Separation of Rat Liver Cytoplasmic Trnas by High Performance Liquid Chromatography

Experiments aimed at developing a chromatographic map of cytoplasmic tRNA population from rat liver are described. HPLC tRNA fractionation by a linear gradient of acetonitrile/ammonium acetate, results in a chromatogram with more than 60 significant peaks, having area peak percentages ranging from 0.001 to 5.0. This paper presents evidence that each peak corresponds to a specific tRNA species. Specificity of tRNA-peak assignment was established by charging total tRNA with a mixture of 19 unlabeled aminoacids plus a radioactive aminoacid, and isolating the radioactive aminoacylated iso-tRNAs by gel electrophoresis, followed by HPLC acetonitrile gradient. The radioactive aminoacyl-tRNA species were eluted as well-behaved peaks. Here we describe the analysis of tRNATrp, which gave 2 well separated radioactive peaks, most likely corresponding to tRNATrp isoacceptor species postulated by the wobble hypothesis. Taken together, the present data validate this procedure for monitoring quantitatively and q...