Skip to main content
NCBI home page
RNA logoLink to RNA
. 1999 Jul;5(7):939–946. doi: 10.1017/s1355838299990210

Ribosomal RNA is the target for oxazolidinones, a novel class of translational inhibitors.

N B Matassova 1, M V Rodnina 1, R Endermann 1, H P Kroll 1, U Pleiss 1, H Wild 1, W Wintermeyer 1
PMCID: PMC1369818  PMID: 10411137

Abstract

Oxazolidinones are antibacterial agents that act primarily against gram-positive bacteria by inhibiting protein synthesis. The binding of oxazolidinones to 70S ribosomes from Escherichia coli was studied by both UV-induced cross-linking using an azido derivative of oxazolidinone and chemical footprinting using dimethyl sulphate. Oxazolidinone binding sites were found on both 30S and 50S subunits, rRNA being the only target. On 16S rRNA, an oxazolidinone footprint was found at A864 in the central domain. 23S rRNA residues involved in oxazolidinone binding were U2113, A2114, U2118, A2119, and C2153, all in domain V. This region is close to the binding site of protein L1 and of the 3' end of tRNA in the E site. The mechanism of action of oxazolidinones in vitro was examined in a purified translation system from E. coli using natural mRNA. The rate of elongation reaction of translation was decreased, most probably because of an inhibition of tRNA translocation, and the length of nascent peptide chains was strongly reduced. Both binding sites and mode of action of oxazolidinones are unique among the antibiotics known to act on the ribosome.

