Valine Transport Research Articles

Role of branched-chain amino acid transport in Bacillus subtilis CodY activity.

CodY is a branched-chain amino acid-responsive transcriptional regulator that controls the expression of several dozen transcription units in Bacillus subtilis. The presence of isoleucine, valine, and leucine in the growth medium is essential for achieving high activity of CodY and for efficient regulation of the target genes. We identified three permeases-BcaP, BraB, and BrnQ-that are responsible for the bulk of isoleucine and valine uptake and are also involved in leucine uptake. At least one more permease is capable of efficient leucine uptake, as well as low-affinity transport of isoleucine and valine. The lack of the first three permeases strongly reduced activity of CodY in an amino acid-containing growth medium. BcaP appears to be the most efficient isoleucine and valine permease responsible for their utilization as nitrogen sources. The previously described strong CodY-mediated repression of BcaP provides a mechanism for fine-tuning CodY activity by reducing the availability of amino acids and for delaying the utilization of isoleucine and valine as nitrogen and carbon sources under conditions of nutrient excess. Bacillus subtilis CodY is a global transcriptional regulator that is activated by branched-chain amino acids (BCAA). Since the level of BCAA achieved by intracellular synthesis is insufficient to fully activate CodY, transport of BCAA from the environment is critical for CodY activation, but the permeases needed for such activation have not been previously identified. This study identifies three such permeases, reports their amino acid transport specificity, and reveals their impact on CodY activation.

Valine partitioning and kinetics between the gastrointestinal tract and hind limbs in lambs with an adult Trichostrongylus colubriformis burden1

Intestinal parasitic infection increases the demand for AA because of increased protein synthesis in the intestine and increased luminal losses of AA, and these increased demands may be supported by increased mobilization of AA from the skeletal muscles. Two experiments were conducted to determine the effects of parasitic infection on valine kinetics within the gastrointestinal tract and hind limbs of lambs fed fresh forages. On d 1, lambs were given 6,000 stage-3 Trichostrongylus colubriformis larvae per day for 6 d (n = 6) or kept as parasite-free controls (n = 6) and fed fresh lucerne (Medicago sativa; Exp. 1) or fresh sulla (Hedysarum coronarium; Exp. 2). On d 48, valine kinetics within the mesenteric- (MDV) and portal-drained viscera (PDV) and hind limbs were obtained by carrying out concurrent infusions of para-amminohippuric acid into the mesenteric vein and indocyanin green into the abdominal aorta (for blood flow), and [3,4-(3)H]valine into the jugular vein and [1-(13)C]valine into the abomasum for 8 h (for kinetics). During the infusions, blood was collected from the mesenteric and portal veins and from the mesenteric artery and vena cava, and plasma was harvested. After the 8-h infusion, lambs were euthanized, ileal digesta were collected, and tissues were sampled from the intestine and muscle (biceps femoris). Tissues, digesta, and plasma were analyzed for valine concentration, specific radioactivity, and isotopic enrichment. In both experiments, intestinal worm burdens on d 48 were greater in parasitized lambs (P = 0.0001 and 0.003). In Exp. 1, parasitic infection increased (P = 0.03) the total valine irreversible loss rate (ILR) in the MDV and PDV. In Exp. 2, luminal ILR of valine in the MDV was reduced (P = 0.01); however, ILR of valine in the PDV was unaffected. Despite these changes within the MDV and PDV, parasitic infection did not affect the ILR of valine within the hind limbs, and valine transport rates were largely unchanged. We suggest that the increased mobilization of AA from the hind limbs that might have occurred in the early phase of inflammation was no longer required when the parasitic infection was established. The MDV and PDV data may indicate that the non-MDV parts of the PDV play an important role in this adaptation, which warrants further study.

Facilitated transport of valine through bulk liquid membranes containing Aliquat 336: A kinetic study

This paper describes a kinetic study, in optimal conditions, of the facilitated counter-transport of valine through bulk liquid membranes using tricapryl methyl ammonium chloride (Aliquat 336) as carrier and chloride as counter-ion. Recovery close to 80% has been obtained after 24 h. The transport kinetic was analysed by means of a model involving two consecutive irreversible first order reactions. The rate constants of the extraction and stripping reactions were determined by numerical analysis. Good agreement between the model and experimental data was observed. A maximum flux of valine transport through the bulk liquid membrane of 0.052/h was obtained.

