Origin of Amino Acids in Datura stramonium Seeds

¹⁴C isotope studies show that the seeds of Datura stramoniumL. can produce a number of amino acids (particularly alanine,glutamate, phenylalanine, and tyrosine) from a supply of sucroseand nitrate. These amino acids can be incorporated into theseed protein. The bulk of the amino acids incorporated into the seed proteinmust, however, be supplied by adult leaves in the proximityof the fruit, either as the amino acids themselves, or theirimmediate precursors. The major free-amino-acid products of Datura leaves are theamides asparagine and glutamine.     

The Inhibitory Effects of Various Amino Acids on the Growth of Barley Seedlings

The effect of fifteen amino acids, supplied singly, on the growthof isolated germinating barley embryos in the presence of nitratehas been studied. The L forms of lysine, arginine, tyrosine,proline, threonine, methionine, leucine, and valine at concentrationsof either 1 or 2 mM have been found to inhibit fresh-weightaccumulation. The inhibition by valine is relieved by furtheraddition of isoleucine and that of leucine by the addition ofboth isoleucine and valine. These interrelations have been interpretedas suggesting that leucine and valine can inhibit acetolactateand acetohydrorybutyrate synthesis. The inhibition of tyrosinecan be relieved by phenylalanine and that of lysine by ornithineor arginine. The possible reasons for these interrelationshipsare discussed.     

Kinetics and Specificity of Amino Acid Uptake by the Duckweed Spirodela polyrhiza (L.) Schleiden

Borstlap, A. G, Meenks, J. L. D., van Eck, W. F. and Bicker,J. T. E. 1986. Kinetics and specificity of amino acid uptakeby the duckweed Spirodela polyrhiza (L.) Schleiden.—J.exp. Bot. 37: 1020–1035. Uptake of ¹⁴C-labelled amino acids by intact, axenically grownplants of Spirodela polyrhiza (L.) Schleiden was investigated.Experiments in which uptake was measured from the decrease inthe amino acid concentration in the medium, indicated that saturableuptake conforms to the sum of two Michaelis-Menten terms, possiblycorresponding with a high-affinity and a low-affinity system.Further experiments with L-leucine, L-glutamic acid, and L-lysine,in which uptake was measured by assaying the amount of ¹⁴ inthe plants, showed the presence of a non-saturable componentin addition to the dual saturable uptake. Uptake of L-glutamic acid precipitously declined between pH4?0 and 6? and that of L-leucine between pH 4?0 and 8? whereasL-lysine uptake was optimal at pH 6?0. No evidence was foundthat the apparent high-affinity and low-affinity systems respondeddifferently to changes in external pH or to the addition ofCCCP. The non-saturable uptake component was not affected bychanges in external pH or by adding CCCP, and might have beendue to free space uptake. Mutual inhibition of uptake was found between acidic and neutralamino acids (L-leucine, L-methionine, L-glutamic acid) and betweenbasic amino acids (L-lysine, L-ornithine). The basic amino acidshad no effect on the uptake of L-leucine, L-methionine and L-glutamicacid, although the uptake of basic amino acids was inhibitedby glutaminc acid and several neutral amino acids. It is suggested that the duckweed has a high-affinity transportsystem for neutral and acidic amino acids, and a distinct high-affinitysystem for basic amino acids. It is argued that the first systemtransports zwitterionic amino acids (z-system), and that thesecond system transports cationic amino acids(y⁺-system). Thespecificity of the low-affinity system is less certain, butthere is some evidence that it is similar to that of their high-affinitycounterparts. Key words: Kinetics, membrane transport, pH-dependency, transport systems, uptake isotherms     

General properties and regulation of arginine transporting systems in Saccharomyces cerevisiae

The transport of arginine-¹⁴C by exponentially growing cellsof Saccharomyces cerevisiae (ATCC 9763) was studied in the presenceof various amino acids, ammonium and urea. Arginine transportwas inhibited when the cells were preincubated with these compoundsfor 1 hr. Little or no inhibition of transport occurred whenthe preincubation period was omitted. Kinetics studies revealedthat arginine was transported by two distinct systems havinghigh and low affinities for this amino acid. At given arginineconcentrations the high affinity system was capable of transportingarginine molecules at approximately seventy times the rate ofthe low affinity system. The general requirements for arginine transport revealed energyand temperature dependencies in addition to sensitivity to anumber of metabolic inhibitors. Transfer of cells to N-freemedium was accompanied by increased rates of transport. Thisincrease was shown for the uptake of ten different amino acids.For L-arginine, this increase was prevented by addition of cycloheximide. Analyses of amino acid pools, after various experimental treatments,failed to reveal any consistent correlation between transportrates and the concentrations of individual amino acids or ammonium. It is concluded that arginine transport of S. cerevisiae isregulated by inhibition and repression. In this respect theavailability of ammonium would appear to be of prime importancein the development of transport activity. (Received December 5, 1975; )     

