Figure 6
VapC-mt11 cleaves tRNAs within the ASL. (A) Primer extension analysis of in vitro synthesized M. tuberculosis tRNAArg27, tRNAPro14, tRNAPro23, tRNAPro35, tRNALeu3, tRNALeu13, tRNALeu15, tRNAGln32 and tRNAGln41 treated with (+) or without (−) recombinant VapC-mt11. G, A, T, and C correspond to DNA-sequencing ladders using the same oligonucleotide as in the primer extension reactions for each tRNA. RNA sequence shown below the gels and major cleavage sites are indicated by the red arrow; alternate weak cleavage sites are indicated by yellow arrow for the three proline tRNAs. Major cleavage products in the gels are depicted by the red arrow head (right). The ASL sequence is illustrated above each tRNA sequencing gel, the anticodon (grey shading) and major cleavage position indicated by red arrow. Bands visible in (−) lanes of tRNAArg27, tRNALeu3 and tRNAGln32 correspond to secondary structure. (B) Diagram of tRNA halves produced by VapC-mt11. tRNA numbering and anticodon sequences of each numbered tRNA from the Lowe lab genomic tRNA database http://gtrnadb.ucsc.edu33. All uncropped images shown in Supplementary Information Fig. 5 (tRNAArg27, tRNAPro14, tRNAPro23, and tRNAPro35) and Supplementary Information Fig. 6 (tRNALeu3, tRNALeu13, tRNALeu15, tRNAGln32, and tRNAGln41).
