotic physical and chemical factors (pH, O2, temperature, the medium agitation rate, etc.). The contribution of chemical degradation was found to increase drastically at pH below 9.0; when air was blown through the medium (irrespective of the pH value); under active agitation of the medium; and when the medium surface interfacing air was increased. Even at elevated pH values (9.0-9.2), suboptimal for bacterial growth, the microbial degradation could account for at most 20–25 mg/1 of CN, regardless of its initial concentration. When CN and SCN were concurrently present in the medium, the former compound was the first to be degraded by microorganisms. The rate of bacterial degradation of SCN under continuous cultivation in a chain of reactors was found to depend on its concentration, the medium flow rate, agitation rate, and the pattern of carbon source supply and could exceed 1 g/(l day). CN and SCN are utilized by bacteria solely as nitrogen sources. The mechanism of CN and SCN degradation by the microbial community is discussed. Deceased.  相似文献   

14.
The ability of spring barley (Hordeum vulgare cv. Akcent) to adjust the composition and function of the photosynthetic apparatus to growth irradiances of 25–1200 μmol m−2 s−1 was studied by gas exchange and chlorophyll a fluorescence measurements and high-performance liquid chromatography. The increased growth irradiance stimulated light- and CO2-saturated rates of CO2 assimilation expressed on a leaf area basis up to 730 μmol m−2 s−1 (HL730), whereas at an irradiance of 1200 μmol m−2 s−1 (EHL1200) both rates decreased significantly. Further, the acclimation to EHL1200 was associated with an extremely high chlorophyll a/b ratio (3.97), a more than doubled xanthophyll cycle pool (VAZ) and a six-fold higher de-epoxidation state of the xanthophyll cycle pigments as compared to barley grown under 25 μmol m−2 s−1 (LL25). EHL1200 plants also exhibited a long-term inhibition of Photosystem II (PS II) photochemical efficiency (F v/F m). Photosynthetic capacity, chlorophyll a/b and VAZ revealed a linear trend of dependence on PS II excitation pressure in a certain range of growth irradiances (100–730 μmol m−2 s−1). The deviation from linearity of these relationships for EHL1200 barley is discussed. In addition, the role of increased VAZ and/or accumulation of zeaxanthin and antheraxanthin in acclimation of barley to high irradiance is studied with respect to regulation of non-radiative dissipation and/or photochemical efficiency within PS II. This revised version was published online in June 2006 with corrections to the Cover Date.  相似文献   

15.
In an experimental site for reforestation of degraded area, three-year-old plants of Bertholletia excelsa Humb. & Bonpl. were subjected to different fertilization treatments: T0 = unfertilized control, T1 = green fertilization (branches and leaves) and T2 = chemical fertilization. Higher net photosynthetic rates (P N) were observed in T1 [13.2±1.0 μmol(CO2) m−2 s−1] compared to T2 [8.0±1.8 μmol(CO2) m−2 s−1] and T0 [4.8±1.3 μmol(CO2) m−2 s−1]. Stomatal conductance (g s), transpiration rate (E) and water use efficiency (WUE) of individuals of T1 and T2 did not differ significantly, however, they were by 88, 55 and 63%, respectively, higher in T1 than in the control. The mean values of variable fluorescence (Fv), performance index (P.I.) and total chlorophyll [Chl (a+b)] were higher in T1. Our results indicate that green fertilization improves photosynthetic structure and function in plants of B. excelsa in young phase.  相似文献   

16.
Selenium shares many chemical facets with sulphur but differs from it in the redox potential, especially of the Se2−/S2− oxidation state. The higher chemical reactivity of the deprotonated selenol has been used by Biology in the synthesis of the amino acid selenocysteine and its DNA-encoded incorporation into specific positions of proteins to enhance their structural role or their activity. Since selenocysteine is a steric isomer of cysteine, numerous control mechanisms have been developed which prevent cross-intrusion of the elements during biosynthesis and insertion. As described in this review, these fidelity steps occur at the genetic, biochemical and physiological level.  相似文献   

