The Experts below are selected from a list of 309 Experts worldwide ranked by ideXlab platform
Otavio Henrique Thiemann - One of the best experts on this subject based on the ideXlab platform.
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Initial steps in Selenocysteine biosynthesis: The interaction between Selenocysteine lyase and selenophosphate synthetase.
International Journal of Biological Macromolecules, 2020Co-Authors: Jessica Fernandes Scortecci, Vitor Hugo Balasco Serrão, Adriano De Freitas Fernandes, Luis G.m. Basso, Raissa F. Gutierrez, Ana Paula Ulian De Araújo, Mario De Oliveira Neto, Otavio Henrique ThiemannAbstract:Abstract The Selenocysteine (Sec) incorporation is a co-translational event taking place at an in-frame UGA-codon and dependent on an organized molecular machinery. Selenium delivery requires mainly two enzymes, the Selenocysteine lyase (CsdB) is essential for Sec recycling and conversion to selenide, further used by the selenophosphate synthetase (SelD), responsible for the conversion of selenide in selenophosphate. Therefore, understanding the catalytic mechanism involved in selenium compounds delivery, such as the interaction between SelD and CsdB (EcCsdB.EcSelD), is fundamental for the further comprehension of the Selenocysteine synthesis pathway and its control. In Escherichia coli, EcCsdB.EcSelD interaction must occur to prevent cell death from the release of the toxic intermediate selenide. Here, we demonstrate and characterize the in vitro EcSelD.EcCsdB interaction by biophysical methods. The EcSelD.EcCsdB interaction occurs with a stoichiometry of 1:1 in presence of Selenocysteine and at a low-nanomolar affinity (~1.8 nM). The data is in agreement with the small angle X-ray scattering model fitted using available structures. Moreover, yeast-2-hybrid assays supported the macromolecular interaction in the cellular environment. This is the first report that demonstrates the interaction between EcCsdB and EcSelD supporting the hypothesis that EcSelD.EcCsdB interaction is necessary to sequester the selenide during the Selenocysteine incorporation pathway in Bacteria.
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The unique tRNA^Sec and its role in Selenocysteine biosynthesis
Amino Acids, 2018Co-Authors: Vitor Hugo Balasco Serrão, Jessica Fernandes Scortecci, Adriano De Freitas Fernandes, Marco Tulio Alves Da Silva, Ivan Rosa Silva, Otavio Henrique ThiemannAbstract:Selenium (Se) is an essential trace element for several organisms and is mostly present in proteins as l -Selenocysteine (Sec or U). Sec is synthesized on its l -seryl–tRNA^Sec to produce Sec–tRNA^Sec molecules by a dedicated Selenocysteine synthesis machinery and incorporated into selenoproteins at specified in-frame UGA codons. UGA–Sec insertion is signaled by an mRNA stem-loop structure called the Selenocysteine Insertion Sequence (SECIS). tRNA^Sec transcription regulation and folding have been described showing its importance to Sec biosynthesis. Here, we discuss structural aspects of Sec–tRNA^Sec and its role in Sec biosynthesis as well as Sec incorporation into selenoproteins. Defects in the Sec biosynthesis or incorporation pathway have been correlated with pathological conditions.
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the unique trna sec and its role in Selenocysteine biosynthesis
Amino Acids, 2018Co-Authors: Jessica Fernandes Scortecci, Vitor Hugo Balasco Serrão, Adriano De Freitas Fernandes, Otavio Henrique Thiemann, Ivan Rosa E Silva, Marco Tulio Alves Da SilvaAbstract:: Selenium (Se) is an essential trace element for several organisms and is mostly present in proteins as L-Selenocysteine (Sec or U). Sec is synthesized on its L-seryl-tRNASec to produce Sec-tRNASec molecules by a dedicated Selenocysteine synthesis machinery and incorporated into selenoproteins at specified in-frame UGA codons. UGA-Sec insertion is signaled by an mRNA stem-loop structure called the Selenocysteine Insertion Sequence (SECIS). tRNASec transcription regulation and folding have been described showing its importance to Sec biosynthesis. Here, we discuss structural aspects of Sec-tRNASec and its role in Sec biosynthesis as well as Sec incorporation into selenoproteins. Defects in the Sec biosynthesis or incorporation pathway have been correlated with pathological conditions.
