scholarly journals Codon-reading specificities of mitochondrial release factors and translation termination at non-standard stop codons

2013 ◽  
Vol 4 (1) ◽  
Author(s):  
Christoffer Lind ◽  
Johan Sund ◽  
Johan Åqvist
Keyword(s):  
Translation Termination ◽  
Stop Codons ◽  
Release Factors ◽  
Codon Reading

Termination of translation in eukaryotes

1995 ◽  
Vol 73 (11-12) ◽  
pp. 1079-1086 ◽  
Author(s):  
Lev L. Kisselev ◽  
Lyudmila Yu. Frolova
Keyword(s):  
Polypeptide Chain ◽  
Translation Termination ◽  
Protein Biosynthesis ◽  
High Rate ◽  
Vitro Assay ◽  
Binding Motifs ◽  
Stop Codons ◽  
Release Factors ◽  
Terminal Domain ◽  
Gtp Binding

Termination of translation is governed in ribosomes by polypeptide chain release factors (pRF and eRF in prokaryotes and eukaryotes, respectively). In prokaryotes, three pRF have been identified and sequenced, while in eukaryotes, only a single eRF has been identified to date. Recently, we have characterized a highly conserved protein family called eRF1. At least, human and Xenopus laevis proteins from this family are active as eRFs in the in vitro assay with any of the three stop codons. No structural similarity has been revealed between any of the three pRFs and eRF1 family. Furthermore, GTP-binding motifs have not been revealed, although translation termination in eukaryotes is a GTP-dependent process. We have demonstrated that in eukaryotes a second eRF exists in addition to eRF1, called eRF3. The eRF3 family has two features in common: presence of GTP-binding motifs and high conservation of the C-terminal domain structure. The C-terminal domain of the X. laevis eRF3 has no RF activity although it stimulates the eRF1 activity considerably at low concentration of the stop codons, conferring GTP dependence to the termination reaction. Without eRF3, the eRF1 activity is entirely GTP independent. Some features of X. laevis eRF3 (C-terminal domain) resemble those of pRF3. The newly identified eRF1 and eRF3 are structurally conserved and distinct from the respective pRF1/2 and pRF3 proteins, pointing to the possibility of different evolution of translation termination machinery in prokaryotes and eukaryotes. Bipartition of the translation termination apparatus probably provides high rate and accuracy of translation termination.Key words: higher eukaryotic polypeptide chain release factors, translation termination, protein biosynthesis.

scholarly journals Mechanisms that ensure speed and fidelity in eukaryotic translation termination

2021 ◽  
Author(s):  
Michael R Lawson ◽  
Laura N Lessen ◽  
Jinfan Wang ◽  
Arjun Prabhakar ◽  
Nicholas C Corsepius ◽  
...  
Keyword(s):  
Single Molecule ◽  
Translation Termination ◽  
Translation System ◽  
Single Molecule Fluorescence ◽  
Translation Elongation ◽  
Eukaryotic Translation ◽  
Stop Codons ◽  
Release Factors ◽  
Molecular Events

Translation termination, which liberates a nascent polypeptide from the ribosome specifically at stop codons, must occur accurately and rapidly. We established single molecule fluorescence assays to track the dynamics of ribosomes and two requisite release factors (eRF1 and eRF3) throughout termination using an in vitro reconstituted yeast translation system. We found that the two eukaryotic release factors bind together to recognize stop codons rapidly and elicit termination via a tightly regulated, multi-step process that resembles tRNA selection during translation elongation. Because the release factors are conserved from yeast to humans, the molecular events that underlie yeast translation termination are likely broadly fundamental to eukaryotic protein synthesis.

scholarly journals N 6-Methyladenosines in mRNAs reduce the accuracy of codon reading by transfer RNAs and peptide release factors

2021 ◽  
Vol 49 (5) ◽  
pp. 2684-2699
Author(s):  
Ka-Weng Ieong ◽  
Gabriele Indrisiunaite ◽  
Arjun Prabhakar ◽  
Joseph D Puglisi ◽  
Måns Ehrenberg
Keyword(s):  
Single Molecule ◽  
Stop Codon ◽  
Product Formation ◽  
Release Factors ◽  
Substrate Complex ◽  
Peptidyl Transfer ◽  
Enzyme Substrate ◽  
Peptide Release ◽  
Accuracy Ratio ◽  
Codon Reading

