A Facile Method for Producing Selenocysteine-Containing Proteins

Takahito Mukai, Anastasia Sevostyanova, Tateki Suzuki, Xian Fu, Dieter Söll

Research output: Contribution to journalArticle

12 Citations (Scopus)

Abstract

Selenocysteine (Sec, U) confers new chemical properties on proteins. Improved tools are thus required that enable Sec insertion into any desired position of a protein. We report a facile method for synthesizing selenoproteins with multiple Sec residues by expanding the genetic code of Escherichia coli. We recently discovered allo-tRNAs, tRNA species with unusual structure, that are as efficient serine acceptors as E. coli tRNASer. Ser-allo-tRNA was converted into Sec-allo-tRNA by Aeromonas salmonicida selenocysteine synthase (SelA). Sec-allo-tRNA variants were able to read through five UAG codons in the fdhF mRNA coding for E. coli formate dehydrogenase H, and produced active FDHH with five Sec residues in E. coli. Engineering of the E. coli selenium metabolism along with mutational changes in allo-tRNA and SelA improved the yield and purity of recombinant human glutathione peroxidase 1 (to over 80 %). Thus, our allo-tRNAUTu system offers a new selenoprotein engineering platform.

Original languageEnglish
Pages (from-to)7215-7219
Number of pages5
JournalAngewandte Chemie - International Edition
Volume57
Issue number24
DOIs
Publication statusPublished - Jun 11 2018
Externally publishedYes

All Science Journal Classification (ASJC) codes

  • Catalysis
  • Chemistry(all)

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