Vitor Hugo Balasco Serrão - One of the best experts on this subject based on the ideXlab platform.
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Initial steps in Selenocysteine biosynthesis: The interaction between Selenocysteine lyase and selenophosphate synthetase.
International Journal of Biological Macromolecules, 2020Co-Authors: Jessica Fernandes Scortecci, Vitor Hugo Balasco Serrão, Adriano De Freitas Fernandes, Luis G.m. Basso, Raissa F. Gutierrez, Ana Paula Ulian De Araújo, Mario De Oliveira Neto, Otavio Henrique ThiemannAbstract:Abstract The Selenocysteine (Sec) incorporation is a co-translational event taking place at an in-frame UGA-codon and dependent on an organized molecular machinery. Selenium delivery requires mainly two enzymes, the Selenocysteine lyase (CsdB) is essential for Sec recycling and conversion to selenide, further used by the selenophosphate synthetase (SelD), responsible for the conversion of selenide in selenophosphate. Therefore, understanding the catalytic mechanism involved in selenium compounds delivery, such as the interaction between SelD and CsdB (EcCsdB.EcSelD), is fundamental for the further comprehension of the Selenocysteine synthesis pathway and its control. In Escherichia coli, EcCsdB.EcSelD interaction must occur to prevent cell death from the release of the toxic intermediate selenide. Here, we demonstrate and characterize the in vitro EcSelD.EcCsdB interaction by biophysical methods. The EcSelD.EcCsdB interaction occurs with a stoichiometry of 1:1 in presence of Selenocysteine and at a low-nanomolar affinity (~1.8 nM). The data is in agreement with the small angle X-ray scattering model fitted using available structures. Moreover, yeast-2-hybrid assays supported the macromolecular interaction in the cellular environment. This is the first report that demonstrates the interaction between EcCsdB and EcSelD supporting the hypothesis that EcSelD.EcCsdB interaction is necessary to sequester the selenide during the Selenocysteine incorporation pathway in Bacteria.
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The unique tRNA^Sec and its role in Selenocysteine biosynthesis
Amino Acids, 2018Co-Authors: Vitor Hugo Balasco Serrão, Jessica Fernandes Scortecci, Adriano De Freitas Fernandes, Marco Tulio Alves Da Silva, Ivan Rosa Silva, Otavio Henrique ThiemannAbstract:Selenium (Se) is an essential trace element for several organisms and is mostly present in proteins as l -Selenocysteine (Sec or U). Sec is synthesized on its l -seryl–tRNA^Sec to produce Sec–tRNA^Sec molecules by a dedicated Selenocysteine synthesis machinery and incorporated into selenoproteins at specified in-frame UGA codons. UGA–Sec insertion is signaled by an mRNA stem-loop structure called the Selenocysteine Insertion Sequence (SECIS). tRNA^Sec transcription regulation and folding have been described showing its importance to Sec biosynthesis. Here, we discuss structural aspects of Sec–tRNA^Sec and its role in Sec biosynthesis as well as Sec incorporation into selenoproteins. Defects in the Sec biosynthesis or incorporation pathway have been correlated with pathological conditions.
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the unique trna sec and its role in Selenocysteine biosynthesis
Amino Acids, 2018Co-Authors: Jessica Fernandes Scortecci, Vitor Hugo Balasco Serrão, Adriano De Freitas Fernandes, Otavio Henrique Thiemann, Ivan Rosa E Silva, Marco Tulio Alves Da SilvaAbstract:: Selenium (Se) is an essential trace element for several organisms and is mostly present in proteins as L-Selenocysteine (Sec or U). Sec is synthesized on its L-seryl-tRNASec to produce Sec-tRNASec molecules by a dedicated Selenocysteine synthesis machinery and incorporated into selenoproteins at specified in-frame UGA codons. UGA-Sec insertion is signaled by an mRNA stem-loop structure called the Selenocysteine Insertion Sequence (SECIS). tRNASec transcription regulation and folding have been described showing its importance to Sec biosynthesis. Here, we discuss structural aspects of Sec-tRNASec and its role in Sec biosynthesis as well as Sec incorporation into selenoproteins. Defects in the Sec biosynthesis or incorporation pathway have been correlated with pathological conditions.
