The bacterial tRNA modifying enzyme tRNAIle2 lysidine synthetase is genetically conserved but catalytically variable

The AUA isoleucine codon is generally rare and used with varying frequency in bacterial genomes. The tRNAIle2 responsible for decoding this trinucleotide must be modified at the wobble position by tRNAIle2 lysidine synthetase (TilS) prior to aminoacylation and accommodation at the ribosome. To test the hypothesis that TilS catalytic efficiency correlates with AUA frequency, we cloned tilS genes from bacteria with varying AUA codon usage. A previous study identified mutations in the Burkholderia cenocepacia tilS gene affecting locations distal to the catalytic domain that caused large fitness increases by enabling earlier exponential growth in minimal growth media. We made comparable mutations in TilS orthologs to better understand the effect of amino acid substitutions at these locations. While the tilS gene is present in nearly all bacteria, orthologs exhibit notable differences in substrate recognition and catalytic efficiency that are not readily correlated with codon usage.