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Sample records for acids phenylalanine tyrosine

  1. Hydration of protonated aromatic amino acids: phenylalanine, tryptophan, and tyrosine.

    PubMed

    Gao, Bing; Wyttenbach, Thomas; Bowers, Michael T

    2009-04-01

    The first steps of hydration of the protonated aromatic amino acids phenylalanine, tryptophan, and tyrosine were studied experimentally employing a mass spectrometer equipped with a drift cell to examine the sequential addition of individual water molecules in equilibrium experiments and theoretically by a combination of molecular mechanics and electronic structure calculations (B3LYP/6-311++G**) on the three amino acid systems including up to five water molecules. It is found that both the ammonium and carboxyl groups offer good water binding sites with binding energies of the order of 13 kcal/mol for the first water molecule. Subsequent water molecules bind less strongly, in the range of 7-11 kcal/mol for the second through fifth water molecules. The ammonium group is able to host up to three water molecules and the carboxyl group one water molecule before additional water molecules bind either to the amino acid side chain as in tyrosine or to already-bound water in a second solvation shell around the ammonium group. Reasons for the surprisingly high water affinity of the neutral carboxyl group, comparable to that of the charge-carrying ammonium group, are found to be high intrinsic hydrophilicity, favorable charge-dipole alignment, and--for the case of multiply hydrated species--favorable dipole-dipole interaction among water molecules and the lack of alternative fully exposed hydration sites.

  2. An experimental and computational investigation into the gas-phase acidities of tyrosine and phenylalanine: three structures for deprotonated tyrosine.

    PubMed

    Bokatzian, Samantha S; Stover, Michele L; Plummer, Chelsea E; Dixon, David A; Cassady, Carolyn J

    2014-11-01

    Using mass spectrometry and correlated molecular orbital theory, three deprotonated structures were revealed for the amino acid tyrosine. The structures were distinguished experimentally by ion/molecule reactions involving proton transfer and trimethylsilyl azide. Gas-phase acidities from proton transfer reactions and from G3(MP2) calculations generally agree well. The lowest energy structure, which was only observed experimentally using electrospray ionization from aprotic solvents, is deprotonated at the carboxylic acid group and is predicted to be highly folded. A second unfolded carboxylate structure is several kcal/mol higher in energy and primarily forms from protic solvents. Protic solvents also yield a structure deprotonated at the phenolic side chain, which experiments find to be intermediate in energy to the two carboxylate forms. G3(MP2) calculations indicate that the three structures differ in energy by only 2.5 kcal/mol, yet they are readily distinguished experimentally. Structural abundance ratios are dependent upon experimental conditions, including the solvent and accumulation time of ions in a hexapole. Under some conditions, carboxylate ions may convert to phenolate ions. For phenylalanine, which lacks a phenolic group, only one deprotonated structure was observed experimentally when electrosprayed from protic solvent. This agrees with G3(MP2) calculations that find the folded and unfolded carboxylate forms to differ by 0.3 kcal/mol.

  3. An Experimental and Computational Investigation into the Gas-Phase Acidities of Tyrosine and Phenylalanine: Three Structures for Deprotonated Tyrosine

    SciTech Connect

    Bokatzian, Samantha S.; Stover, Michele L.; Plummer, Chelsea E.; Dixon, David A.; Cassady, Carolyn J.

    2014-11-06

    Using mass spectrometry and correlated molecular orbital theory, three deprotonated structures were revealed for the amino acid tyrosine. The structures were distinguished experimentally by ion/molecule reactions involving proton transfer and trimethylsilyl azide. Gas-phase acidities from proton transfer reactions and from G3(MP2) calculations generally agree well. The lowest energy structure, which was only observed experimentally using electrospray ionization from aprotic solvents, is deprotonated at the carboxylic acid group and is predicted to be highly folded. A second unfolded carboxylate structure is several kcal/mol higher in energy and primarily forms from protic solvents. Protic solvents also yield a structure deprotonated at the phenolic side chain, which experiments find to be intermediate in energy to the two carboxylate forms. G3(MP2) calculations indicate that the three structures differ in energy by only 2.5 kcal/mol, yet they are readily distinguished experimentally. Structural abundance ratios are dependent upon experimental conditions, including the solvent and accumulation time of ions in a hexapole. Under some conditions, carboxylate ions may convert to phenolate ions. For phenylalanine, which lacks a phenolic group, only one deprotonated structure was observed experimentally when electrosprayed from protic solvent. This agrees with G3(MP2) calculations that find the folded and unfolded carboxylate forms to differ by 0.3 kcal/mol.

