Translational Barrier in Central Region of Encephalomyocarditis Virus Genome

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Abstract
A fractionated cell-free system from [mouse carcinoma] Krebs-2 cells was prepared which contained ribosomes and a high-speed supernatant. When this system was programmed with encephalomyocarditis virus RNA, the synthesis of a precursor of capsid proteins, polypeptide preA, proceeded at a rate not very different from that observed in unfractionated extracts; the synthesis of more distally encoded proteins, in particular polypeptide F, was greatly retarded, if not abolished. A protein was purified from the cytoplasmic extracts of Krebs-2 cells which greatly enhanced production of polypeptide F as well as other noncapsid proteins in the fractionated system. By several criteria, this protein was identified as eukaryotic elongation factor 2 (eEF-2). By using the ADP-ribosylation assay, it was found that the fractionated system contained .apprx. 15% of the amount of eEF-2 present in the unfractionated extracts. The results suggest that changes in the eEF-2 content may affect the elongation rate differently at different regions of the RNA template.