Extraction partielle des proteines de ribosomes de foie de rat par differents cations monovalents: Proprietes des particules obtenues

Abstract

Partial extraction of proteins from rat liver ribosomes by different monovalent cations. Properties of the core particles 1. Rat liver ribosomes were dissociated into subunits by centrifugation through a sucrose layer containing increasing concentrations of various monovalent cation salts. Poly U-directed polyphenylalanine synthesis by subunits was dependent on the nature and concentration of monovalent cations. Subunits prepared with the following salts: RbCl, KCl, NH 4Cl and NaCl had maximum activity when 0.3 M salts were used. The activity of the subunits obtained with 0.3 M RbCl was particularly high. Then activity decreased more or less rapidly according to monovalent cation, when the salt concentration was raised from 0.3 to 0.7 M. The activity of the subunits prepared with other salts (LiCl, CsCl and thallium acetate) continuously decreased with salt concentration without any maximum. When ribosomes were treated with salts at 25° instead of 4°, similar results were obtained, except for NaCl that behaved like the last mentioned salts. 2. Subunits prepared with 0.5–0.7 M monovalent cation salts sedimented in a sucrose gradient a little slower than those obtained with 0.3 M salts or native subunits. In a MgCl 2-rich buffer ability of subunits to reassociate varied in the same manner as activity: it was dependent on the monovalent cation used for preparing them, but diminished in every case when salt concentration was raised beyond 0.3 M. 3. Subunits prepared with 0.5 M salts, unlike the one prepared with 0.3 M salts, had a density and a chemical composition markedly different from that of control ribosomes. They had lost between 15 and 36 % of their proteins and then should be considered as incomplete subunits or core particles. The amount of proteins that were extracted depended on the monovalent cation used. The salts could be classified according to their ability to extract ribosomal proteins as follows: NaCl > LiCl > KCl, RbCl > NH 4Cl, CsCl. Thallium acetate represented a special case for extracting 27 % proteins even at 0.3 M concentration. 4. Extracted proteins and particle proteins were analysed by polyacrylamide gel electrophoresis. In every case it was observed that salts preferentially extracted some proteins, which were acidic or had relatively high molecular weight. Proteins of subribosomal particles prepared with various salts, when compared together, were found to be not exactly the same. 5. Extracted proteins, when added back to the corresponding core particles, greatly increased the ability of particles to reassociate. The reconstituted material had the same density as control ribosomes. However, activity was not restored when inactive core particles were used for these experiments.