Fig. 3: ava2C promotes AUA decoding in translation.
From: A tRNA modification with aminovaleramide facilitates AUA decoding in protein synthesis
a, Schematic representation of the enzymatic synthesis of E. coli tRNAIle2 transcripts bearing t6A37 and L34 or ava2C34. b, In vitro isoleucylation of tRNA transcripts bearing C34 (circles), L34 (rectangles) or ava2C34 (triangles). c, Schematic depiction of tRNA binding to the A-site of the E. coli ribosome. The P-site was occupied by E. coli tRNAGlu, followed by incubation with 32P-labeled tRNA transcripts with different modification statuses. d, Ribosome-binding ability of E. coli tRNAIle2 transcripts bearing C34, L34 or ava2C34 to examine decoding of the AUA (filled bars) or AUG (blank bars) codon at the A-site. The background signal measured in the same reaction without mRNA was subtracted. Data are shown as mean ± s.d. (n = 3 technical replicates). e, Constructs of dual reporters to measure the decoding efficiency of Ile codons (AUA or AUC) in the V. cholerae strain. Ile codons (AUA or AUC) were tandemly inserted at the N-terminus of GFP. GFP signals normalized by mCherry signals reflect the translation efficiency of Ile codons. f, Dual reporter assay evaluating the decoding efficiency of AUC or AUA codons in the V. cholerae strain cultured with 0.2% or 0.02% arabinose. Relative values of GFP/mCherry are shown as mean ± s.d. (n = 3 biological replicates). A multiple two-tailed t test was used for a statistical test. Comparisons are made between arabinose 0.2% and 0.02%. P = 0.44 (AUC reporter) and P = 0.000078 (AUA reporter) after adjustment by the Holm test.
