Influx of Glycylsarcosine and l-Lysyl-l-Lysine into Hamster Jejunum in Vitro

1980 ◽  
Vol 58 (3) ◽  
pp. 221-225 ◽  
Author(s):  
E. Taylor ◽  
D. Burston ◽  
D. M. Matthews
Keyword(s):  
Amino Acid ◽  
Glutamic Acid ◽  
Amino Acid Uptake ◽  
The Other ◽  
Amino Acid Side Chain ◽  
Acid Uptake ◽  
High Concentration ◽  
Simple Diffusion ◽  
Wide Range

1. This paper reports an investigation of whether the dipeptides glycylsarcosine and l-lysyl-l-lysine share a single mediated transport mechanism into hamster jejunum, or whether one of these peptides is taken up in part by a mediated mechanism unavailable to the other. The investigation, using rings of everted jejunum in vitro, was carried out at pH 5 in order to reduce brush border and/or intramedium hydrolysis of lysyl-lysine. 2. The kinetics of uptake of each peptide was studied over a wide range of concentrations. Estimates of the simple diffusion component in uptake of each peptide were made by the method of self-inhibition of transport as previously described. After correction for simple diffusion, uptake of each peptide conformed to Michaelis-Menten kinetics, and values for Kt and Vmax. were obtained. 3. It was found that each peptide was capable, at infinitely high concentration, of complete inhibition of mediated uptake of the other. The inhibitory effect was competitive. We concluded that glycylsarcosine and lysyl-lysine were taken up by a common mediated mechanism (or possibly mechanisms), neither peptide being taken up by a mediated mechanism unavailable to the other. 4. A previous paper showed that l-glutamyl-l-glutamic acid and glycylsarcosine were taken up by a common mediated mechanism, and this paper shows that l-lysyl-l-lysine and glycylsarcosine are taken up by a common mediated mechanism. It is therefore postulated that the neutral dipeptide glycylsarcosine, the acidic dipeptide glutamyl-glutamic acid and the basic dipeptide lysyl-lysine all share a common mediated mechanism for uptake. This suggests that peptide uptake differs from amino acid uptake in that it is indifferent to the net charge on the amino acid side chain(s).

Influx of Two Dipeptides, Glycylsarcosine and l-glutamyl-l-glutamic Acid, into hamster Jejunum in Vitro

1979 ◽  
Vol 56 (1) ◽  
pp. 15-23 ◽  
Author(s):  
D. M. Matthews ◽  
R. H. Gandy ◽  
E. Taylor ◽  
D. Burston
Keyword(s):  
Glutamic Acid ◽  
Brush Border ◽  
Transport Mechanism ◽  
Inhibitory Effect ◽  
The Other ◽  
Peptide Transport ◽  
High Concentration ◽  
Simple Diffusion ◽  
Diffusion Component

