Nucleosome reconstruction by electron spectroscopic imaging of phosphorus

Abstract

The three dimensional structure of a biological macromolecule or complex can be reconstructed from electron micrographs of different macromolecules or complexes lying in random orientation, provided that the relative orientations of individual particles can be determined (figure 1). In the case of the nucleosome core particle, a complex comprising 146 base pairs of DNA and 8 histone proteins, such orientational information can be obtained using a new technique called electron spectroscopic imaging which permits not only electron microscopy of the structure, but also the visualisation at high spatial resolution of the phosphorus distribution within the particle. Since phosphorus is a predominant constituent of DNA and not of protein, this elemental map represents an image of the nucleosomal DNA component. A comparison of a projection of a DNA supercoil model with images of the phosphorus distribution allowed us to orient individual distributions (figure 2). Using a direct convolution back-projection algorithm, the entire nucleosome containing both protein and DNA information was then reconstructed to a resolution of about 1.5 nm.