title: The aminoglycoside resistance methyltransferases from the ArmA/Rmt family operate late in the 30S ribosomal biogenesis pathway.
abstract: Bacterial resistance to 4,6-type aminoglycoside antibiotics, which target the ribosome, has been traced to the ArmA/RmtA family of rRNA methyltransferases. These plasmid-encoded enzymes transfer a methyl group from S-adenosyl-L-methionine to N7 of the buried G1405 in the aminoglycoside binding site of 16S rRNA of the 30S ribosomal subunit. ArmA methylates mature 30S subunits but not 16S rRNA, 50S, or 70S ribosomal subunits or isolated Helix 44 of the 30S subunit. To more fully characterize this family of enzymes, we have investigated the substrate requirements of ArmA and to a lesser extent its ortholog RmtA. We determined the Mg+² dependence of ArmA activity toward the 30S ribosomal subunits and found that the enzyme recognizes both low Mg+² (translationally inactive) and high Mg+² (translationally active) forms of this substrate. We tested the effects of LiCl pretreatment of the 30S subunits, initiation factor 3 (IF3), and gentamicin/kasugamycin resistance methyltransferase (KsgA) on ArmA activity and determined whether in vivo derived pre-30S ribosomal subunits are ArmA methylation substrates. ArmA failed to methylate the 30S subunits generated from LiCl washes above 0.75 M, despite the apparent retention of ribosomal proteins and a fully mature 16S rRNA. From our experiments, we conclude that ArmA is most active toward the 30S ribosomal subunits that are at or very near full maturity, but that it can also recognize more than one form of the 30S subunit.
authors.affiliation: Department of Biochemistry and Molecular Biology, Virginia Commonwealth University, Richmond, Virginia 23298, USA.
authors.forename: Tamara
authors.initials: T
authors.lastname: Zarubica
authors.forename: Matthew R
authors.initials: MR
authors.lastname: Baker
authors.forename: H Tonie
authors.initials: HT
authors.lastname: Wright
authors.forename: Jason P
authors.initials: JP
authors.lastname: Rife
mesh_terms: D040981:Acinetobacter baumannii / Q000201:enzymology / Q000378:metabolism; D000617:Aminoglycosides / Q000494:pharmacology*; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001426:Bacterial Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism; D001665:Binding Sites; D024881:Drug Resistance, Bacterial; D005839:Gentamicins / Q000494:pharmacology; D017665:Hydroxyl Radical / Q000737:chemistry / Q000378:metabolism; D008780:Methyltransferases / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D008958:Models, Molecular; D012336:RNA, Ribosomal, 16S / Q000378:metabolism; D054680:Ribosome Subunits, Small, Bacterial / Q000378:metabolism*; D013379:Substrate Specificity
chemical_list: D000617:Aminoglycosides; D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D005839:Gentamicins; D012336:RNA, Ribosomal, 16S; D017665:Hydroxyl Radical; D008780:Methyltransferases; C100305:kasugamycin