Subcellular distribution of proteolytically generated valine in isolated rat hepatocytes.

1. A small fraction of the intracellular acid-soluble radioactivity in [14C]valine-labelled hepatocytes remained non-extracted even after repeated incubations in isotope-free medium at 37 degrees C. This non-extracted radioactivity was only present when the cells had been labelled under conditions allowing protein synthesis. 2. Approximately two-thirds of the non-extracted radioactivity was recovered in material with a molecualr weight larger than free valine, i.e. presumably acid-soluble peptides. The intracellular contents of these peptides were unaffected by inhibitors of hepatocytic protein degradation, and they would therefore seem to represent primary synthesis products rather than products of proteolysis. 3. The remaining one-third of the radioactivity was free [14C]valine, derived from intracellular protein degradation as indicated by the reduced levels seen in the presence of inhibitors of lysosomal (ammonia, methylamine, leupeptin, chymostatin) and non-lysosomal (chymostatin) proteolysis. The intracellular levels of twelve other amino acids were similarly reduced upon inhibition of protein degradation. 4. The contents of free [14C]valine in subcellular fractions were compatible with a uniform distribution of the amino acid throughout the cell. There was no evidence for any enrichment of [14C]valine in a purified lysosomal fraction, separated from mitochondria by means of isotonic metrizamide gradients. 5. It can be concluded that the non-extracted valine in hepatocytes may represent a steady-state level, maintained by intracellular protein degradation, of amino acid in transit through the cell. The non-extractability, i.e. the maintenance of a concentration gradient towards the extracellular medium, can be calculated to be compatible with a limitation of valine efflux by the concentration-dependent valine transport system. Since no specific subcellular compartmentation is indicated, intracellular valine can provisionally be regarded as a single, uniform pool.

The N-terminal domain of yeast Bap2 permease is phosphorylated dependently on the Npr1 kinase in response to starvation

The Saccharomyces cerevisiae branched-chain amino acid permease Bap2p plays a major role in leucine, isoleucine, and valine transport, and its synthesis is regulated transcriptionally. Bap2p undergoes a starvation-induced degradation depending upon ubiquitination and the functions of N- and C-terminal domains of Bap2p. Here we show that the N-terminal domain of Bap2p is phosphorylated in response to rapamycin treatment when both the N- and C-termini of Bap2p are fused to glutathione S-transferase. The phosphorylation is dependent on Ser/Thr kinase Npr1p. In npr1 cells, Bap2p becomes slightly more susceptible to the rapamycin-induced degradation, suggesting that Npr1p counteracts the degradation system for Bap2p.

Amino Acid Availability Affects Amino Acid Flux and Protein Metabolism in the Porcine Mammary Gland

A kinetic model was used to examine transmembrane flux kinetics of lysine, methionine and valine across the porcine mammary gland (MG) under dietary amino acid (AA) limiting, adequate and excess conditions. Lactating sows (3 per treatment) were offered three diets: lysine-deficient [LD, 4.9 lysine and 9.9 valine (g/kg diet)], adequate (Control, 9.7 and 10.2) and valine-excess (VE, 9.8 and 13.4). On d 18 of lactation, 2-(15)N-lysine, 5-methyl-(2)H(3)-methionine and 1-(13)C-valine were infused into a jugular vein for 20.5 h. Milk and arterial and mammary venous blood samples were collected at 2- and 1-h intervals, respectively. Compared with Control, milk yield and litter growth rate decreased (P < 0.05) in sows fed the LD diet. Model estimates of mammary protein synthesis (PS), breakdown (PB) and net PS decreased (P < 0.05) in sows fed the LD diet. Net uptake of lysine decreased (P < 0.05) in sows fed the LD diet as a result of decreases in inward and outward transport of lysine. Inward transport of methionine tended to be reduced (P < 0.10) in sows fed the LD diet, resulting in a decrease in net methionine uptake. In sows fed the VE diet, PB was reduced (P < 0.05) and PS unchanged compared with Control. Outward transport of valine and net lysine uptake were reduced (P < 0.05), but net valine uptake was unchanged in sows fed the VE diet compared with Control. In conclusion, the kinetic model provided estimates of PS that were similar to empirical measurements of milk protein output and mammary protein accretion. Transport of lysine and methionine by the porcine MG is closely linked to regulation of mammary PS. Lysine availability has little effect on the transmembrane flux of valine.

