TY - JOUR
T1 - In vitro suppression of two different stop codons
AU - Ozer, Eden
AU - Chemla, Yonatan
AU - Schlesinger, Orr
AU - Aviram, Haim Yuval
AU - Riven, Inbal
AU - Haran, Gilad
AU - Alfonta, Lital
N1 - Funding Information:
Mr. Itai Peker and Prof. Ronni Grannek are greatly acknowledged for useful discussions. Mr. Michael Shaferman is acknowledged for technical support. We would also like to acknowledge Prof. Vincent Noiraeux for generously giving us the pBEST plasmid and for useful comments regarding CFPS preparation. Prof. Dudy Bar-Zvi is gratefully acknowledged for his assistance with the FRET performed with a Typhoon Gel reader. This research was supported by an E.R.C. grant number 260647 (LA) Support of Kreitman school for graduate students for PhD fellowships (EO and YC) and of a Merage interdisciplinary PhD fellowship (OS). The work of GH was supported in part by grant no 686/14 from the Israel Science Foundation.
Publisher Copyright:
© 2016 Wiley Periodicals, Inc.
PY - 2017/5/1
Y1 - 2017/5/1
N2 - Proteins play a crucial role in all living organisms, with the 20 natural amino acids as their building blocks. Unnatural amino acids are synthetic derivatives of these natural building blocks. These amino acids have unique chemical or physical properties as a result of their specific side chain residues. Their incorporation into proteins through ribosomal translation in response to one of the stop codons has opened a new way to manipulate and study proteins by enabling new functionalities, thus expending the genetic code. Different unnatural amino acids have different functionalities, hence, the ability to incorporate two different unnatural amino acids, in response to two different stop codons into one protein is a useful tool in protein manipulation. This ability has been achieved previously only in in vivo translational systems, however, with limited functionality. Herein, we report the incorporation of two different unnatural amino acids in response to two different stop codons into one protein, utilizing a cell-free protein synthesis system. Biotechnol. Bioeng. 2017;114: 1065–1073.
AB - Proteins play a crucial role in all living organisms, with the 20 natural amino acids as their building blocks. Unnatural amino acids are synthetic derivatives of these natural building blocks. These amino acids have unique chemical or physical properties as a result of their specific side chain residues. Their incorporation into proteins through ribosomal translation in response to one of the stop codons has opened a new way to manipulate and study proteins by enabling new functionalities, thus expending the genetic code. Different unnatural amino acids have different functionalities, hence, the ability to incorporate two different unnatural amino acids, in response to two different stop codons into one protein is a useful tool in protein manipulation. This ability has been achieved previously only in in vivo translational systems, however, with limited functionality. Herein, we report the incorporation of two different unnatural amino acids in response to two different stop codons into one protein, utilizing a cell-free protein synthesis system. Biotechnol. Bioeng. 2017;114: 1065–1073.
KW - Förster resonance energy transfer
KW - cell-free protein synthesis
KW - dual genetic code expansion
KW - orthogonal translation systems
UR - http://www.scopus.com/inward/record.url?scp=85006100785&partnerID=8YFLogxK
U2 - 10.1002/bit.26226
DO - 10.1002/bit.26226
M3 - Article
AN - SCOPUS:85006100785
VL - 114
SP - 1065
EP - 1073
JO - Biotechnology and Bioengineering
JF - Biotechnology and Bioengineering
SN - 0006-3592
IS - 5
ER -