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Expanding and reprogramming the genetic code

Abstract

Nature uses a limited, conservative set of amino acids to synthesize proteins. The ability to genetically encode an expanded set of building blocks with new chemical and physical properties is transforming the study, manipulation and evolution of proteins, and is enabling diverse applications, including approaches to probe, image and control protein function, and to precisely engineer therapeutics. Underpinning this transformation are strategies to engineer and rewire translation. Emerging strategies aim to reprogram the genetic code so that noncanonical biopolymers can be synthesized and evolved, and to test the limits of our ability to engineer the translational machinery and systematically recode genomes.

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Figure 1: The natural translational machinery performs encoded amino acid polymerization.
Figure 2: Genetic code expansion for ncAA incorporation into proteins in vivo and its use for creating attenuated viruses.
Figure 3: Strategies to go beyond nonsense codons for the incorporation of multiple distinct unnatural monomers.

References

  1. Ambrogelly, A., Palioura, S. & Söll, D. Natural expansion of the genetic code. Nat. Chem. Biol. 3, 29–35 (2007)

    Article  CAS  PubMed  Google Scholar 

  2. Dougherty, D. A. & Van Arnam, E. B. In vivo incorporation of non-canonical amino acids by using the chemical aminoacylation strategy: a broadly applicable mechanistic tool. ChemBioChem 15, 1710–1720 (2014)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Cornish, V. W., Mendel, D. & Schultz, P. G. Probing protein structure and function with an expanded genetic code. Angew. Chem. Int. Edn Engl. 34, 621–633 (1995)

    Article  CAS  Google Scholar 

  4. Bondalapati, S., Jbara, M. & Brik, A. Expanding the chemical toolbox for the synthesis of large and uniquely modified proteins. Nat. Chem. 8, 407–418 (2016)

    Article  CAS  PubMed  Google Scholar 

  5. Chin, J. W. Expanding and reprogramming the genetic code of cells and animals. Annu. Rev. Biochem. 83, 379–408 (2014)

    Article  CAS  PubMed  Google Scholar 

  6. Liu, C. C. & Schultz, P. G. Adding new chemistries to the genetic code. Annu. Rev. Biochem. 79, 413–444 (2010)

    Article  CAS  PubMed  Google Scholar 

  7. Zhang, M. S . et al. Biosynthesis and genetic encoding of phosphothreonine through parallel selection and deep sequencing. Nat. Methods 14, 729–736 (2017). Describes a scalable approach to the discovery of orthgonal synthetases through parallel positive selection and sequencing and a strategy to biosynthesize and encode a key post-translational modification.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Anderson, J. C. & Schultz, P. G. Adaptation of an orthogonal archaeal leucyl-tRNA and synthetase pair for four-base, amber, and opal suppression. Biochemistry 42, 9598–9608 (2003)

    Article  CAS  PubMed  Google Scholar 

  9. Neumann, H., Peak-Chew, S. Y. & Chin, J. W. Genetically encoding N(ε)-acetyllysine in recombinant proteins. Nat. Chem. Biol. 4, 232–234 (2008)

    Article  CAS  PubMed  Google Scholar 

  10. Ellefson, J. W. et al. Directed evolution of genetic parts and circuits by compartmentalized partnered replication. Nat. Biotechnol. 32, 97–101 (2014)

    Article  CAS  PubMed  Google Scholar 

  11. Wang, L., Brock, A., Herberich, B. & Schultz, P. G. Expanding the genetic code of Escherichia coli. Science 292, 498–500 (2001)

    Article  ADS  CAS  PubMed  Google Scholar 

  12. Young, D. D. et al. An evolved aminoacyl-tRNA synthetase with atypical polysubstrate specificity. Biochemistry 50, 1894–1900 (2011)

    Article  CAS  PubMed  Google Scholar 

  13. Cooley, R. B., Karplus, P. A. & Mehl, R. A. Gleaning unexpected fruits from hard-won synthetases: probing principles of permissivity in non-canonical amino acid-tRNA synthetases. ChemBioChem 15, 1810–1819 (2014)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Dumas, A., Lercher, L., Spicer, C. D. & Davis, B. G. Designing logical codon reassignment — Expanding the chemistry in biology. Chem. Sci. 6, 50–69 (2015)

