scholarly journals PROTEIN SYNTHESIS IN ISOLATED CELL NUCLEI

1957 ◽  
Vol 40 (3) ◽  
pp. 451-490 ◽  
Author(s):  
V. G. Allfrey ◽  
A. E. Mirsky ◽  
Syozo Osawa
Keyword(s):  
Amino Acids ◽  
Protein Synthesis ◽  
Amino Acid ◽  
Ribonucleic Acid ◽  
Orotic Acid ◽  
Nuclear Protein ◽  
Acid Uptake ◽  
Amino Acid Incorporation ◽  
Acid Incorporation ◽  
Thymus Tissue

1. Nuclei prepared from calf thymus tissue in a sucrose medium actively incorporate labelled amino acids into their proteins. This is an aerobic process which is dependent on nuclear oxidative phosphorylation. 2. Evidence is presented to show that the uptake of amino acids represents nuclear protein synthesis. 3. The deoxyribonucleic acid of the nucleus plays a role in amino acid incorporation. Protein synthesis virtually ceases when the DNA is removed from the nucleus, and uptake resumes when the DNA is restored. 4. In the essential mechanism of amino acid incorporation, the role of the DNA can be filled by denatured or partially degraded DNA, by DNAs from other tissues, and even by RNA. Purine and pyrimidine bases, monoribonucleotides, and certain dinucleotides are unable to substitute for DNA in this system. 5. When the proteins of the nucleus are fractionated and classified according to their specific activities, one finds the histones to be relatively inert. The protein fraction most closely associated with the DNA has a very high activity. A readily extractable ribonucleoprotein complex is also extremely active, and it is tempting to speculate that this may be an intermediary in nucleocytoplasmic interaction. 6. The isolated nucleus can incorporate glycine into nucleic acid purines, and orotic acid into the pyrimidines of its RNA. Orotic acid uptake into nuclear RNA requires the presence of the DNA. 7. The synthesis of ribonucleic acid can be inhibited at any time by a benzimidazole riboside (DRB) (which also retards influenza virus multiplication (11)). 8. The incorporation of amino acids into nuclear proteins seems to require a preliminary activation of the nucleus. This can be inhibited by the same benzimidazole derivative (DRB) which interferes with RNA synthesis, provided that the inhibitor is present at the outset of the incubation. DRB added 30 minutes later has no effect on nuclear protein synthesis. These results suggest that the activation of the nucleus so that it actively incorporates amino acids into its proteins requires a preliminary synthesis of ribonucleic acid. 9. Together with earlier observations (27, 28) on the incorporation of amino acids by cytoplasmic particulates, these results show that protein synthesis can occur in both nucleus and cytoplasm.

scholarly journals Action of growth hormone in vitro on the net uptake and incorporation into protein of amino acids in muscle from intact rabbits given protein-deficient diets

1976 ◽  
Vol 35 (1) ◽  
pp. 1-10 ◽  
Author(s):  
M. R. Turner ◽  
P. J. Reeds ◽  
K. A. Munday
Keyword(s):  
Amino Acids ◽  
Growth Hormone ◽  
Protein Synthesis ◽  
Amino Acid ◽  
De Novo ◽  
Amino Acid Uptake ◽  
Acid Uptake ◽  
Amino Acid Incorporation ◽  
Acid Incorporation

1. Net amino acid uptake, and incorporation into protein have been measured in vitro in the presence and absence of porcine growth hormone (GH) in muscle from intact rabbits fed for 5 d on low-protein (LP), protein-free (PF) or control diets.2. In muscle from control and LP animals GH had no effect on the net amino acid uptake but stimulated amino acid incorporation into protein, although this response was less in LP animals than in control animals.3. In muscle from PF animals, GH stimulated both amino acid incorporation into protein and the net amino acid uptake, a type of response which also occurs in hypophysectomized animals. The magnitude of the effect of GH on the incorporation of amino acids into protein was reduced in muscle from PF animals.4. The effect of GH on the net amino acid uptake in PF animals was completely blocked by cycloheximide; the uptake effect of GH in these animals was dependent therefore on de novo protein synthesis.5. It is proposed that in the adult the role of growth hormone in protein metabolism is to sustain cellular protein synthesis when there is a decrease in the level of substrate amino acids, similar to that which occurs during a short-term fast or when the dietary protein intake is inadequate.

