Abstract
Escherichia coli has been considered as the most used model bacteria in the majority of studies for several decades. However, a new, faster chassis for synthetic biology is emerging in the form of the fast-growing gram-negative bacterium Vibrio natriegens. Different methodologies, well established in E. coli, are currently being adapted for V. natriegens in the hope to enable a much faster platform for general molecular biology studies. Amongst the vast technologies available for E. coli, genetic code expansion, the incorporation of unnatural amino acids into proteins, serves as a robust tool for protein engineering and biorthogonal modifications. Here we designed and adapted the genetic code expansion methodology for V. natriegens and demonstrate an unnatural amino acid incorporation into a protein for the first time in this organism.
Highlights
Vibrio natriegens (V. natriegens), is a fast-growing gram-negative bacterium identified in the 1960s of the 20th century with a remarkable division time of less than 10 min (Eagon, 1962)
Considering the need to optimize the efficiency of molecular and synthetic biology studies in the lab, V. natriegens presents an unprecedented opportunity to speed-up lab work and emerges as a novel synthetic biology and biotechnological chassis
In order to confirm the orthogonality of the orthogonal translation system (OTS) in V. natriegens, the OTS needs to interact with a specific unnatural amino acids (Uaas) and to have no cross-reactivity with other components: native amino acids, endogenous tRNAs or tRNA-synthetases
Summary
Vibrio natriegens (V. natriegens), is a fast-growing gram-negative bacterium identified in the 1960s of the 20th century with a remarkable division time of less than 10 min (Eagon, 1962). Over 200 Uaas were incorporated into different proteins (Xiao and Schultz, 2016) in different organisms including the primary E. coli system, amongst them are Salmonella (Gan et al, 2016), cyanobacteria (Chemla et al, 2017), Yeast (Hancock et al, 2010), Caenorhabditis elegans (Greiss and Chin, 2011), mouse (Ernst et al, 2016), mammalian cells (Italia et al, 2017), and cellfree systems (Ozer et al, 2017). Genetic code expansion in V. natriegens can serve as an improved and a faster platform for future protein engineering in this new chassis, possibly for much-needed research in the organism itself but mainly as a powerhouse for production of proteins with expanded amino acid repertoire
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