Amino acid and vitamin requirements in mammalian cultured cells

Summary The development of the tissue culture technique has enabled us to cultivate mammalian cells in a way which is similar to that in use with bacterial cells. As such, the nutritional requirements of mammalian cells in culture have been studied with simplicity and exactness. According to Eagle's extensive works it is accepted that cultured cells generally require 13 amino acids, 8 or 9 vitamins, glucose and 6 inorganic salts. However, although some cultured cells have a capacity for the biosynthesis of Eagle's essential nutrients and others require non-essential nutrients.In this review we will discuss the amino acid and vitamin requirements of cultured cells, and a cell line (R-Y121B · cho) which propagates continuously in a chemically defined medium containing 11 amino acids, 7 vitamins, glucose and 6 ionic salts. Arginine, glutamine, tyrosine and choline are synthesized in the R-Y121B · cho cells.     

The components of taurine transport across the rat small intestine A kinetic study

Summary The transport of taurine across adult Sprague-Dawley rat small intestine was studied in vitro using small intestinal strips. The kinetics of the transport mechanism were investigated under both steady-state and influx conditions. Our findings were compatible with the presence of two distinct transport mechanisms; a linear non-carrier mediated component and a saturable carrier mediated component, with almost equal contribution from each. The mediated component was found to be largely Na⁺-dependent and exhibited marked inhibition by B-alanine and structurally related sulfur amino acids.     

Amino acid determination in some edible Mexican insects

Summary The amino acid contents of edible insects from different provinces of Mexico and reference proteins were analysed by reversed-phase high-performance liquid chromatography and ion exchange chromatography. The insect amino acid contents were higher than the adult requirements indicated by the WHO/FAO pattern.     

Evolution of the amino acid substitution in the mammalian myoglobin gene

Summary Multivariate statistical analyses were applied to 16 physical and chemical properties of amino acids. Four of these properties; volume, polarity, isoelectric point (charge), and hydrophobicity were found to explain adequately 96% of the total variance of amino acid attributes. Using these four quantitative measures of amino acid properties, a structural discriminate function in the form of a weighted difference sum of squares equation was developed. The discriminate function is weighted by the location of each particular residue within a given tertiary structure and yields a numerical discriminate or difference value for the replacement of these residues by different amino acids. This resulting discriminate value represents an expression of the perturbation in the local positional environment of a protein when an amino acid substitution occurs. With the use of this structural discriminate function, a residue by residue comparison of the known mammalian myoglobin sequences was carried out in an attempt to elucidate the positions of possible deviations from the known tertiary structure of sperm whale myoglobin. Only 11 of the 153 residue positions in myoglobin demonstrated possible structural deviations. From this analysis, indices of difference were calculated for all amino acid exchanges between the various myoglobins. All comparisons yielded indices of difference that were considerably lower than would be expected if mutations had been fixed at random, even if the organization of the genetic code is taken into consideration. On the basis of these results, it is inferred that some form of selection has acted in the evolution of mammalian myoglobins to favor amino acid substitutions that are compatible with the retention of the original conformation of the protein.     

The role of system A for neutral amino acid transport in the regulation of cell volume

System A is a secondary active, sodium dependent transport system for neutral amino acids. Strictly coupled with Na,KATPase, its activity determines the size of the intracellular amino acid pool, through a complex network of metabolic reaction and exchange fluxes. Many hormones and drugs affect system A activity in specific cell models or tissues. In all the cell models tested thus far the activity of the system is stimulated by amino acid starvation, cell cycle progression, and the incubation under hypertonic conditions. These three conditions produce marked alterations of cell volume. The stimulation of system A activity plays an important role in cell volume restoration, through an expansion of the intracellular amino acid pool. Under normal conditions, system A substrates represent a major fraction of cell compatible osmolytes, organic compounds that exert a protein stabilizing effect. It is, therefore, likely that the activation of system A represents a portion of a more complex response triggered by exposure to stresses of various nature. Since system A transporters have been recently cloned, the molecular bases of these regulatory mechanisms will probably be elucidated in a short time.     

Amino acids as regulators of gene expression: molecular mechanisms

Regulation of gene expression by nutrients in mammals is an important mechanism allowing them to adapt their physiological functions according to the supply of nutrient in the diet. It has been shown recently that amino acids are able to regulate by themselves the expression of numerous genes. CHOP, asparagine synthetase, and IGFBP-1 regulation following AA starvation will be described in this review with special interest in the molecular mechanisms involved.     