Effect of Cycloheximide on In Vitro and In Vivo Protein Synthesis by Certain Tissues of Rats and Pigeons with Special Reference to Muscle

1973 ◽  
Vol 51 (12) ◽  
pp. 933-941 ◽  
Author(s):  
Njanoor Narayanan ◽  
Jacob Eapen
Keyword(s):  
Amino Acids ◽  
Body Weight ◽  
Protein Synthesis ◽  
Amino Acid ◽  
Amino Acid Incorporation ◽  
Contrast Administration ◽  
Acid Incorporation ◽  
Tissue Homogenates

The effect of cycloheximide in vitro and in vivo on the incorporation of labelled amino acids into protein by muscles, liver, kidneys, and brain of rats and pigeons was studied. In vitro incorporation of amino acids into protein by muscle microsomes, myofibrils, and myofibrillar ribosomes was not affected by cycloheximide. In contrast, administration of the antibiotic into intact animals at a concentration of 1 mg/kg body weight resulted in considerable inhibition of amino acid incorporation into protein by muscles, liver, kidneys, and brain. This inhibition was observed in all the subcellular fractions of these tissues during a period of 10–40 min after the administration of the precursor. Tissue homogenates derived from in vivo cycloheximide-treated animals did not show significant alteration in in vitro amino acid incorporation with the exception of brain, which showed a small but significant enhancement.

scholarly journals Transfer of serine into polypeptides and myosin by chromatographic species of seryl-transfer ribonucleic acid

1975 ◽  
Vol 146 (2) ◽  
pp. 395-400 ◽  
Author(s):  
M Nwagwu
Keyword(s):  
Protein Synthesis ◽  
Amino Acid ◽  
Free Amino Acid ◽  
Total Protein ◽  
Ribonucleic Acid ◽  
Free Amino ◽  
Maximum Yield ◽  
Amino Acid Incorporation ◽  
Acid Incorporation

The efficiencies of two chromatographic species of [3-H]seryl-tRNA, namely peaks I and II, in cell-free amino acid incorporation were investigated. The maximum yield of polypeptide seems to be the same for the reaction mixtures containing either peak I or peak II, suggesting that the efficiency of both peaks in total protein synthesis is the same. The efficiency of transfer of serine into myosin heavy subunit (myosin H) by peaks I and II was also investigated. Peak II of [3-H]seryl-tRNA transfers three times as much serine into myosin H as peak I.

EFFECT OF IODIDE AND THYROTROPHIN ON IN VITRO 14C-AMINO ACID INCORPORATION INTO RAT THYROID PROTEINS

1974 ◽  
Vol 76 (2) ◽  
pp. 273-285 ◽  
Author(s):  
Lubomir J. Valenta
Keyword(s):  
Amino Acids ◽  
Protein Synthesis ◽  
Amino Acid ◽  
Relative Concentration ◽  
Total Radioactivity ◽  
Amino Acid Incorporation ◽  
Control Value ◽  
Acid Incorporation ◽  
Rat Thyroid

ABSTRACT Thyroid lobes from rats on normal (NID) or low iodine (LID) intake were incubated for 4 hours in vitro in the presence of 14C-amino acids. The 14C-amino acid incorporation into thyroid protein was significantly higher in thyroids from LID than from NID fed rats, 7.82 ± 1.01 % (mean ± sd) of total radioactivity of the incubation mixture per 100 mg tissue compared to 3.74 ± 0.60 % respectively. Thyrotrophin (TSH) in vitro did not influence the 14C-amino acid incorporation. Iodide in concentration 10−7 m and higher decreased 14C-radioactivity incorporation into protein by 19.40 ± 3.06 and 26.59 ± 4.06 % of the control value for NID and LID rats respectively. This effect of iodide did not depend on iodine organification and was not influenced by the changes of free amino acids pool. There were no significant differences in the relative concentration of 14C-labelled thyroglobulin and total 14C-thyroid protein. Differential fragility demonstrable by unfolding or dissociation was observed between different classes of thyroglobulin. The fragility was increasing from the old non-labelled molecules to newly iodinated and newly synthesized ones. It is concluded that iodide has a direct intrathyroidal blocking effect on thyroid protein synthesis which may contribute to its antigoitrogenic action. The lack of in vitro stimulation of protein synthesis by TSH remains unexplained.

scholarly journals Lack of effect of amino acid concentration on protein synthesis in the perfused rat liver

1976 ◽  
Vol 160 (3) ◽  
pp. 797-801 ◽  
Author(s):  
D E Peavy ◽  
R J Hansen
Keyword(s):  
Amino Acids ◽  
Protein Synthesis ◽  
Amino Acid ◽  
Rat Plasma ◽  
The Other ◽  
Liver Protein ◽  
Amino Acid Incorporation ◽  
Fixed Amount ◽  
Acid Incorporation ◽  
Rat Livers

The effect of increasing the perfusate concentration of amino acids on the incorporation of labelled valine into protein was followed in perfusions of rat livers lasting for 2h. A fixed amount of labelled and unlabelled valine was added to the perfusate as the other amino acids were increased in multiples of the concentrations normally found in rat plasma. Under these conditions no increase in valine incorporation was observed, which appeared to be in conflict with results published by other workers, However, a different method of labelling from that used here was used in the earlier studies. An increasing amount of a labelled amino acid was added as the concentrations of the unlabelled amino acids were increased in the perfusate. An experiment directly comparing to the two labelling methods produced results that indicated that the apparent increase in liver protein synthesis observed by the other workers could have been due to the method of radioisotope addition. It is therefore concluded that increasing the perfusate concentration of amino acids does not increase amino acid incorporation into liver protein.

scholarly journals Nuclear binding of oestradiol-17β and induction of protein synthesis in the rat uterus during postnatal development

