scholarly journals Identification of proteins of the 40 S ribosomal subunit involved in interaction with initiation factor eIF-2 in the quaternary initiation complex by means of monospecific antibodies

FEBS Letters ◽  
1988 ◽  
Vol 233 (1) ◽  
pp. 114-118 ◽  
Author(s):  
Ulrich-Axel Bommer ◽  
Joachim Stahl ◽  
Annemarie Henske ◽  
Gudrun Lutsch ◽  
Heinz Bielka
Keyword(s):  
Ribosomal Subunit ◽  
Initiation Factor ◽  
Initiation Complex ◽  
Monospecific Antibodies

scholarly journals Interaction between membrane functions and protein synthesis in reticulocytes. An elongation-stage inhibitor of protein synthesis extracted from the reticulocyte membrane

1979 ◽  
Vol 180 (2) ◽  
pp. 379-387 ◽  
Author(s):  
D H Wreschner ◽  
M Herzberg
Keyword(s):  
Protein Synthesis ◽  
Cell Membrane ◽  
High Speed ◽  
Ribosomal Subunit ◽  
Initiation Factor ◽  
Membrane Component ◽  
Initiation Complex ◽  
Initiation Factor 2 ◽  
Reticulocyte Lysate ◽  
Inhibitor Of Protein Synthesis

A component of the reticulocyte cell membrane was found to inhibit protein synthesis severely in a reticulocyte lysate system. An investigation into the mode of action of the membrane inhibitor revealed the following facts. (1) The binding of the tertiary initiation complex (methionyl-tRNAfMet-Initiation Factor 2-GTP) to the 40S ribosomal subunit was unaffected by the membrane inhibitor. (2) The membrane component did not interfere with the binding of the 40S initiation complex to the AUG initiation codon and subsequent attachment of the 60S ribosomal subunit. (3) Elongation of the peptide chain, as assayed by peptidyl-puromycin formation, was markedly affected by the membrane inhibitor. Surprisingly, the membrane component caused a considerable increase in peptidyl-puromycin formation. (4) Reticulocyte ribosomes that had been reisolated by high-speed centrifugation, after preincubation with the membrane component, were found to be highly defective when assayed in a cell-free protein-synthesizing system. These results indicated that an extract of the reticulocyte cell membrane inhibited protein synthesis by interacting with the ribosome and thus interfered with the correct functions of the elongation stage of protein synthesis. The implications of this conclusion are discussed in the light of data showing that a highly purified preparation of the membrane inhibitor also displayed an endonucleolytic activity highly specific for 28S RNA.

scholarly journals Translation Initiation Factor eIF3b Contains a Nine-Bladed β-Propeller and Interacts with the 40S Ribosomal Subunit

Structure ◽  
2014 ◽  
Vol 22 (6) ◽  
pp. 923-930 ◽  
Author(s):  
Yi Liu ◽  
Piotr Neumann ◽  
Bernhard Kuhle ◽  
Thomas Monecke ◽  
Stephanie Schell ◽  
...  
Keyword(s):  
Translation Initiation ◽  
Ribosomal Subunit ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
40S Ribosomal Subunit

scholarly journals The Oxazolidinone Linezolid Inhibits Initiation of Protein Synthesis in Bacteria

1998 ◽  
Vol 42 (12) ◽  
pp. 3251-3255 ◽  
Author(s):  
Steve M. Swaney ◽  
Hiroyuki Aoki ◽  
M. Clelia Ganoza ◽  
Dean L. Shinabarger
Keyword(s):  
Protein Synthesis ◽  
Complex Formation ◽  
Antimicrobial Agents ◽  
Ribosomal Subunit ◽  
Multidrug Resistant ◽  
Binary Complex ◽  
Initiation Complex ◽  
Ribosomal Subunits ◽  
Bacterial Translation

ABSTRACT The oxazolidinones represent a new class of antimicrobial agents which are active against multidrug-resistant staphylococci, streptococci, and enterococci. Previous studies have demonstrated that oxazolidinones inhibit bacterial translation in vitro at a step preceding elongation but after the charging ofN-formylmethionine to the initiator tRNA molecule. The event that occurs between these two steps is termed initiation. Initiation of protein synthesis requires the simultaneous presence of N-formylmethionine-tRNA, the 30S ribosomal subunit, mRNA, GTP, and the initiation factors IF1, IF2, and IF3. An initiation complex assay measuring the binding of [3H]N-formylmethionyl-tRNA to ribosomes in response to mRNA binding was used in order to investigate the mechanism of oxazolidinone action. Linezolid inhibited initiation complex formation with either the 30S or the 70S ribosomal subunits fromEscherichia coli. In addition, complex formation withStaphylococcus aureus 70S tight-couple ribosomes was inhibited by linezolid. Linezolid did not inhibit the independent binding of either mRNA or N-formylmethionyl-tRNA toE. coli 30S ribosomal subunits, nor did it prevent the formation of the IF2–N-formylmethionyl-tRNA binary complex. The results demonstrate that oxazolidinones inhibit the formation of the initiation complex in bacterial translation systems by preventing formation of theN-formylmethionyl-tRNA–ribosome–mRNA ternary complex.