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formation of a ternary complex for Selenocysteine biosynthesis in bacteria
Journal of Biological Chemistry, 2015Co-Authors: Ivan Rosa E Silva, Vitor Hugo Balasco Serrão, Livia Regina Manzine, Livia Maria Faim, Marco Tulio Alves Da Silva, Raphaela Makki, Daniel M Saidemberg, Marinonio Lopes Cornelio, Mario Sergio PalmaAbstract:Abstract The synthesis of Selenocysteine-containing proteins (selenoproteins) involves the interaction of Selenocysteine synthase (SelA), tRNA (tRNASec), selenophosphate synthetase (SelD, SPS), a specific elongation factor (SelB), and a specific mRNA sequence known as Selenocysteine insertion sequence (SECIS). Because selenium compounds are highly toxic in the cellular environment, the association of selenium with proteins throughout its metabolism is essential for cell survival. In this study, we demonstrate the interaction of SPS with the SelA-tRNASec complex, resulting in a 1.3-MDa ternary complex of 27.0 ± 0.5 nm in diameter and 4.02 ± 0.05 nm in height. To assemble the ternary complex, SPS undergoes a conformational change. We demonstrated that the glycine-rich N-terminal region of SPS is crucial for the SelA-tRNASec-SPS interaction and selenoprotein biosynthesis, as revealed by functional complementation experiments. Taken together, our results provide new insights into selenoprotein biosynthesis, demonstrating for the first time the formation of the functional ternary SelA-tRNASec-SPS complex. We propose that this complex is necessary for proper Selenocysteine synthesis and may be involved in avoiding the cellular toxicity of selenium compounds.
M J Berry - One of the best experts on this subject based on the ideXlab platform.
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Selenocysteine codons decrease polysome association on endogenous selenoprotein mRNAs
2001Co-Authors: G W Martin, M J BerryAbstract:BACKGROUND: Selenocysteine incorporation has been reported to be inefficient in all systems studied, including Escherichia coli, baculovirus-insect cell systems, rabbit reticulocyte in vitro translation systems, transiently transfected mammalian cells, and intact animals. Nonetheless, full-length selenoproteins containing up to 17 Selenocysteine residues are produced in animals, indicating that the efficiency observed in manipulated systems might not accurately reflect the true efficiency of this process in nature. RESULTS: To begin to address this apparent discrepancy, we have examined the polysome profiles of endogenously expressed selenoprotein mRNAs in a mammalian cell line, and compared them with nonselenoprotein mRNAs. We report that three selenoprotein mRNAs, type 1 deiodinase, glutathione peroxidase and selenoprotein P, are under-loaded with ribosomes, based on their predicted open reading frame sizes. The average numbers of ribosomes per mRNA correspond to the sizes predicted by termination at the UGA Selenocysteine codons. Appropriate loading on the type 1 deiodinase mRNA is seen following substitution of a cysteine codon for the Selenocysteine codon, indicating that the UGA codon confers a translational penalty on the mRNA. Surprisingly, ribosomal loading is also increased by the expression of eukaryotic release factors eRF1 and eRF3. CONCLUSIONS: These results suggest that the presence of a Selenocysteine codon confers a translational penalty on selenoprotein mRNAs, and that increased levels of release factors may alter the kinetics of termination.
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Interplay between termination and translation machinery in eukaryotic selenoprotein synthesis.