Valine Transport and Biodiversity of Leuconostoc Wild Strains from French Raw Milk Cheeses

The rate of L-valine transport in whole cells of Leuconostoc was at the maximum at 30 degrees C, pH 6.0 in the presence of an energy source. Transport was inhibited by 40-55%, in the presence of the ionophores (valinomycin, nigericin or monensin), and uncouplers (carbonyl cyanide-m-chloro-phenylhydrazone or 2,4-dinitrophenol) confirming the previously described delta p-driven branched-chain amino acid transport system described in cytoplasmic membranes (Winters et al., 1991, Appl. Environ. Microbiol., 57, 3350-3354). Sulfhydryl group reagents (p-chloro-mercuribenzoate, iodoacetate and N-ethyl maleimide) all inhibited valine transport by 60-70%, indicating that valine is actively transported at high valine concentration. Three kinetically distinguishable transport systems were identified for each strain using whole cells, confirming results obtained with membranes. L-valine transport Kt and Vmax could be an additional tool to estimate the biodiversity of 18 Leuconostoc strains belonging to the dominant flora of French raw milk cheeses. Kt values varied from 20 to 510 nmol/l for the very high affinity system, from 26 to 427 pmol/l for the high affinity system and from 0.65 to 4.40 mmol/l for the low affinity system. No correlation existed between valine transport rates and a particular strain's ability to acidify milk or complex media, suggesting that valine transport is not a growth-limiting function in species of the genus Leuconostoc.

Cellular uptake of valine by lactating porcine mammary tissue.

The cellular uptake of branched-chain amino acids in mammary tissue is important for understanding their role in milk synthesis in the sow. This study characterized the kinetic properties and substrate specificity of the valine uptake system in the porcine mammary gland. Mammary tissue was collected from lactating sows at slaughter and tissue explants were incubated in media containing isosmotic salt and amino acids of interest, plus [3H]valine tracer. Valine uptake was time-dependent and was dependent on the presence of sodium, as indicated by a reduction in uptake when sodium in the medium was replaced by choline. The valine transport system in porcine mammary tissue had a Km of 0.64 mM, a Vmax of 1.84 mmol-kg cell water(-1) 30 min(-l), and a Kd (diffusion constant) of 1.16 L x kg cell water(-1) x 30 min(-1). Valine uptake was inhibited by leucine and alpha-aminoisobutyric acid and by high concentrations of L-alanine, L-lysine, cycloleucine, L-glutamine, and L-methionine, but not by 2-(methyl-amino)-isobutyric acid. This transport system is the primary system responsible for uptake of valine, and probably other branched-chain amino acids, in lactating sow mammary tissue. Physiological concentrations of valine in the blood are below the Km of the specific valine transport system and well below the diffusion uptake capabilities. The kinetic parameters of this valine transport system should not be limiting to valine uptake for milk protein synthesis. However, competition of valine uptake with branched-chain amino acids, as well as with other amino acids, may affect valine uptake in lactating tissue.

De-regulated assimilation and over-production of amino acids in analogue-resistant mutants of a cyanobacterium, Phormidium uncinatum

Mutant strains of Phormidium uncinatum resistant to fluoro-phenylalanine, aztryptophan, fluorotyrosine and azaleucine accumulated a wide range of amino acids, notably glutamic acid, lysine, tyrosine and phenylalanine, and exhibited de-regulated valine and phenylalanine transport. While acetohydroxy acid synthase in azaleucine-resistant mutants lost valine- and leucine-sensitivity, 3-deoxy-DXXX-arabinoheplulosonate-7-phosphate (DAHP) synthase and prephenate dehydratase in aromatic analogue-resistant strains became phenylalanine-insensitive and shikimate and prephenate dehydrogenases were activated by tyrosine. In addition, activities of nitrate-assimilating enzymes were higher in the mutants, which also exhibited increased nitrogen, protein and phycocyanin contents. The proteins in the mutants were better digested upon enzymatic-treatments and feeding trials than those of the wild type, indicating that they are usable as single-cell protein.

