Russell D W, Spremulli L L
J Biol Chem. 1979 Sep 25;254(18):8796-800.
A wheat germ ribosome dissociation factor, eukaryotic initiation factor 6 (eIF-6), has been purified almost to homogeneity from the 25 to 40% ammonium sulfate fraction of the postribosomal supernatant. This dissociation factor is distinct from initiation factor eIF-3 and its chromatographic properties permit its separation from the known wheat germ initiation factors. Under certain conditions, eIF-6 stimulates the incorporation of amino acids into polypeptides in a partially fractionated wheat germ cell-free system. The eight-step purification procedure developed includes chromatography on DEAE-cellulose, phosphocellulose, Sephadex G-75, and hydroxyapatite and yields a dissociation factor more than 80% pure. The purified factor is composed of a single polypeptide chain with a molecular weight of approximately 23,000 as determined by gel filtration chromatography and by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. It is an acidic protein which is heat labile and is inactivated by treatment with N-ethylmaleimide. The dissociation factor is much more effective in preventing the reassociation of 40 S and 60 S ribosomal subunits than in directly dissociating 80 S ribosomes. Like Escherichia coli IF-3, about 10 pmol of the dissociation factor are required to dissociate 1 pmol of ribosomes.
从小麦胚芽核糖体后上清液的25%至40%硫酸铵级分中已将一种小麦胚芽核糖体解离因子——真核起始因子6（eIF - 6）纯化至几乎同质。这种解离因子与起始因子eIF - 3不同，其色谱性质使其能够与已知的小麦胚芽起始因子分离。在某些条件下，eIF - 6在部分分级的小麦胚芽无细胞体系中刺激氨基酸掺入多肽。所开发的八步纯化程序包括在DEAE - 纤维素、磷酸纤维素、葡聚糖G - 75和羟基磷灰石上进行色谱分离，得到纯度超过80%的解离因子。通过凝胶过滤色谱法和十二烷基硫酸钠 - 聚丙烯酰胺凝胶电泳测定，纯化后的因子由一条分子量约为23,000的单一多肽链组成。它是一种酸性蛋白质，对热不稳定，经N - 乙基马来酰亚胺处理会失活。该解离因子在防止40 S和60 S核糖体亚基重新结合方面比直接解离80 S核糖体更有效。与大肠杆菌IF - 3一样，解离1 pmol核糖体大约需要10 pmol的解离因子。
