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Specific recognition of p53 tetramers by peptides derived from p53 interacting proteins
Ronen Gabizon1, Tobias Brandt, Shahar Sukenik
1Institute of Chemistry, The Hebrew University of Jerusalem, Jerusalem, Israel.
Abstract:
Oligomerization plays a major role in regulating the activity of many proteins, and in modulating their interactions. p53 is a homotetrameric transcription factor that has a pivotal role in tumor suppression. Its tetramerization domain is contained within its C-terminal domain, which is a site for numerous protein-protein interactions. Those can either depend on or regulate p53 oligomerization. Here we screened an array of peptides derived from proteins known to bind the tetrameric p53 C-terminal domain (p53CTD) and identified ten binding peptides. We quantitatively characterized their binding to p53CTD using fluorescence anisotropy. The peptides bound tetrameric p53CTD with micromolar affinities. Despite the high charge of the binding peptides, electrostatics contributed only mildly to the interactions. NMR studies indicated that the peptides bound p53CTD at defined sites. The most significant chemical shift deviations were observed for the peptides WS100B(81-92), which bound directly to the p53 tetramerization domain, and PKCα(281-295), which stabilized p53CTD in circular dichroism thermal denaturation studies. Using analytical ultracentrifugation, we found that several of the peptides bound preferentially to p53 tetramers. Our results indicate that the protein-protein interactions of p53 are dependent on the oligomerization state of p53. We conclude that peptides may be used to regulate the oligomerization of p53.
Insights
Researchers identified peptides that bind to the p53 tetramerization domain, potentially regulating its oligomerization and protein interactions. These findings suggest peptides can modulate tumor suppressor p53 activity.
Area of Science:
- Molecular Biology
- Protein Interactions
- Biochemistry
Background:
- Protein oligomerization is crucial for regulating protein activity and interactions.
- p53 is a homotetrameric transcription factor vital for tumor suppression.
- The C-terminal domain of p53 (p53CTD) contains the tetramerization domain and mediates protein-protein interactions.
Purpose of the Study:
- To screen for peptides that bind to the tetrameric p53 C-terminal domain (p53CTD).
- To quantitatively characterize the binding of these peptides to p53CTD.
- To investigate the potential of these peptides in regulating p53 oligomerization.
Main Methods:
- Peptide screening and selection.
- Fluorescence anisotropy for quantitative binding characterization.
- Nuclear Magnetic Resonance (NMR) spectroscopy for binding site analysis.
- Circular dichroism thermal denaturation for stability studies.
- Analytical ultracentrifugation to assess binding to different p53 oligomeric states.
Main Results:
- Ten peptides were identified that bind to tetrameric p53CTD with micromolar affinities.
- Peptide binding to p53CTD was characterized, with minimal electrostatic contribution despite high peptide charge.
- NMR studies revealed defined binding sites for the peptides on p53CTD.
- Specific peptides, WS100B(81-92) and PKCα(281-295), showed direct binding and stabilization of p53CTD, respectively.
- Several peptides demonstrated preferential binding to p53 tetramers, confirmed by analytical ultracentrifugation.
Conclusions:
- Peptide binding to p53CTD is sequence-specific and influences p53 oligomerization.
- Protein-protein interactions involving p53 are dependent on its oligomeric state.
- Identified peptides offer a potential strategy for regulating p53 oligomerization and function.
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