17.
Butanol, a four-carbon primary alcohol (C4H10O), is an important industrial chemical and has a good potential to be used as a superior biofuel. Bio-based production of butanol from renewable feedstock is a promising and sustainable alternative to substitute petroleum-based fuels. Here, we report the development of a process for butanol production from glycerol, which is abundantly available as a byproduct of biodiesel production. First, a hyper butanol producing strain of Clostridium pasteurianum was isolated by chemical mutagenesis. The best mutant strain, C. pasteurianum MBEL_GLY2, was able to produce 10.8 g l−1 butanol from 80 g l−1 glycerol as compared to 7.6 g l−1 butanol produced by the parent strain. Next, the process parameters were optimized to maximize butanol production from glycerol. Under the optimized batch condition, the butanol concentration, yield, and productivity of 17.8 g l−1, 0.30 g g−1, and 0.43 g l−1 h−1 could be achieved. Finally, continuous fermentation of C. pasteurianum MBEL_GLY2 with cell recycling was carried out using glycerol as a major carbon source at several different dilution rates. The continuous fermentation was run for 710 h without strain degeneration. The acetone–butanol–ethanol productivity and the butanol productivity of 8.3 and 7.8 g l−1 h−1, respectively, could be achieved at the dilution rate of 0.9 h−1. This study reports continuous production of butanol with reduced byproducts formation from glycerol using C. pasteurianum, and thus could help design a bioprocess for the improved production of butanol.  相似文献   

18.
The paper reports the effects of selenium (Se) supply on growth and antioxidant traits of wheat (Triticum aestivum L. cv Han NO.7086) seedlings exposed to enhanced ultraviolet-B (UV-B) stress. Antioxidant responses of seedlings were different depending on the Se concentration. Compared with the control, the lower amount used (0.5 mg Se kg−1 soil) had no significant effect on biomass accumulation. The treatments with 1.0, 2.0, and 3.0 mg Se kg−1 promoted biomass accumulation of wheat seedlings, and the increased amount in biomass was the most at 1.0 mg Se kg−1 treatment. Se treatments with 1.0, 2.0, and 3.0 mg kg−1 also significantly increased activities of peroxidase (POD) and superoxide dismutase (SOD) and reduced the rate of superoxide radical (O2) production and malondialdehyde (MDA) content of wheat seedlings. In addition, anthocyanins and phenolic compounds content in wheat seedlings evidently increased by the treatments with 1.0 and 2.0 mg Se kg−1. The lower Se treatment had no significant effect on MDA content, although it increased activities of antioxidant enzymes (POD, SOD, and catalase activities) and reduced the rate of O2 production in wheat seedlings. These results suggest that optimal Se supply is favorable for the growth of wheat seedlings and that optimal Se supply can reduce oxidative stress of seedlings under enhanced UV-B radiation.  相似文献   

19.
Guo J  Zhou J  Wang D  Xiang X  Yu H  Tian C  Song Z 《Biodegradation》2006,17(4):341-346
Some experiments were conducted to study some electrochemical factors affecting the bacterial reduction (cleavage) of azo dyes, knowledge of which will be useful in the wastewater treatments of azo dyes. A common mixed culture was used as a test organism and the reductions of Acid Yellow 4, 11, 17 and Acid Yellow BIS were studied. It was found that the azo dyes were reduced at different rates, which could be correlated with the reduction potential of the azo compounds in cyclic voltammetric experiments. Acid Yellow BIS (E r − 616.75 mV) was reduced at the highest rate of 0.0284 mol g dry cell weight−1 h−1, Acid Yellow 11 (E r − 593.25 mV) at 0.0245 mol g dry cell weight−1 h−1 and Acid Yellow 4 (E r − 513 mV) at 0.0178 mol g dry cell weight−1 h−1. At the same time, the decolourization rate of Acid Yellow 17 (E r − 627.5 mV) was 0.0238 mol g dry cell weight−1 h−1, which was affected by the nature of chlorine substituent. Reduction of these azo dyes did not occur under aeration conditions. These studies with a common mixed culture indicate that the reduction of azo dyes may be influenced by the chemical nature of the azo compound. The reduction potential is a preliminary tool to predict the decolourization capacity of oxidative and reductive biocatalysts.  相似文献   

20.
Effects of selenium (Se) on growth and some physiological traits of roots in wheat (Triticum aestivum L. cv Han NO.7086) seedlings exposed to enhanced ultraviolet-B (UV-B) stress are reported. Responses of roots were different depending on the Se concentration. Compared with the control, root weight of wheat seedlings treated with 1.0 and 2.0 mg Se kg−1 soil increased by 39.47% and 16.28%, respectively. The lower amount Se (0.5 mg kg−1) and the higher amount Se treatments (3.0 mg kg−1) did not significantly affect on root weight. Se treatments significantly increased root activity, flavonoids and proline content, and activities of peroxidase and superoxide dimutase in wheat roots exposed to enhanced UV-B. In addition, the treatments with 0.5, 1.0, and 2.0 mg Se kg−1 significantly reduced malondialdehyde content and the rate of superoxide radical (O2) production of roots, whereas the higher amount Se treatment only induced a decrease in the rate of O2 production. The results of this study demonstrated that optimal Se supply promoted roots growth of wheat seedlings, and that optimal Se supply could reduce oxidative stress in wheat roots under enhanced UV-B radiation.  相似文献   

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