The human translation initiation multi-factor complex promotes methionyl-tRNA _i binding to the 40S ribosomal subunit

The delivery of Met-tRNAi to the 40S ribosomal subunit is thought to occur by way of a ternary complex (TC) comprising eIF2, GTP and Met-tRNAi. We have generated from purified human proteins a stable multifactor complex (MFC) comprising eIF1, eIF2, eIF3 and eIF5, similar to the MFC reported in yeast and plants. A human MFC free of the ribosome also is detected in HeLa cells and rabbit reticulocytes, indicating that it exists in vivo. In vitro, the MFC-GTP binds Met-tRNAi and delivers the tRNA to the ribosome at the same rate as the TC. However, MFC-GDP shows a greatly reduced affinity to Met-tRNAi compared to that for eIF2-GDP, suggesting that MFC components may play a role in the release of eIF2-GDP from the ribosome following AUG recognition. Since an MFC-Met-tRNAi complex is detected in cell lysates, it may be responsible for Met-tRNAi-40S ribosome binding in vivo, possibly together with the TC. However, the MFC protein components also bind individually to 40S ribosomes, creating the possibility that Met-tRNAi might bind directly to such 40S-factor complexes. Thus, three distinct pathways for Met-tRNAi delivery to the 40S ribosomal subunit are identified, but which one predominates in vivo remains to be elucidated.