Alternative mechanisms of initiating translation of mammalian mRNAs

2005 ◽  
Vol 33 (6) ◽  
pp. 1231-1241 ◽  
Author(s):  
R.J. Jackson
Keyword(s):  
Site Selection ◽  
Mammalian Cells ◽  
Ribosomal Subunit ◽  
Aggregate Size ◽  
Mrna Translation ◽  
Initiation Site ◽  
Initiation Factor ◽  
Viral Mrnas ◽  
Initiation Factors ◽  
Scanning Mechanism

Of all the steps in mRNA translation, initiation is the one that differs most radically between prokaryotes and eukaryotes. Not only is there no equivalent of the prokaryotic Shine–Dalgarno rRNA–mRNA interaction, but also what requires only three initiation factor proteins (aggregate size ∼125 kDa) in eubacteria needs at least 28 different polypeptides (aggregate >1600 kDa) in mammalian cells, which is actually larger than the size of the 40 S ribosomal subunit. Translation of the overwhelming majority of mammalian mRNAs occurs by a scanning mechanism, in which the 40 S ribosomal subunit, primed for initiation by the binding of several initiation factors including the eIF2 (eukaryotic initiation factor 2)–GTP–MettRNAi complex, is loaded on the mRNA immediately downstream of the 5′-cap, and then scans the RNA in the 5′→3′ direction. On recognition of (usually) the first AUG triplet via base-pairing with the Met-tRNAi anticodon, scanning ceases, triggering GTP hydrolysis and release of eIF2–GDP. Finally, ribosomal subunit joining and the release of the other initiation factors completes the initiation process. This sketchy outline conceals the fact that the exact mechanism of scanning and the precise roles of the initiation factors remain enigmatic. However, the factor requirements for initiation site selection on some viral IRESs (internal ribosome entry sites/segments) are simpler, and investigations into these IRES-dependent mechanisms (particularly picornavirus, hepatitis C virus and insect dicistrovirus IRESs) have significantly enhanced our understanding of the standard scanning mechanism. This article surveys the various alternative mechanisms of initiation site selection on mammalian (and other eukaryotic) cellular and viral mRNAs, starting from the simplest (in terms of initiation factor requirements) and working towards the most complex, which paradoxically happens to be the reverse order of their discovery.

scholarly journals A Complementary Mechanism of Bacterial mRNA Translation Inhibition by Tetracyclines

2021 ◽  
Vol 12 ◽  
Author(s):  
Victor Barrenechea ◽  
Maryhory Vargas-Reyes ◽  
Miguel Quiliano ◽  
Pohl Milón
Keyword(s):  
Protein Synthesis ◽  
Translation Initiation ◽  
Ribosomal Subunit ◽  
Mrna Translation ◽  
Resistance Mechanisms ◽  
Dissociation Rate ◽  
Initiation Factor ◽  
Translation Elongation ◽  
Initiation Phase ◽  
Complementary Mechanism

Tetracycline has positively impacted human health as well as the farming and animal industries. Its extensive usage and versatility led to the spread of resistance mechanisms followed by the development of new variants of the antibiotic. Tetracyclines inhibit bacterial growth by impeding the binding of elongator tRNAs to the ribosome. However, a small number of reports indicated that Tetracyclines could also inhibit translation initiation, yet the molecular mechanism remained unknown. Here, we use biochemical and computational methods to study how Oxytetracycline (Otc), Demeclocycline (Dem), and Tigecycline (Tig) affect the translation initiation phase of protein synthesis. Our results show that all three Tetracyclines induce Initiation Factor IF3 to adopt a compact conformation on the 30S ribosomal subunit, similar to that induced by Initiation Factor IF1. This compaction was faster for Tig than Dem or Otc. Furthermore, all three tested tetracyclines affected IF1-bound 30S complexes. The dissociation rate constant of IF1 in early 30S complexes was 14-fold slower for Tig than Dem or Otc. Late 30S initiation complexes (30S pre-IC or IC) exhibited greater IF1 stabilization by Tig than for Dem and Otc. Tig and Otc delayed 50S joining to 30S initiation complexes (30S ICs). Remarkably, the presence of Tig considerably slowed the progression to translation elongation and retained IF1 in the resulting 70S initiation complex (70S IC). Molecular modeling of Tetracyclines bound to the 30S pre-IC and 30S IC indicated that the antibiotics binding site topography fluctuates along the initiation pathway. Mainly, 30S complexes show potential contacts between Dem or Tig with IF1, providing a structural rationale for the enhanced affinity of the antibiotics in the presence of the factor. Altogether, our data indicate that Tetracyclines inhibit translation initiation by allosterically perturbing the IF3 layout on the 30S, retaining IF1 during 70S IC formation, and slowing the transition toward translation elongation. Thus, this study describes a new complementary mechanism by which Tetracyclines may inhibit bacterial protein synthesis.

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