Abstract
AbstractSite-specific incorporation of unnatural amino acids (UAAs) with similar incorporation efficiency to that of natural amino acids (NAAs) and low background activity is extremely valuable for efficient synthesis of proteins with diverse new chemical functions and design of various synthetic auxotrophs. However, such efficient translation systems remain largely unknown in the literature. Here, we describe engineered chimeric phenylalanine systems that dramatically increase the yield of proteins bearing UAAs, through systematic engineering of the aminoacyl-tRNA synthetase and its respective cognate tRNA. These engineered synthetase/tRNA pairs allow single-site and multi-site incorporation of UAAs with efficiencies similar to those of NAAs and high fidelity. In addition, using the evolved chimeric phenylalanine system, we construct a series of E. coli strains whose growth is strictly dependent on exogenously supplied of UAAs. We further show that synthetic auxotrophic cells can grow robustly in living mice when UAAs are supplemented.
Publisher
Springer Science and Business Media LLC
Subject
General Physics and Astronomy,General Biochemistry, Genetics and Molecular Biology,General Chemistry
Reference50 articles.
1. Wang, L., Brock, A., Herberich, B. & Schultz, P. G. Expanding the genetic code of Escherichia coli. Science 292, 498–500 (2001).
2. Liu C. C. & Schultz P. G. Adding new chemistries to the genetic code. Annu. Rev. Biochem. 79, 413–444 (2010)
3. Hu, C., Chan, S. I., Sawyer, E. B., Yu, Y. & Wang, J. Metalloprotein design using genetic code expansion. Chem. Soc. Rev. 43, 6498–6510 (2014).
4. 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).
5. Xiao, H. & Schultz, P. G. At the interface of chemical and biological synthesis: an expanded genetic code. CSH Perspect. Biol. 8, a023945 (2016).
Cited by
32 articles.
订阅此论文施引文献
订阅此论文施引文献，注册后可以免费订阅5篇论文的施引文献，订阅后可以查看论文全部施引文献