Full Text

The Full Text of this article is available as a PDF (379.3 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Allmang C., Mougel M., Westhof E., Ehresmann B., Ehresmann C. Role of conserved nucleotides in building the 16S rRNA binding site of E. coli ribosomal protein S8. Nucleic Acids Res. 1994 Sep 11;22(18):3708–3714. doi: 10.1093/nar/22.18.3708. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bogdanov A. A., Chichkova N. V., Kopylov A. M., Mankin A. S., Skripkin E. A. Surface topography of ribosomal RNA. Methods Enzymol. 1988;164:440–456. doi: 10.1016/s0076-6879(88)64060-2. [DOI] [PubMed] [Google Scholar]
  3. Borowski C., Rodnina M. V., Wintermeyer W. Truncated elongation factor G lacking the G domain promotes translocation of the 3' end but not of the anticodon domain of peptidyl-tRNA. Proc Natl Acad Sci U S A. 1996 Apr 30;93(9):4202–4206. doi: 10.1073/pnas.93.9.4202. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Brickner S. J., Hutchinson D. K., Barbachyn M. R., Manninen P. R., Ulanowicz D. A., Garmon S. A., Grega K. C., Hendges S. K., Toops D. S., Ford C. W. Synthesis and antibacterial activity of U-100592 and U-100766, two oxazolidinone antibacterial agents for the potential treatment of multidrug-resistant gram-positive bacterial infections. J Med Chem. 1996 Feb 2;39(3):673–679. doi: 10.1021/jm9509556. [DOI] [PubMed] [Google Scholar]
  5. Calogero R. A., Pon C. L., Canonaco M. A., Gualerzi C. O. Selection of the mRNA translation initiation region by Escherichia coli ribosomes. Proc Natl Acad Sci U S A. 1988 Sep;85(17):6427–6431. doi: 10.1073/pnas.85.17.6427. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Daly J. S., Eliopoulos G. M., Willey S., Moellering R. C., Jr Mechanism of action and in vitro and in vivo activities of S-6123, a new oxazolidinone compound. Antimicrob Agents Chemother. 1988 Sep;32(9):1341–1346. doi: 10.1128/aac.32.9.1341. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Douthwaite S. Interaction of the antibiotics clindamycin and lincomycin with Escherichia coli 23S ribosomal RNA. Nucleic Acids Res. 1992 Sep 25;20(18):4717–4720. doi: 10.1093/nar/20.18.4717. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Ehresmann C., Moine H., Mougel M., Dondon J., Grunberg-Manago M., Ebel J. P., Ehresmann B. Cross-linking of initiation factor IF3 to Escherichia coli 30S ribosomal subunit by trans-diamminedichloroplatinum(II): characterization of two cross-linking sites in 16S rRNA; a possible way of functioning for IF3. Nucleic Acids Res. 1986 Jun 25;14(12):4803–4821. doi: 10.1093/nar/14.12.4803. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Eustice D. C., Feldman P. A., Slee A. M. The mechanism of action of DuP 721, a new antibacterial agent: effects on macromolecular synthesis. Biochem Biophys Res Commun. 1988 Feb 15;150(3):965–971. doi: 10.1016/0006-291x(88)90723-1. [DOI] [PubMed] [Google Scholar]
  10. Eustice D. C., Feldman P. A., Zajac I., Slee A. M. Mechanism of action of DuP 721: inhibition of an early event during initiation of protein synthesis. Antimicrob Agents Chemother. 1988 Aug;32(8):1218–1222. doi: 10.1128/aac.32.8.1218. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Ford C. W., Hamel J. C., Wilson D. M., Moerman J. K., Stapert D., Yancey R. J., Jr, Hutchinson D. K., Barbachyn M. R., Brickner S. J. In vivo activities of U-100592 and U-100766, novel oxazolidinone antimicrobial agents, against experimental bacterial infections. Antimicrob Agents Chemother. 1996 Jun;40(6):1508–1513. doi: 10.1128/aac.40.6.1508. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Kalurachchi K., Uma K., Zimmermann R. A., Nikonowicz E. P. Structural features of the binding site for ribosomal protein S8 in Escherichia coli 16S rRNA defined using NMR spectroscopy. Proc Natl Acad Sci U S A. 1997 Mar 18;94(6):2139–2144. doi: 10.1073/pnas.94.6.2139. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Kruse T. A., Siboska G. E., Clark B. F. Photosensitized crosslinking of tRNA to the P-, A- and R-sites of Escherichia coli ribosomes. Biochimie. 1982 Apr;64(4):279–284. doi: 10.1016/s0300-9084(82)80495-1. [DOI] [PubMed] [Google Scholar]
