scholarly journals The parable of the caveman and the Ferrari: protein synthesis and the RNA world

2017 ◽  
Vol 372 (1716) ◽  
pp. 20160187 ◽  
Author(s):  
Harry F. Noller
Keyword(s):  
Protein Synthesis ◽  
Ribosomal Rna ◽  
Rna World ◽  
Bond Formation ◽  
Peptide Bond ◽  
Peptide Bond Formation ◽  
Ribonucleoprotein Particle ◽  
Supportive Role

The basic steps of protein synthesis are carried out by the ribosome, a very large and complex ribonucleoprotein particle. In keeping with its proposed emergence from an RNA world, all three of its core mechanisms—aminoacyl-tRNA selection, catalysis of peptide bond formation and coupled translocation of mRNA and tRNA—are embodied in the properties of ribosomal RNA, while its proteins play a supportive role. This article is part of the themed issue ‘Perspectives on the ribosome’.

scholarly journals A Single Polyphosphate Kinase-Based NTP Regeneration System Driving Cell-Free Protein Synthesis

2019 ◽  
Author(s):  
Po-Hsiang Wang ◽  
Kosuke Fujishima ◽  
Samuel Berhanu ◽  
Yutetsu Kuruma ◽  
Tony Z. Jia ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Bond Formation ◽  
Peptide Bond ◽  
Functional Expression ◽  
Protein Yield ◽  
Polyphosphate Kinase ◽  
Peptide Bond Formation ◽  
Regeneration System ◽  
Free Protein ◽  
Cell Free Protein Synthesis

<p>Current reconstituted cell-free protein synthesis systems regenerate the ATP and GTP required for peptide bond formation using phosphocreatine and three coupled kinase reactions. Here, we replace the three-kinase system with a single polyphosphate kinase, resulting in a comparable protein yield and enabling functional expression of a heat-sensitive enzyme at 37°C.</p>

scholarly journals Origins of the Mechanochemical Coupling of Peptide Bond Formation to Protein Synthesis

2018 ◽  
Vol 140 (15) ◽  
pp. 5077-5087 ◽  
Author(s):  
Benjamin Fritch ◽  
Andrey Kosolapov ◽  
Phillip Hudson ◽  
Daniel A. Nissley ◽  
H. Lee Woodcock ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Bond Formation ◽  
Peptide Bond ◽  
Peptide Bond Formation ◽  
Mechanochemical Coupling

A Single Polyphosphate Kinase-Based NTP Regeneration System Driving Cell-Free Protein Synthesis

2019 ◽  
Author(s):  
Po-Hsiang Wang ◽  
Kosuke Fujishima ◽  
Samuel Berhanu ◽  
Yutetsu Kuruma ◽  
Tony Z. Jia ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Bond Formation ◽  
Peptide Bond ◽  
Functional Expression ◽  
Protein Yield ◽  
Polyphosphate Kinase ◽  
Peptide Bond Formation ◽  
Regeneration System ◽  
Free Protein ◽  
Cell Free Protein Synthesis

<p>Current reconstituted cell-free protein synthesis systems regenerate the ATP and GTP required for peptide bond formation using phosphocreatine and three coupled kinase reactions. Here, we replace the three-kinase system with a single polyphosphate kinase, resulting in a comparable protein yield and enabling functional expression of a heat-sensitive enzyme at 37°C.</p>

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.

scholarly journals STUDIES ON THE MODE OF ACTION OF DIPHTHERIA TOXIN

1967 ◽  
Vol 126 (5) ◽  
pp. 923-939 ◽  
Author(s):  
Ronald S. Goor ◽  
A. M. Pappenheimer ◽  
Elizabeth Ames
Keyword(s):  
Protein Synthesis ◽  
Diphtheria Toxin ◽  
Bond Formation ◽  
Peptide Bond ◽  
Peptide Bond Formation ◽  
Amino Acid Incorporation ◽  
Quantitative Studies ◽  
Acid Incorporation ◽  
Reversible Reactions ◽  
Sufficient Concentration

Inhibition of soluble transferase II activity in cell-free systems by diphtheria toxin and NAD can be prevented or reversed in the presence of a sufficient concentration of nicotinamide. Quantitative studies on inhibition of peptide bond formation in cell-free extracts by toxin and NAD have indicated that two successive reversible reactions are involved. First, toxin and NAD interact mole for mole to form a relatively dissociable complex. This toxin-NAD complex then reacts with transferase II to form an enzymatically inactive product that is but slightly dissociated. In the presence of sufficient nicotinamide, however, the latter complex can be broken down to yield active transferase II once more. Based on the above model, an equation has been derived that accurately predicts the per cent inhibition of amino acid incorporation in cell-free systems at any given toxin and NAD level. The observed inhibition appears to be independent of the sensitivity to toxin of the cell species from which the extracts were derived, and depends only on the toxin and NAD concentrations. Although the model satisfactorily explains inhibition of peptide bond formation by toxin in cell-free systems, further assumptions are needed to explain how still lower concentrations of toxin are able to arrest protein synthesis completely in the living cell.

Ribosome structural dynamics in translocation: yet another functional role for ribosomal RNA

2017 ◽  
Vol 50 ◽  
Author(s):  
Harry F. Noller ◽  
Laura Lancaster ◽  
Srividya Mohan ◽  
Jie Zhou
Keyword(s):  
Protein Synthesis ◽  
Structural Dynamics ◽  
Ribosomal Rna ◽  
Messenger Rna ◽  
Peptide Bond ◽  
Peptide Bond Formation ◽  
Detailed Structural Analysis ◽  
Ribonucleoprotein Complexes ◽  
Elongation Phase ◽  
Polypeptide Chains

AbstractRibosomes are remarkable ribonucleoprotein complexes that are responsible for protein synthesis in all forms of life. They polymerize polypeptide chains programmed by nucleotide sequences in messenger RNA in a mechanism mediated by transfer RNA. One of the most challenging problems in the ribosome field is to understand the mechanism of coupled translocation of mRNA and tRNA during the elongation phase of protein synthesis. In recent years, the results of structural, biophysical and biochemical studies have provided extensive evidence that translocation is based on the structural dynamics of the ribosome itself. Detailed structural analysis has shown that ribosome dynamics, like aminoacyl-tRNA selection and catalysis of peptide bond formation, is made possible by the properties of ribosomal RNA.

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