March 2006
Volume 5
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March 2006
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Contents
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How Selenium Makes Its Way into Protein as Selenocysteine, the 21st Amino Acid in the Genetic Code
The biosynthesis of Sec, unlike the 20 other amino acids in the genetic code, occurs on its tRNA, and it is the pathway by which the element selenium makes its way into protein. A stem-loop structure in the 3′-untranslated region of selenium-containing (selenoprotein) genes is responsible for recoding UGA for Sec, which circumvents the normal function of UGA as a stop codon in protein synthesis. The stem-loop structure in the selenoprotein genes is recognized by a specific factor, designated SBP2, that forms a complex with Sec tRNA[Ser]Sec and its specific elongation factor and inserts Sec into protein in response to UGA. Although many factors dedicated to the insertion of selenium into protein as Sec have been identified, the biosynthesis of Sec in eukaryotes and the role of phosphoseryl-tRNA[Ser]Sec have not been resolved. Using a comparative genomics approach that searched completely sequenced archaeal genomes for a kinase-like protein with the pattern of occurrence similar to that of components of the Sec insertion machinery, we detected a candidate gene for mammalian phosphoseryl-tRNA[Ser]Sec kinase (pstk). Mouse pstk was cloned, and the gene product (PSTK) was expressed and characterized. PSTK specifically phosphorylated the seryl moiety on seryl-tRNA[Ser]Sec and in addition had a requirement for ATP and Mg++. Proteins with homology to mammalian PSTK occur in the fruit fly, Drosophila, the nematode, Caenorhabditis elegans, and in the archaea, Methanopyrus kandleri and Methanococcus jannaschii. This suggests a conservation of its function across archaea and eukaryotes that synthesize selenoproteins, and the absence of this function in bacteria, plants, and yeast. The fact that PSTK has been highly conserved in evolution suggests that it plays an important role in selenoprotein biosynthesis and/or regulation. The recent identification of the means by which cysteine (Cys) is synthesized on its tRNA in some archaea provides an excellent model of how Sec is biosynthesized on its tRNA. Cys RNACys is aminoacylated by phosphoserine to form phosphoseryl-tRNACys that in turn is converted to cysteyl-tRNACys by an enzyme that replaces the phosphate on serine with an activated form of sulfur (Sauerwald A et al. Science 307: 1969–72, 2005). Since phosphoserine is attached to tRNA[Ser]Sec, it would seem to be the most likely intermediate in Sec biosynthesis wherein selenium would be activated by selenophosphate synthetase, an enzyme previously identified in mammals. This pathway of Sec biosynthesis is shown in Figure 1. Interestingly, Sec tRNA[Ser]Sec has a dual role of serving as the carrier molecule for the biosynthesis of Sec and as the adaptor molecule for decoding UGA for the insertion of Sec into protein. Figure 1. Proposed pathway of Sec biosynthesis on its tRNA in mammalian cells. Serine (Ser, shown in blue as an oblong circle) is attached to tRNA[Ser]Sec (shown in green as a cloverleaf structure) by seryl-tRNA synthetase (SRS) to form seryl-tRNA[Ser]Sec (shown in blue as serine attached to tRNA[Ser]Sec) and is then phosphorylated by phosphoseryl-tRNA kinase (PSTK) to form the intermediate phosphoseryl-tRNA[Ser]Sec (P, shown in red as a circle attached to serine). The phosphate on phosphoseryl-tRNA[Ser]Sec is then replaced by the selenium donor that is likely activated by selenophosphate synthetase (SPS), and the compound is converted to selenocysteyl-tRNA[Ser]Sec (Sec, shown in gold as an oblong circle attached to tRNA[Ser]Sec) by Sec synthase (SST). It should also be noted that selenium is an essential micronutrient in the diet of mammals. Numerous health benefits have been associated with selenium, such as preventing cancer and heart disease, delaying the aging process, and delaying the onset of AIDS in HIV-positive patients, as well as beneficial roles in male reproduction, immune function, and development. Most, if not all, of these health benefits are due to selenoproteins. Thus, it is of paramount importance to determine how this element makes its way into protein. The identification and characterization of PSTK provides a major step in establishing the pathway of Sec biosynthesis.
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1970, a kinase activity that phosphorylated a minor species of seryl-tRNA to
form phosphoseryl-tRNA was observed in rooster liver (Maenpaa PH and Bernfield
MR. Proc Natl Acad Sci U S A 67: 688–94, 1970), and a minor species
of seryl-tRNA that decoded the termination codon UGA was observed in bovine
and chicken livers (Hatfield D and Portugal FH. Proc Natl Acad Sci U S A
67: 1200–06, 1970). The seryl-tRNA in both cases was subsequently identified
by us as selenocysteine (Sec) 