In vitro synthesis of glutathione peroxidase from selenite Translational incorporation of selenocysteine

Wayne C. Hawkes, Al L. Tappel

Research output: Contribution to journal › Article

28 Citations (Scopus)

Abstract

The synthesis of glutathione peroxidase from [75Se]selenite was studied in slices and cell-free extracts from rat liver. The incorporation of [75Se]selenocysteine at the active site was detected by carboxymethylation and hydrolysis of partially purified glutathione peroxidase (glutathione:hydrogen peroxide oxidoreductase, EC 1.11.1.9) in the presence of [3H]selenocysteine and subsequent amino acid analysis. The synthesis of glutathione peroxidase in slices was inhibited by cycloheximide or puromycin and 75Se was incorporated from [75Se]selenite into free selenocysteine and selenocysteyl tRNA. Increasing concentrations of selenocystine caused a progressive dilution of the 75Se and a corresponding decrease in glutathione peroxidase labeling. In cell-free systems, [75Se]selenocysteyl tRNA was the best substrate for glutathione peroxidase synthesis. These results indicate the existence in rat liver of the de novo synthesis of free selenocysteine and a translational pathway of selenocysteine incorporation into glutathione peroxidase.

Original languageEnglish (US)
Pages (from-to)225-234
Number of pages10
JournalBBA - Gene Structure and Expression
Volume739
Issue number2
DOIs
StatePublished - Mar 10 1983

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Selenocysteine
Selenious Acid
Glutathione Peroxidase
Glutathione
Liver
Rats
Puromycin
Cell-Free System
Cycloheximide
Transfer RNA
Cell Extracts
Labeling
Dilution
In Vitro Techniques
Hydrolysis
Catalytic Domain
Amino Acids
Substrates

Keywords

  • Glutathione peroxidase
  • Selenocysteine, Selenite, Protein synthesis
  • tRNA

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Genetics
  • Structural Biology
  • Medicine(all)

Cite this

In vitro synthesis of glutathione peroxidase from selenite Translational incorporation of selenocysteine. / Hawkes, Wayne C.; Tappel, Al L.

In: BBA - Gene Structure and Expression, Vol. 739, No. 2, 10.03.1983, p. 225-234.

Research output: Contribution to journal › Article

Hawkes, WC & Tappel, AL 1983, 'In vitro synthesis of glutathione peroxidase from selenite Translational incorporation of selenocysteine', BBA - Gene Structure and Expression, vol. 739, no. 2, pp. 225-234. https://doi.org/10.1016/0167-4781(83)90033-7
Hawkes WC, Tappel AL. In vitro synthesis of glutathione peroxidase from selenite Translational incorporation of selenocysteine. BBA - Gene Structure and Expression. 1983 Mar 10;739(2):225-234. https://doi.org/10.1016/0167-4781(83)90033-7
Hawkes, Wayne C. ; Tappel, Al L. / In vitro synthesis of glutathione peroxidase from selenite Translational incorporation of selenocysteine. In: BBA - Gene Structure and Expression. 1983 ; Vol. 739, No. 2. pp. 225-234.
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N2 - The synthesis of glutathione peroxidase from [75Se]selenite was studied in slices and cell-free extracts from rat liver. The incorporation of [75Se]selenocysteine at the active site was detected by carboxymethylation and hydrolysis of partially purified glutathione peroxidase (glutathione:hydrogen peroxide oxidoreductase, EC 1.11.1.9) in the presence of [3H]selenocysteine and subsequent amino acid analysis. The synthesis of glutathione peroxidase in slices was inhibited by cycloheximide or puromycin and 75Se was incorporated from [75Se]selenite into free selenocysteine and selenocysteyl tRNA. Increasing concentrations of selenocystine caused a progressive dilution of the 75Se and a corresponding decrease in glutathione peroxidase labeling. In cell-free systems, [75Se]selenocysteyl tRNA was the best substrate for glutathione peroxidase synthesis. These results indicate the existence in rat liver of the de novo synthesis of free selenocysteine and a translational pathway of selenocysteine incorporation into glutathione peroxidase.

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