Raman active diyne-girder conformationally constrained p53 stapled peptides bind to MDM2 for visualisation without...

Area of Science:

• Medicinal Chemistry
• Biochemistry
• Molecular Biology

Background:

• Peptide stapling stabilizes alpha-helical peptides, improving their conformation and properties.
• The p53-MDM2/MDMX interaction is a key target for cancer therapeutics.

Purpose of the Study:

• To apply a novel diyne-girder stapling method to stabilize the p53 transactivation domain's binding region for MDM2/MDMX.
• To evaluate the resulting stapled peptide's affinity, selectivity, and potential for cellular imaging.

Main Methods:

• Incorporation of an unnatural amino acid to create an i, i+7 bridge length for peptide stapling.
• Circular dichroism spectroscopy to confirm alpha-helicity.
• Fluorescence polarization assays to assess binding affinity to MDM2 and MDMX.
• Molecular modeling and docking to investigate binding interactions.
• Raman spectroscopy for cellular visualization.

Main Results:

• A highly alpha-helical stapled peptide (peptide 4) was successfully synthesized using the diyne-girder approach.
• Peptide 4 demonstrated nanomolar binding affinity for MDM2 with significant selectivity over MDMX (>100-fold).
• Molecular modeling indicated that the diyne-girder's constrained geometry influences selectivity by altering hydrophobic interactions.
• Exploration of the diyne-girder for cellular visualization via Raman spectroscopy in the cell-silent region was successful.

Conclusions:

• The diyne-girder stapling strategy effectively stabilizes alpha-helical peptides, enhancing binding affinity and selectivity.
• Peptide 4 is a potent and selective inhibitor of MDM2, with potential as a therapeutic agent.
• The diyne-girder offers a unique platform for cellular tracking of stapled peptides using Raman spectroscopy without exogenous labels.