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Related Experiment Videos

Evidence for tertiary structure in natural single stranded RNAs in solution.

S Ghribi1, M C Maurel, M Rougee

  • 1Institut J. Monod, Laboratoire de Photobiologie Moléculaire, Paris, France.

Nucleic Acids Research
|February 11, 1988
PubMed
Summary

Ethidium bromide binding to transfer RNA (tRNA) reveals that its three-dimensional structure restricts intercalation. This restriction is dependent on salt concentration and requires specific folding, unlike double-helical nucleic acids.

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Area of Science:

  • Molecular Biology
  • Biochemistry
  • Structural Biology

Background:

  • Ethidium bromide is a fluorescent intercalating agent used to study nucleic acid structure.
  • Transfer RNA (tRNA) possesses a complex three-dimensional (3D) structure crucial for its function.
  • The interaction of small molecules with nucleic acids can be influenced by ionic strength and structural conformation.

Purpose of the Study:

  • To investigate the binding of ethidium bromide to tRNA and other RNA molecules.
  • To determine how ionic strength and tRNA structure affect ethidium bromide intercalation.
  • To elucidate the structural requirements for intercalation restriction in single-stranded RNAs.

Main Methods:

  • Binding isotherms of ethidium bromide to various tRNA species were measured at different ionic strengths (Mg2+ and Na+).

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  • The effect of strategic modifications on tRNA structure was assessed for intercalation.
  • Intercalation restriction was examined in other RNA types, including ribosomal RNA, viral RNA, mRNA, and synthetic copolymers.

    • Main Results:

    • The number of ethidium bromide intercalation sites on tRNA decreased significantly at high salt concentrations (form I) compared to low salt conditions (form III).
    • Modifications to tRNA's 3D folding regions hindered full intercalation restriction.
    • Maximal intercalation restriction occurred at salt concentrations exceeding those required for full conversion to form I, suggesting a requirement beyond simple structural change.

    Conclusions:

    • Intercalation restriction in tRNA is dependent on its native 3D structure and requires physical coupling between folded regions and helical arms.
    • Unlike double-helical RNA or DNA, tRNA exhibits this restriction, highlighting the role of its unique tertiary structure.
    • Three-dimensional folding is a common feature of single-stranded RNAs in solution under physiological conditions, influencing their interaction with intercalating agents.