ABSTRACT
The incorporation of unnatural amino acids are attractive methods for improving or bringing new and novel functions in peptides and proteins. Cell-free protein synthesis using the Protein Synthesis Using Recombinant Elements (PURE) system is an attractive platform for efficient unnatural amino acid incorporation. In this work, we further adapted and modified the One Pot PURE for a robust and modular system of enzymatic single site-specific of unnatural amino acid. We demonstrated the flexibility of this system through the introduction of two orthogonal aminoacyl tRNA synthetases and the suppression of two distinctive stop codons.
Competing Interest Statement
The authors have declared no competing interest.
ABBREVIATIONS
- PURE
- Protein Synthesis Using Recombinant Elemtns
- CFPS
- Cell-free protein synthesis
- IVTT
- In vitro transcription and translation
- aaRS
- aminoacyl-tRNA synthetease
- ncAA
- noncanonical amino acids
- L-DOPA
- levodopa
- ProgK
- and N-ε-propagyloxycarbonyl-L-lysine
- OFP
- orange fluorescent protein
- PylRS
- pyrrolysyl-tRNA synthetase
- RF-1
- Release Factor-1
- RF-2
- Release Factor -2
- M. Barkeri
- Methanosacrina Barkeri
- M. Alvus
- Methanomethylophilus Alvus
- sfGFP
- superfolder green fluorescent protein
Copyright
The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC-ND 4.0 International license.
Posted July 28, 2023.
Changes in coding and efficiency through modular modifications to a One Pot PURE system for in vitro transcription & translation