Abstract We used quench flow to study how N6-methylated adenosines (m6A) affect the accuracy ratio between kcat/Km (i.e. association rate constant (ka) times probability (Pp) of product formation after enzyme-substrate complex formation) for cognate and near-cognate substrate for mRNA reading by tRNAs and peptide release factors 1 and 2 (RFs) during translation with purified Escherichia coli components. We estimated kcat/Km for Glu-tRNAGlu, EF-Tu and GTP forming ternary complex (T3) reading cognate (GAA and Gm6AA) or near-cognate (GAU and Gm6AU) codons. ka decreased 10-fold by m6A introduction in cognate and near-cognate cases alike, while Pp for peptidyl transfer remained unaltered in cognate but increased 10-fold in near-cognate case leading to 10-fold amino acid substitution error increase. We estimated kcat/Km for ester bond hydrolysis of P-site bound peptidyl-tRNA by RF2 reading cognate (UAA and Um6AA) and near-cognate (UAG and Um6AG) stop codons to decrease 6-fold or 3-fold by m6A introduction, respectively. This 6-fold effect on UAA reading was also observed in a single-molecule termination assay. Thus, m6A reduces both sense and stop codon reading accuracy by decreasing cognate significantly more than near-cognate kcat/Km, in contrast to most error inducing agents and mutations, which increase near-cognate at unaltered cognate kcat/Km.

scholarly journals The Origin and Evolution of Release Factors: Implications for Translation Termination, Ribosome Rescue and Quality Control Pathways

2019 ◽  
Author(s):  
A.Maxwell Burroughs ◽  
L Aravind
Keyword(s):  
Quality Control ◽  
Translation Termination ◽  
Last Universal Common Ancestor ◽  
Origin And Evolution ◽  
Release Factors ◽  
Universal Common Ancestor ◽  
Translation Apparatus ◽  
Ph Domains ◽  
Evolutionary Trajectories ◽  
Hydrolysis Of

The evolution of release factors catalyzing the hydrolysis of the final peptidyl-tRNA bond and the release of the polypeptide from the ribosome has been a longstanding paradox. While the components of the translation apparatus are generally well-conserved across extant life, structurally-unrelated release factor peptidyl hydrolases (RF-PHs) emerged in the stems of the bacterial and archaeo-eukaryotic lineages. We analyze the diversification of RF-PH domains within the broader evolutionary framework of the translation apparatus. Thus, we reconstruct the possible state of translation termination in the Last Universal Common Ancestor with possible tRNA-like terminators. Further, evolutionary trajectories of the several auxiliary release factors in ribosome quality control (RQC) and rescue pathways point to multiple independent solutions to this problem and frequent transfers between superkingdoms including the recently-characterized ArfT, which is more widely-distributed across life than previously appreciated. The eukaryotic RQC system was pieced together from components with disparate provenance, which include the long-sought Vms1/ANKZF1 RF-PH of bacterial origin. We also uncover an under-appreciated evolutionary driver of innovation in rescue pathways: effectors deployed in biological conflicts that target the ribosome. At least three rescue pathways (centered on the prfH/RFH, baeRF-1, and C12orf65 RF-PH domains), were likely innovated in response to such conflicts.

scholarly journals Metabolic stress promotes stop-codon readthrough and phenotypic heterogeneity

2020 ◽  
Vol 117 (36) ◽  
pp. 22167-22172
Author(s):  
Hong Zhang ◽  
Zhihui Lyu ◽  
Yongqiang Fan ◽  
Christopher R. Evans ◽  
Karl W. Barber ◽  
...  
Keyword(s):  
Phenotypic Diversity ◽  
Stop Codon ◽  
Single Cells ◽  
Metabolic Stress ◽  
Aminoglycoside Antibiotic ◽  
Bacterial Cells ◽  
Excess Carbon ◽  
Stop Codons ◽  
Release Factors ◽  
Stop Codon Readthrough

