Towards understanding selenocysteine incorporation into bacterial proteins
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Niels Fischer
Abstract
In bacteria, UGA stop codons can be recoded to direct the incorporation of selenocysteine into proteins on the ribosome. Recoding requires a selenocysteine incorporation sequence (SECIS) downstream of the UGA codon, a specialized translation factor SelB, and the non-canonical Sec-tRNASec, which is formed from Ser-tRNASec by selenocysteine synthase, SelA, using selenophosphate as selenium donor. Here we describe a rapid-kinetics approach to study the mechanism of selenocysteine insertion into proteins on the ribosome. Labeling of SelB, Sec-tRNASec and other components of the translational machinery allows direct observation of the formation or dissociation of complexes by monitoring changes in the fluorescence of single dyes or fluorescence resonance energy transfer between two fluorophores. Furthermore, the structure of SelA was studied by electron cryomicroscopy (cryo-EM). We report that intact SelA from the thermophilic bacterium Moorella thermoacetica (mthSelA) can be vitrified for cryo-EM using a controlled-environment vitrification system. Two-dimensional image analysis of vitrified mthSelA images shows that SelA can adopt the wide range of orientations required for high-resolution structure determination by cryo-EM. The results indicate that mthSelA forms a homodecamer that has a ring-like structure with five bilobed wings, similar to the structure of the E. coli complex determined previously.
©2007 by Walter de Gruyter Berlin New York
Articles in the same Issue
- Selenoproteins – biochemistry and clinical relevance
- Selenium in mammalian spermiogenesis
- Selenium in chemistry and biochemistry in comparison to sulfur
- Molecular biology of glutathione peroxidase 4: from genomic structure to developmental expression and neural function
- Physiological role of phospholipid hydroperoxide glutathione peroxidase in mammals
- Activation of the glutathione peroxidase 2 (GPx2) promoter by β-catenin
- Effect of age on sexually dimorphic selenoprotein expression in mice
- Post-translational processing of selenoprotein P: implications of glycosylation for its utilisation by target cells
- Selenoproteins of the thyroid gland: expression, localization and possible function of glutathione peroxidase 3
- Towards understanding selenocysteine incorporation into bacterial proteins
- Glutathione- and thioredoxin-related enzymes are modulated by sulfur-containing chemopreventive agents
- B- and T-cell-specific inactivation of thioredoxin reductase 2 does not impair lymphocyte development and maintenance
- Effect of selenium on thioredoxin reductase activity in Txnrd1 or Txnrd2 hemizygous mice
- Influence of pH and flanking serine on the redox potential of S-S and S-Se bridges of Cys-Cys and Cys-Sec peptides
- An essential role for Pin1 in Xenopus laevis embryonic development revealed by specific inhibitors
- Glucocorticoid receptor-mediated expression of kallikrein 10 in human breast cancer cell lines
Articles in the same Issue
- Selenoproteins – biochemistry and clinical relevance
- Selenium in mammalian spermiogenesis
- Selenium in chemistry and biochemistry in comparison to sulfur
- Molecular biology of glutathione peroxidase 4: from genomic structure to developmental expression and neural function
- Physiological role of phospholipid hydroperoxide glutathione peroxidase in mammals
- Activation of the glutathione peroxidase 2 (GPx2) promoter by β-catenin
- Effect of age on sexually dimorphic selenoprotein expression in mice
- Post-translational processing of selenoprotein P: implications of glycosylation for its utilisation by target cells
- Selenoproteins of the thyroid gland: expression, localization and possible function of glutathione peroxidase 3
- Towards understanding selenocysteine incorporation into bacterial proteins
- Glutathione- and thioredoxin-related enzymes are modulated by sulfur-containing chemopreventive agents
- B- and T-cell-specific inactivation of thioredoxin reductase 2 does not impair lymphocyte development and maintenance
- Effect of selenium on thioredoxin reductase activity in Txnrd1 or Txnrd2 hemizygous mice
- Influence of pH and flanking serine on the redox potential of S-S and S-Se bridges of Cys-Cys and Cys-Sec peptides
- An essential role for Pin1 in Xenopus laevis embryonic development revealed by specific inhibitors
- Glucocorticoid receptor-mediated expression of kallikrein 10 in human breast cancer cell lines
