Oxazolidinone Resistance Mutations in 23S rRNA of Escherichia coli Reveal the Central Region of Domain V as the Primary Site of Drug Action
- 1 October 2000
- journal article
- research article
- Published by American Society for Microbiology in Journal of Bacteriology
- Vol. 182 (19), 5325-5331
- https://doi.org/10.1128/jb.182.19.5325-5331.2000
Abstract
Oxazolidinone antibiotics inhibit bacterial protein synthesis by interacting with the large ribosomal subunit. The structure and exact location of the oxazolidinone binding site remain obscure, as does the manner in which these drugs inhibit translation. To investigate the drug-ribosome interaction, we selected Escherichia coli oxazolidinone-resistant mutants, which contained a randomly mutagenized plasmid-borne rRNA operon. The same mutation, G2032 to A, was identified in the 23S rRNA genes of several independent resistant isolates. Engineering of this mutation by site-directed mutagenesis in the wild-type rRNA operon produced an oxazolidinone resistance phenotype, establishing that the G2032A substitution was the determinant of resistance. Engineered U and C substitutions at G2032, as well as a G2447-to-U mutation, also conferred resistance to oxazolidinone. All the characterized resistance mutations were clustered in the vicinity of the central loop of domain V of 23S rRNA, suggesting that this rRNA region plays a major role in the interaction of the drug with the ribosome. Although the central loop of domain V is an essential integral component of the ribosomal peptidyl transferase, oxazolidinones do not inhibit peptide bond formation, and thus these drugs presumably interfere with another activity associated with the peptidyl transferase center.Keywords
This publication has 32 references indexed in Scilit:
- The 3D arrangement of the 23 S and 5 S rRNA in the Escherichia coli 50 S ribosomal subunit based on a cryo-electron microscopic reconstruction at 7.5 Å resolutionJournal of Molecular Biology, 2000
- Resistance mutations in 23 S rRNA identify the site of action of the protein synthesis inhibitor linezolid in the ribosomal peptidyl transferase centerJournal of Molecular Biology, 1999
- Mutations in the Peptidyl Transferase Center of 23 S rRNA Reveal the Site of Action of Sparsomycin, a Universal Inhibitor of TranslationJournal of Molecular Biology, 1996
- Pseudoknot in Domain II of 23 S rRNA is Essential for Ribosome FunctionJournal of Molecular Biology, 1995
- Specific structural probing of plasmid-coded ribosomal RNAs from Escherichia coliBiochimie, 1991
- Defining the Structural Requirements for a Helix in 23 S Ribosomal RNA that Confers Erythromycin ResistanceJournal of Molecular Biology, 1989
- The mechanism of action of DuP 721, a new antibacterial agent: Effects on macromolecular synthesisBiochemical and Biophysical Research Communications, 1988
- Improved M13 phage cloning vectors and host strains: nucleotide sequences of the M13mpl8 and pUC19 vectorsGene, 1985
- Gene organization and primary structure of a ribosomal RNA operon from Escherichia coliJournal of Molecular Biology, 1981
- Ribosome-catalysed reaction of puromycin with a formylmethionine-containing oligonucleotideJournal of Molecular Biology, 1967