2001Co-Authors: Elisabeth Grundner-Culemann, G W Martin, R Tujebajeva, John W. Harney, M J BerryAbstract:Termination of translation in eukaryotes is catalyzed by eRF1, the stop codon recognition factor, and eRF3, an eRF1 and ribosome-dependent GTPase. In selenoprotein mRNAs, UGA codons, which typically specify termination, serve an alternate function as sense codons. Selenocysteine incorporation involves a unique tRNA with an anticodon complementary to UGA, a unique elongation factor specific for this tRNA, and cis-acting secondary structures in selenoprotein mRNAs, termed SECIS elements. To gain insight into the interplay between the Selenocysteine insertion and termination machinery, we investigated the effects of overexpressing eRF1 and eRF3, and of altering UGA codon context, on the efficiency of selenoprotein synthesis in a transient transfection system. Overexpression of eRF1 does not increase termination at naturally occurring Selenocysteine codons. Surprisingly, Selenocysteine incorporation is enhanced. Overexpression of eRF3 did not affect incorporation efficiency. Coexpression of both factors reproduced the effects with eRF1 alone. Finally, we show that the nucleotide context immediately upstream and downstream of the UGA codon significantly affects termination to incorporation ratios and the response to eRF overexpression. Implications for the mechanisms of Selenocysteine incorporation and termination are discussed.
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Selenoprotein P expression, purification, and immunochemical characterization
2000Co-Authors: Rosa M. Tiyebajeva, John W. Harney, M J BerryAbstract:Most selenoproteins contain a single Selenocysteine residue per polypeptide chain, encoded by an in-frame UGA codon. Selenoprotein P is unique in that its mRNA encodes 10-12 Selenocysteine residues, depending on species. In addition to the high number of Selenocysteines, the protein is cysteine- and histidine-rich. The function of selenoprotein P has remained elusive, in part due to the inability to express the recombinant protein. This has been attributed to presumed inefficient translation through the Selenocysteine/stop codons. Herein, we report for the first time the expression of recombinant rat selenoprotein P in a transiently transfected human epithelial kidney cell line, as well as the endogenously expressed protein from HepG2 and Chinese hamster ovary cells. The majority of the expressed protein migrates with the predicted 57-kDa size of full-length glycosylated selenoprotein P. Based on the histidine-rich nature of selenoprotein P, we have purified the recombinant and endogenously expressed proteins using nickel-agarose affinity chromatography. We show that the recombinant rat and endogenous human proteins react in Western blotting and immunoprecipitation assays with commercial anti-histidine antibodies. The ability to express, purify, and immunochemically detect the recombinant protein provides a foundation for investigating the functions and efficiency of expression of this intriguing protein.
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SECIS-SBP2 interactions dictate Selenocysteine incorporation efficiency and selenoprotein hierarchy
2000Co-Authors: S. C. Low, Elisabeth Grundner-Culemann, John W. Harney, M J BerryAbstract:Selenocysteine incorporation at UGA codons requires cis-acting mRNA secondary structures and several specialized trans-acting factors. The latter include a Selenocysteine-specific tRNA, an elongation factor specific for this tRNA and a SECIS-binding protein, SBP2, which recruits the elongation factor to the selenoprotein mRNA. Overexpression of selenoprotein mRNAs in transfected cells results in inefficient Selenocysteine incorporation due to limitation of one or more of these factors. Using a transfection-based competition assay employing overexpression of selenoprotein mRNAs to compete for selenoprotein synthesis, we investigated the ability of the trans-acting factors to overcome competition and restore Selenocysteine incorporation. We report that co-expression of SBP2 overcomes the limitation produced by selenoprotein mRNA overexpression, whereas selenocysteyl-tRNA and the Selenocysteine-specific elongation factor do not. Competition studies indicate that once bound to SECIS elements, SBP2 does not readily exchange between them. Finally, we show that SBP2 preferentially stimulates incorporation directed by the seleno protein P and phospholipid hydroperoxide glutathione peroxidase SECIS elements over those of other selenoproteins. The mechanistic implications of these findings for the hierarchy of selenoprotein synthesis and nonsense-mediated decay are discussed.
