A complex attenuator regulates inducible resistance to macrolides, lincosamides, and streptogramin type B antibiotics in Streptococcus sanguis

Abstract

Macrolide-lincosamide-streptogramin B resistance specified by Streptococcus sanguis plasmid pAM77 involves an adenine methylase, whose synthesis, demonstrable both phenotypically and by analysis of methionine-labeled proteins made in Bacillus subtilis minicells, is inducible by erythromycin, lincomycin, and streptogramin type B antibiotics. Localization of the methylase structural gene, including its control region in DNA fragments obtained with restriction endonucleases, has been deduced from DNA blot experiments with characterized target and probe DNAs from other streptococci, combined with DNA sequence analysis and comparison of the putative streptococcal methylase sequence with that of a cognate methylase in staphylococcal plasmid pE194. The streptococcal methylase migrates electrophoretically in polyacrylamide gels with the mobility of a 29,000-dalton protein. The sequence organization of the putative streptococcal methylase mRNA leader sequence partially resembles its staphylococcal counterpart and can support a similar mechanism of secondary structure rearrangement leading to methylase synthesis. The deduced 5' leader sequence preceding the pAM77 methylase structural gene sequence comprises approximately 155 nucleotides within which one can identify a putative control peptide 36 amino acid residues in length (in contrast to 19 in the pE194 peptide) and at least 14 possible classes of overlapping inverted complementary repeat sequences (in contrast to 3 in the pE194 control region), one of which can sequester the sequence AGGAG 7 nucleotides upstream from the putative (methionine) start codon of the streptococcal methylase. Comparison of the pAM77 and pE194 methylase amino acid sequences and their respective nucleotide sequences shows 51% conservation of amino acid residues (124 of 244) and 59% conservation of nucleotide residues (433 of 738), which suggests a common origin for the two methylase structural gene sequences. Differences in mRNA base composition associated with conserved amino acid residues occur mostly in the third nucleotide ("wobble") position of codons and may reflect adaptation of methylase genes to optimal expression in host cells with differing codon use patterns.