Abstract
During the translation cycle, a cognate deacylated tRNA can only move together with the codon into the E site. We here present the first structure of a cognate tRNA bound to the ribosomal E site resulting from translocation by EF-G, in which an entire L1 stalk (L1 protein and L1 rRNA) interacts with E-site tRNA (E-tRNA), representing an authentic ribosome elongation complex. Our results revealed that the Watson-Crick base pairing is formed at the first and second codon-anticodon positions in the E site in the ribosome elongation complex, whereas the codon-anticodon interaction in the third position is indirect. Analysis of the observed conformations of mRNA and E-tRNA suggests that the ribosome intrinsically has the potential to form codon-anticodon interaction in the E site, independently of the mRNA configuration. We also present a detailed description of the biologically relevant position of the entire L1 stalk and its interacting cognate E-tRNA, which provides a better understanding of the structural basis for translation elongation. Furthermore, to gain insight into translocation, we report the positioning of protein L6 contacting EF-G, as well as the conformational change of the C-terminal tail of protein S13 in the decoding center.
Highlights
Proteins are synthesized by the ribosome in a process called translation
The binding of EF-GNGTP to ribosome in this ratcheted state catalyzes translocation, which involves movement of the anticodon stem-loops (ASLs) of tRNAs and the mRNA with respect to 30S subunit leading to the posttranslocational state where the ribosome preserves a peptidyl-tRNA and a deacylated tRNA in the P and E sites, respectively
Codon-anticodon Interaction in the Ribosomal E Site We here report the crystal structure of the 70 S ribosome with an entire L1 stalk and its interacting cognate E-site tRNA (E-tRNA), which is originally translocated from the P site with the aid of EF-G (Fig. 1A)
Summary
Proteins are synthesized by the ribosome in a process called translation. In bacteria, translation comprising initiation, elongation, and termination, involves four GTPase factors, initiation factor 2 (IF2), elongation factors Tu and G (EF-Tu/G), and peptide release factor 3 (RF3), respectively, reviewed in [1]. EF-Tu delivers aminoacyl tRNA to the ribosomal A site, peptidyl transfer from the P to the A site occurs, resulting in a pretranslocational state where the ribosome has a deacylated tRNA in the P site and a peptidyl-tRNA in the A site. The binding of EF-GNGTP to ribosome in this ratcheted state ( called hybrid state) catalyzes translocation, which involves movement of the ASLs of tRNAs and the mRNA with respect to 30S subunit leading to the posttranslocational state where the ribosome preserves a peptidyl-tRNA and a deacylated tRNA in the P and E sites, respectively. After GTP hydrolysis, the conformational change of EF-G renders it incompatible with ribosome binding, resulting in a rapid release. In the presence of the antibiotic fusidic acid, EF-G can be trapped in the ribosome [5]
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