Interaction of the anticancer p28 peptide with p53-DBD as studied by fluorescence, FRET, docking and MD simulations

Abstract

BackgroundThe p28 peptide, derived from the blue copper protein Azurin, exerts an anticancer action due to interaction with the tumor suppressor p53, likely interfering with its down-regulators. Knowledge of both the kinetics and topological details of the interaction, could greatly help to understand the peptide anticancer mechanism. MethodsFluorescence and Förster resonance energy transfer (FRET) were used to determine both the binding affinity and the distance between the lone tryptophan (FRET donor) of DNA Binding Domain (DBD) of p53 and the Iaedens dye (FRET acceptor) bound to the p28 peptide. Docking, Molecular Dynamic simulations and free energy binding calculations were used to single out the best complex model, compatible with the distance measured by FRET. ResultsTryptophan fluorescence quenching provided a 105 M−1 binding affinity for the complex. Both FRET donor fluorescence quenching and acceptor enhancement are consistent with a donor-acceptor distance of about 2.6 nm. Docking and molecular dynamics simulations allowed us to select the best complex, enlightening the contact regions between p28 and DBD. Conclusionsp28 binds to DBD partially engaging the L1 loop, at the same region of the p53 down-regulator COP1, leaving however the DNA binding site available for functional interactions. General significanceElucidation of the DBD-p28 complex gets insights into the functional role of p28 in regulating the p53 anticancer activity, also offering new perspectives to design new drugs able to protect the p53 anticancer function.