Preparation and Crystallization of Nucleosome Core Particle

Publisher Summary The role of sample preparation and purification cannot be overestimated in experimental structural biology studies, especially for the crystallization of macromolecules. Purity, homogeneity, conformational state, and sample yield are major considerations in the preparation and purification of biological materials for structural studies. This chapter delineates some of the techniques used to produce large quantities of crystallizable material for chromatin studies. The scale of the procedures can be adapted to suit the situation. The first protocol described is for the isolation of histones from eukaryotic sources. Subsequent topics include isolation and purification of defined sequence DNA from recombinant sources. Reconstitution of nucleosome core particles (NCP) from histone octamer and defined sequence DNA is detailed, followed by descriptions for crystallizing NCP.