UC San Diego

p53 tumor suppressor protein plays a central role in responding to DNA damage by activating DNA repair, cell cycle arrest, senescence or apoptosis. In resting cells, MDM2 negatively regulates p53 by both blocking p53- coactivator interactions, as well as promoting p53 ubiquitin-dependent proteasomal degradation. In stressed cells, p53 transcriptional activity is positively regulated by its own C-terminal regulatory domain through non-specific DNA interactions that enhance specific DNA binding through the DNA binding domain. Mutations in p53 DNA binding domain and upregulation of MDM2 are involved in 50% of cancer cases. Understanding p53 and MDM2 at molecular level is crucial to design pharmacological targets to fight cancerous cells. However, structures of full-length of p53 and MDM2 have not yet been determined. The first goal of this thesis is to understand the DNA binding properties and crystal structure of the two most C -terminal domains of p53 (p53CTD) that include the tetramerization and regulatory domains. Our analytical centrifugation results indicated that p53 CTD binds to specific and non-specific DNA and that it changes from tetrameric state to dimeric state upon DNA binding. Our second goal is to understand the oligomerization state of MDM2 using AUC and transmission electron microscopy. Our results indicated that recombinant full-length MDM2 self- assembles to forms spherical particles of different sizes formed by different oligomerization