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Identification of a selenocysteyl-tRNA(Ser) in mammalian cells that recognizes the nonsense codon, UGA
The presence of a unique opal suppressor seryl-tRNA in higher vertebrates which is converted to phosphoseryl-tRNA has been known for several years, but its function has been uncertain (see Hatfield, D. (1985) Trends Biochem. Sci. 10, 201-204 for review). In the present study, we demonstrate that thi...
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Published in: | The Journal of biological chemistry 1989-06, Vol.264 (17), p.9724 |
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Main Authors: | , , , , |
Format: | Article |
Language: | English |
Subjects: | |
Online Access: | Get full text |
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Summary: | The presence of a unique opal suppressor seryl-tRNA in higher vertebrates which is converted to phosphoseryl-tRNA has been
known for several years, but its function has been uncertain (see Hatfield, D. (1985) Trends Biochem. Sci. 10, 201-204 for
review). In the present study, we demonstrate that this tRNA species also occurs in vivo as selenocysteyl-tRNA(Ser) suggesting
that it functions both as a carrier molecule upon which selenocysteine is synthesized and as a direct selenocysteine donor
to a growing polypeptide chain in response to specific UGA codons. [75Se]Seleno[3H]cysteyl-tRNA(Ser) formed by administering
75Se and [3H]serine to rat mammary tumor cells (TMT-081-MS) in culture was isolated from the cell extract. The amino acid
attached to the tRNA was identified as selenocysteine following its deacylation and reaction with iodoacetate and 3-bromopropionate.
The resulting alkyl derivatives co-chromatographed on an amino acid analyzer with authentic carboxymethylselenocysteine and
carboxyethylselenocysteine. Seryl-tRNA(Ser) and phosphoseryl-tRNA(Ser) (Hatfield, D., Diamond, A., and Dudock, B. (1982) Proc.
Natl. Acad. Sci. U. S. A. 79, 6215-6219), which co-migrate on a reverse phase chromatographic column with selenocysteyl-tRNA(Ser),
were also identified in extracts of TMT-018-MS cells. Hence, we propose that a metabolic pathway for selenocysteine synthesis
in mammalian cells is the conversion of seryl-tRNA(Ser) via phosphoseryl-tRNA(Ser) to selenocysteyl-tRNA(Ser). In a ribosomal
binding assay selenocysteyl-tRNA(Ser) recognizes UGA but not any of the serine codons. Selenocysteyl-tRNA(Ser) is deacylated
more readily than seryl-tRNA(Ser) (i.e. 58% deacylation during 15 min at pH 8.0 and 37 degrees C as compared to 41%). |
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ISSN: | 0021-9258 1083-351X |