Accurate protein synthesis is a tightly controlled biological process with multiple quality control steps safeguarded by aminoacyl-transfer RNA (tRNA) synthetases and the ribosome. Reduced translational accuracy leads to various physiological changes in both prokaryotes and eukaryotes. Termination of translation is signaled by stop codons and catalyzed by release factors. Occasionally, stop codons can be suppressed by near-cognate aminoacyl-tRNAs, resulting in protein variants with extended C termini. We have recently shown that stop-codon readthrough is heterogeneous among single bacterial cells. However, little is known about how environmental factors affect the level and heterogeneity of stop-codon readthrough. In this study, we have combined dual-fluorescence reporters, mass spectrometry, mathematical modeling, and single-cell approaches to demonstrate that a metabolic stress caused by excess carbon substantially increases both the level and heterogeneity of stop-codon readthrough. Excess carbon leads to accumulation of acid metabolites, which lower the pH and the activity of release factors to promote readthrough. Furthermore, our time-lapse microscopy experiments show that single cells with high readthrough levels are more adapted to severe acid stress conditions and are more sensitive to an aminoglycoside antibiotic. Our work thus reveals a metabolic stress that promotes translational heterogeneity and phenotypic diversity.

Structural basis for ArfA–RF2-mediated translation termination on mRNAs lacking stop codons

Nature ◽  
2016 ◽  
Vol 541 (7638) ◽  
pp. 546-549 ◽  
Author(s):  
Paul Huter ◽  
Claudia Müller ◽  
Bertrand Beckert ◽  
Stefan Arenz ◽  
Otto Berninghausen ◽  
...  
Keyword(s):  
Translation Termination ◽  
Structural Basis ◽  
Stop Codons

Reassigning stop codons via translation termination: How a few eukaryotes broke the dogma

BioEssays ◽  
2016 ◽  
Vol 39 (3) ◽  
pp. 1600213 ◽  
Author(s):  
Elena Alkalaeva ◽  
Tatiana Mikhailova
Keyword(s):  
Translation Termination ◽  
Stop Codons

scholarly journals Distinct Paths To Stop Codon Reassignment by the Variant-Code Organisms Tetrahymena and Euplotes

2006 ◽  
Vol 26 (2) ◽  
pp. 438-447 ◽  
Author(s):  
Joe Salas-Marco ◽  
Hua Fan-Minogue ◽  
Adam K. Kallmeyer ◽  
Lawrence A. Klobutcher ◽  
Philip J. Farabaugh ◽  
...  
Keyword(s):  
Stop Codon ◽  
Translation Termination ◽  
Yeast Cells ◽  
Universal Genetic Code ◽  
Stop Codons ◽  
Codon Recognition ◽  
Ciliate Species ◽  
Stop Codon Recognition ◽  
Euplotes Octocarinatus

ABSTRACT The reassignment of stop codons is common among many ciliate species. For example, Tetrahymena species recognize only UGA as a stop codon, while Euplotes species recognize only UAA and UAG as stop codons. Recent studies have shown that domain 1 of the translation termination factor eRF1 mediates stop codon recognition. While it is commonly assumed that changes in domain 1 of ciliate eRF1s are responsible for altered stop codon recognition, this has never been demonstrated in vivo. To carry out such an analysis, we made hybrid proteins that contained eRF1 domain 1 from either Tetrahymena thermophila or Euplotes octocarinatus fused to eRF1 domains 2 and 3 from Saccharomyces cerevisiae. We found that the Tetrahymena hybrid eRF1 efficiently terminated at all three stop codons when expressed in yeast cells, indicating that domain 1 is not the sole determinant of stop codon recognition in Tetrahymena species. In contrast, the Euplotes hybrid facilitated efficient translation termination at UAA and UAG codons but not at the UGA codon. Together, these results indicate that while domain 1 facilitates stop codon recognition, other factors can influence this process. Our findings also indicate that these two ciliate species used distinct approaches to diverge from the universal genetic code.

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