UPTAKE SPECIFICITY FOR ORGANIC SUBSTRATES BY THE MARINE DIATOM MELOSIRA NUMMULOIDES1

Melosira nummuloides, clone Mel-3, shows a very high specificity with regard to its ability to take up organic substrates. Amino acids supplied in the medium at 1 X 10^-4 M are taken up at initial rates of the same order of magnitude as that of photoassirnilation of COj. However, sugars, sugar alcohols, or organic acids supplied at the same concentration are not taken up. The mechanism for uptake of amino acids appears to require energy, since tlie uptake of the amino acid analog α-aminoisobutyric acid is strongly inhibited by 2 f-dinitrophenul. The uptake mechanism does not appear to be inducible. The ability of M. numinuloides to utilize amino acids as a nitrogen source is quite restricted. Arginine, ghttamine, asparagine, proline, and glutamic acid were good nitrogen sources. Seventeen other amino acids, including α-aminoisobutyric acid, were unsatisfactory for growth, although they were rapidly taken up from the medium.     

The Regulation of Nitrite Reductase Level in Lemna minor L.

The regulation of nitrite reductase in Lemna minor has beenstudied. The evidence indicates that in nitrate-fed plants nitrateitself is the inducer of nitrite reductase. The enzyme is subjectto end-product repression by ammonia and various amino acids.Nitrate reductase is also repressed by a similar range of compounds.Most of the repressors tested are more effective when nitraterather than nitrite is supplied as the inducer. The effectsof cyclo-heximide, D-threo-chloramphenicol and lincomycin onthe induction by nitrate and nitrite suggest that both enzymesare synthesized on cytoplasmic ribosomes. The mechanism of repressionby ammonia and amino acids is discussed.     

Nitrate Assimilation in Higher Plants with Special Reference to the Cocklebur (Xanthium pennsylvanicum Wallr.)

A soluble NADH-dependent nitrate reductase is described forthe shoot system of Xanthium. Young leaves and immature stemtissues contain high levels of the enzyme. They are relativelyrich in free amino acids and amides but store little free nitrate.The specific activity of the enzyme is lower in fully expandedleaves, although these leaves exhibit higher rates of fixationof carbon in photosynthesis than do younger leaves. Neithernitrate nor free amino acids accumulate in the mesophyll ofthe leaf. Older parts of the stem axis accumulate large amountsof soluble nitrogen, almost entirely as free nitrate. Reservesof nitrate in the shoot and root are rapidly depleted if nitrateis removed from the external medium. Nitrate reductase is apparently absent from roots of Xanthium.This finding is supported by analyses of bleeding sap from nitrate-fedplants which show that 95 per cent of the nitrogen exportedfrom roots is present as free nitrate. However, roots are capableof synthesizing and exporting large amounts of amino nitrogenif supplied with reduced nitrogen such as urea or ammonium. A scheme is presented summarizing the main features of the metabolismof nitrate in Xanthium and this is compared with the situationin nitrate-fed plants of the field pea (Pisum arvense L.), aspecies previously shown to be capable of reducing nitrate inits root system.     

Uptake of Lysine by Wild-Type and S-(2-Aminoethyl)-L-cysteine-Resistant Suspension-Cultured Cells of Triticum aestivum

The kinetics of uptake of L-lysine in wheat (Triticum aestivumcv. Chinese Spring) were analyzed in wild-type cells and inAEC-1 variant cells that are resistant to S-(2-aminoethyl)-L-cysteine(AEC). Uptake of lysine by AEC-1 cells was considerably slowerthan that by the wild-type cells. In the presence of carbonylcyanidem-chlorophenylhydrazone, the rates of uptake by both types ofcell were reduced to a similar linear component. Fitting theuptake data to one linear (diffusional) component and one Michaelis-Menten(active) system showed that, as compared to wild-type cells,AEC-1 cells have a reduced V_max and an increased K_m with respectto the active component, byt they have a similar diffusionalcomponent. Inhibition experiments with various amino acids indicatedthat the active component represents a carrier specific forbasic amino acids, which was competitively inhibited by AEC.The AEC-1 cells also showed reduced uptake of several neutraland acidic amino acids, but the rate of uptake of 3-O-methylglucosewas somewhat higher than that by wild-type cells. (Received May 16, 1989; Accepted September 4, 1989)     