    Article  CAS  PubMed  Google Scholar 

  15. Santoro, S. W., Anderson, J. C., Lakshman, V. & Schultz, P. G. An archaebacteria-derived glutamyl-tRNA synthetase and tRNA pair for unnatural amino acid mutagenesis of proteins in Escherichia coli. Nucleic Acids Res. 31, 6700–6709 (2003)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Tes¸ileanu, T., Colwell, L. J. & Leibler, S. Protein sectors: statistical coupling analysis versus conservation. PLOS Comput. Biol. 11, e1004091 (2015)

    Article  CAS  Google Scholar 

  17. Iraha, F. et al. Functional replacement of the endogenous tyrosyl-tRNA synthetase-tRNATyr pair by the archaeal tyrosine pair in Escherichia coli for genetic code expansion. Nucleic Acids Res. 38, 3682–3691 (2010)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Hughes, R. A. & Ellington, A. D. Rational design of an orthogonal tryptophanyl nonsense suppressor tRNA. Nucleic Acids Res. 38, 6813–6830 (2010)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Italia, J. S . et al. An orthogonalized platform for genetic code expansion in both bacteria and eukaryotes. Nat. Chem. Biol. 13, 446–450 (2017). Evolution of the E. coli TrpRS–tRNA pair in E. coli and transfer of this pair to mammalian cells for genetic code expansion.

    Article  CAS  PubMed  Google Scholar 

  20. Ernst, R. J. et al. Genetic code expansion in the mouse brain. Nat. Chem. Biol. 12, 776–778 (2016)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Han, S. et al. Expanding the genetic code of Mus musculus. Nat. Commun. 8, 14568 (2017)

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  22. Elliott, T. S. et al. Proteome labeling and protein identification in specific tissues and at specific developmental stages in an animal. Nat. Biotechnol. 32, 465–472 (2014)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Elliott, T. S., Bianco, A., Townsley, F. M., Fried, S. D. & Chin, J. W. Tagging and enriching proteins enables cell-specific proteomics. Cell Chem. Biol. 23, 805–815 (2016)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Stone, S. E., Glenn, W. S., Hamblin, G. D. & Tirrell, D. A. Cell-selective proteomics for biological discovery. Curr. Opin. Chem. Biol. 36, 50–57 (2017)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Schmied, W. H., Elsässer, S. J., Uttamapinant, C. & Chin, J. W. Efficient multisite unnatural amino acid incorporation in mammalian cells via optimized pyrrolysyl tRNA synthetase/tRNA expression and engineered eRF1. J. Am. Chem. Soc. 136, 15577–15583 (2014)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Amiram, M. et al. Evolution of translation machinery in recoded bacteria enables multi-site incorporation of nonstandard amino acids. Nat. Biotechnol. 33, 1272–1279 (2015)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Zheng, Y., Lewis, T. L., Jr, Igo, P., Polleux, F. & Chatterjee, A. Virus-enabled optimization and delivery of the genetic machinery for efficient unnatural amino acid mutagenesis in mammalian cells and tissues. ACS Synth. Biol. 6, 13–18 (2017)

    Article  CAS  PubMed  Google Scholar 

  28. Chatterjee, A., Sun, S. B., Furman, J. L., Xiao, H. & Schultz, P. G. A versatile platform for single- and multiple-unnatural amino acid mutagenesis in Escherichia coli. Biochemistry 52, 1828–1837 (2013)

    Article  CAS  PubMed  Google Scholar 

  29. Wang, K., Neumann, H., Peak-Chew, S. Y. & Chin, J. W. Evolved orthogonal ribosomes enhance the efficiency of synthetic genetic code expansion. Nat. Biotechnol. 25, 770–777 (2007)

    Article  CAS  PubMed  Google Scholar 

  30. Neumann, H., Wang, K., Davis, L., Garcia-Alai, M. & Chin, J. W. Encoding multiple unnatural amino acids via evolution of a quadruplet-decoding ribosome. Nature 464, 441–444 (2010)

