Does azurin bind to the transactivation domain of p53? A Trp phosphorescence study

The bacterial redox protein azurin has been shown to be able to enter into cancer cells and induce apoptosis by stabilizing p53. Although the formation of a complex between the two proteins has been demonstrated, little is known about their binding features. We investigated the interaction between the transcription activation domain of p53 (p53(1-63)) and Pseudomonas aeruginosa azurin using fluorescence and phosphorescence spectroscopic techniques. Trp phosphorescence lifetime measurements revealed conformational changes in azurin induced by the interaction with p53(1-63). Acrylamide quenching of Trp phosphorescence also indicated a significant increase in the overall flexibility of azurin upon binding to p53(1-63). We show that azurin binds to the N-terminal region of p53 with a dissociation constant in the 5–10 μM range. No change in the fluorescence and phosphorescence emission of p53(1-63) was detected in the presence of azurin. This result indicated that no Trp residue of p53(1-63) is located in the interaction site with azurin and therefore suggested that the azurin binding site does not overlap that of MDM2, the protein that plays a crucial role in the p53 regulation. The present results may assist in the design of novel cancer treatments based on p53 stabilization by azurin. [Display omitted] ► N-transactivation domain of p53 interacts with azurin by inducing structural changes. ► The estimated dissociation constant of the complex ranges from 5 to 10 μM. ► The binding site of azurin on p53 seems to differ from that of MDM2.