Dissolved Organic Matter as a Potential Source of Nutrition for Marine Organisms

The general features of accumulation of amino acids by marineinvertebrates are outlined. The wide distribution of this abilityis noted. Data from observations of this process for periodsof several days are presented for the polychaete worm, Doruilleaarticulata. Measurements of the constituents in the pool offree amino acids of this same organism are presented togetherwith data concerning the rate of appearance of radioactivityin the medium after permitting animals to accumulate amino acids.These data permit tentative estimates of rates at which aminoacids are lost to the medium. These apparent "leakage" ratesare low when compared with rates of uptake. The thermodynamicwork necessary to move material against the concentration differencesinvolved is calculated. The fraction of metabolic energy neededis small. The possible biological significance of uptake ofsmall organic compounds is discussed. It is concluded that uptakeof amino acids occurs sufficiently generally and at such ratesthat it should be included in any analysis of the pathways wherebymaterial is acquired from the environment by marine organisms.     

Free amino acid levels and the regulation of nitrate uptake in maize cell suspension cultures

The ability of individual amino acids to regulate nitrate uptakeand induction was studied in a Zea mays embryo cell line grownin suspension culture. The maize cells exhibited a marked preferencefor absorbing amino acids over nitrate when both were presentin culture medium. The addition of an individual amino acid(2 mM glutamine, glycine, aspartic acid, or arginine) to theculture medium with 1 mM nitrate completely inhibited nitrateuptake and resulted in a cycle of low levels of nitrate influxfollowed by efflux to the growth medium. Glutamine was readilyabsorbed by the cells and was particularly effective in supportingoptimum cell growth in the absence of an inorganic nitrogensource as compared to the three other amino acids evaluated.However, neither glutamine nor any of the remaining 19 proteinaceousamino acids appeared to be solely responsible for regulationof nitrate uptake and induction. The ability of amino acidsto regulate nitrate uptake and assimilation appears to be morerelated to their overall levels in the cell rather than to anaccumulation of a specific amino acid. Key words: Amino acids, nitrate uptake, maize, regulation, cell suspension culture     

Mobilization of Endosperm Reserves and Uptake of Sucrose and Valine by the Cotyledons of Euphorbia lathyris L.

Upon germination, the endosperm triacylglycerols and proteinswere converted to sucrose and amino acids. During early postgerminativegrowth, the rate of sucrose and amino acid production exceededthe rate of uptake by the cotyledons. As a result, the levelsof total amino acid and sucrose in the endosperm increased;maximum levels were reached at 7 d and 10 d after imbibition(DAI), respectively. Intact seedlings were used to measure thedevelopment of valine, arginine, glutamic acid, and sucroseuptake rate throughout the course of endosperm depletion. Maximumamino acid uptake rates were measured at around 9 DAI, the highestuptake rate for sucrose was obtained at 12 DAI (just beforedepletion of the endosperm). The daily increase of sucrose andamino acid uptake could be manipulated, by replacing the endospermwith a pre-incubation solution during 1 d. The increase in sucroseuptake in vitro was equal to that measured with intact seedlingswhen the cotyledons were pre-incubated in 10 mol m^–3 sucrose.Higher sucrose concentrations reduced the increase of sucroseuptake; at 300 mol m^–3 sucrose (corresponding to the meanendosperm sucrose concentration) sucrose uptake after pre-incubationwas even lower than before. This reduction was largely counteractedwhen the pre-incubation solution was supplemented with minerals.The development of the valine uptake was hardly affected bysucrose, but was inhibited by several amino acids. Key words: Euphorbia lathyris seedling, sucrose uptake, amino acid uptake, reserve mobilization     

