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A Facile Method for Producing Selenocysteine‐Containing Proteins
Author(s) -
Mukai Takahito,
Sevostyanova Anastasia,
Suzuki Tateki,
Fu Xian,
Söll Dieter
Publication year - 2018
Publication title -
angewandte chemie international edition
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 5.831
H-Index - 550
eISSN - 1521-3773
pISSN - 1433-7851
DOI - 10.1002/anie.201713215
Subject(s) - selenocysteine , selenoprotein , transfer rna , escherichia coli , biochemistry , chemistry , genetic code , formate dehydrogenase , serine , amino acid , cysteine , enzyme , gene , rna , glutathione , glutathione peroxidase , cofactor
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 tRNA Ser . 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 FDH H 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‐tRNA UTu system offers a new selenoprotein engineering platform.