## 3 Structural Snapshots of tRNA Conformation Plasticity during Translation
### Decoding  
For the incorporation of the correct aa-tRNA, the ribosome has to distinguish cognate ternary complexes (formed by aa-tRNA\({}^{\text{aa}}\). EF-Tu-GTP in prokaryotes) from the near-cognate and non-cognate species. The decoding process is based on a kinetic discrimination mechanism, and can be divided into two stages, GTPase activation and accommodation, which are separated by the irreversible hydrolysis of GTP by EF-Tu [104; 105; 106; 107; 108; 109]. This approach, based on a kinetic proofreading mechanism, grants multiple chances to reject the incorrect tRNA species [110]. Structurally, decoding relies on the mRNA codon and the tRNA anticodon's Watson-Crick complementarily, which is monitored by nucleotides G530, A1492 and A1493 from 16S rRNA [111; 112], as well as 30S conformational changes upon cognate codon-anticodon match [113], although it should be noted that some controversy raised on this subject recently [114; 115; 116]. All in all, however, it is clear that the ribosome has an active role in the accuracy of the whole process, using induced fit mechanisms (a combination of local and global conformational changes) to exclusively accelerate the forward rates of GTPase activation and accommodation for cognate species and thus, control the fidelity of tRNA selection (for more details, see [115; 117; 118; 119; 120] and references therein). Distinct transient states in the pathway of aa-tRNA delivery to the A site have been described, initially by kinetic methods that follow the fluorescence signal of strategically placed proflavin molecules [20] (see Figure 4d), later also via smFRET [26; 27], showing fluorescence changes in the anticodon region and the D loop of tRNA along the pathway. Essential structural information into the aa-tRNA incorporation process has been obtained through the analysis of one of such intermediates [15]. This state has been analyzed by cryoEM in ribosomal complexes bound with different ternary complexes in the presence of the antibiotic kirromycin [121; 122; 123; 124; 125; 126; 127; 128], as well as X-ray [129; 130] (Figure 4). When bound to kirromycin, elongation factor EF-Tu mimics the GTPase-activated configuration, albeit in a post-hydrolysis scenario [20]. The crystal structure of the ternary complex bound to the ribosome along with the antibiotic paromomycin and GTP analog GDPCP on the other hand, has suggested how GTPase activation might occur on the ribosome [131]. In all these structures, the aa-tRNA molecule adopts a distorted configuration, the distortion being located between the anticodon- and D-stem loop regions. The structures show that the tRNA deformation and the resulting spatial relationship with EF-Tu and the ribosome is responsible for the correct cognate codon recognition that activates the factor itself for GTP hydrolysis (see Figure 5 and corresponding legend for more details). Computational simulations have suggested that a corridor formed by conserved residues is responsible for directing the CCA end toward the PTC during accommodation [57], and that the process relies on reversible structural fluctuations of the accommodating aa-tRNA [60], very much in the same vein as during initial selection [132]. Figure 4: The A/T state of tRNA binding. (**a**) Comparison of the A/T tRNA (PDB 4V5G [129]) with the A/A tRNA that mimics the accommodated state (PDB 4V5D [133]); (**b**) Major changes are observed in the ASL region, as the helical twist is reduced after base pair 30:40. Also, the helical strands split at nucleotides 25 to 45 and 26 to 44.