Function of Three Protein Factors and Ribosomal Subunits in the Initiation of Protein Synthesis in E. coli

Nature ◽  
1968 ◽  
Vol 219 (5158) ◽  
pp. 1016-1021 ◽  
Author(s):  
M. REVEL ◽  
J. C. LELONG ◽  
G. BRAWERMAN ◽  
F. GROS
Keyword(s):  
Protein Synthesis ◽  
Ribosomal Subunits ◽  
E Coli

Structural Role of rRNAs on the Ribosomal Subunits from E. Coli by Scanning Transmission Electron Microscopy

1981 ◽  
Vol 39 ◽  
pp. 424-425
Author(s):  
M. Boublik ◽  
N. Robakis ◽  
J.S. Wall
Keyword(s):  
Three Dimensional ◽  
Uranyl Acetate ◽  
Dimensional Structure ◽  
Electron Microscopic ◽  
Ribosomal Subunits ◽  
E Coli ◽  
Freeze Dried ◽  
16S Rna ◽  
Scanning Transmission ◽  
And Function

The three-dimensional structure and function of biological supramolecular complexes are, in general, determined and stabilized by conformation and interactions of their macromolecular components. In the case of ribosomes, it has been suggested that one of the functions of ribosomal RNAs is to act as a scaffold maintaining the shape of the ribosomal subunits. In order to investigate this question, we have conducted a comparative TEM and STEM study of the structure of the small 30S subunit of E. coli and its 16S RNA.The conventional electron microscopic imaging of nucleic acids is performed by spreading them in the presence of protein or detergent; the particles are contrasted by electron dense solution (uranyl acetate) or by shadowing with metal (tungsten). By using the STEM on freeze-dried specimens we have avoided the shearing forces of the spreading, and minimized both the collapse of rRNA due to air drying and the loss of resolution due to staining or shadowing. Figure 1, is a conventional (TEM) electron micrograph of 30S E. coli subunits contrasted with uranyl acetate.

scholarly journals Development and comparison of cell-free protein synthesis systems derived from typical bacterial chassis

2021 ◽  
Vol 8 (1) ◽  
Author(s):  
Liyuan Zhang ◽  
Xiaomei Lin ◽  
Ting Wang ◽  
Wei Guo ◽  
Yuan Lu
Keyword(s):  
Protein Synthesis ◽  
Protein Production ◽  
Influential Factors ◽  
Competent Cell ◽  
Free Protein ◽  
Application Range ◽  
E Coli ◽  
Short Time ◽  
Cell Free Protein Synthesis

AbstractCell-free protein synthesis (CFPS) systems have become an ideal choice for pathway prototyping, protein production, and biosensing, due to their high controllability, tolerance, stability, and ability to produce proteins in a short time. At present, the widely used CFPS systems are mainly based on Escherichia coli strain. Bacillus subtilis, Corynebacterium glutamate, and Vibrio natriegens are potential chassis cells for many biotechnological applications with their respective characteristics. Therefore, to expand the platform of the CFPS systems and options for protein production, four prokaryotes, E. coli, B. subtilis, C. glutamate, and V. natriegens were selected as host organisms to construct the CFPS systems and be compared. Moreover, the process parameters of the CFPS system were optimized, including the codon usage, plasmid synthesis competent cell selection, plasmid concentration, ribosomal binding site (RBS), and CFPS system reagent components. By optimizing and comparing the main influencing factors of different CFPS systems, the systems can be optimized directly for the most influential factors to further improve the protein yield of the systems. In addition, to demonstrate the applicability of the CFPS systems, it was proved that the four CFPS systems all had the potential to produce therapeutic proteins, and they could produce the receptor-binding domain (RBD) protein of SARS-CoV-2 with functional activity. They not only could expand the potential options for in vitro protein production, but also could increase the application range of the system by expanding the cell-free protein synthesis platform.

scholarly journals LOCALIZED MUTAGENESIS FOR THE ISOLATION OF TEMPERATURE-SENSITIVE MUTANTS OF ESCHERICHIA COLI AFFECTED IN PROTEIN SYNTHESIS