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Expression and characterization of nonmammalian selenoprotein P in the zebrafish, Danio rerio
2000Co-Authors: R Tujebajeva, David G. Ransom, John W. Harney, M J BerryAbstract:BACKGROUND: Selenoprotein P is a protein of considerable intrigue, due to its unusual composition and requirements for its biosynthesis. Whereas most selenoproteins contain a single Selenocysteine residue, the human, bovine and rodent selenoprotein P genes encode proteins containing 10-12 Selenocysteines. Selenoprotein P genes have, to date, only been reported in mammals, and the function of the protein remains elusive. RESULTS: Herein, we report the identification and characterization of nonmammalian selenoprotein P in the zebrafish Danio rerio. Sequencing of the cDNA revealed the presence of 17 Selenocysteine codons, the highest number reported in any protein. Two histidine-rich regions present in the mammalian selenoprotein P sequences are conserved in the zebrafish protein, and two SECIS elements are present in the 3' untranslated region. Whole-mount in situ hybridization of zebrafish embryos revealed high levels of expression of selenoprotein P mRNA in fertilized eggs and in the yolk sac of developing embryos. Transient transfection of the cDNA in mammalian cells resulted in efficient expression of the full-length secreted selenoprotein. A single N-glycosylation site is predicted, and shown to be utilized. CONCLUSIONS: Discovery of selenoprotein P in the zebrafish opens a previously unavailable avenue for genetic investigation of the functions of this unusual protein.
August Böck - One of the best experts on this subject based on the ideXlab platform.
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Encyclopedia of Inorganic and Bioinorganic Chemistry - Selenium Proteins Containing Selenocysteine
Encyclopedia of Inorganic and Bioinorganic Chemistry, 2011Co-Authors: August BöckAbstract:Selenium is incorporated in the form of the nonstandard amino acid Selenocysteine into selected proteins of organisms belonging to all three lines of descent. The majority of these proteins, which contain Selenocysteine in the active site, catalyze oxidation–reduction reactions and are involved in numerous biochemical and regulatory processes, which are indispensable for the organism. Because of the higher chemical reactivity of Selenocysteine in comparison to cysteine, selenoenzymes display a greatly increased rate of catalysis. Apart from its biochemical function, Selenocysteine is also unique since its incorporation is DNA-encoded and its cotranslational insertion follows a route independent in many features from the path of insertion of the 20 classical amino acids. Selenocysteine, therefore, can be considered the 21st amino acid. Keywords: selenoproteins; selenoenzymes; nonstandard amino acid; tRNASec; translation factor SelB; Selenocysteine synthesis; selenophosphate
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Selenomethionine and Selenocysteine Double Labeling Strategy for Crystallographic Phasing
Structure, 2003Co-Authors: Marie-paule Strub, August Böck, Jean-frédéric Sanchez, Jean Marc Strub, André Aumelas, Christian DumasAbstract:Abstract A protocol for the quantitative incorporation of both selenomethionine and Selenocysteine into recombinant proteins overexpressed in Escherichia coli is described. This methodology is based on the use of a suitable cysteine auxotrophic strain and a minimal medium supplemented with selenium-labeled methionine and cysteine. The proteins chosen for these studies are the cathelin-like motif of protegrin-3 and a nucleoside-diphosphate kinase. Analysis of the purified proteins by electrospray mass spectrometry and X-ray crystallography revealed that both cysteine and methionine residues were isomorphously replaced by Selenocysteine and selenomethionine. Moreover, Selenocysteines allowed the formation of unstrained and stable diselenide bridges in place of the canonical disulfide bonds. In addition, we showed that NDP kinase contains a Selenocysteine adduct on Cys122. This novel selenium double-labeling method is proposed as a general approach to increase the efficiency of the MAD technique used for phase determination in protein crystallography.
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Selenocysteine inserting tRNAs: an overview
Fems Microbiology Reviews, 1999Co-Authors: Stephane Commans, August BöckAbstract:One of the recent discoveries in protein biosynthesis was the finding that Selenocysteine, the 21st amino acid, is cotranslationally inserted into polypeptides under the direction of a UGA codon assisted by a specific structural signal in the mRNA. The key to Selenocysteine biosynthesis and insertion is a special tRNA species, tRNASec. The formation of Selenocysteine from serine represents an interesting tRNA-mediated amino acid transformation. tRNASec (or the gene encoding it) has been found over all three domains of life. It displays a number of unique features that designate it a Selenocysteine-inserting tRNA and differentiate it from canonical elongator tRNAs. Although there are still some uncertainties concerning the precise secondary and tertiary structures of eukaryal tRNASec, the major identity determinant for Selenocysteine biosynthesis and insertion appears to be the 13 bp long extended acceptor arm. In addition the core of the 3D structure of these tRNAs is different from that of class II tRNAs like tRNASer. The biological implications of these structural differences still remain to be fully understood.
