Comprised of eight core histones wrapped around at least 147 base pairs of DNA, nucleosomes are the fundamental unit the chromatin fiber from which long arrays are built to compact genetic... Show moreComprised of eight core histones wrapped around at least 147 base pairs of DNA, nucleosomes are the fundamental unit the chromatin fiber from which long arrays are built to compact genetic information into the cell nucleus. Structural proteins, such as linker histones (LH) and centromere proteins (CENP), interact with the DNA to dictate the exact architecture of the fiber which can directly influence the regulation of epigentic processes. However, the mechanisms by which structural proteins affect these processes are poorly understood. In this thesis, I will explore the various way in which LHs and CENP-N affect nucleosome and, by extension, chromatin fiber dynamics. First, I present a series of simulations of nucleosomes bound to LHs, otherwise known as chromatosomes, with the globular domain of two LH variants, generic H1 (genGH1) and H1.0 (GH1.0), to determine how their differences influence chromatosome structures, energetics and dynamics. These simulations highlight the thermodynamic basis for different LH binding motifs, and details their physical and chemical effects on chromatosomes. Second, I examine how well the findings above translate from mono-nucleosomes to poly-nucleosome arrays. I present a series of molecular dynamics simulations of octa-nucleosome arrays, based on a cryo-EMstructure of the 30-nm chromatin fiber, with and without the globular domains of the H1 LH to determine how they influence fiber structures and dynamics. These simulations highlight the effects of LH binding on the internal dynamics and global structure of poly- nucleosome arrays, while providing physical insight into a mechanism of chromatin compaction. Third, I took a brief departure from LHs to study the effects that the centromere protein N (CENP-N) has on the poly-nucleosome systems. I present a series of molecular dynamics simulations of CENP-N and di-nucleosome complexes based on cryo- EM and crystal structures provided by Keda Zhou and Karolin Luger. Simulations were conducted with nucleosomes in complex with one, two, and no CENP-Ns. This work, in collaboration with the Karolin Luger Group (University of Colorado – Boulder) and the Aaron Straight Group (Stanford University), represents the first atomistic simulations of this novel complex, providing the foundation for a plethora of future research opportunities exploring centromeric chromatin the effect that its structure and dynamics have on epigenetics. Lastly, I return to the chromatosome to study how DNA sequence affects the free energy surface and detailed mechanism of LH transitions between binding modes. I used umbrella sampling simulations to produce PMFs of chromatosomes wrapped in three different DNA sequences: Widom 601, poly-AT, and poly-CG. This work, my final in the series, represents a culmination of my studies furthering the understanding of biophysical phenomena surrounding LHs and how they can be extrapolated towards epigentic mechanisms. I was able to report on the first PMFs illustrating a previously unknown transition and describe the transition mechanism as it depends on DNA sequence. Show less
