title: The aminoglycoside resistance methyltransferase Sgm impedes RsmF methylation at an adjacent rRNA nucleotide in the ribosomal A site.
abstract: Ribosome-targeting antibiotics block protein synthesis by binding at functionally important regions of the bacterial rRNA. Resistance is often conferred by addition of a methyl group at the antibiotic binding site within an rRNA region that is already highly modified with several nucleotide methylations. In bacterial rRNA, each methylation requires its own specific methyltransferase enzyme, and this raises the question as to how an extra methyltransferase conferring antibiotic resistance can be accommodated and how it can gain access to its nucleotide target within a short and functionally crowded stretch of the rRNA sequence. Here, we show that the Sgm methyltransferase confers resistance to 4,6-disubstituted deoxystreptamine aminoglycosides by introducing the 16S rRNA modification m(7)G1405 within the ribosomal A site. This region of Escherichia coli 16S rRNA already contains several methylated nucleotides including m(4)Cm1402 and m(5)C1407. Modification at m(5)C1407 by the methyltransferase RsmF is impeded as Sgm gains access to its adjacent G1405 target on the 30S ribosomal subunit. An Sgm mutant (G135A), which is impaired in S-adenosylmethionine binding and confers lower resistance, is less able to interfere with RsmF methylation on the 30S subunit. The two methylations at 16S rRNA nucleotide m(4)Cm1402 are unaffected by both the wild-type and the mutant versions of Sgm. The data indicate that interplay between resistance methyltransferases and the cell's own indigenous methyltransferases can play an important role in determining resistance levels.
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Southern Denmark, DK-5230 Odense M, Denmark.
authors.forename: Sonja
authors.initials: S
authors.lastname: Cubrilo
authors.forename: Fedora
authors.initials: F
authors.lastname: Babić
authors.forename: Stephen
authors.initials: S
authors.lastname: Douthwaite
authors.forename: Gordana
authors.initials: G
authors.lastname: Maravić Vlahovicek
mesh_terms: D000617:Aminoglycosides / Q000494:pharmacology; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D024881:Drug Resistance, Bacterial / Q000502:physiology*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000378:metabolism; D008745:Methylation; D008780:Methyltransferases / Q000235:genetics / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D009690:Nucleic Acid Conformation; D012329:RNA, Bacterial / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012335:RNA, Ribosomal / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012336:RNA, Ribosomal, 16S / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012270:Ribosomes / Q000187:drug effects / Q000378:metabolism; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013379:Substrate Specificity
chemical_list: D000617:Aminoglycosides; D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D012329:RNA, Bacterial; D012335:RNA, Ribosomal; D012336:RNA, Ribosomal, 16S; D011994:Recombinant Proteins; C446079:16S rRNA - m7G methyltransferase; D008780:Methyltransferases; C078392:sgm protein, Micromonospora zionensis