    Article  ADS  CAS  PubMed  Google Scholar 

  31. Wang, K. et al. Optimized orthogonal translation of unnatural amino acids enables spontaneous protein double-labelling and FRET. Nat. Chem. 6, 393–403 (2014)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Park, H. S. et al. Expanding the genetic code of Escherichia coli with phosphoserine. Science 333, 1151–1154 (2011)

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  33. Fan, C., Ip, K. & Söll, D. Expanding the genetic code of Escherichia coli with phosphotyrosine. FEBS Lett. 590, 3040–3047 (2016)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Mukai, T. et al. Codon reassignment in the Escherichia coli genetic code. Nucleic Acids Res. 38, 8188–8195 (2010)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Johnson, D. B. et al. RF1 knockout allows ribosomal incorporation of unnatural amino acids at multiple sites. Nat. Chem. Biol. 7, 779–786 (2011)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Lajoie, M. J. et al. Genomically recoded organisms expand biological functions. Science 342, 357–360 (2013)

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  37. Wu, I. L. et al. Multiple site-selective insertions of noncanonical amino acids into sequence-repetitive polypeptides. ChemBioChem 14, 968–978 (2013)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Chatterjee, A., Lajoie, M. J., Xiao, H., Church, G. M. & Schultz, P. G. A bacterial strain with a unique quadruplet codon specifying non-native amino acids. ChemBioChem 15, 1782–1786 (2014)

    Article  CAS  PubMed  Google Scholar 

  39. Mukai, T. et al. Highly reproductive Escherichia coli cells with no specific assignment to the UAG codon. Sci. Rep. 5, 9699 (2015)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Zheng, Y. et al. Performance of optimized noncanonical amino acid mutagenesis systems in the absence of release factor 1. Mol. Biosyst. 12, 1746–1749 (2016)

    Article  CAS  PubMed  Google Scholar 

  41. Rogerson, D. T. et al. Efficient genetic encoding of phosphoserine and its nonhydrolyzable analog. Nat. Chem. Biol. 11, 496–503 (2015)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. Richardson, S. M. et al. Design of a synthetic yeast genome. Science 355, 1040–1044 (2017)

    Article  ADS  CAS  PubMed  Google Scholar 

  43. Davis, L. & Chin, J. W. Designer proteins: applications of genetic code expansion in cell biology. Nat. Rev. Mol. Cell Biol. 13, 168–182 (2012)

    Article  CAS  PubMed  Google Scholar 

  44. Minnihan, E. C., Seyedsayamdost, M. R., Uhlin, U. & Stubbe, J. Kinetics of radical intermediate formation and deoxynucleotide production in 3-aminotyrosine-substituted Escherichia coli ribonucleotide reductases. J. Am. Chem. Soc. 133, 9430–9440 (2011)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Yang, Y. et al. Genetically encoded protein photocrosslinker with a transferable mass spectrometry-identifiable label. Nat. Commun. 7, 12299 (2016)

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  46. Coin, I. et al. Genetically encoded chemical probes in cells reveal the binding path of urocortin-I to CRF class B GPCR. Cell 155, 1258–1269 (2013)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  47. Lang, K. & Chin, J. W. Cellular incorporation of unnatural amino acids and bioorthogonal labeling of proteins. Chem. Rev. 114, 4764–4806 (2014)

    Article  CAS  PubMed  Google Scholar 

  48. Chatterjee, A., Guo, J., Lee, H. S. & Schultz, P. G. A genetically encoded fluorescent probe in mammalian cells. J. Am. Chem. Soc. 135, 12540–12543 (2013)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  49. Luo, J. et al. Genetically encoded optochemical probes for simultaneous fluorescence reporting and light activation of protein function with two-photon excitation. J. Am. Chem. Soc. 136, 15551–15558 (2014)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  50. Lang, K. & Chin, J. W. Bioorthogonal reactions for labeling proteins. ACS Chem. Biol. 9, 16–20 (2014)

    Article  CAS  PubMed  Google Scholar 

  51. Lukinavicˇius, G. et al. A near-infrared fluorophore for live-cell super-resolution microscopy of cellular proteins. Nat. Chem. 5, 132–139 (2013)