  4. An experimental and computational investigation into the gas-phase acidities of tyrosine and phenylalanine: three structures for deprotonated tyrosine.

    PubMed

    Bokatzian, Samantha S; Stover, Michele L; Plummer, Chelsea E; Dixon, David A; Cassady, Carolyn J

    2014-11-01

    Using mass spectrometry and correlated molecular orbital theory, three deprotonated structures were revealed for the amino acid tyrosine. The structures were distinguished experimentally by ion/molecule reactions involving proton transfer and trimethylsilyl azide. Gas-phase acidities from proton transfer reactions and from G3(MP2) calculations generally agree well. The lowest energy structure, which was only observed experimentally using electrospray ionization from aprotic solvents, is deprotonated at the carboxylic acid group and is predicted to be highly folded. A second unfolded carboxylate structure is several kcal/mol higher in energy and primarily forms from protic solvents. Protic solvents also yield a structure deprotonated at the phenolic side chain, which experiments find to be intermediate in energy to the two carboxylate forms. G3(MP2) calculations indicate that the three structures differ in energy by only 2.5 kcal/mol, yet they are readily distinguished experimentally. Structural abundance ratios are dependent upon experimental conditions, including the solvent and accumulation time of ions in a hexapole. Under some conditions, carboxylate ions may convert to phenolate ions. For phenylalanine, which lacks a phenolic group, only one deprotonated structure was observed experimentally when electrosprayed from protic solvent. This agrees with G3(MP2) calculations that find the folded and unfolded carboxylate forms to differ by 0.3 kcal/mol. PMID:25299802

  5. Acute oral administration of a tyrosine and phenylalanine-free amino acid mixture reduces exercise capacity in the heat.

    PubMed

    Tumilty, Les; Davison, Glen; Beckmann, Manfred; Thatcher, Rhys

    2013-06-01

    Acute tyrosine administration is associated with increased exercise capacity in the heat. To explore whether reduced plasma tyrosine and phenylalanine (tyrosine precursor) is associated with impaired exercise capacity in the heat, eight healthy, moderately trained male volunteers, unacclimated to exercise in the heat, performed two tests in a crossover design separated by at least 7 days. In a randomised, double-blind fashion, subjects ingested 500 mL flavoured, sugar-free water containing amino acids [(TYR-free; isoleucine 15 g, leucine 22.5 g, valine 17.5 g, lysine 17.5 g, methionine 5 g, threonine 10 g, tryptophan 2.5 g)] to lower the ratio of plasma tyrosine plus phenylalanine:amino acids competing for blood-brain barrier uptake (CAA), a key determinant of brain uptake, or a balanced mixture (BAL; TYR-free plus 12.5 g tyrosine and 12.5 g phenylalanine). One hour later, subjects cycled to exhaustion at 63 ± 5 % [Formula: see text]O2peak in 30 °C and 60 % relative humidity. Pre-exercise ratio of plasma tyrosine plus phenylalanine:ΣCAA declined 75 ± 5 % from rest in TYR-free (P < 0.001), but was unchanged in BAL (P = 0.061). Exercise time was shorter in TYR-free (59.8 ± 19.0 min vs. 66.2 ± 16.9 min in TYR-free and BAL respectively; P = 0.036). Heart rate (P = 0.298), core (P = 0.134) and skin (P = 0.384) temperature, RPE (P > 0.05) and thermal sensation (P > 0.05) were similar at exhaustion in both trials. These data indicate that acutely depleting plasma catecholamine precursors:ΣCAA is associated with reduced submaximal exercise capacity in the heat.

  6. Multicenter bond index analysis of influence of metal cations on the aromaticity of aromatic amino acids: Phenylalanine and tyrosine

    NASA Astrophysics Data System (ADS)

    Pakiari, A. H.; Farrokhnia, M.; Azami, S. M.

