The nucleosome core particle (NCP) is the basic building block of chromatin, which is a compact, yet dynamic structure that packages DNA. The NCP consists of a positively charged histone octameric core, surrounded by negatively charged nucleosomal DNA which is wrapped ~ 1.7 times around the core. There are many levels of structure in chromatin organization, ranging from the packaging of DNA around the NCP to compact chromatin fibers with diameters ~ 30 nm to denser chromatin fibers in metaphase chromosome with diameters ~ 1.5 µm. DNA binding proteins such as transcription factors and chromatin remodelers need to bind nucleosomal DNA. Thus, in order for transcription to occur, the nucleosomal DNA needs to unwrap fully or partially from the histone core. Modification of the histone core, commonly known as post-translational modification (PTM), can allow for easier access to nucleosomal DNA through modifications in the structure and dynamics of the NCP. Covalent modifications of either histone proteins or DNA often control gene activity and they are the most important epigenetic markers. Furthermore, mutations in chromatin components, such as the NCP, are also found to be commonly involved in diseases such as cancer. The innovative aspect of this proposal is the development of free energy methods in molecular dynamics to characterize complex reaction coordinates in hierarchical protein-nucleic acid assemblies. The PI will develop new computational methods/reaction coordinates to be used in free energy methods, validate atomistic force fields with local collaborators, and develop further methods to predict the impact of single amino acid mutations on nucleosome stability. We expect that the results from these computational investigations can add additional insight into the rational design of experimental investigations into fundamental chromatin structure. The PI’s laboratory will utilize advanced sampling methods in molecular dynamics to characterize the stability of the nucleosome core particle. The role of nucleic acid sequence, PTM, and oncogenic mutations on stability will be elucidated. Force fields for histone tails will be validated through comparison with NMR studies with local collaborators. New methodology to predict the effect of oncogenic mutations on NCP stability will be developed. Results will be tested by collaborators at MSKCC. The computational force fields and methodologies developed during the course of this proposed work could be used to characterize the interactions of nucleic acids and proteins in other macromolecular assemblies, for example, viruses