Fractionation of Shope papilloma virus in cesium chloride density gradients

Abstract

Virus particles prepared from extracts of glycerinated Shope papilloma tissue of wild cottontail rabbits by means of three alternate low and high speed sedimentations were not uniform in the electron microscope. Variations in size and electron density were seen in negative-contrast preparations. When such particles were centrifuged to equilibrium in a cesium chloride density gradient they separated into four layers corresponding to densities of about 1.29, 1.32, 1.33, and 1.34. Ultraviolet absorption studies, infectivity tests, and the homogeneity of the particles were consistent with the thesis that the most dense particles contained the highest proportion of deoxyribonucleic acid (DNA) and were probably the complete virus. The intermediate layers appeared to have less DNA, and the top layer little or none. In electron micrographs the most dense particles were uniform. Their surfaces were composed of an estimated 60 to 70 capsomeres. The particles in the intermediate layers showed various degrees of emptiness at their centers. The lightest particles appeared to be loose aggregates of capsomeres. When the particles were exposed to fresh cesium chloride, they appeared to become more fragile, many of them disintegrating into free capsomeres on the electron microscope grid. These subunits of the protein coat had a structure suggestive of hollow cylinders or cups. Subunits of the viral core could not be definitely characterized in density gradients or visualized by means of the electron microscope.