    Article  CAS  Google Scholar 

  52. Uttamapinant, C. et al. Genetic code expansion enables live-cell and super-resolution imaging of site-specifically labeled cellular proteins. J. Am. Chem. Soc. 137, 4602–4605 (2015)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  53. Kipper, K. et al. Application of noncanonical amino acids for protein labeling in a genomically recoded Escherichia coli. ACS Synth. Biol. 6, 233–255 (2017)

    Article  CAS  PubMed  Google Scholar 

  54. Peng, T. & Hang, H. C. Site-specific bioorthogonal labeling for fluorescence imaging of intracellular proteins in living cells. J. Am. Chem. Soc. 138, 14423–14433 (2016)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  55. Baumdick, M. et al. EGF-dependent re-routing of vesicular recycling switches spontaneous phosphorylation suppression to EGFR signaling. eLife 4, e12223 (2015)

    Article  PubMed  PubMed Central  Google Scholar 

  56. Sakin, V. et al. A versatile tool for live-cell imaging and super-resolution nanoscopy studies of HIV-1 Env distribution and mobility. Cell. Chem. Biol. 24, 635–645 (2017)

    Article  CAS  PubMed  Google Scholar 

  57. Xue, L., Prifti, E. & Johnsson, K. A general strategy for the semisynthesis of ratiometric fluorescent sensor proteins with increased dynamic range. J. Am. Chem. Soc. 138, 5258–5261 (2016)

    Article  CAS  PubMed  Google Scholar 

  58. Sachdeva, A., Wang, K., Elliott, T. & Chin, J. W. Concerted, rapid, quantitative, and site-specific dual labeling of proteins. J. Am. Chem. Soc. 136, 7785–7788 (2014)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  59. Xiao, H. et al. Genetic incorporation of multiple unnatural amino acids into proteins in mammalian cells. Angew. Chem. Int. Edn Engl. 52, 14080–14083 (2013)

    Article  CAS  Google Scholar 

  60. Li, J. et al. Palladium-triggered deprotection chemistry for protein activation in living cells. Nat. Chem. 6, 352–361 (2014)

    Article  CAS  PubMed  Google Scholar 

  61. Li, J., Jia, S. & Chen, P. R. Diels–Alder reaction-triggered bioorthogonal protein decaging in living cells. Nat. Chem. Biol. 10, 1003–1005 (2014)

    Article  CAS  PubMed  Google Scholar 

  62. Zhang, G. et al. Bioorthogonal chemical activation of kinases in living systems. ACS Cent. Sci. 2, 325–331 (2016)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  63. Wang, J. et al. Palladium-triggered chemical rescue of intracellular proteins via genetically encoded allene-caged tyrosine. J. Am. Chem. Soc. 138, 15118–15121 (2016)

    Article  CAS  PubMed  Google Scholar 

  64. Baker, A. S. & Deiters, A. Optical control of protein function through unnatural amino acid mutagenesis and other optogenetic approaches. ACS Chem. Biol. 9, 1398–1407 (2014)

    Article  CAS  PubMed  Google Scholar 

  65. Hemphill, J., Borchardt, E. K., Brown, K., Asokan, A. & Deiters, A. Optical control of CRISPR/Cas9 gene editing. J. Am. Chem. Soc. 137, 5642–5645 (2015)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  66. Nguyen, D. P. et al. Genetic encoding of photocaged cysteine allows photoactivation of TEV protease in live mammalian cells. J. Am. Chem. Soc. 136, 2240–2243 (2014)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  67. Walker, O. S. et al. Photoactivation of mutant isocitrate dehydrogenase 2 reveals rapid cancer-associated metabolic and epigenetic changes. J. Am. Chem. Soc. 138, 718–721 (2016)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  68. Bose, M., Groff, D., Xie, J., Brustad, E. & Schultz, P. G. The incorporation of a photoisomerizable amino acid into proteins in E. coli. J. Am. Chem. Soc. 128, 388–389 (2006)

