The isolation of polysomes from rat liver. Contamination of polysomes by ribonucleoprotein of nuclear origin

Abstract

Polysomes isolated by methods involving Triton X-100 were contaminated by cosedimenting rapidly labeled material to a considerably greater extent than if the detergent sodium deoxycholate was used. A method for obtaining uncontaminated polysomes by using Triton X-100 was developed and criteria for purity were established. The contaminating material sedimented on the average somewhat slower than the polysomes. The sedimentation rate was not influenced by EDTA. Nuclear ribonucleoprotein particles containing RNA of DNA-like base composition have a sedimentation rate and a buoyant density similar to those of the contaminating material. They were not attacked by EDTA nor by nonionic detergents, whereas sodium deoxycholate reduced the particles to slowly sedimenting material by removal of protein from their RNA moiety. Nonionic detergents were able to protect against the action of sodium deoxycholate. Rapidly labeled material leaked out from the nuclei during the homogenization procedure. It cosedimented with the polysomes in sucrose gradients without being actually bound to them. It is suggested that the radioactive material contaminating polysome preparations is of nuclear origin. Part of it may have been present in the cytoplasm in vivo.