Ensemble cryo-EM elucidates the mechanism of translation fidelity

• Published in: Nature (London) Vol. 546; no. 7656; pp. 113 - 117
• Main Authors: Loveland, Anna B., Demo, Gabriel, Grigorieff, Nikolaus, Korostelev, Andrei A.
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 01-06-2017; Nature Publishing Group
• Subjects: 101/28; 631/337/1645; 631/337/574/1789; 631/45/500; 631/535/1258/1259; Accommodation; Anticodon - chemistry; Anticodon - genetics; Anticodon - ultrastructure; Anticodons; Binding; Binding sites; Codon - chemistry; Codon - genetics; Codon - ultrastructure; Codons; Conformation; Cryoelectron Microscopy; Decoding; Docks; Elongated structure; Elongation; Elongation factor EF-Tu; Escherichia coli - chemistry; Escherichia coli - genetics; Escherichia coli - ultrastructure; GTP Phosphohydrolases - metabolism; GTP Phosphohydrolases - ultrastructure; Guanosine triphosphate; Guanosine Triphosphate - metabolism; High resolution; Humanities and Social Sciences; Hydrolysis; Intermediates; Messenger RNA; Methods; Microscopy; Models, Molecular; Molecular biology; Molecular structure; multidisciplinary; Observations; Peptide Elongation Factor Tu - metabolism; Peptide Elongation Factor Tu - ultrastructure; Physiological aspects; Protein Biosynthesis; Protein Domains; Proteins; Ribonucleic acid; Ribosome Subunits - chemistry; Ribosome Subunits - metabolism; Ribosome Subunits - ultrastructure; Ribosomes; Ribosomes - chemistry; Ribosomes - metabolism; Ribosomes - ultrastructure; Ricin; RNA; RNA sequencing; RNA, Ribosomal, 16S - genetics; RNA, Ribosomal, 16S - metabolism; RNA, Ribosomal, 16S - ultrastructure; RNA, Transfer, Amino Acyl - genetics; RNA, Transfer, Amino Acyl - metabolism; RNA, Transfer, Amino Acyl - ultrastructure; Science; Transfer RNA; Translation (Genetics); tRNA
• ISSN: 0028-0836; 1476-4687
• DOI: 10.1038/nature22397

Gene translation depends on accurate decoding of mRNA, the structural mechanism of which remains poorly understood. Ribosomes decode mRNA codons by selecting cognate aminoacyl-tRNAs delivered by elongation factor Tu (EF-Tu). Here we present high-resolution structural ensembles of ribosomes with cognate or near-cognate aminoacyl-tRNAs delivered by EF-Tu. Both cognate and near-cognate tRNA anticodons explore the aminoacyl-tRNA-binding site (A site) of an open 30S subunit, while inactive EF-Tu is separated from the 50S subunit. A transient conformation of decoding-centre nucleotide G530 stabilizes the cognate codon–anticodon helix, initiating step-wise ‘latching’ of the decoding centre. The resulting closure of the 30S subunit docks EF-Tu at the sarcin–ricin loop of the 50S subunit, activating EF-Tu for GTP hydrolysis and enabling accommodation of the aminoacyl-tRNA. By contrast, near-cognate complexes fail to induce the G530 latch, thus favouring open 30S pre-accommodation intermediates with inactive EF-Tu. This work reveals long-sought structural differences between the pre-accommodation of cognate and near-cognate tRNAs that elucidate the mechanism of accurate decoding.
Structural ensembles of the 70S ribosome bound to cognate or near-cognate charged tRNAs in complex with EF-Tu illustrate the crucial role of the nucleotide G530 in decoding of mRNA, and demonstrate that translational fidelity results from direct control of GTPase by the decoding centre.
Decoding faithful gene translation
The issue of fidelity in translating a DNA sequence into protein is central to the functioning of the ribosome. The insertion of even a single incorrect amino acid can potentially alter protein function. However, the structural understanding of translation fidelity remains unclear. Andrei Korostelev and colleagues have determined a series of structures of the 70S ribosome bound to cognate or near-cognate charged tRNAs. These structures illustrate the critical role of two nucleotides in the 16S rRNA, G530 and A1492, in forming a latch. The interaction of these nucleotides locks down the cognate tRNA by shifting the 30S domain and positioning the GTPase, elongation factor Tu, in an active conformation for hydrolysis.