Mechanism of action of an inhibitor from Proteus vulgaris on cell-free protein synthesis by Escherichia coli B

1969 ◽  
Vol 15 (2) ◽  
pp. 159-164
Author(s):  
J. J. McEvoy ◽  
W. E. Inniss
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Protein Synthesis ◽  
Mechanism Of Action ◽  
Ribonucleic Acid ◽  
Proteus Vulgaris ◽  
Flash Evaporation ◽  
Free System ◽  
Messenger Ribonucleic Acid ◽  
Escherichia Coli B

An inhibitory substance(s) has been found in S-30 fractions from Proteus vulgaris which prevented an Escherichia coli B cell-free system from incorporating a mixture of 14C-amino acids, L-phenylalanine-14C, or L-lysine-14C into protein, as directed by natural messenger ribonucleic acid, polyuridylic acid, or polyadenylic acid respectively. Similar results were obtained when the inhibitor was isolated from S-100 fractions by using dialysis, concentration of the dialysate by flash evaporation, hydrolysis, evaporation to dryness, dissolution to the original volume in distilled water, and neutralization. The effect of the inhibitor on the various individual reactions involved in protein synthesis was examined. No effect was found on the activation of amino acids as determined by the formation of L-phenylalanine-14C hydroxamate isolated chromatographically or by adenosine triphosphate – pyrophosphate exchange. Also no inhibition of L-phenylalanine-14C attachment to transfer ribonucleic acid occurred. However, ribosome-dependent reactions were markedly inhibited. The mechanism of action of the inhibitor appeared to be the prevention of binding of phenylalanyl-transfer ribonucleic acid to the ribosomes.

scholarly journals Aminoacyltransferase I-catalysed binding of phenylalanyl-transfer ribonucleic acid to muscle ribosomes from normal and diabetic rats

1971 ◽  
Vol 124 (3) ◽  
pp. 537-541 ◽  
Author(s):  
D. P. Leader ◽  
I. G. Wool ◽  
J. J. Castles
Keyword(s):  
Escherichia Coli ◽  
Skeletal Muscle ◽  
Amino Acids ◽  
Protein Synthesis ◽  
Ribonucleic Acid ◽  
Diabetic Rats ◽  
The Difference ◽  
Escherichia Coli B

The aminoacyltransferase I-catalysed binding of phenylalanyl-tRNA (unfractionated Escherichia coli B tRNA acylated with radioactive phenylalanine and 19 non-radioactive amino acids) to skeletal-muscle ribosomes from diabetic rats was less than that to ribosomes from normal rats when the Mg2+ concentration was low (7.5mm); whereas just the reverse was true when the concentration of the cation was higher (15mm). Thus the Mg2+ dependency of aminoacyltransferase I-catalysed binding of phenylalanyl-tRNA to ribosomes from normal and diabetic rats paralleled the effect of Mg2+ concentration on synthesis of polyphenylalanine reported before. During incubation at 7.5mm-Mg2+ phenylalanyl-tRNA was bound only to ribosomes bearing nascent peptidyl-tRNA. There are fewer such ribosomes in a preparation from the muscle of diabetic animals because diabetic animals synthesize less protein in vivo. Thus the difference in polyphenylalanine synthesis in vitro is adequately explained by the difference in enzyme-catalysed binding of phenylalanyl-tRNA to ribosomes, however, the basis of the difference in protein synthesis in vivo is still unknown.

Propriétés physiques et chimiques des tryptophanases de cinq espèces d'Entérobactériaceae

1975 ◽  
Vol 21 (6) ◽  
pp. 828-833 ◽  
Author(s):  
C. Simard ◽  
A. Mardini ◽  
L. M. Bordeleau
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Molecular Weight ◽  
Sedimentation Coefficient ◽  
Species Differentiation ◽  
Proteus Vulgaris ◽  
Amino Acids Composition ◽  
Propriétés Physiques ◽  
Tryptophan Content ◽  
Escherichia Coli B

The molecular weight, sedimentation coefficient, and amino acids composition were determined on five tryptophanases (TPases) from Escherichia coli B and E. aurescens, Shigella alkalescens, and Proteus vulgaris and P. morganii. These TPases have identical sedimentation profile and coefficient (9.6 S), and the same molecular weight (220 000). Each enzyme is constituted of four identical subunits having a molecular weight of 55 000. The amino acids composition of these TPases is very similar, with the exception of P. morganii and P. vulgaris TPases which present significative variations in basic amino acids and tryptophan content. The species differentiation of the coli group cannot be made on their TPase characteristics only, contrary to P. morganii and P. vulgaris which can be differentiated between them and from the coli group. [Journal translation]

scholarly journals Cell-Free Protein Synthesis Using S30 Extracts from Escherichia coli RFzero Strains for Efficient Incorporation of Non-Natural Amino Acids into Proteins

