Figure 4.

In vivo methylation of m¹A2142 by episomally expressed recombinant Bmt2. To confirm the functionality of episomally expressed Bmt2 and analyse the function of Bmt2 in methylating 25S rRNA at position 2142, the plasmid pSH20 and pPK468 was transformed in the Δbmt2 strain. The 25S rRNA was isolated from the strains carrying these plasmids and processed for HPLC analysis. (A) RP–HPLC chromatogram of the Δbmt2 strain carrying pPK468 plasmid. (B) RP–HPLC chromatogram of the Δbmt2 strain harbouring pSH20 plasmid. It became apparent from the chromatograms that the recombinant Bmt2 expressed from pSH20 plasmid was able to methylate the N¹ of A2142 of 25S rRNA, as the area corresponding to the m¹A peak increases to almost double amount compared with strain carrying pPK468.