Leucine Affects the Metabolism of Valine by Isolated Perfused Rat Hearts: Relation to Branched-Chain Amino Acid Antagonism

This study was conducted to determine the effects of different concentrations of leucine on the transport, transamination and oxidation of valine and on incorporation of valine into heart proteins in the isolated perfused rat heart. Valine metabolism was studied in rat hearts perfused with medium containing glucose and graded levels of L-leucine. In transport studies L-phenylalanine was also tested. Uptake of L-[1-14C]valine (0.2 mmol/L) was significantly reduced (∼50%) by inclusion of 0.2 mmol/L phenylalanine or leucine, and by ∼70% by inclusion of 1.0 mmol/L phenylalanine or leucine in the perfusate. Transamination of valine decreased by 37 and 48%, and oxidation of valine by 53 and 71%, respectively, when 0.2 or 1.0 mmol/L leucine was included in the perfusate. Tissue concentrations of valine decreased by 43, 48 and 62% in the presence of 0.2, 0.5 and 1.0 mmol/L leucine, respectively; tissue concentrations of leucine, glutamate and alanine increased ∼11-fold, 1.2-fold and 0.5-fold, respectively, when 1.0 mmol/L leucine was present in the perfusate. Addition of 0.2–1.0 mmol/L leucine did not affect incorporation of valine into heart proteins. We conclude that 1) competition among large neutral amino acids for transport into heart occurs at physiological concentrations of these amino acids in plasma; 2) inhibition of valine uptake by leucine can limit the rate of valine catabolism in heart; and 3) depletion of tissue valine concentration by an excess of leucine did not affect the rate of protein synthesis.

Effects of starvation on valine and alanine transport across the intestinal mucosal border in sea bass, Dicentrarchus labrax.

Kinetics of intestinal transport of L-alanine and L-valine (substrates of the A-system and the L-system, respectively, in mammals) across the brush-border membrane in sea bass, Dicentrarchus labrax, were studied on intact mucosa using a short-term uptake technique. When fish were starved for 4-8 weeks, total influx (mucosa-to-cell) of valine fell owing to disappearance or modification of the diffusion component. The maximum influx rate of saturable component increased but its affinity (reflected by the Michaelis constant) decreased. Alanine transport by Na(+)-dependent and diffusion pathways was unchanged after starvation. Fasting also induced an almost 20% decrease in the length of intestinal microvilli.

Immunoaffinity purification and reconstitution of sodium-coupled branched-chain amino acid carrier of Pseudomonas aeruginosa.

The gene product of braB encoding the Na+(Li+)-coupled carrier protein for L-leucine, L-isoleucine, and L-valine (LIV-II carrier) of Pseudomonas aeruginosa PML strain was identified and overexpressed using a T7 RNA polymerase/promoter plasmid system. The gene product was pulse-labeled with [35S]methionine as a protein of an apparent Mr of 34,000 on a sodium dodecyl sulfate-polyacrylamide gel. Cell membranes overproducing the LIV-II carrier were solubilized with n-dodecyl beta-D-maltopyranoside. The carrier protein was purified from the detergent extract by two purification steps: (i) immunoaffinity column chromatography using purified polyclonal antibody directed against synthetic 13-mer peptide corresponding to the carboxyl terminus region of the carrier and (ii) subsequent DEAE-cellulose column chromatography. The detergent was replaced by n-octyl beta-D-glucopyranoside prior to the first elution and phospholipid was present during purification. Proteoliposomes reconstituted with the purified LIV-II carrier exhibited Na+ or Li+ concentration gradient-driven transport of leucine, isoleucine, and valine. These results show that the LIV-II carrier was purified to be in a functional form.

Sequence and structural similarities between the leucine-specific binding protein and leucyl-tRNA synthetase of Escherichia coli.

A role for the leucyl-tRNA synthetase (EC 6.1.1.4) has been established for regulating the transport of leucine across the inner membrane of Escherichia coli by the leucine, isoleucine, valine (LIV-I) transport system. This transport system is mediated by interactions of periplasmic binding proteins with a complex of membrane-associated proteins, and transcription of the high-affinity branched-chain amino acid transport system genes is repressed by growth of E. coli on high levels of leucine. We now report results from sequence comparisons and structural modeling studies, which indicate that the leucine-specific binding protein, one of the periplasmic components of the LIV-I transport system, contains a 121-residue stretch, representing 36% of the mature protein, which displays both sequence and structural similarities to a region within the putative nucleotide-binding domain of leucyl-tRNA synthetase. Early fusion events between ancestral genes for the leucine-specific binding protein and leucyl-tRNA synthetase could account for the similarity and suggest that processes of aminoacylation and transport for leucine in E. coli may be performed by evolutionarily interrelated proteins.

