Abstract In eukaryotic translation, the termination and recycling phases are linked to subsequent initiation by persistence of several factors. These comprise the large eIF3 complex, eIF3j (Hcr1 in yeast) and the ATP-binding cassette protein ABCE1 (Rli1 in yeast). The ATPase is mainly active as a recycling factor, but it can remain bound to the dissociated 40S subunit until formation of 43S pre-initiation complexes. However, its functional role and native architectural context remains largely enigmatic. Here, we present an architectural inventory of native yeast and human ABCE1-containing pre-initiation complexes by cryo-EM. We found that ABCE1 was mostly associated with early 43S but also later 48S phases of initiation. It directly interacted with eIF3j via its unique iron-sulfur cluster domain and adopted a novel hybrid conformation, which was ATPase-inhibited and stabilized by an unknown factor bound between the nucleotide binding sites. Moreover, the native human samples provided a near-complete molecular picture of the architecture and sophisticated interaction network of the 43S-bound eIF3 complex and also the eIF2 ternary complex containing the initiator tRNA.

Structural inventory of native ribosomal ABCE1-43S pre-initiation complexes