2019 ◽  
Vol 20 (3) ◽  
pp. 492 ◽  
Author(s):  
Jiro Adachi ◽  
Kazushige Katsura ◽  
Eiko Seki ◽  
Chie Takemoto ◽  
Mikako Shirouzu ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Protein Synthesis ◽  
Trna Synthetase ◽  
Translation Efficiency ◽  
Free System ◽  
Free Protein ◽  
Cell Free System ◽  
Natural Amino Acids ◽  
Cell Free Protein Synthesis

Cell-free protein synthesis is useful for synthesizing difficult targets. The site-specific incorporation of non-natural amino acids into proteins is a powerful protein engineering method. In this study, we optimized the protocol for cell extract preparation from the Escherichia coli strain RFzero-iy, which is engineered to lack release factor 1 (RF-1). The BL21(DE3)-based RFzero-iy strain exhibited quite high cell-free protein productivity, and thus we established the protocols for its cell culture and extract preparation. In the presence of 3-iodo-l-tyrosine (IY), cell-free protein synthesis using the RFzero-iy-based S30 extract translated the UAG codon to IY at various sites with a high translation efficiency of >90%. In the absence of IY, the RFzero-iy-based cell-free system did not translate UAG to any amino acid, leaving UAG unassigned. Actually, UAG was readily reassigned to various non-natural amino acids, by supplementing them with their specific aminoacyl-tRNA synthetase variants (and their specific tRNAs) into the system. The high incorporation rate of our RFzero-iy-based cell-free system enables the incorporation of a variety of non-natural amino acids into multiple sites of proteins. The present strategy to create the RFzero strain is rapid, and thus promising for RF-1 deletions of various E. coli strains genomically engineered for specific requirements.

scholarly journals Translational activity of messenger ribonucleic acid isolated from unstimulated and phytohaemagglutinin-activated lymphocytes

1979 ◽  
Vol 184 (2) ◽  
pp. 277-282 ◽  
Author(s):  
D M Wallace ◽  
R Jagus ◽  
C R Benzie ◽  
J E Kay
Keyword(s):  
Protein Synthesis ◽  
Ribonucleic Acid ◽  
Wheat Germ ◽  
Lower Efficiency ◽  
Free System ◽  
Activated Lymphocytes ◽  
Messenger Ribonucleic Acid ◽  
Translational Activity ◽  
Routine Assessment ◽  
Degree Of Purification

Purified cytoplasmic poly(A)+ RNA isolated from unstimulated pig lymphocytes has the same ability to direct translation in a range of cell-free systems as the corresponding mRNA from 20h phytohaemagglutinin-activated lymphocytes. Additional methylation of the mRNA is not required for maximum protein synthesis in the wheat-germ cell-free system. Misleading results are obtained if the mRNA preparations used are not adequately purified, and a method suitable for routine assessment of the degree of purification achieved is described. Cell-free protein-synthesizing systems from unstimulated lymphocytes translate added lymphocyte mRNA with lower efficiency than do comparable systems from phytohaemagglutinin-activated lymphocytes, whatever the source of the mRNA used.

MICROBIOLOGICAL STUDIES ON THE MECHANISM OF ACTION OF SPARSOMYCIN

1966 ◽  
Vol 12 (4) ◽  
pp. 595-604 ◽  
Author(s):  
Edward R. Bannister ◽  
Dale E. Hunt ◽  
Robert F. Pittillo
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Protein Synthesis ◽  
Mechanism Of Action ◽  
Rna Synthesis ◽  
Primary Site ◽  
Cross Resistance ◽  
E Coli

A primary site of sparsomycin attack in Escherichia coli appears to be inhibition of synthesis of protein, which occurs at concentrations of sparsomycin that do not affect DNA or RNA synthesis. Sparsomycin interferes with the normal excretion of amino acids by E. coli. Some cross-resistance was observed between a culture resistant to sparsomycin and cultures resistant to other inhibitors of protein synthesis.

Sign in / Sign up

Export Citation Format

Share Document