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Related Experiment Videos

Binding interaction between Tet(M) and the ribosome: requirements for binding

K A Dantley1, H K Dannelly, V Burdett

  • 1Department of Microbiology, Duke University Medical Center, Durham, North Carolina 27710, USA.

Journal of Bacteriology
|August 8, 1998
PubMed
Summary

The Tet(M) protein confers tetracycline resistance by interacting with ribosomes and releasing the antibiotic. This interaction requires GTP and overlaps with the binding site of elongation factor G (EF-G).

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Area of Science:

  • Molecular Biology
  • Biochemistry
  • Microbiology

Background:

  • Tetracycline resistance is often mediated by proteins like Tet(M).
  • The Tet(M) protein's mechanism involves interaction with the ribosome and GTP hydrolysis.
  • Understanding this interaction is key to deciphering antibiotic resistance.

Purpose of the Study:

  • To elucidate the molecular mechanism of Tet(M)-mediated tetracycline resistance.
  • To investigate the interaction between Tet(M) protein and the ribosome.
  • To identify the specific conditions and factors influencing Tet(M)-ribosome complex formation.

Main Methods:

  • Gel filtration assay using radioactively labeled Tet(M) protein.
  • Investigation of Tet(M) binding to ribosomes in the presence of different guanine nucleotides (GTP, GDP, 5'-guanylyl imido diphosphate).

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  • Competition assays with thiostrepton and elongation factors (EF-G, EF-Tu).

    • Main Results:

    • Tet(M)-ribosome complex formation is promoted by GTP and 5'-guanylyl imido diphosphate, but not GDP.
    • Thiostrepton inhibits stable Tet(M)-ribosome complex formation.
    • Tet(M) and elongation factor G (EF-G) bind to overlapping sites on the ribosome.

    Conclusions:

    • Tet(M) interaction with the ribosome is GTP-dependent.
    • The binding site of Tet(M) on the ribosome overlaps with that of EF-G, suggesting a competitive or mutually exclusive interaction.
    • This study clarifies the mechanism of Tet(M) action in conferring tetracycline resistance at a molecular level.