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Mol Biosyst. 2016 May 24;12(6):1746-9. doi: 10.1039/c6mb00070c.

Performance of optimized noncanonical amino acid mutagenesis systems in the absence of release factor 1.

Author information

1
Department of Chemistry, Boston College, 2609 Beacon Street, Chestnut Hill, MA 02467, USA. abhishek.chatterjee@bc.edu.
2
Department of Genetics, Harvard Medical School, 77 Avenue Louis Pasteur, Boston, MA 02115, USA and Department of Biochemistry, University of Washington, Seattle, WA 98195, USA.
3
Department of Genetics, Harvard Medical School, 77 Avenue Louis Pasteur, Boston, MA 02115, USA.

Abstract

Site-specific incorporation of noncanonical amino acids (ncAAs) into proteins expressed in E. coli using UAG-suppression competes with termination mediated by release factor 1 (RF1). Recently, unconditional deletion of RF1 was achieved in a genomically recoded E. coli (C321), devoid of all endogenous UAG stop codons. Here we evaluate the efficiency of ncAA incorporation in this strain using optimized suppression vectors. Even though the absence of RF1 does not benefit the suppression efficiency of a single UAG codon, multi-site incorporation of a series of chemically distinct ncAAs was significantly improved.

PMID:
27027374
DOI:
10.1039/c6mb00070c
[Indexed for MEDLINE]

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