Binding of Amphipathic Cell Penetrating Peptide p28 to Wild Type and Mutated p53 as studied by Raman, Atomic Force and Surface Plasmon Resonance spectroscopies

• Published in: Biochimica et biophysica acta. General subjects Vol. 1861; no. 4; pp. 910 - 921
• Main Authors: Signorelli, Sara, Santini, Simona, Yamada, Tohru, Bizzarri, Anna Rita, Beattie, Craig W., Cannistraro, Salvatore
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01.04.2017
• Subjects: Amino Acid Sequence; Atomic Force Spectroscopy; binding capacity; Binding Sites - genetics; Cell Line, Tumor; Cell penetrating peptide; Cell-Penetrating Peptides - metabolism; correlation; DNA-binding domains; DNA-Binding Proteins - genetics; DNA-Binding Proteins - metabolism; Docking; Humans; Hydrophobic and Hydrophilic Interactions; hydrophobic bonding; hydrophobicity; Microscopy, Atomic Force - methods; Models, Molecular; Molecular Dynamics Simulation; mutants; Mutation - genetics; Mutations; neoplasms; p53; Peptide Fragments - genetics; Peptide Fragments - metabolism; point mutation; Protein Binding - genetics; Protein Structure, Secondary; Raman spectroscopy; surface plasmon resonance; Surface Plasmon Resonance - methods; Tumor Suppressor Protein p53 - genetics; Tumor Suppressor Protein p53 - metabolism; Mutations; Docking; Atomic Force Spectroscopy; Cell penetrating peptide; Raman spectroscopy; p53
• DOI: 10.1016/j.bbagen.2017.01.022
• ISSN: 0304-4165

Mutations within the DNA binding domain (DBD) of the tumor suppressor p53 are found in >50% of human cancers and may significantly modify p53 secondary structure impairing its function. p28, an amphipathic cell-penetrating peptide, binds to the DBD through hydrophobic interaction and induces a posttranslational increase in wildtype and mutant p53 restoring functionality. We use mutation analyses to explore which elements of secondary structure may be critical to p28 binding. Molecular modeling, Raman spectroscopy, Atomic Force Spectroscopy (AFS) and Surface Plasmon Resonance (SPR) were used to identify which secondary structure of site-directed and naturally occurring mutant DBDs are potentially altered by discrete changes in hydrophobicity and the molecular interaction with p28. We show that specific point mutations that alter hydrophobicity within non-mutable and mutable regions of the p53 DBD alter specific secondary structures. The affinity of p28 was positively correlated with the β-sheet content of a mutant DBD, and reduced by an increase in unstructured or random coil that resulted from a loss in hydrophobicity and redistribution of surface charge. These results help refine our knowledge of how mutations within p53-DBD alter secondary structure and provide insight on how potential structural alterations in p28 or similar molecules improve their ability to restore p53 function. Raman spectroscopy, AFS, SPR and computational modeling are useful approaches to characterize how mutations within the p53DBD potentially affect secondary structure and identify those structural elements prone to influence the binding affinity of agents designed to increase the functionality of p53. [Display omitted] •A structural investigation of p53 containing single point mutations is proposed.•The interaction between mutants p53 and the anticancer peptide p28 is investigated.•Structural changes in p53 are mutation position-dependent.•Alteration in p53 structure affects the binding of p28 in a position dependent way.