Vivat, V. | bioRxiv Science

bioRxiv2026

Thermodynamic Insights into the WT and Y220C TP53 DBDs Reveals that the Oncogenic Y220C Variant is a Loss of Function Mutation for Zn2+-binding at Physiological Temperature

We present evidence of previously unrecognized allosteric connectivity across the TP53 DNA binding domain (DBD). Specifically, we have found evidence of explicit influence on the Zn2+-binding site from the region surrounding the hotspot Y220C mutation. This allosteric connectivity is intertwined with a temperature-dependent destabilization of Zn2+ binding in both the WT and Y220C DBDs. Our studies indicate that the Y220C mutation exacerbates this temperature-dependent destabilization of Zn2+-binding to result in overall destabilization of the Y220C variant. We provide detailed thermodynamic evidence that Rezatapopt, a small molecule reactivator of the Y220C DBD, engages Y220C through two distinct thermodynamic pathways and restores WT-level Zn2+-affinity to this oncogenic variant. A series of thermodynamic models describing the WT and Y220C conformational landscapes, as well as the Rezatapopt binding mechanisms, are proposed.

biochemistry↗

bioRxiv2018

Interplay of protein disorder in retinoic acid receptor heterodimer and its corepressor regulates gene expression

The retinoic acid receptors (RARs) form heterodimers with retinoid X receptors (RXRs) and control gene transcription in response to ligand binding and via allosteric activation of the C-termini helix (helix H12) of its ligand-binding domain. Herein we show that in the absence of ligand, helices H12 of RXR and RAR are disordered. The selective RAR agonist, Am580, induces folding of H12, whereas in the presence of the inverse agonist BMS493, H12 stays mostly disordered. These results substantiate a link between the structural dynamics of H12 and RXR/RAR heterodimer biological functions, and highlight disordered-to-order transition as an essential mechanism for retinoic acid mediated regulation. Unliganded RAR exerts a strong repressive activity allowed by the recruitment of transcriptional corepressors and establishment of a corepressor complex in the promoter region of target genes. The human regulatory complex of the RAR bound to the full-length interaction domain of the corepressor N-CoR was studied by integrating several experimental (SAXS, X-ray crystallography, NMR, CD, AUC) and computational data. Unexpectedly, we found that, while mainly intrinsically disordered, the N-CoR presents partially evolutionary conserved structured regions that are involved in transient intramolecular contacts. In the presence of RXR/RAR, we show that N-CoR exploits its multivalency to form a multi-site complex that diplays an equilibrium between different conformational states. This conformational equilibrium is modulated by cognate ligands, RAR point mutation and RXR H12 deletion. Now, we can state that, in addition to NR conformation and ligand-induced allosteric changes, intrinsic disorder is substantially embedded in the synergetic regulation of RXR/RAR activity and its resulting abilities to communicate with the intracellular components.

biophysics↗