Elongation factor Tu ternary complex binds to small ribosomal subunits in a functionally active state

Biochemistry ◽  
1984 ◽  
Vol 23 (25) ◽  
pp. 6171-6178 ◽  
Author(s):  
Jerome A. Langer ◽  
Frances Jurnak ◽  
James A. Lake
Keyword(s):  
Ternary Complex ◽  
Elongation Factor ◽  
Active State ◽  
Elongation Factor Tu ◽  
Ribosomal Subunits

scholarly journals The mechanism of error induction by the antibiotic viomycin provides insight into the fidelity mechanism of translation

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Mikael Holm ◽  
Chandra Sekhar Mandava ◽  
Måns Ehrenberg ◽  
Suparna Sanyal
Keyword(s):  
Ternary Complex ◽  
Elongation Factor ◽  
Translation System ◽  
Elongation Factor Tu ◽  
Active Conformation ◽  
Cell Growth Inhibition ◽  
Gtp Hydrolysis ◽  
Steady State Kinetics ◽  
Initial Selection ◽  
Insight Into

Applying pre-steady state kinetics to an Escherichia-coli-based reconstituted translation system, we have studied how the antibiotic viomycin affects the accuracy of genetic code reading. We find that viomycin binds to translating ribosomes associated with a ternary complex (TC) consisting of elongation factor Tu (EF-Tu), aminoacyl tRNA and GTP, and locks the otherwise dynamically flipping monitoring bases A1492 and A1493 into their active conformation. This effectively prevents dissociation of near- and non-cognate TCs from the ribosome, thereby enhancing errors in initial selection. Moreover, viomycin shuts down proofreading-based error correction. Our results imply a mechanism in which the accuracy of initial selection is achieved by larger backward rate constants toward TC dissociation rather than by a smaller rate constant for GTP hydrolysis for near- and non-cognate TCs. Additionally, our results demonstrate that translocation inhibition, rather than error induction, is the major cause of cell growth inhibition by viomycin.

scholarly journals A recent intermezzo at the Ribosome Club

2017 ◽  
Vol 372 (1716) ◽  
pp. 20160185 ◽  
Author(s):  
Michael Y. Pavlov ◽  
Anders Liljas ◽  
Måns Ehrenberg
Keyword(s):  
Messenger Rna ◽  
Ternary Complex ◽  
Elongation Factor ◽  
Transfer Rna ◽  
Elongation Factor Tu ◽  
Transfer Rnas ◽  
30S Subunit ◽  
Translation Accuracy ◽  
Codon Selection ◽  
Two Sides

Two sets of ribosome structures have recently led to two different interpretations of what limits the accuracy of codon translation by transfer RNAs. In this review, inspired by this intermezzo at the Ribosome Club, we briefly discuss accuracy amplification by energy driven proofreading and its implementation in genetic code translation. We further discuss general ways by which the monitoring bases of 16S rRNA may enhance the ultimate accuracy ( d -values) and how the codon translation accuracy is reduced by the actions of Mg 2+ ions and the presence of error inducing aminoglycoside antibiotics. We demonstrate that complete freezing-in of cognate-like tautomeric states of ribosome-bound nucleotide bases in transfer RNA or messenger RNA is not compatible with recent experiments on initial codon selection by transfer RNA in ternary complex with elongation factor Tu and GTP. From these considerations, we suggest that the sets of 30S subunit structures from the Ramakrishnan group and 70S structures from the Yusupov/Yusupova group may, after all, reflect two sides of the same coin and how the structurally based intermezzo at the Ribosome Club may be resolved simply by taking the dynamic aspects of ribosome function into account. This article is part of the themed issue ‘Perspectives on the ribosome’.

scholarly journals Elongation factor-Tu can repetitively engage aminoacyl-tRNA within the ribosome during the proofreading stage of tRNA selection

2020 ◽  
Vol 117 (7) ◽  
pp. 3610-3620 ◽  
Author(s):  
Justin C. Morse ◽  
Dylan Girodat ◽  
Benjamin J. Burnett ◽  
Mikael Holm ◽  
Roger B. Altman ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Ternary Complex ◽  
Bond Formation ◽  
Critical Role ◽  
Ribosomal Subunit ◽  
Elongation Factor ◽  
Peptide Bond ◽  
Elongation Factor Tu ◽  
Peptide Bond Formation ◽  
Gtp Hydrolysis

