p53 is a tumor suppressor protein involved in regulation and progression of the cell cycle, apoptosis, and genomic stability through several mechanisms.1 Homotetrameric p53 translocates to the nucleus to bind its DNA recognition motif (RRRCWWGYYY: R, G/A; W, A/T; Y, C/T) and activate downstream signaling pathways.2,3 Mutations to the TP53 gene are the most common single mutations in cancer (>50%), and fall into two main categories: contact and conformational mutations. Contact mutations affect p53’s ability to recognize and bind target DNA sequences. Conformational mutations alter p53’s folded state, often affecting its ability to oligomerize and translocate to the nucleus to activate target genes.4 In the past couple decades, efforts have been made to target p53 mutants by stabilizing the WT conformation or restoring DNA-binding ability, though only a handful of molecules have made it to phase II/III clinical trials, and none have been approved as drugs.5,6,7 With the recent development of tyrosine-selective chemistry and pursuit of histidine-selective chemistry, we aim to use computational models to identify potential drug targets for these mutations.8,9 For this study, we will be using a X base pair DNA sequence from cyclin-dependent kinase inhibitor 1A (CDKN1A) containing a p53 recognition sequence. We chose to study this model due to CDKN1A’s critical role in the cellular response to DNA damage and availability of the crystal structure of p53 bound to it.10
Our objective is to elucidate the structural and functional implications of mutations in p53’s DNA binding domain (residues 102-292) observed in cancer. We aim to use ESMFold to generate predicted structures of the p53 mutants C176Y, H193Y, R175H, R273C and Y220C. We will then use PyMOL to align each mutant with WTp53 and calculate the RMSD between the predicted mutant structures and known wildtype as an indication of conformational change for the monomer. With protein-protein interaction-predicting software, we can predict the tetrameric structures of WT and mutant p53, which can then be used to perform DNA docking simulations using pyDockDNA. Potential DNA contact mutants are expected to have significant structural differences in their predicted protein-DNA interface while maintaining similar tetrameric structures relative to WT. Conformational mutants are hypothesized to have significant structural differences at the monomeric and/or tetrameric state, which is expected to distort the predicted protein-DNA interface.