A sequence element that tunes Escherichia coli tRNAAlaGGC to ensure accurate decoding

Ledoux, Sarah; Olejniczak, Mikołaj; Uhlenbeck, Olke C.

doi:10.1038/nsmb.1581

Cited by 81 publications

(95 citation statements)

References 47 publications

(69 reference statements)

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“…Any changes to this pair increase near-cognate tRNA incorporation (49,50). Our structural results, along with structural studies of 30S-extended ASL complexes (46), imply that insertions that perturb the structural integrity of the anticodon loop of suppressor tRNAs may direct recoding by destabilization of the 32-38 pairing.…”

Section: Discussionmentioning

confidence: 80%

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Structural insights into +1 frameshifting promoted by expanded or modification-deficient anticodon stem loops

Maehigashi

Dunkle

Miles

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

104

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Maintenance of the correct reading frame on the ribosome is essential for accurate protein synthesis. Here, we report structures of the 70S ribosome bound to frameshift suppressor tRNA SufA6 and N1-methylguanosine at position 37 (m 1 G37) modification-deficient anticodon stem loop Pro , both of which cause the ribosome to decode 4 rather than 3 nucleotides, resulting in a +1 reading frame. Our results reveal that decoding at +1 suppressible codons causes suppressor tRNA SufA6 to undergo a rearrangement of its 5′ stem that destabilizes U32, thereby disrupting the conserved U32-A38 base pair. Unexpectedly, the removal of the m 1 G37 modification of tRNA Pro also disrupts U32-A38 pairing in a structurally analogous manner. The lack of U32-A38 pairing provides a structural correlation between the transition from canonical translation and a +1 reading of the mRNA. Our structures clarify the molecular mechanism behind suppressor tRNA-induced +1 frameshifting and advance our understanding of the role played by the ribosome in maintaining the correct translational reading frame.near cognate | processivity T he three polymerase reactions of DNA replication, RNA transcription, and protein translation are essential to life and all involve a delicate balance between speed and fidelity. The bacterial ribosome decodes three mRNA nucleotides into a single amino acid at a rate of ∼20 residues/s with high fidelity (10 3

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Section: Discussionmentioning

confidence: 80%

“…Instead, what seems to promote +1 decoding by ASL SufA6 is the lack of a 32-38 interaction, which has previously been shown to be important for correct tRNA selection (49,50). The nucleotide identity of the 32-38 pair is finely tuned to both the Fig.…”

Section: Discussionmentioning

confidence: 99%

Structural insights into +1 frameshifting promoted by expanded or modification-deficient anticodon stem loops

Maehigashi

Dunkle

Miles

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

104

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“…On the other hand, the rates of GTP hydrolysis are remarkably similar (within a factor of 3) for different aa-tRNAs on their respective cognate codons; the same is true for the rates of peptide bond formation [40 -44]. This uniformity is achieved by tuning of the tRNA sequence and structure according to the nature of the amino acid [45]. Single mismatches at any position of the codon -anticodon complex result in slower forward reactions and a uniformly 1000-fold faster dissociation of ternary complexes from the ribosome (figure 2).…”

Section: Kinetic Discrimination Of Incorrect Trnasmentioning

confidence: 99%

“…This suggests that the network of interactions in the decoding centre [28] has evolved to yield a similar accuracy threshold, independent of the position of mismatches in the codon-anticodon duplex. Limited data obtained with Phe-tRNA Phe , Trp-tRNA Trp and Ala-tRNA Ala suggest that error frequencies are also similar for the near-cognate codons of other aa-tRNAs [30,40,44,45]. Uniform rates of decoding cognate and near-cognate codons presumably reflect the selective pressure that maintains similar translational accuracy for all codons.…”

Section: Kinetic Discrimination Of Incorrect Trnasmentioning

confidence: 99%

“…Although the tRNA is distorted also in the near-cognate A/T state [36], subtle changes in the orientation of tRNA and EF-Tu could interfere with GTPase activation by preventing A2662 from properly placing His84 into the active site [48]. In this framework, tRNA mutants that strongly activate GTP hydrolysis on a near-cognate codon [40,45] appear to have found their own unique conformational solution to dock EF-Tu on the sarcin-ricin loop. Nevertheless, other contacts in the codon recognition complex may specifically affect the stringency of decoding, e.g.…”

Section: The Crucial Forward Steps Of Decodingmentioning

confidence: 99%

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Evolutionary optimization of speed and accuracy of decoding on the ribosome

