Figure 6.
RNA cleavage assay using the non-canonical hBH motif. (A) The predicted secondary structure of the substrate mini-helix RNA, mimicking a tRNA intron with an hBH motif located at the non-canonical position 59/60 in ARMAN-2 pre-tRNACys (GCA) (12). Intron cleavage activity of the mini-helix RNA by AFU EndA, APE EndA and fMKA EndA at 70°C for 15 min (B), at 70°C for 30 min (C), and at 70°C for 1 h. (D) Reaction mixtures were separated on 15% polyacrylamide/7 M urea gels. In each gel, ‘M’ (first lane) and ‘N’ (second lane) indicate the molecular markers of fragmented RNA and control (no enzyme), respectively. The cleavage products are shown using schematic models at the right-hand side of the gel.