Initiation Factor eIF2γ Promotes eIF2–GTP–Met-tRNA(i)(Met) Ternary Complex Binding to the 40S Ribosome

In contrast to elongation factor EF-Tu, which delivers aminoacyl-tRNAs to the ribosomal A-site, eukaryotic initiation factor eIF2 binds initiator Met-tRNA(i)(Met) to the P-site of the 40S ribosomal subunit. We used directed hydroxyl radical probing experiments to map the binding of Saccharomyces cerevisiae eIF2 on the ribosome and on Met-tRNA(i)(Met). Our results identify a key binding-interface between domain III of eIF2γ and 18S rRNA helix h44 on the 40S subunit. Moreover, we showed that eIF2γ primarily contacts the acceptor stem of Met-tRNA(i)(Met). Whereas the analogous domain III of EF-Tu contacts the T-stem of tRNAs, biochemical analyses demonstrated that eIF2γ domain III is important for ribosome, but not Met-tRNA(i)(Met), binding. Thus despite their structural similarity, eIF2 and EF-Tu bind tRNAs in substantially different manners, and we propose that the tRNA-binding domain III of EF-Tu has acquired a new ribosome-binding function in eIF2γ.