DOMAIN-V OF 23S RIBOSOMAL-RNA CONTAINS ALL THE STRUCTURAL ELEMENTS NECESSARY FOR RECOGNITION BY THE ERME METHYLTRANSFERASE

The ErmE methyltransferase from the erythromycin-producing actinomycet e Saccharopolyspora erythraea dimethylates the N-6 position of adenine 2058 in domain V of 23S rRNA, This modification confers resistance to erythromycin and to other macrolide, lincosamide, and streptogramin B antibiotics. We investigated what structural elements in 23S rRNA are required for specific recognition by the ErmE methyltransferase. The ermE gene was cloned into R1 plasmid derivatives, providing a means of inducible expression in Escherichia coli. Expression of the methyltra nsferase in vivo confers resistance to erythromycin and clindamycin. T he degree of resistance corresponds to the level of ermE expression. I n turn, ermE expression also correlates with the proportion of 23S rRN A molecules that are dimethylated at adenine 2058. The methyltransfera se was isolated in an active, concentrated form from E. coli, and the enzyme efficiently modifies 23S rRNA in vitro. Removal of most of the 23S rRNA structure, so that only domain V (nucleotides 2000 to 2624) r emains, does not affect the efficiency of modification by the methyltr ansferase. In addition; modification still occurs after the rRNA terti ary structure has been disrupted by removal of magnesium ions. We conc lude that the main features that are specifically recognized by the Er mE methyltransferase are displayed within the primary and secondary st ructures of 23S rRNA domain V.