    2008-05-01

    In order to provide insight into the influence of metal cations on the aromaticity of amino acids, evaluation of six-center delocalization indices is accomplished in the context of quantum theory of atoms in molecules (QTAIM). Aromaticity of two amino acids, phenylalanine and tyrosine, is investigated as typical amino acids containing aromatic ring in their isolated state and complexed by some metal cations. The results showed that the metal cations affect the most important three connectivities differently. Also, it is shown that the existence of metal cations can increase two-center delocalization in certain parts of the aromatic rings.

  7. The cysteine, total sulfur amino acid, tyrosine, phenylalanine + tyrosine, and non-essential amino acid maintenance requirements of broiler breeders.

    PubMed

    Ekmay, R D; Mei, S J; Sakomura, N K; Coon, C N

    2016-06-01

    Two hundred and fifty Cobb-Vantress broiler breeders were used to determine the maintenance requirement and efficiency of utilization of dietary Cys, Tyr, and non-essential amino acids (AA) in a 21-day experiment. The breeders were fed crystalline amino acid diets containing graded levels of Cys or Tyr representing 0, 10, 20, 30, and 40% of their suggested requirement level with all other amino acids maintained at 40% of their suggested requirement level. To determine the non-essential AA maintenance requirement, graded levels of non-essential AA were provided by glutamic acid to represent 12, 19, 26, 33, and 40% of the ideal level of glutamic acid with all other amino acids maintained at their maintenance requirement level. The total sulfur amino acid (TSAA) and Phe + Tyr requirements were calculated by combining Cys and Tyr results, respectively, with previously determined Met and Phe, respectively. The slope of Cys, Tyr, and non-essential AA accretion regression line indicated that 29% Cys, 24% TSAA, 21% Tyr, 20% Phe + Tyr, and 9% non-essential AA of crystalline amino acids were retained. The Cys requirement for zero protein accretion was calculated to be 30.48 mg/d or 17.006 mg/ kgBW(0.75)/d or 75.426 mg/kgCP/d. The TSAA requirement for zero accretion was calculated to be 132.25 mg/b/d, 71.48 mg/kgBW(0.75)/d, and 307.55 mg/kgCP/d. The Tyr requirement for zero protein accretion was calculated to be 65.907 mg/d or 37.233 mg/ kgBW(0.75)/d or 175.566 mg/kgCP/d. The Phe + Tyr requirement for zero protein accretion was calculated to be 352.18 mg/b/d, 190.37 mg/kgBW(0.75)/d, and 749.33 mg/kgCP/d. The non-essential AA requirement for zero protein accretion was calculated to be 3715.194 mg/d or 2003.155 mg/kgBW(0.75)/d or 9452.954 mg/kgCP/d.

  8. Dissociations of copper(II)-containing complexes of aromatic amino acids: radical cations of tryptophan, tyrosine, and phenylalanine.

    PubMed

    Siu, Chi-Kit; Ke, Yuyong; Guo, Yuzhu; Hopkinson, Alan C; Siu, K W Michael

    2008-10-14

    The dissociations of two types of copper(II)-containing complexes of tryptophan (Trp), tyrosine (Tyr), or phenylalanine (Phe) are described. The first type is the bis-amino acid complex, [Cu(II)(M)(2)].(2+), where M = Trp, Tyr, or Phe; the second [Cu(II)(4Cl-tpy)(M)].(2+), where 4Cl-tpy is the tridendate ligand 4'-chloro-2,2':6',2''-terpyridine. Dissociations of the Cu(ii) bis-amino acid complexes produce abundant radical cation of the amino acid, M.(+), and/or its secondary products. By contrast, dissociations of the 4Cl-tpy-bearing ternary complexes give abundant M.(+) only for Trp. Density functional theory (DFT) calculations show that for Tyr and Phe, amino-acid displacement reactions by H(2)O and CH(3)OH (giving [Cu(II)(4Cl-tpy)(H(2)O)].(2+) and [Cu(II)(4Cl-tpy)(CH(3)OH)].(2+)) are energetically more favorable than dissociative electron transfer (giving M.(+) and [Cu(I)(4Cl-tpy)](+)). The fragmentation pathway common to all these [Cu(II)(4Cl-tpy)(M)].(2+) ions is the loss of NH(3). DFT calculations show that the loss of NH(3) proceeds via a "phenonium-type" intermediate. Dissociative electron transfer in [Cu(II)(4Cl-tpy)(M-NH(3))].(2+) results in [M-NH(3)].(+). The [Phe-NH(3)] (+) ion dissociates facilely by eliminating CO(2) and giving a metastable phenonium-type ion that rearranges readily into the styrene radical cation.