Amino acid profiles in some scented rice varieties

Summary Twelve scented (basmati) and one non-scented variety were analysed for their amino acid composition. The essential amino acid profiles of scented varieties when compared with non-scented, revealed that these varieties exhibited higher values, which ranged from 2.82 to 4.86 gm/100 gm protein for lysine, 1.92 to 3.13 for methionine, 1.67 to 4.23 for tyrosine, 3.65 to 4.91 for phenylalanine, 5.50 to 8.95 for leucine, 2.25 to 3.40 for isoleucine, 2.84 to 3.46 for threonine, 3.36 to 5.33 for valine. When these values were compared to FAO recommended standards, it was observed that most of the scented varieties had comparable or superior values, while varieties such as, Type 3, Basmati sufaid 100, Likitimachi, Randhunipagalu and Basmati 370 showed superior lysine, phenylalanine, leucine, and methionine content. These observations suggest that the scented varieties posses better amino acid profiles and exhibit superior nutritional qualities, which could be utilised in breeding varieties with improved amino acid composition.     

Amino acid studies in transient acute polymorphic psychosis

Summary Data are reviewed on the amino acid metabolism in patients suffering from transient acute polymorphic psychoses according to ICD-10. The psychotic episodes of many of these patients are characterized by distorted sensory perceptions and intense emotional states; plasma amino acid analysis revealed a disturbed serine-glycine metabolism.In remitted patients oral loading with serine induced the characteristic dysperceptions and psychedelic symptoms. Plasma concentrations of serine and methionine were decreased, and the concentration of taurine was increased. Fibroblast experiments suggest that the activities of the serine metabolizing enzymes serine hydroxymethyltransferase and cystathionine-synthase are increased in these patients.The determination of plasma amino acid concentrations proved to be useful in discriminating these patients. The prevalence of this transient psychosis in a psychiatric in-patient population ranged between 1.4 and 3.6%.     

Amino acid limited growth of starter cultures in milk

The specific growth rates of several Streptococcus cremoris strains were 10–40% lower in milk than in other growth in media. The growth rates in milk increased when an amino acid mixture or casein was added, whereas, when milk was diluted, the specific growth rate of the streptococci decreased. This decrease could be overcome by bringing the casein concentration in the diluted milk back to the normal value (3%). This indicates that casein-hydrolysis proceeded at a rate too low for the streptococci to reach their potential maximum specific growth rates in milk so that growth in milk is essentially amino acid-limited. This was subsequently demonstrated for S. cremoris by continuous cultivation in media with low casein concentrations. At a low dilution rate casein hydrolysis was fast enough to supply the cells with enough amino acids and lactose was growth-limiting, whereas at higher dilution rates amino acids became growth-limiting. In cultures exponentially growing in milk the concentration of free amino acids was measured to determine which amino acid(s) was(were) absent and could possibly limit growth. A number of essential amino acids (leucine, methionine, glutamate and in some cases phenylalanine) were not detected and addition of these, together, stimulated the growth of S. cremoris in milk. The amino acids leucine and phenylalanine appeared to play a particularly important role in this stimulation. These two are, supposedly, the first amino acids that become limiting during growth in milk. The effect of competition for casein and amino acids by different organisms was studied in continuous cultures. At different dilution rates different strains became dominant in these mixed cultures, suggesting that differences in apparent affinity constants (K_S) for casein, leucine and glutamate existed between the strains.     

Effect of Cortico-Striate Pathway Lesion on the Activities of Enzymes Involved in Synthesis and Metabolism of Amino Acid Neurotransmitters in the Striatum

The activities of several enzymes involved in the metabolism of aspartate and glutamate were measured in striatal (nucleus caudatus and putamen) homogenates 2-3, 6-7, and 35-40 days following frontoparietal and frontal cortical ablation. The activity of glutamine synthetase (GS) was substantially increased (46-48%) on the operated side 6-7 days following the lesion whereas smaller changes were observed at 2-3 and 35-40 days after lesion. In contrast, decreased levels of glutaminase and malate dehydrogenase (MDH) were observed by 6-7 days while no significant change was found at either 2-3 or 35-40 after the lesion. The activities of glutamate dehydrogenase (GDH) and glutamate decarboxylase (GAD) were elevated after 35-40 days whereas no changes in the levels of either GDH or aspartate aminotransferase (ASAT) were found at 2-3 or 6-7 days after the fronto-parietal decortication. When only the frontal cortex was removed quantitatively similar changes were observed in striatal GS and glutaminase activity. The content of glutamate and glutamine in the denervated striatum followed qualitatively the changes in glutaminase and GS. The results indicate that the degeneration of cortico-striatal terminals causes a profound glial reaction in the striatum, and both glutaminase and MDH are present in relatively high concentrations in the corticostriatal terminals.     

