title: Evolving tRNA(Sec) for efficient canonical incorporation of selenocysteine.
abstract: Bacterial selenocysteine incorporation occurs in response to opal stop codons and is dependent on the presence of a selenocysteine insertion sequence (SECIS) element, which recruits the selenocysteine specific elongation factor and tRNA(Sec) needed to reassign the UGA codon. The SECIS element is a stem-loop RNA structure immediately following the UGA codon and forms part of the coding sequence in bacterial selenoproteins. Although the site specific incorporation of selenocysteine is of great interest for protein engineering, the sequence constraints imposed by the adjoining SECIS element severely limit its use. We have evolved an E. coli tRNA(Sec) that is compatible with the canonical translation machinery and can suppress amber stop codons to incorporate selenocysteine with high efficiency. This evolved tRNA(Sec) allows the production of new recombinant selenoproteins containing structural motifs such as selenyl-sulfhydryl and diselenide bonds.
authors.affiliation: Institute for Cellular and Molecular Biology, University of Texas at Austin , Austin, Texas 78712, United States.
authors.forename: Ross
authors.initials: R
authors.lastname: Thyer
authors.forename: Scott A
authors.initials: SA
authors.lastname: Robotham
authors.forename: Jennifer S
authors.initials: JS
authors.lastname: Brodbelt
authors.forename: Andrew D
authors.initials: AD
authors.lastname: Ellington
mesh_terms: D004926:Escherichia coli / Q000737:chemistry*; D008958:Models, Molecular; D015202:Protein Engineering; D012343:RNA, Transfer / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D017279:Selenocysteine / Q000737:chemistry* / Q000378:metabolism; D001618:beta-Lactamases / Q000378:metabolism
chemical_list: D017279:Selenocysteine; D012343:RNA, Transfer; D001618:beta-Lactamases