1. This paper reports an investigation of whether the dipeptides glycylsarcosine and l-glutamyl-l-glutamic acid share a single mediated transport mechanism into hamster jejunum, or whether one of these peptides is transported in part by a transport mechanism unavailable to the other. It describes the kinetics of influx of glycylsarcosine and of l-glutamyl-l-glutamic acid into rings of everted hamster jejunum in vitro, incubations being carried out at pH 5 in order to minimize brush-border and intra-medium hydrolysis of l-glutamyl-l-glutamic acid, so that peptide transport rather than a mixture of peptide transport and transport of free glutamic acid was studied. With glycylsarcosine, brush-border and intra-medium hydrolysis are negligibly small. 2. Estimates of the simple diffusion component in transport of each peptide were made by treating each of the substrates as a competitive inhibitor of its own mediated transport (assuming that mediated transport conforms to simple Michaelis-Menten kinetics), extrapolating the observed inhibitory effect over a range of concentrations to an infinitely high concentration of inhibitor, and estimating the transport component remaining at such a concentration. This component in transport would be expected to represent transport by simple diffusion, and this assumption was supported by the observation that for glycylsarcosine the uninhibitable component in transport was linearly proportional to substrate concentration; with l-glutamyl-l-glutamic acid the observations were too few to provide this demonstration. Estimates of apparent Kt and Vmax. for mediated transport of both peptides are given. Before correction for simple diffusion, linearizing plots were clearly biphasic for both peptides; after correction for simple diffusion, they became linear, providing no evidence for transport of either peptide by more than one mediated transport system, though not excluding the possibility of multiple systems. 3. Measurement of influx of [14C]Gly-Sar over a range of concentrations both alone and in the presence of a constant concentration of Glu-Glu showed that after correction for the non-mediated component in influx of Gly-Sar (simple diffusion), influx of this peptide conformed to Michaelis-Menten kinetics and the inhibitory effect of Glu-Glu on influx of Gly-Sar appeared to be competitive. The extent of inhibition corresponded well with that predicted from the Kt values of the two peptides. 4. Measurement of influx of [14C]Gly-Sar (1 mmol/l) in the presence of a range of concentrations of Glu-Glu, with extrapolation of the inhibitory effect of Glu-Glu to an infinitely high concentration of this peptide, showed that at such a concentration mediated influx of Gly-Sar was completely abolished, influx being reduced to the simple diffusion component in total influx of [14C]Gly-Sar. Measurement of influx of [14C]Glu-Glu (1 mmol/l) in the presence of a range of concentrations of Gly-Sar, with extrapolation of the inhibitory effect of Gly-Sar to an infinitely high concentration of this peptide, showed that at such a concentration mediated influx of Glu-Glu was completely abolished, influx being reduced to the simple diffusion component in total influx of [14C]Glu-Glu. 5. The results are compatible with the conclusion that Gly-Sar and Glu-Glu are taken up by the absorptive cells by a single mediated mechanism. They do not exclude the possibility that these peptides are taken up by multiple common mechanisms, but they do appear to exclude the possibility that at the substrate concentration used (1 mmol/l) there is appreciable uptake of one of the peptides by a system unavailable to the other.

THE METABOLISM OF ANIMAL TISSUES CULTIVATED IN VITRO: III. AMINO ACID METABOLISM OF STRAIN L CELLS IN COMPLETELY SYNTHETIC MEDIA

1958 ◽  
Vol 36 (1) ◽  
pp. 771-782 ◽  
Author(s):  
Arthur E. Pasieka ◽  
Helen J. Morton ◽  
Joseph F. Morgan
Keyword(s):  
Amino Acid ◽  
Glutamic Acid ◽  
Culture Medium ◽  
Synthetic Medium ◽  
Amino Acid Content ◽  
Amino Acid Uptake ◽  
Acid Uptake ◽  
L Cells ◽  
Synthetic Media

Strain L cells, of mouse fibroblastic origin, have been cultivated in vitro in completely synthetic medium M 150 and in various modifications of this medium. The amino acid changes in the nutrient medium during cell cultivation have been studied by paper chromatography. A characteristic pattern of amino acid uptake and accumulation in the medium has been found. No change in the alanine concentration was observed but omission of alanine from the culture medium resulted in its accumulation in appreciable amounts. Omission of glutamic acid did not alter the pattern of amino acid changes by the cells. Omission of glutamine increased the uptake of amino acids and prevented amino acid accumulation. Omission of both glutamic acid and glutamine resulted in a virtual cessation of amino acid changes in the culture medium. Strain L cells decreased the adenine content of the medium and produced small amounts of hypoxanthine. These changes were not affected by alterations in the amino acid content of the medium. Omission of glutamic acid and glutamine from the culture medium did not cause an appreciable decrease in cell population or apparent degeneration of the cultures over a 30-day period.

