Abstract
The protein p73 acts as a transcription factor, resulting in tumour suppression. MDM2, an oncogenic protein, can negatively influence p73-mediated apoptosis by binding to p73 transactivation domains (TAD). Inhibition of the protein-protein interaction between p73 and oncogenic proteins is an attractive strategy for promoting p73-mediated apoptosis. Herein, we describe the use of a modified p73-TAD peptide for the FRET-based assay of the binding of p73-TAD to MDM2. The FRET probe, equipped with 1-naphthylamine (λex =330 nm, λem =445 nm), serves as a FRET acceptor, and the tryptophan of the protein acts as FRET donor (λex =280 nm, λem =340 nm). Sensitized emission from the FRET probe was observed upon excitation of the protein-FRET-probe complex at the excitation wavelength of Trp. Furthermore, addition of the MDM2 inhibitor Nutiln-3 drastically reduced the FRET signal, thus indicating that the FRET probe competes with Nutiln-3 for MDM2 binding. The developed FRET binding assay might be applicable in high-throughput screening of novel drugs that inhibit interactions between p73 and MDM2.
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
More From: Chembiochem : a European journal of chemical biology
Disclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.