1973 ◽  
Vol 136 (1) ◽  
pp. 25-33 ◽  
Author(s):  
Dalia Sömjen ◽  
G. Sömjen ◽  
R. J. B. King ◽  
A. M. Kaye ◽  
H. R. Lindner
Keyword(s):  
Protein Synthesis ◽  
Amino Acid ◽  
Postnatal Development ◽  
Total Protein ◽  
Nuclear Protein ◽  
Amino Acid Incorporation ◽  
Rat Uterus ◽  
Nuclear Binding ◽  
Acid Incorporation ◽  
Old Rats

1. The uterine response to a single injection of oestradiol-17β during postnatal development of the rat was studied with respect to (i) nuclear binding of oestradiol-17β; (ii) induction of the synthesis of a specific cytoplasmic protein (‘induced protein’ of Gorski); (iii) rate of incorporation of 3H-labelled amino acids into total protein and into nuclear acid-soluble and acid-insoluble protein; and (iv) rate of [3H]thymidine incorporation into DNA. 2. Specific nuclear binding of oestradiol-17β could be demonstrated even at birth. Administration of oestradiol-17β in vivo caused a significant increase in the number of nuclear binding sites in rats aged 10 days or older. 3. A rapid method is described for the detection of the ‘induced protein’, based on cellulose acetate electrophoresis. Induction of this protein could be demonstrated at the age of 10, 15 and 20 days, but not in 5-day-old rats. 4. In 20-day-old rats the rate of 3H-labelled amino acid incorporation into protein increased by 3h after oestradiol administration. Incorporation into the different protein fractions reached peak values asynchronously: at 3–4h for acid-insoluble nuclear protein, at 6h for total protein and at about 12h for acid-soluble protein. 5. Treatment with oestradiol failed to stimulate amino acid incorporation into protein in 5- or 10-day-old rats; at the age of 15 to 30 days the hormone caused a significant increase in incorporation into total protein and into both types of nuclear protein. 6. Since the capacity for nuclear binding of oestradiol and for synthesis of the induced protein is demonstrable in the rat uterus before it acquires the ability to respond to the hormone with enhanced general protein synthesis and DNA synthesis, it appears that nuclear binding and the synthesis of the induced protein may be necessary but not sufficient conditions for the trophic action of oestradiol.

scholarly journals Transferability of N-terminal mutations of pyrrolysyl-tRNA synthetase in one species to that in another species on unnatural amino acid incorporation efficiency

Amino Acids ◽  
2020 ◽  
Author(s):  
Thomas L. Williams ◽  
Debra J. Iskandar ◽  
Alexander R. Nödling ◽  
Yurong Tan ◽  
Louis Y. P. Luk ◽  
...  
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Genetic Code ◽  
Unnatural Amino Acids ◽  
Trna Synthetase ◽  
Unnatural Amino Acid ◽  
Amino Acid Incorporation ◽  
Acid Incorporation ◽  
Genetic Code Expansion ◽  
Incorporation Efficiency

AbstractGenetic code expansion is a powerful technique for site-specific incorporation of an unnatural amino acid into a protein of interest. This technique relies on an orthogonal aminoacyl-tRNA synthetase/tRNA pair and has enabled incorporation of over 100 different unnatural amino acids into ribosomally synthesized proteins in cells. Pyrrolysyl-tRNA synthetase (PylRS) and its cognate tRNA from Methanosarcina species are arguably the most widely used orthogonal pair. Here, we investigated whether beneficial effect in unnatural amino acid incorporation caused by N-terminal mutations in PylRS of one species is transferable to PylRS of another species. It was shown that conserved mutations on the N-terminal domain of MmPylRS improved the unnatural amino acid incorporation efficiency up to five folds. As MbPylRS shares high sequence identity to MmPylRS, and the two homologs are often used interchangeably, we examined incorporation of five unnatural amino acids by four MbPylRS variants at two temperatures. Our results indicate that the beneficial N-terminal mutations in MmPylRS did not improve unnatural amino acid incorporation efficiency by MbPylRS. Knowledge from this work contributes to our understanding of PylRS homologs which are needed to improve the technique of genetic code expansion in the future.

scholarly journals FRUCTOSE-AMINO ACIDS IN LIVER: STIMULI OF AMINO ACID INCORPORATION IN VITRO

1955 ◽  
Vol 215 (1) ◽  
pp. 111-124 ◽  
Author(s):  
Henry Borsook ◽  
Adolph Abrams ◽  
Peter H. Lowy
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Amino Acid Incorporation ◽  
Acid Incorporation

scholarly journals The Involvement of Soluble Ribonucleic Acid in Amino Acid Incorporation by a Maize System

1964 ◽  
Vol 239 (6) ◽  
pp. 1762-1768 ◽  
Author(s):  
Rusty J. Mans ◽  
Caridad Maria Purcell ◽  
G. David Novelli
Keyword(s):  
Amino Acid ◽  
Ribonucleic Acid ◽  
Amino Acid Incorporation ◽  
Acid Incorporation
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