Amino acid influx across the mucosal border of the rat intestine in vivo

The lack of an in vivo method for measuring influx of amino acid into the mucosa has prevented a systematic comparison of characteristics of amino acid influx in vivo with prior in vitro studies. We developed and validated a technique for measuring amino acid influx in vivo. The mucosa is exposed briefly to labelled amino acid perfused luminally at a rapid rate and tissue uptake is measured. The brief exposure period insures that amino acid is confined to the segment. The rapid perfusion rate minimizes concentration of endogenous Na⁺ in the lumen and permits Na-dependency for α-aminoisobutyric acid influx to be demonstrated in vivo for the first time. We also demonstrated the inhibitory effect of K⁺ and competition by glycine on α-aminoisobutyric acid influx in vivo. The saturation kinetics for l-leucine in vivo and in vitro were compared under varying perfusion rates and with and without stirring with air. Under optimal conditions of agitation (rapid perfusion and bubbling with air), the apparent Michaelis constant ($\text{K}{}_{\mathrm{<mtext>t</mtext>}}$) is decreased to be almost equal to that determined under comparable influx conditions in vitro. These studies demonstrate no major difference between characteristics of amino acid transport under more physiologic in vivo conditions as compared with prior in vitro studies.     

Amino acid flooding doses for measuring rates of protein synthesis

Summary The development, advantages and disadvantages of using the amino acid flooding dose technique to determinein vivo rates of protein synthesis are examined in this review. A discussion of the use of this procedure in animals greater than 5 kg is included. The flooding dose procedure reduces the disparity between isotope enrichment in different amino acid precursor pools, which should theoretically improve the precision and accuracy of protein synthesis measurements. However, the possibility must be considered that the large doses of amino acids injected or infused in conjunction with this technique may influence protein turnover due to attendant metabolic effects. Therefore, a judicious choice of an amino acid and an evaluation of the experimental parameters involved in this procedure are required to optimize the accuracy of results obtained.Scientific Paper No. 710. Agriculture Canada, Lacombe Research Station, Bag Service 5000, Lacombe, Alberta T0C 1S0.     

Amino acid modifications in canine, equine and porcine pituitary growth hormones, identified by peptide-mass mapping

Modified amino acid residues in porcine, canine and equine growth hormones purified from pituitary glands were characterised by tryptic mapping and high-performance liquid chromatography with on-line coupled electrospray ionisation mass spectrometry (HPLC–ESI-MS) detection. Hormones from all three species showed the same changes. Conversion of Asp¹²⁸ to iso-Asp¹²⁸ was a component of native hormones, while deamidation of Asn¹² and Asn⁹⁸ to Asp and iso-Asp, oxidation of Met⁴, and cyclisation to the pyroglutamyl derivative of Gln¹³⁹, probably occurred in vitro, during isolation, storage or hydrolysis. Porcine and canine hormones had indistinguishable protein fingerprints, confirming the assumption, based on their cDNA sequences, that their mature primary structures are identical.     

Molecular regulation of amino acid biosynthesis in plants

Summary Our understanding of amino acid biosynthesis in plants has grown by leaps and bounds in the last decade. It appears that most of the amino acid biosynthesis takes place in the chloroplast. Recent demonstration of glutamine synthetase and DAHP synthase in the vascular tisuue has added a new dimension in the complexity of the nitrogen cycle in plants. Isolation of various genes and transformation of plants with the modified forms of the genes are providing tools for understanding the regulation of various pathways. Plant transformation approaches are also going to provide the food of the future with an improved amino acid composition.     

良种茶树芽叶中氨基酸研究

本文报导利用日立－ＥＧ型氨基酸自动分析仪测定的六个茶树良种芽叶中十九种氨基酸组成情况,并以此为依据对良好的品质风格进行了探讨。