THE METABOLISM OF ANIMAL TISSUES CULTIVATED IN VITRO: III. AMINO ACID METABOLISM OF STRAIN L CELLS IN COMPLETELY SYNTHETIC MEDIA

1958 ◽  
Vol 36 (7) ◽  
pp. 771-782 ◽  
Author(s):  
Arthur E. Pasieka ◽  
Helen J. Morton ◽  
Joseph F. Morgan
Keyword(s):  
Amino Acid ◽  
Glutamic Acid ◽  
Culture Medium ◽  
Synthetic Medium ◽  
Amino Acid Content ◽  
Amino Acid Uptake ◽  
Acid Uptake ◽  
L Cells ◽  
Synthetic Media

Strain L cells, of mouse fibroblastic origin, have been cultivated in vitro in completely synthetic medium M 150 and in various modifications of this medium. The amino acid changes in the nutrient medium during cell cultivation have been studied by paper chromatography. A characteristic pattern of amino acid uptake and accumulation in the medium has been found. No change in the alanine concentration was observed but omission of alanine from the culture medium resulted in its accumulation in appreciable amounts. Omission of glutamic acid did not alter the pattern of amino acid changes by the cells. Omission of glutamine increased the uptake of amino acids and prevented amino acid accumulation. Omission of both glutamic acid and glutamine resulted in a virtual cessation of amino acid changes in the culture medium. Strain L cells decreased the adenine content of the medium and produced small amounts of hypoxanthine. These changes were not affected by alterations in the amino acid content of the medium. Omission of glutamic acid and glutamine from the culture medium did not cause an appreciable decrease in cell population or apparent degeneration of the cultures over a 30-day period.

The mechanisms of amino acid uptake byBrugia pahangi in vitro

1983 ◽  
Vol 69 (2) ◽  
pp. 247-253 ◽  
Author(s):  
R. E. Howells ◽  
A. M. Mendis ◽  
P. G. Bray
Keyword(s):  
Amino Acid ◽  
Amino Acid Uptake ◽  
Acid Uptake

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.

The uptake of amino acids by mouse cells (strain LS) during growth in batch culture and chemostat culture: the influence of cell growth rate

1967 ◽  
Vol 168 (1013) ◽  
pp. 421-438 ◽  
Keyword(s):  
Amino Acids ◽  
Growth Rate ◽  
Amino Acid ◽  
Cell Growth ◽  
Glutamic Acid ◽  
Batch Culture ◽  
Amino Acid Uptake ◽  
Essential Amino Acids ◽  
Growth Yields ◽  
Acid Uptake

The uptake of thirteen essential amino acids by mouse LS cells in suspension culture was determined by bacteriological assay methods. Chemostat continuous-flow cultures were used to determine the effect of different cell growth rates on the quantitative amino acid requirements for growth. The growth yields of the cells ( Y = g cell dry weight produced/g amino acid utilized) were calculated for each of the essential amino acids. A mixture of the non-essential amino acids, serine, alanine and glycine increased the cell yield from the essential amino acids. The growth yields from nearly all the essential amino acids in batch culture were increased when glutamic acid was substituted for the glutamine in the medium. The growth yields from the amino acids in batch culture were much less at the beginning than at the end of the culture. The highest efficiencies of conversion of amino acids to cell material were obtained by chemostat culture. When glutamic acid largely replaced the glutamine in the medium the conversion of amino acid nitrogen to cell nitrogen was 100 % efficient (that is, the theoretical yield was obtained) at the optimum growth rate (cell doubling time, 43 h). The maximum population density a given amino acid mixture will support can be calculated from the data. It is concluded that in several routinely used tissue culture media the cell growth is limited by the amino acid supply. In batch culture glutamine was wasted by (1) its spontaneous decomposition to pyrrolidone carboxylic acid and ammonia, and (2) its enzymic breakdown to glutamic acid and ammonia, but also glutamine was used less efficiently than glutamic acid. Study of the influence of cell growth rate on amino acid uptake rates per unit mass of cells indicated that a marked change in amino acid metabolism occurred at a specific growth rate of 0.4 day -1 (cell doubling time, 43 h). With decrease in specific growth rate below 0.4 day -1 there was a marked stimulation of amino acid uptake rate per cell and essential amino acids were consumed increasingly for functions other than synthesis of cell material.

