## 3 Structural Snapshots of tRNA Conformation Plasticity during Translation
### Decoding
The well-known 27:43 base pair mutation (highlighted in pink) results in an error-prone phenotype [134], as it probably weakens the ASL stem and thus, facilitates the strand separation that leads to the final distorted form of the incoming tRNA; (**c**) Comparison of the A/T tRNA (PDB 4V5G [129]) with the EF-Tu bound form (TC) in the absence of the ribosome (PDB 1 TTT [11]); (**d**) A gallery of mutations/manipulations in the tRNA molecule that affect the accuracy of decoding are highlighted: Cross-linking of nucleotides 8 and 13 [135], and mutations at 9:12:23 [136] and 24:11 [136; 137; 138]. Note that contrary to previous assumptions, subsequent structural analysis by [PERSON] and coworkers [130] discarded the idea that the so-called Hirsh mutation influences the flexibility/deformability of the tRNA. This framework also puts into context previous kinetic experiments: see proflavin insertions at positions 16 and 17, e.g., see [20]. (Figure adapted from [129], Copyright 2009 Am. Assoc. Adv. Sci.). Recent cryoEM structures of mammalian decoding complexes in the presence of GMPPNP have shown that as it is the case in bacteria, the key events in decoding are also the interactions of the ASL with the decoding center (A1824/A1825 of 18S rRNA) and the signaling of this event to the GTPase center of EF-Tu's eukaryotic counterpart eEF1A [146]. The data also included structural information on the codon sampling conformation prior to codon recognition that is lacking in bacteria (Figure 6). Both  
Figure 5: Graphic representation of the sequential stages during tRNA incorporation. (**a**) Distortion of the aa-tRNA during the initial, reversible codon sampling stage; (**b**) After codon recognition, the tRNA is stabilized in an A/T form. The distorted A/T state is known to be critical for the subsequent activation of GTP hydrolysis on the ribosome [138,139]. Accompanying global changes of the 30S subunit (mostly comprising the shoulder region in 16S rRNA) lead to the conformational change of domain II’s \(\beta\) loop; (**c**) which causes the distortion of the acceptor end; (**d**) Concomitant disruption of the contacts with switch I region of EF-Tu relocate the catalytic residue His84 [140] into the proper orientation. A2662 of the sarcin-ricin loop (SRL) of the 23S rRNA is also involved. Note that some details about the GTPase activation mechanism are still being discussed (see [131,141,142] for more details); Release of Pi is coupled to the (**e**) disordering of the switch I loop and (**f**) subsequent conformational change of EF-Tu [143]; (**g**) Finally, EF-Tu is released and the tRNA molecule relaxes its conformation to accommodate in the vacant A/A site. All in all, the different decoding stages are governed by the dynamic nature of the tRNA molecule within the A site [27]. Note that as the sequence and structure of each aa-tRNA is tuned according to the nature of the amino acid they carry, as well as to the codon-anticodon strength, the different cognate tRNA species display unvarying decoding properties (_i.e._, uniform rates of acceptance by the ribosome) [144,145]. (Figure adapted from [115], Copyright 2013 Annual Reviews). A/T substates (initial recognition and GTPase activated) were differentiated due to the configuration of the decoding center, as well as eEF1A's interaction pattern with the universally conserved sarcin ricin loop/SRL, which is an essential part of the factor binding and activating site (H95 of 28S rRNA) [147]. The ASL was visualized buried deeper in the decoding center once the codon was recognized. Conformational changes in the shoulder and head regions of the 40S subunit after the stabilization of the codon-anticodon duplex, similar to those described in the case of the bacterial domain closure rearrangement [113] were also detected. Nonetheless, the tRNA was visualized in the distorted configuration in both stages. Figure 6: Comparison of mammalian and bacterial ternary complexes. CryoEM maps of eukaryotic (**a**) codon sampling and (**b**) GTPase activation steps.