  14. Lill R., Robertson J. M., Wintermeyer W. Binding of the 3' terminus of tRNA to 23S rRNA in the ribosomal exit site actively promotes translocation. EMBO J. 1989 Dec 1;8(12):3933–3938. doi: 10.1002/j.1460-2075.1989.tb08574.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Lin A. H., Murray R. W., Vidmar T. J., Marotti K. R. The oxazolidinone eperezolid binds to the 50S ribosomal subunit and competes with binding of chloramphenicol and lincomycin. Antimicrob Agents Chemother. 1997 Oct;41(10):2127–2131. doi: 10.1128/aac.41.10.2127. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Moazed D., Noller H. F. Chloramphenicol, erythromycin, carbomycin and vernamycin B protect overlapping sites in the peptidyl transferase region of 23S ribosomal RNA. Biochimie. 1987 Aug;69(8):879–884. doi: 10.1016/0300-9084(87)90215-x. [DOI] [PubMed] [Google Scholar]
  17. Moazed D., Noller H. F. Interaction of tRNA with 23S rRNA in the ribosomal A, P, and E sites. Cell. 1989 May 19;57(4):585–597. doi: 10.1016/0092-8674(89)90128-1. [DOI] [PubMed] [Google Scholar]
  18. Mougel M., Eyermann F., Westhof E., Romby P., Expert-Bezançon A., Ebel J. P., Ehresmann B., Ehresmann C. Binding of Escherichia coli ribosomal protein S8 to 16 S rRNA. A model for the interaction and the tertiary structure of the RNA binding site. J Mol Biol. 1987 Nov 5;198(1):91–107. doi: 10.1016/0022-2836(87)90460-8. [DOI] [PubMed] [Google Scholar]
  19. Mueller F., Brimacombe R. A new model for the three-dimensional folding of Escherichia coli 16 S ribosomal RNA. I. Fitting the RNA to a 3D electron microscopic map at 20 A. J Mol Biol. 1997 Aug 29;271(4):524–544. doi: 10.1006/jmbi.1997.1210. [DOI] [PubMed] [Google Scholar]
  20. Mueller F., Brimacombe R. A new model for the three-dimensional folding of Escherichia coli 16 S ribosomal RNA. II. The RNA-protein interaction data. J Mol Biol. 1997 Aug 29;271(4):545–565. doi: 10.1006/jmbi.1997.1211. [DOI] [PubMed] [Google Scholar]
  21. Powers T., Noller H. F. Hydroxyl radical footprinting of ribosomal proteins on 16S rRNA. RNA. 1995 Apr;1(2):194–209. [PMC free article] [PubMed] [Google Scholar]
  22. Rodnina M. V., Fricke R., Wintermeyer W. Transient conformational states of aminoacyl-tRNA during ribosome binding catalyzed by elongation factor Tu. Biochemistry. 1994 Oct 11;33(40):12267–12275. doi: 10.1021/bi00206a033. [DOI] [PubMed] [Google Scholar]
  23. Rodnina M. V., Semenkov Y. P., Wintermeyer W. Purification of fMet-tRNA(fMet) by fast protein liquid chromatography. Anal Biochem. 1994 Jun;219(2):380–381. doi: 10.1006/abio.1994.1282. [DOI] [PubMed] [Google Scholar]
  24. Rodnina M. V., Wintermeyer W. GTP consumption of elongation factor Tu during translation of heteropolymeric mRNAs. Proc Natl Acad Sci U S A. 1995 Mar 14;92(6):1945–1949. doi: 10.1073/pnas.92.6.1945. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Said B., Cole J. R., Nomura M. Mutational analysis of the L1 binding site of 23S rRNA in Escherichia coli. Nucleic Acids Res. 1988 Nov 25;16(22):10529–10545. doi: 10.1093/nar/16.22.10529. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Shinabarger D. L., Marotti K. R., Murray R. W., Lin A. H., Melchior E. P., Swaney S. M., Dunyak D. S., Demyan W. F., Buysse J. M. Mechanism of action of oxazolidinones: effects of linezolid and eperezolid on translation reactions. Antimicrob Agents Chemother. 1997 Oct;41(10):2132–2136. doi: 10.1128/aac.41.10.2132. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Slee A. M., Wuonola M. A., McRipley R. J., Zajac I., Zawada M. J., Bartholomew P. T., Gregory W. A., Forbes M. Oxazolidinones, a new class of synthetic antibacterial agents: in vitro and in vivo activities of DuP 105 and DuP 721. Antimicrob Agents Chemother. 1987 Nov;31(11):1791–1797. doi: 10.1128/aac.31.11.1791. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Stade K., Rinke-Appel J., Brimacombe R. Site-directed cross-linking of mRNA analogues to the Escherichia coli ribosome; identification of 30S ribosomal components that can be cross-linked to the mRNA at various points 5' with respect to the decoding site. Nucleic Acids Res. 1989 Dec 11;17(23):9889–9908. doi: 10.1093/nar/17.23.9889. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Stern S., Moazed D., Noller H. F. Structural analysis of RNA using chemical and enzymatic probing monitored by primer extension. Methods Enzymol. 1988;164:481–489. doi: 10.1016/s0076-6879(88)64064-x. [DOI] [PubMed] [Google Scholar]
  30. Wollenzien P. L. Isolation and identification of RNA cross-links. Methods Enzymol. 1988;164:319–329. doi: 10.1016/s0076-6879(88)64052-3. [DOI] [PubMed] [Google Scholar]

Articles from RNA are provided here courtesy of The RNA Society

RESOURCES