  9. Tyrosine, phenylalanine, and tryptophan in gastroesophageal malignancy: a systematic review.

    PubMed

    Wiggins, Tom; Kumar, Sacheen; Markar, Sheraz R; Antonowicz, Stefan; Hanna, George B

    2015-01-01

    Gastroesophageal cancer has a rapidly increasing incidence worldwide and reliable biomarkers are urgently required to facilitate earlier diagnosis and improve survival. The aromatic amino acids tyrosine, phenylalanine, and tryptophan represent potential biomarkers and their relation to gastroesophageal cancer will be evaluated in this review. An electronic literature search was performed to identify all published research relating to the measurement of tyrosine, phenylalanine, or tryptophan in the biofluids or tissues of patients with gastroesophageal cancer. Sixteen studies were included in this systematic review. Six studies investigated serum concentrations, which all found decreased concentrations of these aromatic amino acids, except one study that found increased phenylalanine. Five studies reported increased concentrations within gastric content of these patients and two reported increased urinary concentrations. Tissue concentrations of these aromatic amino acids were increased in three studies. Tyrosine, phenylalanine, and tryptophan represent potential biomarkers of gastroesophageal cancer, and further research is necessary to definitively establish the mechanism responsible for altered concentrations of these compounds in patients with gastroesophageal cancer.

  10. Single-channel studies on linear gramicidins with altered amino acid sequences. A comparison of phenylalanine, tryptophane, and tyrosine substitutions at positions 1 and 11.

    PubMed Central

    Mazet, J L; Andersen, O S; Koeppe, R E

    1984-01-01

    The relation between chemical structure and permeability characteristics of transmembrane channels has been investigated with the linear gramicidins (A, B, and C), where the amino acid at position 1 was chemically replaced by phenylalanine, tryptophane or tyrosine. The purity of most of the compounds was estimated to be greater than 99.99%. The modifications resulted in a wide range of conductance changes in NaCl solutions: sixfold from tryptophane gramicidin A to tyrosine gramicidin B. The conductance changes induced by a given amino acid substitution at position 1 are not the same as at position 11. The only important change in the Na+ affinity was observed when the first amino acid was tyrosine. No major conformational changes of the polypeptide backbone structure could be detected on the basis of experiments with mixtures of different analogues and valine gramicidin A (except possibly with tyrosine at position 1), as all the compounds investigated could form hybrid channels with valine gramicidin A. The side chains are not in direct contact with the permeating ions. The results were therefore interpreted in terms of modifications of the energy profile for ion movement through the channel, possibly due to an electrostatic interaction between the dipoles of the side chains and ions in the channel. Images FIGURE 1 FIGURE 2 FIGURE 3 PMID:6201199

  11. Determination of Phenylalanine and Tyrosine by High Performance Liquid Chromatography-Tandem Mass Spectrometry.

    PubMed

    Peat, Judy; Garg, Uttam

    2016-01-01

    Hyperphenylalaninemia/phenylketonuria (PKU) is one of the most common inborn errors of amino acid metabolism affecting about 1:15,000 infants in the United States. PKU is an autosomal recessive disorder that if untreated results in mental retardation. The most common cause of PKU is deficiency of the enzyme phenylalanine hydroxylase that converts phenylalanine to tyrosine. Tyrosine deficiency results in impaired synthesis of catecholamines and thyroxine. Less commonly, it can result from defects in the synthesis or regeneration of tetrahydrobiopterin (BH4), an essential cofactor for the enzyme phenylalanine hydroxylase. Increased phenylalanine and decreased tyrosine in blood are used in the diagnosis and follow-up of patients with PKU. LC/MS/MS method is described for the quantification of phenylalanine and tyrosine.

