Adoption of a Turn Conformation Drives the Binding Affinity of p53 C-Terminal Domain Peptides to 14-3-3σ

The interaction between the adapter protein 14-3-3σ and transcription factor p53 is important for preserving the tumor-suppressor functions of p53 in the cell. A phosphorylated motif within the C-terminal domain (CTD) of p53 is key for binding to the amphipathic groove of 14-3-3. This motif is unique among 14-3-3 binding partners, and the precise dynamics of the interaction is not yet fully understood. Here, we investigate this interaction at the molecular level by analyzing the binding of different length p53 CTD peptides to 14-3-3σ using ITC, SPR, NMR, and MD simulations. We observed that the propensity of the p53 peptide to adopt turn-like conformation plays an important role in the binding to the 14-3-3σ protein. Our study contributes to elucidate the molecular mechanism of the 14-3-3-p53 binding and provides useful insight into how conformation properties of a ligand influence protein binding

Design of a Chemical Probe for the Bromodomain and Plant Homeodomain Finger-Containing (BRPF) Family of Proteins

The bromodomain and plant homeodomain finger-containing (BRPF) family are scaffolding proteins important for the recruitment of histone acetyltransferases of the MYST family to chromatin. Here, we describe <b>NI-57</b> (<b>16</b>) as new pan-BRPF chemical probe of the bromodomain (BRD) of the BRPFs. Inhibitor <b>16</b> preferentially bound the BRD of BRPF1 and BRPF2 over BRPF3, whereas binding to BRD9 was weaker. Compound <b>16</b> has excellent selectivity over nonclass IV BRD proteins. Target engagement of BRPF1B and BRPF2 with <b>16</b> was demonstrated in nanoBRET and FRAP assays. The binding of <b>16</b> to BRPF1B was rationalized through an X-ray cocrystal structure determination, which showed a flipped binding orientation when compared to previous structures. We report studies that show <b>16</b> has functional activity in cellular assays by modulation of the phenotype at low micromolar concentrations in both cancer and inflammatory models. Pharmacokinetic data for <b>16</b> was generated in mouse with single dose administration showing favorable oral bioavailabilit