A fast and effective method for purification of elongation factor 1 alpha from rabbit liver

Fast, efficient and gentle method of elongation factor 1 alpha (EF-1 alpha) purification from rabbit liver has been developed. Isolation procedure consists of three steps: gel-filtration of postmithohondrial supernatant on Sephacryl S-400, anion(DEAE-Cellulose)- and cation(SP-Sepharose)- exchange chromatographies. The procedure takes two-three days. Minimal number of the purification steps and the only one buffer usage during entire purification provide the high speed and efficiency of the method. Purity of EF-1 alpha preparation obtained was more than 90% as judged from DS-Na polyacrylamide gel electrophoresis. EF-1 alpha was demonstrated to be highly active in three different assays: the [3H]GDP-binding, the stimulation of [14C]phenylalanyl-tRNA binding to poly(U)-programmed 80S ribosomes and the stimulation of poly[14C]phenylalanine synthesis on poly(U)-programmed 80S ribosomes in the presence of EF-2. Since EF-1 alpha may exist in a cell as either GDP- or GTP-bound form the nature of nucleotide bound was studied using high pressure liquid chromatography (HPLC) analysis. Each molecule of EF-1 alpha appears to contain one molecule of GDP.