Wohlgemuth

Pohl

Mittelstaet

et al. 2011

Phil. Trans. R. Soc. B

132

159

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Speed and accuracy of protein synthesis are fundamental parameters for the fitness of living cells, the quality control of translation, and the evolution of ribosomes. The ribosome developed complex mechanisms that allow for a uniform recognition and selection of any cognate aminoacyl-tRNA (aa-tRNA) and discrimination against any near-cognate aa-tRNA, regardless of the nature or position of the mismatch. This review describes the principles of the selection-kinetic partitioning and induced fit-and discusses the relationship between speed and accuracy of decoding, with a focus on bacterial translation. The translational machinery apparently has evolved towards high speed of translation at the cost of fidelity.Keywords: ribosome; protein synthesis; translation fidelity; tRNA; error frequency; mRNA decoding SPEED AND ACCURACY OF TRANSLATIONProtein synthesis on the ribosome is a fundamentally important process that consumes a large part of the energy resources of the cell. Ribosomes are universal macromolecular machines built of two subunits, the small subunit (30S subunit in bacteria), where mRNA decoding takes place, and the large subunit (50S subunit in bacteria), which harbours the catalytic site for peptide bond formation. The decoding and peptidyl transferase centres consist of RNA, suggesting that the ribosome originates from the RNA world. Intuitively, one would expect that the ribosome has evolved to produce proteins with maximum speed and accuracy at minimum metabolic cost. The aim of this review is to discuss the mechanisms by which this is achieved and potential limits to the optimization of the ribosome performance. Protein synthesis entails four major phases: initiation, elongation, termination and recycling. During initiation, the ribosome selects an mRNA and, assisted by initiation factors, places the initiator tRNA on the appropriate start codon in the P site. In the subsequent elongation phase, amino acids are added to the growing peptide in a cyclic process. Aminoacyl-tRNAs (aa-tRNAs) enter the ribosome in a tight complex with elongation factor Tu (EF-Tu) and guanosine-5 0 -triphosphate (GTP). Following the recognition of the codon by the anticodon of aa-tRNA and GTP hydrolysis by EF-Tu, aa-tRNA is accommodated in the A site of the 50S subunit and takes part in peptide bond formation. The rate of protein elongation in bacteria is between 4 and 22 amino acids per second at 378C [1-5]; thus, a protein of an average length of 330 amino acids [6] is completed in about 10-80 s. The times required for initiation, termination and ribosome recycling (around 1 s each [3]) are short enough to make elongation rate-limiting for protein synthesis [7]. Translation of a particular codon depends on both the nature and abundance of the respective tRNAs, particularly on the non-random use of synonymous codons and the availability of the respective isoacceptor tRNAs [8]. The overall rate of translation is limited by the codon-specific rates of cognate ternary complex delivery to the A site and is further attenuated ...

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Breaking the Degeneracy of Sense Codons – How Far Can We Go?

Jones,

Hartman

2024

Israel Journal of Chemistry

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Genetic code expansion aims to incorporate non‐canonical amino acids (ncAAs) into biological systems, enhancing protein functionality or enabling the in vitro selection of peptides from diverse mRNA displayed libraries. Typically, genetic code expansion has involved reassignment of stop codons to ncAAs through orthogonal translation systems. This review instead focuses on efforts to expand the genetic code by breaking the redundancy of sense codons in vitro and in vivo. In vivo, orthogonal aminoacyl‐tRNA synthetase (AARS)/tRNA/AA systems are able to compete with endogenous machinery, enabling partial to full codon reassignment. Recent approaches, like genome recoding, offer potential solutions to reduce competition. In vitro studies utilize cell extract‐based or reconstituted translation systems, allowing precise control of codon usage via gene design and tRNA addition, making breaking of sense degeneracy easier. In these systems several unsplit codon boxes have been successfully reassigned multiple to ncAAs. These efforts showcase both the successes and challenges in achieving orthogonality and selective codon decoding and point towards a future where the 64 codons can encode more than 30 monomers, enabling new advances in synthetic biology and drug discovery.

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A sequence element that tunes Escherichia coli tRNAAlaGGC to ensure accurate decoding

Cited by 81 publications

References 47 publications

Structural insights into +1 frameshifting promoted by expanded or modification-deficient anticodon stem loops

Structural insights into +1 frameshifting promoted by expanded or modification-deficient anticodon stem loops

Evolutionary optimization of speed and accuracy of decoding on the ribosome

Breaking the Degeneracy of Sense Codons – How Far Can We Go?

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