Toward In Situ Dynamics of DNA-Bound Full-Length p53 Tetramer.

p53 is the most important tumor suppressor in humans as well as the most frequently mutated gene found in human cancers, with ∼50% of all human tumors bearing p53 missense mutations that leave p53 inactive. Restoring p53 activity has been shown to lead to tumor regression even in advanced tumors in mouse models and thus is among the most attractive potential strategies for novel cancer therapy. Full-length p53 (fl-p53) consists of 393 residues and multiple domains, some of which are folded while others are disordered. Using the crystal structures of folded domains and integrative molecular modeling techniques for disordered domains, we generated the first wild-type (WT) fl-p53 tetramer model bound to DNA. When solvated, the system size was ∼500 K atoms, challenging extensive sampling. Using the Anton2 supercomputer for microsecond timescale simulations in explicit solvent and the rigorous Markov state model (MSM) framework, we elucidated the conformational landscape of wild-type p53 as well as two of the p53 hotspot cancer mutants, Y220C and G245S, in a physiological DNA-bound, full-length tetramer context. In the simulated timescale, DNA-bound fl-p53 tetramer bent DNA and formed a compact complex with interactions between the N-terminal and DNA-binding domains (DBDs) and C-terminal domains (CTDs) with DNA. The WT fl-p53 tetramer also sampled a unique quaternary DBD organization that is not accessed by the cancer mutants. The free energy landscapes indicated differential dynamics for the inner and outer p53 DBDs due to the dimer-dimer interface. The dynamics of the druggable L1/S3 pocket was also closely monitored. Ultimately, MSMs identified an underexplored loop 6 (L6) cryptic pocket and captured the effect of p53 tetramerization and cancer mutations.