The genetic code is the recipe for life, and provides the instructions for how to make proteins, generally using just 20 amino acids. But certain groups of microbes have an expanded genetic code, in which one or two additional amino acids are inserted into the protein—a finding that has been leveraged for bioengineering.

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Numerous genetic codes developed during the evolution of Eukaryotes and three are known in Bacteria, yet no alternative genetic code has been established for Archaea. Some bacterial and archaeal proteins include selenocysteine or pyrrolysine, the 21 st and 22 nd amino acids, but no evidence establishes the adoption of a genetic code in which a stop codon universally encodes either amino acid. Here, we used proteomics to confirm the prediction that certain Archaea consistently incorporate pyrrolysine at TAG codons, supporting a new archaeal genetic code which we designate Genetic Code 34. This genetic code has 62 sense codons encoding 21 amino acids, and only two stop codons. In contrast with monophyletic genetic code distributions in bacteria, Code 34 occurs sporadically. This, combined with evidence for lateral gene transfer of the code change machinery and anticipated barriers to code reversal, suggests Code 34 arose independently in multiple lineages. TAG codon distribution patterns in Code 34 genomes imply a wide range in time since code switch. We identified many new enzymes containing Pyl residues, raising questions about potential roles of this amino acid in protein structure and function. We used five new PylRS/tRNA Pyl pairs from Code 34 archaea to introduce new-to-nature pyrrolysine analogs into proteins in E. coli , demonstrating their utility for genetic code expansion.

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Multiple genetic codes developed during the evolution of eukaryotes and bacteria, yet no alternative genetic code is known for archaea. We used proteomics to confirm our prediction that certain archae...

Multiple genetic codes developed during the evolution of eukaryotes and bacteria, yet no alternative genetic code is known for archaea. We used proteomics to confirm our prediction that certain archaea consistently incorporate pyrrolysine (Pyl) at TAG ...

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Application to the UW-Madison Evolution Seminar Series - Early Career Scientist Awards.