Genetics ◽  
1979 ◽  
Vol 91 (2) ◽  
pp. 215-227
Author(s):  
W Scott Champney
Keyword(s):  
Escherichia Coli ◽  
Protein Synthesis ◽  
Temperature Sensitive ◽  
Chromosomal Locus ◽  
Ribosomal Subunits ◽  
Prolonged Incubation ◽  
Temperature Sensitive Mutants ◽  
Inhibition Of Growth ◽  
Localized Mutagenesis

ABSTRACT Two variations of the method of localized mutagenesis were used to introduce mutations into the 72 min region of the Escherichia coli chromosome. Twenty temperature-sensitive mutants, with linkage to markers in this region, have been examined. Each strain showed an inhibition of growth in liquid medium at 44°, and 19 of the mutants lost viability upon prolonged incubation at this temperature. A reduction in the rate of in vivo RNA and protein synthesis was observed for each mutant at 44°, relative to a control strain. Eleven of the mutants were altered in growth sensitivity or resistance to one or more of three ribosomal antibiotics. The incomplete assembly of ribosomal subunits was detected in nine strains grown at 44°. The characteristics of these mutants suggest that many of them are altered in genes for translational or transcriptional components, consistent with the clustering of these genes at this chromosomal locus.

Recycling of the initiation factor IF-3 on 30 S ribosomal subunits of E. coli

FEBS Letters ◽  
1973 ◽  
Vol 31 (3) ◽  
pp. 273-276 ◽  
Author(s):  
C. Vermeer ◽  
R.J. de Kievit ◽  
W.J. van Alphen ◽  
L. Bosch
Keyword(s):  
Initiation Factor ◽  
Ribosomal Subunits ◽  
E Coli

Author response for "A rational approach to improving titer in E. coli ‐based cell‐free protein synthesis reactions"

2020 ◽  
Author(s):  
Noelle Colant ◽  
Beatrice Melinek ◽  
Jaime Teneb ◽  
Stephen Goldrick ◽  
William Rosenberg ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Author Response ◽  
Rational Approach ◽  
Free Protein ◽  
E Coli ◽  
Synthesis Reactions ◽  
Cell Free Protein Synthesis

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.

Relationship between glutamine concentration and protein synthesis in rat skeletal muscle

1988 ◽  
Vol 255 (2) ◽  
pp. E166-E172 ◽  
Author(s):  
M. M. Jepson ◽  
P. C. Bates ◽  
P. Broadbent ◽  
J. M. Pell ◽  
D. J. Millward
Keyword(s):  
Protein Synthesis ◽  
Protein Deficiency ◽  
Synthesis Rate ◽  
Protein Synthesis Rate ◽  
Glutamine Concentration ◽  
E Coli ◽  
Protein Group ◽  
Highly Correlated ◽  
Rna Concentration

Muscle glutamine concentration ([GLN]) and protein synthesis rate (Ks) have been examined in vivo in well-fed, protein-deficient, starved, and endotoxemic rats. With protein deficiency (8 or 5% casein diet), [GLN] fell from 7.70 to 5.58 and 3.56 mmol/kg in the 8 and 5% diet groups, with Ks falling from 15.42 to 9.1 and 6.84%/day. Three-day starvation reduced [GLN] and Ks to 2.38 mmol/kg and 5.6%/day, respectively. In all these groups food intakes and insulin were generally well maintained (except in the starved group), whereas free 3,5,3'-triiodothyronine (T3) was depressed in the starved and 5% protein group. The E. coli lipopolysaccharide endotoxin (3 mg/kg) reduced [GLN] to 5.85 and 4.72 mmol/kg and Ks to 10.5 and 9.10%/day in two well-fed groups. Insulin levels were increased, and free T3 levels fell. Combined protein deficiency and endotoxemia further reduced [GLN] and Ks to 1.88 mmol/kg and 4.01%/day, respectively, in the 5% protein rats. Changes in both ribosomal activity (KRNA) and concentration (RNA/protein) contributed to the fall in Ks in malnutrition and endotoxemia, although reductions in the RNA concentration were most marked with protein deficiency and reductions in the KRNA dominated the response to the endotoxin. The changes in [GLN] and Ks were highly correlated as were [GLN] and both KRNA and the RNA concentration, and these relationships were unique to glutamine. These relationships could reflect sensitivity of glutamine transport and protein synthesis to the same regulatory influences, and the particular roles of insulin and T3 are discussed, as well as any direct influence of glutamine on protein synthesis.

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