Transport of leucine, isoleucine and valine by luminal membrane vesicles from rabbit proximal tubule.

1. Transport of L- and D-isomers of leucine, isoleucine and valine by luminal membrane vesicles prepared from either the convoluted part (pars convoluta) or the straight part (pars recta) of rabbit proximal tubule was studied by a rapid filtration technique and by a spectrophotometric method using a potential-sensitive carbocyanine dye. 2. Both types of renal membrane vesicle take up the amino acids in a Na(+)-dependent, H(+)-independent and electrogenic manner. The L-isomers are transported with higher affinities than their corresponding D-forms, of which only D-leucine is taken up to a significant extent. 3. Membrane vesicles prepared from pars convoluta take up the L-amino acids by a single and common system. Filtration studies showed that the Km values for L-leucine and L-valine transport are, on average, 0.23 and 0.83 mM, respectively. The values of KA (the concentration of amino acid producing a half-maximal optical response) are comparable to those of Km, namely 0.18 mM for L-leucine and 0.60 mM for L-valine. KA for L-isoleucine transport was found to be 0.19 mM. D-Leucine is taken up by the same system but with a much lower affinity (KA = 7.2 mM). 4. Membrane vesicles prepared from pars recta possess two, and probably common, transport systems for the L-isomers of the amino acids. The average Michaelis-Menten constants were as follows: L-leucine, K1m = 0.17 mM, K2m = 6.5 mM; L-valine, K1m = 0.19 mM, K2m = 11.5 mM. The KA values were: L-leucine, K1A = 0.12 mM, K2A = 7.4 mM; L-valine, K1A = 0.18 mM, K2A = 10.0 mM; L-isoleucine, K1A = 0.17 mM, K2A = 9.0 mM. D-Leucine is taken up by a low-affinity system only (KA = 6.5 mM), which seems to be the same as the low-affinity system transporting the L-forms of the amino acids.

Separation of L‐valine from fermentation broths using a supported liquid membrane

A carrier-mediated counter transport process is proposed to separate and to purify an amino acid produced by microbial fermentation. The case of L-valine permeation through a liquid membrane, constituted by a solution of Aliquat 336 in decanol and supported by a hydrophobic microporous membrane, is reported. A mathematical model was developed to estimate distribution coefficients and permeabilities and to predict the influence of hydrodynamic and pH conditions on supported liquid membrane (SLM) performances. Optimum conditions for the transport and the concentration of valine were achieved with synthetic aqueous valine solutions. Series of experiments on fermentation broths, where molasses and biomass contents were varied, permitted pointing out the role of the broth composition on the kinetics and yields of separation. The selectivity of transport of valine by an Aliquat 336/decanol liquid membrane was about 10 toward molasses dyes, 100 toward glucose, and beyond 1000 toward sucrose. This allowed us to achieve the recovery and one step of purification of the product in a single operation. The stability of the Aliquat 336/decanol liquid membrane was sufficient to ensure a selective transport of valine during a continuous run lasting 18 days.

Dietary Amino Acid Analogues and Transport of Lysine or Valine across the Blood-Brain Barrier in Rats

Studies were undertaken to determine if dietary disproportions of amino acids would alter flux into brain of the amino acid present in the diet in a growth-limiting concentration. Rats were adapted to a lysine-limiting diet before receiving a meal of this control diet, alone or with added lysine or homoarginine (a competitor for lysine transport) or both, before intravenous infusion of [14C]lysine. The brain-to-plasma radioactivity ratio was lower in rats fed extra lysine or homoarginine than in rats fed the control diet, whereas lysine flux and brain lysine concentration were high in rats fed extra lysine alone. Flux and concentration were lower in rats fed homoarginine + lysine than in rats fed extra lysine alone. Other rats were fed a valine-limiting diet containing added valine, norleucine (a competitor for valine transport) or both, before [14C]valine was infused. Valine flux and brain valine concentrations were higher in rats fed extra valine than in control rats, whereas flux was lower in the group fed norleucine alone. Valine flux was higher in rats fed norleucine + valine than in the rats fed norleucine alone. Our studies show that dietary disproportions of amino acids can alter the flux of specific amino acids across the blood-brain barrier.

