Frolova L, Le Goff X, Zhouravleva G, Davydova E, Philippe M, Kisselev L
Département de Biologie et Génétique du Développement, CNRS URA 256, Université de Rennes 1, France.
RNA. 1996 Apr;2(4):334-41.
Termination of translation in eukaryotes is governed by two polypeptide chain release factors, eRF1 and eRF3 on the ribosome. eRF1 promotes stop-codon-dependent hydrolysis of peptidyl-tRNA, and eRF3 interacts with eRF1 and stimulates eRF1 activity in the presence of GTP. Here, we have demonstrated that eRF3 is a GTP-binding protein endowed with a negligible, if any, intrinsic GTPase activity that is profoundly stimulated by the joint action of eRF1 and the ribosome. Separately, neither eRF1 nor the ribosome display this effect. Thus, eRF3 functions as a GTPase in the quaternary complex with ribosome, eRF1, and GTP. From the in vitro uncoupling of the peptidyl-tRNA and GTP hydrolyses achieved in this work, we conclude that in ribosomes both hydrolytic reactions are mediated by the formation of the ternary eRF1-eRF3-GTP complex. eRF1 and the ribosome form a composite GTPase-activating protein (GAP) as described for other G proteins. A dual role for the revealed GTPase complex is proposed: in " GTP state," it controls the positioning of eRF1 toward stop codon and peptidyl-tRNA, whereas in "GDP state," it promotes release of eRFs from the ribosome. The initiation, elongation, and termination steps of protein synthesis seem to be similar with respect to GTPase cycles.
真核生物中翻译的终止由核糖体上的两种多肽链释放因子eRF1和eRF3控制。eRF1促进肽基 - tRNA的依赖于终止密码子的水解，并且eRF3与eRF1相互作用，并在GTP存在的情况下刺激eRF1的活性。在这里，我们已经证明eRF3是一种GTP结合蛋白，其内在GTPase活性（如果有的话）可以忽略不计，而eRF1和核糖体的联合作用能显著刺激这种活性。单独来看，eRF1和核糖体都不会产生这种效应。因此，eRF3在与核糖体、eRF1和GTP形成的四元复合物中作为一种GTPase发挥作用。根据这项工作中实现的肽基 - tRNA水解和GTP水解的体外解偶联，我们得出结论，在核糖体中，这两种水解反应都是由三元eRF1 - eRF3 - GTP复合物的形成介导的。eRF1和核糖体形成了一种复合GTPase激活蛋白（GAP），就像其他G蛋白所描述的那样。我们提出了所揭示的GTPase复合物的双重作用：在“GTP状态”下，它控制eRF1朝向终止密码子和肽基 - tRNA的定位，而在“GDP状态”下，它促进eRFs从核糖体上释放。蛋白质合成的起始步骤、延伸步骤和终止步骤在GTPase循环方面似乎是相似的。
