Applied Microbiology and Biotechnology

, Volume 103, Issue 7, pp 2947–2958 | Cite as

Incorporation of non-standard amino acids into proteins: challenges, recent achievements, and emerging applications

  • Xing Jin
  • Oh-Jin Park
  • Seok Hoon HongEmail author


The natural genetic code only allows for 20 standard amino acids in protein translation, but genetic code reprogramming enables the incorporation of non-standard amino acids (NSAAs). Proteins containing NSAAs provide enhanced or novel properties and open diverse applications. With increased attention to the recent advancements in synthetic biology, various improved and novel methods have been developed to incorporate single and multiple distinct NSAAs into proteins. However, various challenges remain in regard to NSAA incorporation, such as low yield and misincorporation. In this review, we summarize the recent efforts to improve NSAA incorporation by utilizing orthogonal translational system optimization, cell-free protein synthesis, genomically recoded organisms, artificial codon boxes, quadruplet codons, and orthogonal ribosomes, before closing with a discussion of the emerging applications of NSAA incorporation.


Non-standard amino acids Genetic code expansion Synthetic biology Orthogonal translational system 



This study was funded by the National Institute of Allergy and Infectious Diseases of the National Institutes of Health (R15AI130988).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical approval

This article does not contain any studies with human participants or animals performed by any of the authors.


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© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Department of Chemical and Biological EngineeringIllinois Institute of TechnologyChicagoUSA
  2. 2.Department of Biological and Chemical EngineeringYanbian University of Science and TechnologyYanjiPeople’s Republic of China

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