Structural states of the nucleosome

Nucleosome structure was investigated using biochemical methods, spectroscopic electron microscopy, methods of conformational characterization and newly developed three-dimensional image reconstruction techniques. The results of a study of the effects of ionic strength on nucleosome structure indicated several different ionic-strength dependent conformational states. The results of an additional study of nucleosome structure with gene expression indicated an unfolded structure induced by gene-expression specific post-translational modifications.;Our analysis of nucleosome structure using conformational characterization by principal component analysis indicated that in low ionic environments the particle is highly elongated and adopts a prolate ellipsoidal conformation in an ionic environment of 10 mM NaCl. A spheroidal form was observed at 30 mM NaCl. Results also indicated an oblate conformation at 150 mM NaCl. Above 400 mM NaCl the particle became prolate again and highly extended. Measurements at higher ionic environments were consistent with a disassociation of the protein components of the nucleosome.;Nucleosome structure was additionally investigated using quaternion-assisted three-dimensional image reconstruction techniques. These methods were first applied in an investigation of the structure of the 54 kDa signal sequence binding protein of signal recognition particle, a macromolecular complex involved in protein targeting. Results indicated a two-domain structure consistent with biochemical and genetic studies of the 54 kDa protein. The application of similar reconstruction methods to nucleosomes prepared in 10, 30 and 150 mM NaCl, indicated prolate ellipsoidal, spheroidal, and oblate ellipsoidal conformations for the particle, respectively. These results concurred with principal component analyses and were consistent with previous electron microscopic studies, crystallographic and neutron scattering analyses of the nucleosome.;The quaternion-assisted image reconstruction methods were subsequently applied in an investigation of nucleosomes associated with transcriptionally active genes. Hyperacetylated nucleosomes were purified from active genes using chromatographic methods and imaged using spectroscopic electron microscopy. Three-dimensional image reconstruction indicated a C-shaped and extended conformation which appeared unfolded with an asymmetric mass distribution. The open structure was consistent with other physico-biochemical investigations of hyperacetylated nucleosome particles.;In summary, the results of our investigation of nucleosome structure with ionic environment and gene expression indicated many conformations of nucleosome structure and a dynamic nature for the particle that is in accord with genetic and biochemical studies.