X-Ray Diffraction Characterization of the Dense Phases Formed by Nucleosome Core Particles

Abstract

Multiple dense phases of nucleosome core particles (NCPs) were formed in controlled ionic conditions (15–160 mM monovalent salt, no divalent ions), under osmotic pressures ranging from 4.7 × 10 5 to 2.35 × 10 6 Pa. We present here the x-ray diffraction analysis of these phases. In the lamello-columnar phase obtained at low salt concentration (<25 mM), NCPs stack into columns that align to form bilayers, kept separated from one another by a layer of solvent. NCPs form a monoclinic lattice in the plane of the bilayer. For high salt concentration (>50 mM), NCPs order into either a two-dimensional columnar hexagonal phase or into three-dimensional orthorhombic (quasi-hexagonal) crystals. The lamellar and hexagonal (or quasi-hexagonal) organizations coexist in the intermediate salt range; their demixing requires a long time. For an applied pressure P = 4.7 10 5 Pa, the calculated NCPs concentration ranges from ∼280 to 320 mg/ml in the lamello-columnar phase to 495 to 585 mg/ml in the three-dimensional orthorhombic phase. These concentrations cover the concentration of the living cell.