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Purification of Ubiquitinated p53 Proteins from Mammalian Cells
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The electrostatic surface of MDM2 modulates the specificity of its interaction with phosphorylated and

Christopher John Brown1, Deepa Srinivasan, Lee Hui Jun

  • 1Laboratory of Cell Cycle Control, Institute of Molecular and Cell Biology, Singapore.

Cell Cycle (Georgetown, Tex.)
|February 8, 2008
PubMed

Insights

Phosphorylation of p53 at Thr18 enhances its binding to MDM2, a process influenced by MDM2

Area of Science:

  • Biochemistry
  • Molecular Biology
  • Protein-protein interactions

Background:

  • The interaction between tumor suppressor p53 and its negative regulator MDM2 is crucial for cell cycle control.
  • Post-translational modifications of p53, such as phosphorylation, can significantly alter its binding affinity to MDM2.

Purpose of the Study:

  • To investigate the role of p53 phosphorylation at Thr18 in its binding to MDM2.
  • To explore the influence of MDM2's electrostatic properties on this interaction.
  • To validate computational predictions regarding the impact of MDM2 surface mutations on p53 binding.

Main Methods:

  • Fluorescence anisotropy measurements using FAM-labelled p53 peptides.
  • Site-directed mutagenesis of key residues on the MDM2 surface.

Main Results:

  • p53 peptide binding to MDM2 was dependent on phosphorylation at Thr18.
  • The electrostatic properties of MDM2 modulated the binding affinity.
  • Mutating three key residues on MDM2 enhanced binding to phosphorylated p53 peptides, consistent with computational models.

Conclusions:

  • Phosphorylation of p53 at Thr18 is a critical determinant for its interaction with MDM2.
  • MDM2's surface electrostatics play a significant role in modulating this interaction.
  • Targeting specific MDM2 residues could offer a strategy to enhance p53-MDM2 binding.

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