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A FAMILY OF S-METHYLMETHIONINE-DEPENDENT THIOL/SELENOL METHYLTRANSFERASES : ROLE IN SELENIUM TOLERANCE AND EVOLUTIONARY RELATION
Journal of Biological Chemistry, 1999Co-Authors: Bernhard Neuhierl, Martin Thanbichler, Friedrich Lottspeich, August BöckAbstract:Abstract Several plant species can tolerate high concentrations of selenium in the environment, and they accumulate organoselenium compounds. One of these compounds is Se-methylSelenocysteine, synthesized by a number of species from the genus Astragalus (Fabaceae), like A. bisulcatus. An enzyme has been previously isolated from this organism that catalyzes methyl transfer fromS-adenosylmethionine to Selenocysteine. To elucidate the role of the enzyme in selenium tolerance, the cDNA coding for Selenocysteine methyltransferase from A. bisulcatus was cloned and sequenced. Data base searches revealed the existence of several apparent homologs of hitherto unassigned function. The gene for one of them, yagD from Escherichia coli, was cloned, and the protein was overproduced and purified. A functional analysis showed that the YagD protein catalyzes methylation of homocysteine, selenohomocysteine, and Selenocysteine withS-adenosylmethionine and S-methylmethionine as methyl group donors. S-Methylmethionine was now shown to be also the physiological methyl group donor for the A. bisulcatus Selenocysteine methyltransferase. A model system was set up in E. coli which demonstrated that expression of the plant and, although to a much lesser degree, of the bacterial methyltransferase gene increases selenium tolerance and strongly reduces unspecific selenium incorporation into proteins, provided thatS-methylmethionine is present in the medium. It is postulated that the Selenocysteine methyltransferase under selective pressure developed from anS-methylmethionine-dependent thiol/selenol methyltransferase.
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The Selenocysteine-Inserting tRNA Species: Structure and Function
tRNA, 1995Co-Authors: Christian Baron, August BöckAbstract:The occurrence of the amino acid Selenocysteine in proteins was first demonstrated for protein A of glycine reductase from Clostridium sticklandii in 1976, and questions were immediately raised on its mechanism of incorporation. At that time, the universality of the 20 proteinogenic amino acids was taken for granted, as was the fact that the 64 codons of the "universal" genetic code are assigned either to code for one of these 20 amino acids or to serve as termination signals. Thus, it seemed unlikely that Selenocysteine would be considered as a classical amino acid. In principle, the definition of such a 21st amino acid would require (i) that its incorporation proceeds via a cotranslational mechanism, (ii) that it is directed by a specific codon, and (iii) that a specific tRNA mediates its transport to the ribosome. This chapter illustrates that Selenocysteine fulfills these criteria. It first describes the unusual structural properties of tRNASec, and then discusses the unique pathway of Selenocysteine insertion that has been worked out for Escherichia coli, which has finally led to the proposal of a model for the co-translational incorporation process at the ribosome. The chapter further compares the pathway established in E. coli with the current knowledge on the mammalian system. Finally, it addresses the interesting question of the evolution of the pathway for the incorporation of Selenocysteine that differs from that of the 20 standard amino acids in many respects.
Vadim N Gladyshev - One of the best experts on this subject based on the ideXlab platform.
