Skip to main content
NCBI home page
Antimicrobial Agents and Chemotherapy logoLink to Antimicrobial Agents and Chemotherapy
. 1986 Apr;29(4):681–686. doi: 10.1128/aac.29.4.681

Effects of toluene permeabilization and cell deenergization on tetracycline resistance in Escherichia coli.

L M McMurry, M Hendricks, S B Levy
PMCID: PMC180466  PMID: 3010853

Abstract

Resistance to tetracycline (Tcr) mediated by Tn10 and related Tcr determinants involves an inner membrane protein, TET (similar but not identical for different determinants), and a proton motive force-dependent efflux of tetracycline which keeps the drug away from its intracellular target, the ribosome (L. M. McMurry, R. E. Petrucci, Jr., and S. B. Levy, Proc. Natl. Acad. Sci. USA 77:3974-3977, 1980). However, the amount of tetracycline accumulated by bacteria does not always correlate with their resistance levels, suggesting that an additional resistance mechanism may be present. When we permeabilized susceptible and resistant Tn10-bearing cells with toluene, we found that protein synthesis in the two strains became equally sensitive to tetracycline. Therefore, the protein synthesis machinery was not a source of resistance, and an intact membrane was required for resistance. To determine whether resistance was entirely dependent on energy, we measured susceptibility to tetracycline after inhibition of proton motive force by starvation and specific inhibitors. An 80 to 90% loss of Tcr (measured by protein synthesis) resulted from partial deenergization of resistant cells. A remaining resistance (10- to 20-fold greater than that of susceptible cells) could not be eliminated by further deenergization. These findings indicated that, to a major extent, expression of Tn10 resistance required energy, presumably for tetracycline efflux. They also suggested the existence of a small component of Tcr having little or no energy dependence. Whether this component depends on tetracycline efflux or some other mechanism is not known, but presumably both high- and low-energy components of resistance reflect activity of TET protein.

Full text

PDF
681

Selected References

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

  1. Berger E. A. Different mechanisms of energy coupling for the active transport of proline and glutamine in Escherichia coli. Proc Natl Acad Sci U S A. 1973 May;70(5):1514–1518. doi: 10.1073/pnas.70.5.1514. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Cox G. B., Gibson F. Studies on electron transport and energy-linked reactions using mutants of Escherichia coli. Biochim Biophys Acta. 1974 Apr 30;346(1):1–25. doi: 10.1016/0304-4173(74)90010-x. [DOI] [PubMed] [Google Scholar]
  3. Curiale M. S., McMurry L. M., Levy S. B. Intracistronic complementation of the tetracycline resistance membrane protein of Tn10. J Bacteriol. 1984 Jan;157(1):211–217. doi: 10.1128/jb.157.1.211-217.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Deutscher M. P. Preparation of cells permeable to macromolecules by treatment with toluene: studies of transfer ribonucleic acid nucleotidyltransferase. J Bacteriol. 1974 May;118(2):633–639. doi: 10.1128/jb.118.2.633-639.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Halegoua S., Hirashima A., Inouye M. Puromycin-resistant biosynthesis of a specific outer-membrane lipoprotein of Escherichia coli. J Bacteriol. 1976 Apr;126(1):183–191. doi: 10.1128/jb.126.1.183-191.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Hillen W., Klock G., Kaffenberger I., Wray L. V., Reznikoff W. S. Purification of the TET repressor and TET operator from the transposon Tn10 and characterization of their interaction. J Biol Chem. 1982 Jun 10;257(11):6605–6613. [PubMed] [Google Scholar]
  7. Hillen W., Schollmeier K. Nucleotide sequence of the Tn10 encoded tetracycline resistance gene. Nucleic Acids Res. 1983 Jan 25;11(2):525–539. doi: 10.1093/nar/11.2.525. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Jorgensen R. A., Reznikoff W. S. Organization of structural and regulatory genes that mediate tetracycline resistance in transposon Tn10. J Bacteriol. 1979 Jun;138(3):705–714. doi: 10.1128/jb.138.3.705-714.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Kashket E. R., Barker S. L. Effects of potassium ions on the electrical and pH gradients across the membrane of Streptococcus lactis cells. J Bacteriol. 1977 Jun;130(3):1017–1023. doi: 10.1128/jb.130.3.1017-1023.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Levy S. B., McMurry L. Detection of an inducible membrane protein associated with R-factor-mediated tetracycline resistance. Biochem Biophys Res Commun. 1974 Feb 27;56(4):1060–1068. doi: 10.1016/s0006-291x(74)80296-2. [DOI] [PubMed] [Google Scholar]
  11. McMurry L. M., Aronson D. A., Levy S. B. Susceptible Escherichia coli cells can actively excrete tetracyclines. Antimicrob Agents Chemother. 1983 Oct;24(4):544–551. doi: 10.1128/aac.24.4.544. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. McMurry L. M., Cullinane J. C., Levy S. B. Transport of the lipophilic analog minocycline differs from that of tetracycline in susceptible and resistant Escherichia coli strains. Antimicrob Agents Chemother. 1982 Nov;22(5):791–799. doi: 10.1128/aac.22.5.791. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. McMurry L., Levy S. B. Two transport systems for tetracycline in sensitive Escherichia coli: critical role for an initial rapid uptake system insensitive to energy inhibitors. Antimicrob Agents Chemother. 1978 Aug;14(2):201–209. doi: 10.1128/aac.14.2.201. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. McMurry L., Petrucci R. E., Jr, Levy S. B. Active efflux of tetracycline encoded by four genetically different tetracycline resistance determinants in Escherichia coli. Proc Natl Acad Sci U S A. 1980 Jul;77(7):3974–3977. doi: 10.1073/pnas.77.7.3974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Moyed H. S., Bertrand K. P. Mutations in multicopy Tn10 tet plasmids that confer resistance to inhibitory effects of inducers of tet gene expression. J Bacteriol. 1983 Aug;155(2):557–564. doi: 10.1128/jb.155.2.557-564.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Tritton T. R. Ribosome-tetracycline interactions. Biochemistry. 1977 Sep 6;16(18):4133–4138. doi: 10.1021/bi00637a029. [DOI] [PubMed] [Google Scholar]
  17. Wray L. V., Jr, Jorgensen R. A., Reznikoff W. S. Identification of the tetracycline resistance promoter and repressor in transposon Tn10. J Bacteriol. 1981 Aug;147(2):297–304. doi: 10.1128/jb.147.2.297-304.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. de Smet M. J., Kingma J., Witholt B. The effect of toluene on the structure and permeability of the outer and cytoplasmic membranes of Escherichia coli. Biochim Biophys Acta. 1978 Jan 4;506(1):64–80. doi: 10.1016/0005-2736(78)90435-2. [DOI] [PubMed] [Google Scholar]

Articles from Antimicrobial Agents and Chemotherapy are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES