Abstract
BackgroundFrom bacteria to eukarya, the specific recognition of the amino-acylated initiator tRNA by the universally conserved translational GTPase eIF5B/IF2 is one of the most central interactions in the process of translation initiation. However, the molecular details, particularly also in the context of ribosomal initiation complexes, are only partially understood.ResultsA reinterpretation of the 6.6 Å resolution cryo-electron microscopy (cryo-EM) structure of the eukaryal 80S initiation complex using the recently published crystal structure of eIF5B reveals that domain IV of eIF5B forms extensive interaction interfaces with the Met-tRNAi, which, in contrast to the previous model, directly involve the methionylated 3′ CCA-end of the acceptor stem. These contacts are mediated by a conserved surface area, which is homologous to the surface areas mediating the interactions between IF2 and fMet-tRNAfMet as well as between domain II of EF-Tu and amino-acylated elongator tRNAs.ConclusionsThe reported observations provide novel direct structural insight into the specific recognition of the methionylated acceptor stem by eIF5B domain IV and demonstrate its universality among eIF5B/IF2 orthologs in the three domains of life.
Highlights
From bacteria to eukarya, the specific recognition of the amino-acylated initiator tRNA by the universally conserved translational GTPase eIF5B/IF2 is one of the most central interactions in the process of translation initiation
Cryo-EM structures of bacterial 30S pre-initiation complex (pre-IC) and 70S IC containing GTP/GDPNP-bound IF2 show how this interaction mutually stabilizes fMet-tRNAfMet and IF2 in conformations that allow the efficient association of the 50S subunit [16,17]
In the cteIF5B ortholog the β hairpin formed by β strands 3 and 4 and the loop following strand β5 (L5) contain 9 and 5 additional amino acids, respectively (Figure 1A/B)
Summary
The specific recognition of the amino-acylated initiator tRNA by the universally conserved translational GTPase eIF5B/IF2 is one of the most central interactions in the process of translation initiation. In bacteria and eukarya this process follows significantly different mechanisms, highlighted by different numbers of auxiliary protein factors (initiation factors or IFs) that are employed by bacterial (three IFs) or eukaryal cells (at least 12 eIFs) for correct ribosome assembly [1] Two of these factors, a/eIF1A/IF1 and the translational GTPase a/eIF5B/IF2, are universally conserved in the three domains of life [2]. Domain IV of IF2 exhibits a marked structural homology to domain II of EF-Tu that, together with the G domain, forms the universally conserved structural core among translational GTPases [13] and in EF-Tu constitutes part of the binding pocket for the amino-acylated acceptor arm of elongator tRNAs [12,14,15] Based on this observation it was suggested that IF2 domain IV and EF-Tu domain II use similar interfaces for their interactions with the tRNA [12]. None of these structures were determined at sufficiently high resolution to give any detailed insight into the interaction that would allow a correlation with the biochemical data
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