The substrate for ribosomes actively engaged in protein synthesis is a ternary complex of elongation factor Tu (EF-Tu), aminoacyl-tRNA (aa-tRNA), and GTP. EF-Tu plays a critical role in mRNA decoding by increasing the rate and fidelity of aa-tRNA selection at each mRNA codon. Here, using three-color single-molecule fluorescence resonance energy transfer imaging and molecular dynamics simulations, we examine the timing and role of conformational events that mediate the release of aa-tRNA from EF-Tu and EF-Tu from the ribosome after GTP hydrolysis. Our investigations reveal that conformational changes in EF-Tu coordinate the rate-limiting passage of aa-tRNA through the accommodation corridor en route to the peptidyl transferase center of the large ribosomal subunit. Experiments using distinct inhibitors of the accommodation process further show that aa-tRNA must at least partially transit the accommodation corridor for EF-Tu⋅GDP to release. aa-tRNAs failing to undergo peptide bond formation at the end of accommodation corridor passage after EF-Tu release can be reengaged by EF-Tu⋅GTP from solution, coupled to GTP hydrolysis. These observations suggest that additional rounds of ternary complex formation can occur on the ribosome during proofreading, particularly when peptide bond formation is slow, which may serve to increase both the rate and fidelity of protein synthesis at the expense of GTP hydrolysis.

Ternary complex between elongation factor Tu•GTP and Phe-tRNAPhe

Biochimie ◽  
1993 ◽  
Vol 75 (12) ◽  
pp. 1159-1166 ◽  
Author(s):  
C. Förster ◽  
S. Limmer ◽  
S. Ribeiro ◽  
R. Hilgenfeld ◽  
M. Sprinzl
Keyword(s):  
Ternary Complex ◽  
Elongation Factor ◽  
Elongation Factor Tu

Elongation factor Tu, a GTPase triggered by codon recognition on the ribosome: mechanism and GTP consumption

1995 ◽  
Vol 73 (11-12) ◽  
pp. 1221-1227 ◽  
Author(s):  
Marina V. Rodnina ◽  
Tillmann Pape ◽  
Rainer Fricke ◽  
Wolfgang Wintermeyer
Keyword(s):  
Ternary Complex ◽  
Elongation Factor ◽  
Peptide Bond ◽  
Elongation Factor Tu ◽  
Gtp Hydrolysis ◽  
Time Resolved ◽  
Codon Recognition ◽  
Fluorescence Measurements ◽  
A Site

The mechanism of elongation factor Tu (EF-Tu) catalyzed aminoacyl-tRNA (aa-tRNA) binding to the A site of the ribosome was studied. Two types of complexes of EF-Tu with GTP and aa-tRNA, EF-Tu∙GTP∙aa-tRNA (ternary) and (EF-Tu∙GTP)2∙aa-tRNA (quinternary), can be formed in vitro depending on the conditions. On interaction with the ribosomal A site, generally only one molecule of GTP is hydrolysed per aa-tRNA bound and peptide bond formed. The second GTP molecule from the quinternary complex is hydrolyzed only during translation of an oligo(U) tract in the presence of EF-G. The first step in the interaction between the ribosome and the ternary complex is the codon-independent formation of an initial complex. In the absence of codon recognition, the aa-tRNA–EF-Tu complex does not enter further steps of A site binding and remains in the initial binding state. Despite the rapid formation of the initial complex, the rate constant of GTP hydrolysis in the noncognate complex is four orders of magnitude lower compared with the cognate complex. This, together with the results of time-resolved fluorescence measurements, suggests that codon recognition by the ternary complex on the ribosome initiates a series of structural rearrangements that result in a conformational change of EF-Tu, presumably involving the effector region, which, in turn, triggers GTP hydrolysis and the subsequent steps of A site binding.Key words: translation, A site, codon recognition, fluorescence, stopped-flow.

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