Playing with the Molecules of Life

Abstract

Our understanding of the complex processes of living organisms at the molecular level is growing exponentially. This knowledge, together with a powerful arsenal of tools for manipulating the structures of macromolecules, is allowing chemists to harness and reprogram the cellular machinery. Here we review one example in which the genetic code itself has been expanded with new building blocks that allow us to probe and manipulate the structures and functions of proteins in ways previously unimaginable

Resurrecting the Bacterial Tyrosyl-tRNA Synthetase/tRNA Pair for Expanding the Genetic Code of Both E. coli and Eukaryotes

“…(Chin, 2017; Dumas et al, 2015; Italia et al, 2017b; Mukai et al, 2017; Young and Schultz, 2018) To co-translationally incorporate an Uaa, it is encoded by a repurposed nonsense codon, which is suppressed by an orthogonal (i.e., does not cross-react with its host counterparts) Uaa-selective aminoacyl-tRNA synthetase (aaRS)/tRNA pair. (Chin, 2017; Dumas et al, 2015; Italia et al, 2017b; Mukai et al, 2017; Young and Schultz, 2018) Eukaryote or archaea derived aaRS/tRNA pairs are orthogonal in bacteria and used for Uaa incorporation, while those derived from bacteria are typically orthogonal in eukaryotes (Figure 1A). (Chin, 2017; Dumas et al, 2015; Italia et al, 2017b; Mukai et al, 2017; Young and Schultz, 2018) To create Uaa-specific variants of aaRS/tRNA pairs through directed evolution, two cell-based selection systems have been developed, using Escherichia coli(Santoro et al, 2002; Wang et al, 2001) or Saccharomyces cerevisiae (yeast)(Chin et al, 2003a; Chin et al, 2003b) as selection hosts, for engineering pairs that are orthogonal in bacteria or eukaryotes, respectively.…”

Expansion of the genetic code via expansion of the genetic alphabet

“…Almost 20 years ago, Peter Schultz increased the diversity available to living organisms by expanding the genetic code using the amber stop codon (UAG) to encode ncAAs in Escherichia coli [••2,••3]. This landmark accomplishment was achieved using a tRNA–amino acid tRNA synthetase (aaRS) pair from Methanococcus jannaschii, in which the tRNA was recoded to suppress the stop codon and the aaRS was evolved to charge the tRNA with an ncAA.…”

Synthesis at the interface of virology and genetic code expansion

“…[1–3] This technology has experienced remarkable progress over the last decade, facilitating its application to various domains of life. [1–3] Using this approach, over 150 different Uaas with novel sidechains have been genetically encoded to date, providing an exciting new toolset in the arsenal of synthetic biology.…”