title: Response and adaptation of Escherichia coli to suppression of the amber stop codon.
abstract: Some extant organisms reassign the amber stop codon to a sense codon through evolution, and suppression of the amber codon with engineered tRNAs has been exploited to expand the genetic code for incorporating non-canonical amino acids (ncAAs) in live systems. However, it is unclear how the host cells respond and adapt to such amber suppression. Herein we suppressed the amber codon in Escherichia coli with an orthogonal tRNA/synthetase pair and cultured the cells under such a pressure for about 500 generations. We discovered that E. coli quickly counteracted the suppression with transposon insertion to inactivate the orthogonal synthetase. Persistent amber suppression evading transposon inactivation led to global proteomic changes with a notable up-regulation of a previously uncharacterized protein (YdiI) for which we identified an unexpected function of expelling plasmids. These results should be valuable for understanding codon reassignment in genetic code evolution and for improving the efficiency of ncAA incorporation.
authors.affiliation: Jack H. Skirball Center for Chemical Biology and Proteomics, The Salk Institute for Biological Studies, 10010 N. Torrey Pines Road, La Jolla, CA 92037 (USA); College of Veterinary Medicine, China Agricultural University, 2 Yuan Ming Yuan West Road, Beijing 100193 (China).
authors.forename: Qian
authors.initials: Q
authors.lastname: Wang
authors.forename: Tingting
authors.initials: T
authors.lastname: Sun
authors.forename: Jianfeng
authors.initials: J
authors.lastname: Xu
authors.forename: Zhouxin
authors.initials: Z
authors.lastname: Shen
authors.forename: Steven P
authors.initials: SP
authors.lastname: Briggs
authors.forename: Demin
authors.initials: D
authors.lastname: Zhou
authors.forename: Lei
authors.initials: L
authors.lastname: Wang
mesh_terms: D000596:Amino Acids / Q000235:genetics; D018388:Codon, Terminator / Q000235:genetics*; D004926:Escherichia coli / Q000166:cytology / Q000235:genetics*; D005815:Genetic Code; D008025:Ligases / Q000235:genetics / Q000378:metabolism; D015202:Protein Engineering; D040901:Proteomics; D012343:RNA, Transfer / Q000235:genetics
chemical_list: D000596:Amino Acids; D018388:Codon, Terminator; D012343:RNA, Transfer; D008025:Ligases
keywords: amber codon suppression; amino acids; expansion of the genetic code; mutagenesis; synthetic biology