In Vitro Reconstitution of Yeast Translation System Capable of Synthesizing Long Polypeptide and Recapitulating Programmed Ribosome Stalling.

The rates of translation elongation or termination in eukaryotes are modulated through cooperative molecular interactions involving mRNA, the ribosome, aminoacyl- and nascent polypeptidyl-tRNAs, and translation factors. To investigate the molecular mechanisms underlying these processes, we developed an in vitro translation system from yeast, reconstituted with purified translation elongation and termination factors, utilizing CrPV IGR IRES-containing mRNA, which functions in the absence of initiation factors. The system is capable of synthesizing not only short oligopeptides but also long reporter proteins such as nanoluciferase.