Unraveling the Structural Changes in the DNA-Binding Region of Tumor Protein p53 (TP53) upon Hotspot Mutation p53 Arg248 by Comparative Computational Approach

The vital tissue homeostasis regulator p53 forms a tetramer when it binds to DNA and regulates the genes that mediate essential biological processes such as cell-cycle arrest, senescence, DNA repair, and apoptosis. Missense mutations in the core DNA-binding domain (109-292) simultaneously cause the loss of p53 tumor suppressor function and accumulation of the mutant p53 proteins that are carcinogenic. The most common p53 hotspot mutation at codon 248 in the DNA-binding region, where arginine (R) is substituted by tryptophan (W), glycine (G), leucine (L), proline (P), and glutamine (Q), is reported in various cancers. However, it is unclear how the p53 Arg248 mutation with distinct amino acid substitution affects the structure, function, and DNA binding affinity. Here, we characterized the pathogenicity and protein stability of p53 hotspot mutations at codon 248 using computational tools PredictSNP, Align GVGD, HOPE, ConSurf, and iStable. We found R248W, R248G, and R248P mutations highly deleterious and destabilizing. Further, we subjected all five R248 mutant-p53-DNA and wt-p53-DNA complexes to molecular dynamics simulation to investigate the structural stability and DNA binding affinity. From the MD simulation analysis, we observed increased RMSD, RMSF, and Rg values and decreased protein-DNA intermolecular hydrogen bonds in the R248-p53-DNA than the wt-p53-DNA complexes. Likewise, due to high SASA values, we observed the shrinkage of proteins in R248W, R248G, and R248P mutant-p53-DNA complexes. Compared to other mutant p53-DNA complexes, the R248W, R248G, and R248P mutant-p53-DNA complexes showed more structural alteration. MM-PBSA analysis showed decreased binding energies with DNA in all five R248-p53-DNA mutants than the wt-p53-DNA complexes. Henceforth, we conclude that the amino acid substitution of Arginine with the other five amino acids at codon 248 reduces the p53 protein's affinity for DNA and may disrupt cell division, resulting in a gain of p53 function. The proposed study influences the development of rationally designed molecular-targeted treatments that improve p53-based therapeutic outcomes in cancer.

Automated summary

The study reveals the impact of specific amino acid substitutions on the structure, function, and DNA-binding affinity of p53 protein, which is a vital regulator of tissue homeostasis. The findings suggest that these substitutions reduce the protein's affinity for DNA and may disrupt cell division, leading to a gain of p53 function and promoting cancer development.