Metabolism of Exogenous Malonate by Coffee Leaf Tissue

When [l-¹⁴C]-malonate was supplied to discs cut from matureleaves of Coffea arabica, ¹⁴CO₂ was released (approximately12% of the total CO₂ respired) and organic acids of the Krebscycle, uronic acids, sugars and amino acids became radioactive.There was no incorporation of MC into either lipids or phenoliccompounds. The formation of glucose from malonate has not beenobserved in other studies with plant tissues. The synthesisof labelled glucose together with an active pentose phosphatepathway that is stimulated by malonate explains the accumulationof radioactive phosphogluconate in the leaf discs. Tentativeproposals are made for pathways to account for the results obtained. Key words: Coffee leaves, Malonate metabolism, Pentose phosphate pathway     

Effect of the Osmotic Environment on Assimilate Transport in Isolated Developing Embryos of Maize (Zea mays)

Embryos obtained from developing kernels of maize (Zea maysL.) were incubated in a bathing medium, to measure the effectof the osmotic environment on the balance between uptake andrelease of assimilates by the embryo. Net efflux of sucroseand amino acids from the embryo decreased with increasing mannitolconcentration in the bathing medium and net uptake of [¹⁴C]valine increased with increasing mannitol concentration. Therole of a high osmolality of the seed apoplast in seed developmentis discussed Zea mays, maize, embryo, seed development, assimilate transport, turgor-sensitive transport     

Nitrogenous nutrition of the potentially toxic diatom Pseudonitzschia pungens f. multiseries Hasle

The potentially toxic diatom Pseudonitzschia pungens f. multiserieswas grown on different sources of nitrogen in batch cultures.Ammonium did not support growth at concentrations >200 µM,and even lowered the growth rate, when it was supplied in additionto growth-saturating nitrate concentrations. This seemed tobe a combined effect of inhibition of nitrate uptake and directammonia toxicity. Urea, glutamine and nitrite were used readilyby P.pungens.     

Nitrogen and Nitrate Accumulation in Species Having Different Relationships Between Nitrate Uptake and Reduction

Species differ in the relationship of nitrate reductase activityto nitrate uptake. In Capsicum annuum different diurnal patternsof leaf nitrate reductase activity and nitrate uptake have beenreported. As a consequence, the relationship of free nitratein the plant to nitrate supplied has a higher level of significancethan has reduced nitrogen to nitrate supplied. In Zea mays ithas been reported that leaf nitrate reductase activity respondsdirectly to nitrate translocation to the leaf and in this speciesthe relationship of greatest significance is reduced nitrogencontent to nitrate supplied. In both species, and also in Cucumis melo, the proportion oftotal plant free nitrate and reduced nitrogen in the roots decreases,and in the stem increases, with increasing nitrate supplied. The accumulation of free nitrate in leaves is accompanied bya quantitatively different relationship between reduced nitrogenand dry weight compared to leaves not accumulating nitrate. Capsicum annuum. L., Cucumis melo L., melon, Zea mays L., maize, sweet corn, nitrate reductase, nitrate uptake     

NATURE OF THE INHIBITION OF PROTEIN SYNTHESIS BY ETHIONINE AND AMINO ACID UPTAKE IN EARLY SEA URCHIN EMBRYOS

The inhibition of protein synthesis by ethionine reported previously was found to be apparent, and ethionine inhibited only amino acid uptake like other usual amino acids. Even under such strong inhibition of the uptake, the syntheses of protein and DNA remained almost undiminished. The uptake of amino acid mixture by sea urchin embryos in the early cleavage stage was found to be carried out by active transport, since it was temperature-sensitive and was inhibited by 2,4-dinitrophenol. The uptake of an amino acid mixture or of single amino acids, e.g., valine, leucine and phenylalanine, was inhibited nonspecifically by an excess amount of other single amino acids added exogenously. Reflecting the inhibition of amino acid uptake, in vivo incorporation of amino acids into the protein fraction was apparently inhibited by excess amounts of other amino acids. As far as tested, the inhibition seems to be nonspecific and competitive for all amino acid species. The uptakes of leucine and phenylalanine were inhibited mutually by competition, with almost the same Km and Ki.     

