The tylosin resistance gene tlrB of Streptomyces fradiae encodes a methyltransferase that targets G748 in 23S rRNA

tlrB is one of four resistance genes encoded in the operon for biosynthesis of the macrolide tylosin in antibiotic-producing strains of Streptomyces f radiae. Introduction of tlrB into Streptomyces lividans similarly confers t ylosin resistance. Biochemical analysis of the rRNA from the two Streptomyc es species indicates that in vivo TlrB modifies nucleotide G748 within heli x 35 of 23S rRNA. Purified recombinant TlrB retains its activity and specif icity in vitro and modifies G748 in 23S rRNA as well as in a 74 nucleotide RNA containing helix 35 and surrounding structures. Modification is depende nt on the presence of the methyl group donor, S-adenosyl methionine. Analys is of the 74-mer RNA substrate by biochemical and mass spectrometric method s shows that TlrB adds a single methyl group to the base of G748. Homologue s of TlrB in other bacteria have been revealed through database searches, i ndicating that TlrB is the first member to be described in a new subclass o f rRNA methyltransferases that are implicated in macrolide drug resistance.