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Selenocysteine Biosynthesis and the Replacement of Selenocysteine with Cysteine in the Pathway
Selenium, 2011Co-Authors: Xue-ming Xu, Vadim N Gladyshev, Anton A. Turanov, Bradley A. Carlson, Dolph L HatfieldAbstract:The biosynthetic pathway of Selenocysteine (Sec), the 21st amino acid in the genetic code, has been established in eukaryotes and archaea using comparative genomic and experimental approaches. In addition, cysteine (Cys) was found to arise in place of Selenocysteine in thioredoxin reductase (TR) in NIH 3T3 cells and in mice. An analysis of the Selenocysteine biosynthetic pathway demonstrated that replacement of selenide with sulfide in generating the active cysteine donor, thiophosphate, resulted in cysteine being donated to the acceptor molecule, which is likely dehydroalanyl-tRNA[Ser]Sec, yielding Cys-tRNA[Ser]Sec. The identification of the pathways for biosynthesis of Selenocysteine and cysteine in mammals is discussed in this chapter.
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a 4 Selenocysteine 2 Selenocysteine insertion sequence secis element methionine sulfoxide reductase from metridium senile reveals a non catalytic function of Selenocysteines
Journal of Biological Chemistry, 2011Co-Authors: Alexey V Lobanov, Stefano M Marino, Alaattin Kaya, Javier Seravalli, Dolph L Hatfield, Vadim N GladyshevAbstract:Abstract Selenocysteine (Sec) residues occur in thiol oxidoreductase families, and functionally characterized selenoenzymes typically have a single Sec residue used directly for redox catalysis. However, how new Sec residues evolve and whether non-catalytic Sec residues exist in proteins is not known. Here, we computationally identified several genes with multiple Sec insertion sequence (SECIS) elements, one of which was a methionine-R-sulfoxide reductase (MsrB) homolog from Metridium senile that has four in-frame UGA codons and two nearly identical SECIS elements. One of the UGA codons corresponded to the conserved catalytic Sec or Cys in MsrBs, whereas the three other UGA codons evolved recently and had no homologs with Sec or Cys in these positions. Metabolic 75Se labeling showed that all four in-frame UGA codons supported Sec insertion and that both SECIS elements were functional and collaborated in Sec insertion at each UGA codon. Interestingly, recombinant M. senile MsrB bound iron, and further analyses suggested the possibility of binding an iron-sulfur cluster by the protein. These data show that Sec residues may appear transiently in genes containing SECIS elements and be adapted for non-catalytic functions.
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redox regulation of cell signaling by Selenocysteine in mammalian thioredoxin reductases
Journal of Biological Chemistry, 1999Co-Authors: Yalin Wu, Dolph L Hatfield, Francesca Zappacosta, Kuan Teh Jeang, Vadim N GladyshevAbstract:Abstract The intracellular generation of reactive oxygen species, together with the thioredoxin and glutathione systems, is thought to participate in redox signaling in mammalian cells. The activity of thioredoxin is dependent on the redox status of thioredoxin reductase (TR), the activity of which in turn is dependent on a Selenocysteine residue. Two mammalian TR isozymes (TR2 and TR3), in addition to that previously characterized (TR1), have now been identified in humans and mice. All three TR isozymes contain a Selenocysteine residue that is located in the penultimate position at the carboxyl terminus and which is encoded by a UGA codon. The generation of reactive oxygen species in a human carcinoma cell line was shown to result in both the oxidation of the Selenocysteine in TR1 and a subsequent increase in the expression of this enzyme. These observations identify the carboxyl-terminal Selenocysteine of TR1 as a cellular redox sensor and support an essential role for mammalian TR isozymes in redox-regulated cell signaling.
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Selenocysteine containing proteins in mammals
Journal of Biomedical Science, 1999Co-Authors: Vadim N Gladyshev, Dolph L HatfieldAbstract:Since the recent discovery of Selenocysteine as the 21st amino acid in protein, the field of selenium biology has rapidly expanded. Twelve mammalian selenoproteins have been characterized to date and each contains Selenocysteine that is incorporated in response to specific UGA code words. These selenoproteins have different cellular functions, but in those selenoproteins for which the function is known, Selenocysteine is located at the active center. The presence of Selenocysteine at critical sites in naturally occurring selenoproteins provides an explanation for the important role of selenium in human health and development. This review describes known mammalian selenoproteins and discusses recent developments and future directions in the selenium field.
