155] Aminoacyl transfer to ribosomal protein in the rat liver system

Abstract

Publisher Summary This chapter discusses the aminoacyl transfer to ribosomal protein in the rat liver system. The incorporation of amino acids from aminoacyl-soluble RNA (sRNA) into ribosomal protein requires two protein factors, which can be obtained from the soluble portion of the cell, GTP, magnesium ions, a monovalent cation, such as potassium or ammonium, and a sulfhydryl compound. Resolution of the aminoacyltransferring factors from rat liver has been obtained by ammonium sulfate fractionation and gel filtration. The method presented in the chapter describes the partial purification and the resolution of the two transfer factors from rat liver by molecular sieve chromatography on Sephadex G-200 columns; this procedure results in higher recoveries of activity and greater purification of the transferases and is more reproducible than the ammonium sulfatefractionation method. For routine analysis of Sephadex columns, resolved transferases I and II can be easily detected by assaying in low (4 m M ) glutathione reaction mixtures. However, when high (20 m M ) glutathione solutions are used, small amounts of transferase II in the ribosomal preparations (particularly with ribosomes other than those washed with NH 4 C1) are activated and the complementary effect of added transferase II is not as obvious.