Selective alteration in blood-brain barrier and insulin transport in iron-deficient rats.

Nutritional iron deficiency induced in rats causes a significant reduction in level of brain nonheme iron and is accompanied by selective reduction of dopamine D2 receptor Bmax. Our previous studies have clearly demonstrated that these alterations can be restored to normal by supplementation with ferrous sulfate; however, neither brain nonheme iron level nor dopamine D2 receptor Bmax can be increased beyond control values even after long-term iron therapy. The possibility that iron deficiency can induce the breakdown of the blood-brain barrier (BBB) was examined. A 70 and 100% increase in brain uptake index (BUI) for L-glucose and insulin, respectively, were noted in iron-deficient rats. However, the BUI for valine was decreased by 40%, and those for L-norepinephrine and glycine were unchanged. In addition, it was demonstrated that in normal rats insulin is transported into the brain. The data show that iron deficiency selectively affects the integrity of the BBB for insulin, glucose, and valine transport. Whether the effect of iron deficiency on the BBB is at the level of the capillary endothelial cell tight junction is not yet known. However, this study has shown that an important nutritional disorder (iron-deficiency anemia) has a profound effect on the BBB and brain function.

Relation of growth of Streptococcus lactis and Streptococcus cremoris to amino acid transport.

The maximum specific growth rate of Streptococcus lactis and Streptococcus cremoris on synthetic medium containing glutamate but no glutamine decreases rapidly above pH 7. Growth of these organisms is extended to pH values in excess of 8 in the presence of glutamine. These results can be explained by the kinetic properties of glutamate and glutamine transport (B. Poolman, E. J. Smid, and W. N. Konings, J. Bacteriol. 169:2755-2761, 1987). At alkaline pH the rate of growth in the absence of glutamine is limited by the capacity to accumulate glutamate due to the decreased availability of glutamic acid, the transported species of the glutamate-glutamine transport system. Kinetic analysis of leucine and valine transport shows that the maximal rate of uptake of these amino acids by the branched-chain amino acid transport system is 10 times higher in S. lactis cells grown on synthetic medium containing amino acids than in cells grown in complex broth. For cells grown on synthetic medium, the maximal rate of transport exceeds by about 5 times the requirements at maximum specific growth rates for leucine, isoleucine, and valine (on the basis of the amino acid composition of the cell). The maximal rate of phenylalanine uptake by the aromatic amino acid transport system is in small excess of the requirement for this amino acid at maximum specific growth rates. Analysis of the internal amino acid pools of chemostat-grown cells indicates that passive influx of (some) aromatic amino acids may contribute to the net uptake at high dilution rates.

The third general transport system for branched-chain amino acids in Salmonella typhimurium.

Some properties of the LIV-III transport system, a common factor in the entry of branched-chain amino acids in Salmonella typhimurium, have been studied using a double mutant, KA266 (livA brnQ), which is defective in transport via either the LIV-I or LIV-II system. The LIV-III system operates with a low affinity (Km: >15μM) for L-isoleucine, L-leucine, and L-valine. Isoleucine transport via the LIV-III system is inhibited competitively by leucine or valine, but non-competitively by L-cysteine. Uptake of branched-chain amino acids into cells is partially repressed when the bacteria are grown in the presence of 2mM glycyl-L-leucine or more. In the wild type, transport activity of the LIV-III system for isoleucine and leucine is not detected. The LIV-II system appears to show a low affinity for valine similar to the LIV-III system, hence the combined activity of these systems is expressed as the activity of the LIV-III system. The defect in transport via the LIV-II system leads to a great reduction in the transport of valine in the LIV-III system.

Interference by methionine on valine uptake in Acremonium chrysogenum.

The incorporation of L-[U-14C]valine into delta-(L-alpha-aminoadipyl)-L-cysteinyl-D-valine (ACV), a direct biosynthetic precursor of penicillins and cephalosporins, was studied. When DL-methionine was added to Acremonium chrysogenum culture broths, no labeled ACV was found, while a large amount of radioactive ACV was detected when methionine was not present. DL-Norleucine, a nonsulfur analog of methionine, also inhibited the synthesis of radioactive ACV to some degree. This effect was due to the inhibition of valine transport by methionine and norleucine.