Specific domains in yeast eukaryotic Initiation Factor (eIF) 4G bias the RNA unwinding specificity of eIF4F towards duplexes with a 5′‐overhang

Abstract

Translation initiation factor 4A (eIF4A) is an essential protein in yeast Saccharomyces cerevisiae, and plays a crucial role in the recruitment of mRNA to the 43S preinitiation complex. While the human ortholog of the protein has been extensively characterized, much remains to be elucidated regarding the biochemical properties of the yeast protein. Based on its conserved helicase motifs, eIF4A is believed to unwind secondary structures in the 5′‐untranslated region (UTR) of the mRNA to permit efficient translation. In our present study, we have sought to characterize the RNA binding and unwinding activities of yeast eIF4A by itself and in the context of the eIF4F complex, comprised of eIF4A, eIF4E, and eIF4G. Our results show that the ATPase activity of eIF4A is stimulated >250‐fold when it binds to eIF4E•eIF4G. Unlike its human cousin, the yeast eIF4F complex preferentially unwinds duplexes with a 5′‐overhang. Experiments with deletion mutants of eIF4G indicate that RNA‐binding domains of eIF4G co‐operate to impart this 5′‐dependence to the eIF4F complex.