    Article  CAS  PubMed  Google Scholar 

  69. Hoppmann, C., Maslennikov, I., Choe, S. & Wang, L. In situ formation of an azo bridge on proteins controllable by visible light. J. Am. Chem. Soc. 137, 11218–11221 (2015)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  70. Tsai, Y.-H., Essig, S., James, J. R., Lang, K. & Chin, J. W. Selective, rapid and optically switchable regulation of protein function in live mammalian cells. Nat. Chem. 7, 554–561 (2015)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  71. Elsässer, S. J., Ernst, R. J., Walker, O. S. & Chin, J. W. Genetic code expansion in stable cell lines enables encoded chromatin modification. Nat. Methods 13, 158–164 (2016)

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  72. Si, L . et al. Generation of influenza A viruses as live but replication-incompetent virus vaccines. Science 354, 1170–1173 (2016). Describes the creation of an amber suppression-dependent influenza A virus and the use of the resulting attenuated virus for immunization.

    Article  ADS  CAS  PubMed  Google Scholar 

  73. Wang, Z. A. et al. A genetically encoded allysine for the synthesis of proteins with site-specific lysine dimethylation. Angew. Chem. Int. Edn Engl. 56, 212–216 (2017)

    Article  CAS  Google Scholar 

  74. Wang, Z. A. et al. A versatile approach for site-specific lysine acylation in proteins. Angew. Chem. Int. Edn Engl. 56, 1643–1647 (2017)

    Article  CAS  Google Scholar 

  75. Hoppmann, C. et al. Site-specific incorporation of phosphotyrosine using an expanded genetic code. Nat. Chem. Biol. 13, 842–844 (2017)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  76. Luo, X. et al. Genetically encoding phosphotyrosine and its nonhydrolyzable analog in bacteria. Nat. Chem. Biol. 13, 845–849 (2017)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  77. Wright, T. H. et al. Posttranslational mutagenesis: a chemical strategy for exploring protein side-chain diversity. Science 354, aag1465 (2016)

    Article  PubMed  CAS  Google Scholar 

  78. Yang, A. et al. A chemical biology route to site-specific authentic protein modifications. Science 354, 623–626 (2016)

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  79. Xiao, H. & Schultz, P. G. At the interface of chemical and biological synthesis: an expanded genetic code. Cold Spring Harb. Perspect. Biol. 8, a023945 (2016)

    Google Scholar 

  80. Tian, F. et al. A general approach to site-specific antibody drug conjugates. Proc. Natl Acad. Sci. USA 111, 1766–1771 (2014)

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  81. VanBrunt, M. P. et al. Genetically encoded azide containing amino acid in mammalian cells enables site-specific antibody-drug conjugates using click cycloaddition chemistry. Bioconjug. Chem. 26, 2249–2260 (2015)

    Article  CAS  PubMed  Google Scholar 

  82. Kularatne, S. A. et al. Recruiting cytotoxic T cells to folate-receptor-positive cancer cells. Angew. Chem. Int. Edn Engl. 52, 12101–12104 (2013)

    Article  CAS  Google Scholar 

  83. Ma, J. S. et al. Versatile strategy for controlling the specificity and activity of engineered T cells. Proc. Natl Acad. Sci. USA 113, E450–E458 (2016)

    CAS  PubMed  PubMed Central  Google Scholar 

  84. Wang, N. et al. Construction of a live-attenuated HIV-1 vaccine through genetic code expansion. Angew. Chem. Int. Edn Engl. 53, 4867–4871 (2014)

    Article  CAS  Google Scholar 

  85. Lin, S. et al. Site-specific engineering of chemical functionalities on the surface of live hepatitis D virus. Angew. Chem. Int. Edn Engl. 52, 13970–13974 (2013)

    Article  CAS  Google Scholar 

  86. Loughran, G. et al. Evidence of efficient stop codon readthrough in four mammalian genes. Nucleic Acids Res. 42, 8928–8938 (2014)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  87. Xuan, W. & Schultz, P. G. A strategy for creating organisms dependent on noncanonical amino acids. Angew. Chem. Int. Edn Engl. 56, 9170–9173 (2017)

    Article  CAS  Google Scholar 

  88. Mandell, D. J. et al. Biocontainment of genetically modified organisms by synthetic protein design. Nature 518, 55–60 (2015). Makes the function of essential genes dependent on both amber suppression and the identity of a noncanonical amino acid incorporated by amber suppression.

