9 Ribosomal Subunit Joining

Abstract

Ribosomal subunit joining is the final stage in translation initiation in eukaryotes and follows formation of a 48S initiation complex at the initiation codon of an mRNA. Biochemical studies have identified individual steps leading to 48S complex formation on the majority of mRNAs (Chapter 2). First, a 43S complex is assembled from eukaryotic initiation factor (eIF) 1A, eIF3, an [eIF2/GTP/initiator Met-tRNA_i^Met] ternary complex, and a 40S ribosomal subunit. The 5′-terminal cap is bound by eIF4F, which with eIFs 4A and 4B unwinds RNA structure in the 5′-nontranslated region (5′NTR) and facilitates binding of the 43S complex to the 5′ end of the mRNA. eIF1 and eIF1A are required for 43S complexes to locate the initiation codon (Pestova et al. 1998), presumably by scanning until base-pairing is established between the first AUG triplet and the anticodon of tRNA_i^Met. The resulting 48S complex arrested at the initiation codon is stable and after fractionation by sucrose density gradient centrifugation or gel filtration remains associated with factors that include eIFs 1, 1A, 3 and the [eIF2/GTP/Met-tRNA_i^Met] complex (Benne and Hershey 1978; Trachsel and Staehelin 1978; Peterson et al. 1979a; Thomas et al. 1980; Pestova et al. 2000; Chapter 2).

Joining of a 60S ribosomal subunit to the 48S complex is associated with two linked events: hydrolysis of GTP bound to eIF2 in the 48S complex and displacement of factors. Replacement of GTP by GMPPNP (a nonhydrolyzable analog) does not impair ternary or 48S complex formation but prevents the 48S complex from joining a 60S...