A synthetic tRNA for EF-Tu mediated selenocysteine incorporation in vivo and in vitro.
FEBS Lett
; 589(17): 2194-9, 2015 Aug 04.
Article
em En
| MEDLINE
| ID: mdl-26160755
ABSTRACT
Incorporation of selenocysteine (Sec) in bacteria requires a UGA codon that is reassigned to Sec by the Sec-specific elongation factor SelB and a conserved mRNA motif (SECIS element). These requirements severely restrict the engineering of selenoproteins. Earlier, a synthetic tRNASec was reported that allowed canonical Sec incorporation by EF-Tu; however, serine misincorporation limited its scope. We report a superior tRNASec variant (tRNAUTuX) that facilitates EF-Tu dependent stoichiometric Sec insertion in response to UAG both in vivo in Escherichia coli and in vitro in a cellfree protein synthesis system. We also demonstrate recoding of several sense codons in a SelB supplemented cell-free system. These advances in Sec incorporation will aid rational design and directed evolution of selenoproteins.
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Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Biossíntese de Proteínas
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RNA de Transferência Aminoácido-Específico
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Fator Tu de Elongação de Peptídeos
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Selenocisteína
Idioma:
En
Ano de publicação:
2015
Tipo de documento:
Article