DESCRIPTION (provided by applicant): The goal of this project is to simulate conformational changes caused by phosporylation during the interaction of cell cycle checkpoint kinase, CHK2), with Protein Phosphatase 2A (PP2A) to elucidate signaling mechanisms during the DMA damage response. The research may identify therapeutic targets as outlined in the National Cancer Institute Strategic Plan. This interdisciplinary project is a collaboration between the Keyes laboratory, Boston University (computational chemistry) and the Monteiro laboratory, H. Lee Moffitt Cancer Center & Research Institute (molecular biology). The project has four specific aims. Aim 1 - Formulate, implement, and build a robust algorithm to sample the binding energy landscape of CHK2 and PP2A (Year 1). The Replica Exchange Statistical Temperature Molecular Dynamics (RESTMD) algorithm, which was derived in the Keyes laboratory, will be integrated into the Chemistry at HARvard Macromolecular Mechanics (CHARMM) package by adapting the original replica exchange module (Dr. Robert Best, University of Cambridge). To ascertain effectiveness, the RESTMD-CHARMM will be benchmarked against other sampling algorithms. Aim 2 - Model CHK2's SQ/TQ region (Years 2&3). Using RESTMD-CHARMM, the CHK2's unstructured domain will be modeled to determine the structural basis of the CHK2-PP2A interaction. The final package, data, and results will be made widely available to the biophysical community. Aim 3 - Identify the ammo acid residues of PP2A B' that mediate the interaction (Years 2&3). A large region of PP2A B'a (aa89-322) has been experimentally determined as the minimal binding region to CHK2. In order to narrow down the residues in binding, we will perform molecular dynamic analysis to generate a prediction of the amino acid residues involved. We will validate the prediction of the minimal region by experimentally testing mutations of the target amino acid residues. Aim 4 - Simulate the molecular binding of CHK2 and PP2A to determine the extent to which DNA damage-induced phosphorylation modulates complex formation and dissociation (Year 3). The CHK2-PP2A complex dissociates following DNA damage. Using the RESTMD-CHARMM package, the CHK2-PP2A energy landscape will be mapped. The molecular dynamics simulation will be compared to experimental data provided by the Monteiro laboratory. This research will advance cancer care by providing unprecedented insights into the fundamental genetic processes and determinants of breast cancer susceptibility.