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  89. Mills, J. H. et al. Computational design of an unnatural amino acid dependent metalloprotein with atomic level accuracy. J. Am. Chem. Soc. 135, 13393–13399 (2013)

    Article  CAS  PubMed  Google Scholar 

  90. Mills, J. H. et al. Computational design of a homotrimeric metalloprotein with a trisbipyridyl core. Proc. Natl Acad. Sci. USA 113, 15012–15017 (2016). Design of metalloproteins with an expanded genetic code.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  91. Pearson, A. D. et al. Trapping a transition state in a computationally designed protein bottle. Science 347, 863–867 (2015)

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  92. Xiao, H. et al. Exploring the potential impact of an expanded genetic code on protein function. Proc. Natl Acad. Sci. USA 112, 6961–6966 (2015)

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  93. Tack, D. S. et al. Addicting diverse bacteria to a noncanonical amino acid. Nat. Chem. Biol. 12, 138–140 (2016)

    Article  CAS  PubMed  Google Scholar 

  94. Hammerling, M. J. et al. Bacteriophages use an expanded genetic code on evolutionary paths to higher fitness. Nat. Chem. Biol. 10, 178–180 (2014)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  95. Lopez, G. & Anderson, J. C. Synthetic auxotrophs with ligand-dependent essential genes for a BL21(DE3) biosafety strain. ACS Synth. Biol. 4, 1279–1286 (2015)

    Article  CAS  PubMed  Google Scholar 

  96. Forster, A. C. et al. Programming peptidomimetic syntheses by translating genetic codes designed de novo. Proc. Natl Acad. Sci. USA 100, 6353–6357 (2003)

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  97. Iwane, Y. et al. Expanding the amino acid repertoire of ribosomal polypeptide synthesis via the artificial division of codon boxes. Nat. Chem. 8, 317–325 (2016)

    Article  CAS  PubMed  Google Scholar 

  98. Anderson, J. C. et al. An expanded genetic code with a functional quadruplet codon. Proc. Natl Acad. Sci. USA 101, 7566–7571 (2004)

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  99. Malyshev, D. A. et al. A semi-synthetic organism with an expanded genetic alphabet. Nature 509, 385–388 (2014)

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  100. Zhang, Y . et al. A semisynthetic organism engineered for the stable expansion of the genetic alphabet. Proc. Natl Acad. Sci. USA 114, 1317–1322 (2017). The maintenance of an orthogonal base pair in E. coli.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  101. Ngo, J. T. & Tirrell, D. A. Noncanonical amino acids in the interrogation of cellular protein synthesis. Acc. Chem. Res. 44, 677–685 (2011)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  102. Zeng, Y., Wang, W. & Liu, W. R. Towards reassigning the rare AGG codon in Escherichia coli. ChemBioChem 15, 1750–1754 (2014)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  103. Ostrov, N . et al. Design, synthesis, and testing toward a 57-codon genome. Science 353, 819–822 (2016). Replaces 50-kb sections of the E. coli genome in ten independent strains using variable synonymous codon replacements.

    Article  ADS  CAS  PubMed  Google Scholar 

  104. Wang, K. et al. Defining synonymous codon compression schemes by genome recoding. Nature 539, 59–64 (2016). Describes a powerful strategy for genome replacement in E. coli and its application to deciphering the best synonymous substitutions for codons targeted for removal from the genome.