Growth and 2, 4-Diaminobutyric Acid Composition of Flatpea (Lathyrus sylvestris L.) as Influenced by Mineral- and Symbiotically Fixed-Nitrogen

Flatpea (Lathyrus sylvestris L. cv. ‘Lathco’) plants,grown hydroponically, were inoculated with rhizobia (Rhizobiumleguminosarum strain 92F2), supplied with nitrate nitrogen (2meq 1^–1), or provided with both rhizobia and . Nodulated plants supplied with had the highest biomass and the highest level of free amino acids in leavesand roots. Nitrogen fixation was depressed 80% in inoculatedplants supplemented with . Compared to plants whose nitrogen requirement was satisfied either partially orfully by , plants that were totally dependent upon biological nitrogen fixation had fewer lateral shoots andlower concentrations of nitrogen, soluble protein, and totalfree amino acids in the shoots, especially in the leaves. Thesymbiotic association between plants and rhizobia for nitrogenfixation was not essential to the production and accumulationof 2,4-diaminobutyric acid (A₂bu) and other non-protein aminoacids in flatpea. Levels of A₂bu, and most of the other freeamino acids quantified, were significantly lower in the nodulatedplants grown in the absence of than in the inoculated or noninoculated plants supplied with . Arginine and glutamic acid were exceptions in that levels ofthese two amino acids were higher in roots bearing effectivenodules. Plant responses to the different treatments are attributedto the amount of nitrogen available to the plant, rather thanthe method or form of nitrogen supplied. Key words: 2,4-Diaminobutyric acid, Lathyrus, nitrogen     

Changes of anabolic and energetic metabolism in the recovery of Rhodotorula gracilis from nitrogen starvation

We studied the changes occurring in the adenylate system andrelated metabolic parameters during the shift from rest to activegrowth, by feeding ammonium ions to N-starved Rhodotorula graciliscells. The addition of ammonium induces an early, rapid dropin ATP level and in energy charge, decrease in RQ, a gradualincrease in O₂ uptake and a rapid increase in the synthesesof amino acids, protein and RNA. CHI at a protein-synthesisblocking concentration also blocks the early decrease in ATPlevel. The data suggest that growth inhibition, due to N-starvationis not determined by the phosphorylation state of the adenylatesystem and that the observed behavior of the adenylate poolis a consequence of the onset of macromolecular synthesis. (Received June 1, 1977; )     

Growth Inhibition in Thiobacillus neapolitanus by Histidine, Methionine, Phenylalanine, and Threonine

Thiobacillus neapolitanus, a strict chemoautotroph, is sensitive to the addition of 10(-4)m methionine, histidine, threonine, or phenylalanine to the thiosulfate medium on which it grows. When histidine, threonine, or phenylalanine are added at the time of inoculation, spontaneous mutants tolerant to the three amino acids are selected. These mutants appear to result from a single genetic change; of 18 independently isolated histidine-tolerant mutants, all are also tolerant to phenylalanine and threonine. The uptake of (14)C-phenylalanine into exponentially growing cells of one such mutant is negligible in contrast with the uptake observed in the phenylalanine-sensitive parent. The addition of methionine to the medium slows growth, but spontaneous mutants are not selected. Inhibition of growth by these amino acids is observed only under conditions of amino acid imbalance; the addition of an equimolar mixture of 16 amino acids, in which each component is present at a concentration of 10(-3)m, causes no inhibition. Histidine and threonine inhibition may be released by equimolar amounts of any one of seven amino acids: serine, alanine, glycine, leucine, valine, tryptophan, or tyrosine; histidine inhibition is also released by isoleucine, and threonine inhibition by methionine. None of the inhibiting amino acids inhibits oxidation of thiosulfate in cell suspensions. A group of hexoses, pentoses, and Krebs cycle intermediates were tested for inhibition of growth or release of inhibition by histidine, phenylalanine, or threonine, but no effects, either inhibition or relief of inhibition, were found.     

EFFECTS OF EXOGENOUS AMINO ACIDS ON THE FLOWER AND FROND PRODUCTION IN DUCKWEED, LEMNA GIBBA G3

1. Twenty usual amino acids examined were shown to be dividedinto two groups with respect to their actions on the flowerformation (A) and frond multiplication (B) in a long-day duckweed,L. gibba G3. Amino acids of the first group (e.g., arginine)inhibited A without preventing B, and those of the second group(e.g., lysine) inhibited both A and B. The inhibition of flowerformation was always the greatest when amino acid was appliedat the induction period. 2. The floral inhibition by arginine applied at the inductionperiod was partially or wholly reversed by the simultaneousaddition of other amino acid (especially lysine) or by one additionallong day. The inhibitions by lysine, however, were not reversedby arginine. 3. It was discussed that the terminal step(s) of photoperiodicinduction process might depend largely on the relative in vivoconcentrations of amino acids. (Received January 28, 1964; )