scholarly journals Rationally Design of a Proline Transporter Inhibitor with Anti-Trypanosoma Cruzi Activity

2019 ◽  
Author(s):  
Lucia Fargnoli ◽  
Esteban A. Panozzo-Zénere ◽  
Lucas Pagura ◽  
María Julia Barisón ◽  
Julia A. Cricco ◽  
...  
Keyword(s):  
Amino Acid ◽  
Trypanosoma Cruzi ◽  
Chagas Disease ◽  
Variable Region ◽  
Amino Acid Uptake ◽  
Acid Uptake ◽  
Cell Infection ◽  
Atp Production ◽  
First Case

L-Proline is an important amino acid for the pathogenic protists belonging to <i>Trypanosoma</i> and <i>Leishmania </i>genera. In <i>Trypanosoma cruzi</i>, the etiological agent of Chagas disease, this amino acid is involved in fundamental biological processes such as ATP production, differentiation of the insect and intracellular stages, the host cell infection and the resistance to a variety of stresses, including nutritional and osmotic as well as oxidative imbalance. In this study, we explore the L-Proline uptake as a chemotherapeutic target for <i>T. cruzi</i>. For this, we propose a novel rational to design inhibitors containing this amino acid as a recognizable motif. This rational consists of conjugating the amino acid (proline in this case) to a linker and a variable region able to block the transporter. We obtained a series of sixteen 1,2,3-triazolyl-proline derivatives through alkylation and copper(I)-catalyzed azide-alkyne cycloaddition (click chemistry) for <i>in vitro</i> screening against <i>T. cruzi </i>epimastigotes, trypanocidal activity and proline uptake. We successfully obtained inhibitors that are able to interfere with the amino acid uptake, which validated the first example of a rationally designed chemotherapeutic agent targeting a metabolite's transport. Additionally, we designed and prepared fluorescent analogues of the inhibitors that were successfully taken up by <i>T. cruzi</i>, allowing following up their intracellular fate. In conclusion, we successfully designed and produced a series of metabolite uptake inhibitors. This is one of few examples of rationally designed amino acid transporter inhibitor, being the first case where the strategy is applied on the development of chemotherapy against Chagas disease. This unprecedented development is remarkable having in mind that only a small percent of the metabolite transporters has been studied at the structural and/or molecular level.

THE METABOLISM OF ANIMAL TISSUES CULTIVATED IN VITRO: II. AMINO ACID METABOLISM OF CHICK EMBRYONIC KIDNEY, CHICK EMBRYONIC LIVER, AND MONKEY KIDNEY CORTEX CULTURES

1958 ◽  
Vol 36 (1) ◽  
pp. 171-184 ◽  
Author(s):  
Arthur E. Pasieka ◽  
Helen J. Morton ◽  
Joseph F. Morgan
Keyword(s):  
Tissue Culture ◽  
Amino Acid ◽  
Culture Medium ◽  
Synthetic Medium ◽  
Amino Acid Uptake ◽  
Monkey Kidney ◽  
Kidney Cortex ◽  
Acid Uptake ◽  
Embryonic Liver

Freshly-explanted chick embryonic kidney, chick embryonic liver, and trypsinized monkey kidney cortex cells have been cultivated in vitro in completely synthetic medium M 150. The amino acid changes in the nutrient medium during cultivation of these tissues have been studied by paper chromatography. A characteristic pattern of amino acid uptake and accumulation in the used culture medium has been demonstrated with each type of tissue culture. It has also been shown that, while the amino acid changes in the medium are different with each type of tissue culture, all cultures examined removed adenine from the medium and liberated small amounts of material thought to be hypoxanthine.

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