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  105. Napolitano, M. G. et al. Emergent rules for codon choice elucidated by editing rare arginine codons in Escherichia coli. Proc. Natl Acad. Sci. USA 113, E5588–E5597 (2016)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  106. Lau, Y. H. et al. Large-scale recoding of a bacterial genome by iterative recombineering of synthetic DNA. Nucleic Acids Res. 45, 6971–6980 (2017)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  107. Neumann, H., Slusarczyk, A. L. & Chin, J. W. De novo generation of mutually orthogonal aminoacyl-tRNA synthetase/tRNA pairs. J. Am. Chem. Soc. 132, 2142–2144 (2010)

    Article  CAS  PubMed  Google Scholar 

  108. Chatterjee, A., Xiao, H. & Schultz, P. G. Evolution of multiple, mutually orthogonal prolyl-tRNA synthetase/tRNA pairs for unnatural amino acid mutagenesis in Escherichia coli. Proc. Natl Acad. Sci. USA 109, 14841–14846 (2012)

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  109. Guo, J., Wang, J., Anderson, J. C. & Schultz, P. G. Addition of an alpha-hydroxy acid to the genetic code of bacteria. Angew. Chem. Int. Edn Engl. 47, 722–725 (2008)

    Article  CAS  Google Scholar 

  110. Kobayashi, T., Yanagisawa, T., Sakamoto, K. & Yokoyama, S. Recognition of non-alpha-amino substrates by pyrrolysyl-tRNA synthetase. J. Mol. Biol. 385, 1352–1360 (2009)

    Article  CAS  PubMed  Google Scholar 

  111. Tan, Z., Forster, A. C., Blacklow, S. C. & Cornish, V. W. Amino acid backbone specificity of the Escherichia coli translation machinery. J. Am. Chem. Soc. 126, 12752–12753 (2004)

    Article  CAS  PubMed  Google Scholar 

  112. Fujino, T., Goto, Y., Suga, H. & Murakami, H. Reevaluation of the d-amino acid compatibility with the elongation event in translation. J. Am. Chem. Soc. 135, 1830–1837 (2013)

    Article  CAS  PubMed  Google Scholar 

  113. Maini, R. et al. Protein synthesis with ribosomes selected for the incorporation of β-amino acids. Biochemistry 54, 3694–3706 (2015)

    Article  CAS  PubMed  Google Scholar 

  114. Maini, R. et al. Ribosome-mediated incorporation of dipeptides and dipeptide analogues into proteins in vitro. J. Am. Chem. Soc. 137, 11206–11209 (2015)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  115. Dedkova, L. M., Fahmi, N. E., Golovine, S. Y. & Hecht, S. M. Construction of modified ribosomes for incorporation of d-amino acids into proteins. Biochemistry 45, 15541–15551 (2006)

    Article  CAS  PubMed  Google Scholar 

  116. Melo Czekster, C., Robertson, W. E., Walker, A. S., Söll, D. & Schepartz, A. In vivo biosynthesis of a β-amino acid-containing protein. J. Am. Chem. Soc. 138, 5194–5197 (2016)

    Article  CAS  PubMed  Google Scholar 

  117. Englander, M. T. et al. The ribosome can discriminate the chirality of amino acids within its peptidyl-transferase center. Proc. Natl Acad. Sci. USA 112, 6038–6043 (2015)

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  118. Terasaka, N., Hayashi, G., Katoh, T. & Suga, H. An orthogonal ribosome-tRNA pair via engineering of the peptidyl transferase center. Nat. Chem. Biol. 10, 555–557 (2014)

    Article  CAS  PubMed  Google Scholar 

  119. Fried, S. D., Schmied, W. H., Uttamapinant, C. & Chin, J. W. Ribosome subunit stapling for orthogonal translation in E. coli. Angew. Chem. Int. Edn Engl. 54, 12791–12794 (2015)

    Article  CAS  Google Scholar 

  120. Orelle, C. et al. Protein synthesis by ribosomes with tethered subunits. Nature 524, 119–124 (2015)

    Article  ADS  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

I thank A. Chatterjee, K. Wang, and P. G. Schultz for suggestions and edits to earlier versions of the manuscript and V. Beranek, and K. Wang for assistance with figures. Work in my laboratory is supported by the Medical Research Council, UK (MC_U105181009 and MC_UP_A024_1008) and an ERC Advanced Grant (SGCR).

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Correspondence to Jason W. Chin.

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Reviewer Information Nature thanks A. C. Forster, A. Schepartz and the other anonymous reviewer(s) for their contribution to the peer review of this work.

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Chin, J. Expanding and reprogramming the genetic code. Nature 550, 53–60 (2017). https://doi.org/10.1038/nature24031

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