  12. Vibrational analysis of amino acids and short peptides in hydrated media. VIII. Amino acids with aromatic side chains: L-phenylalanine, L-tyrosine, and L-tryptophan.

    PubMed

    Hernández, Belén; Pflüger, Fernando; Adenier, Alain; Kruglik, Sergei G; Ghomi, Mahmoud

    2010-11-25

    Four out of the 20 natural α-amino acids (α-AAs) contain aromatic rings in their side chains. In a recent paper (J. Phys. Chem. B 2010, 114, 9072-9083), we have analyzed the structural and vibrational features of l-histidine, one of the potent elements of this series. Here, we report on the three remaining members of this family, i.e., l-phenylalanine, l-tyrosine, and l-tryptophan. Their solution (H(2)O and D(2)O) Raman scattering and Fourier transform infrared absorption attenuated total reflection (FT-IR ATR) spectra were measured at room temperature from the species corresponding to those existing at physiological conditions. Because of the very low water solubility of tyrosine, special attention was paid to avoid any artifact concerning the report of the vibrational spectra corresponding to nondissolved powder of this AA in aqueous solution. Finally, we could obtain for the first time the Raman and FT-IR spectra of tyrosine at very low concentration (2.3 mM) upon long accumulation time. To clarify this point, those vibrational spectra of tyrosine recorded either in the solid phase or in a heterogeneous state, where dissolved and nondissolved species of this AA coexist in aqueous solution, are also provided as Supporting Information . To carry out a discussion on the general geometrical and vibrational behavior of these AAs, we resorted to quantum mechanical calculations at the DFT/B3LYP/6-31++G* level, allowing (i) determination of potential energy surfaces of these AAs in a continuum solvent as a function of the torsion angles χ(1) and χ(2), defining the conformation of each aromatic side chain around C(α)-C(β) and C(β)-C(γ) bonds, respectively; (ii) analysis of geometrical features of the AAs surrounded by clusters of n explicit (n = 5-7) water molecules interacting with the backbone and aromatic rings; and (iii) assignment of the observed vibrational modes by means of the theoretical data provided by the lowest energy conformers of explicitly

  13. The conversion of phenylalanine to tyrosine in man. Direct measurement by continuous intravenous tracer infusions of L-(ring-/sup 2/H5)phenylalanine and L-(1-/sup 13/C) tyrosine in the postabsorptive state

    SciTech Connect

    Clarke, J.T.; Bier, D.M.

    1982-10-01

    Steady state phenylalanine and tyrosine turnover and the rate of conversion of phenylalanine of tyrosine in vivo were determined in 6 healthy postabsorptive adult volunteers. Continuous infusions of tracer amounts of L-(ring-/sup 2/H5)phenylalanine were determined intravenously for 13-14 hr. After 9-10 hr, a priming dose followed by a continuous infusion of L-(1-/sup 13/C)tyrosine was added and maintained, along with the (/sup 2/H5)phenylalanine infusion, for 4 hr. Venous plasma samples were obtained before the initiation of each infusion and every 30 min during the course of the combined (/sup 2/H5)phenylalanine and (/sup 13/C)tyrosine infusion for determination of isotopic enrichments of (/sup 2/H5)phenylalanine, (/sup 13/C)tyrosine, and (/sup 2/H4)tyrosine by gas chromatograph-mass spectrometric analysis of the N-trifluoroacetyl-, methyl ester derivatives of the amino acids. Calculated from the observed enrichments, free phenylalanine and tyrosine turnover rates were 36.1 +/- 5.1 mumole . kg-1 . h-1 and 39.8 +/- 3.5 mumole . kg-1 . h-1, respectively. Phenylalanine was converted to tyrosine at the rate of 5.83 +/- 0.59 mumole . kg-1 . h-1, accounting for approximately 16% of either the phenylalanine or the tyrosine flux. The results indicate that the normal basal steady state phenylalanine hydroxylase activity in vivo in man is lower than that obtained from phenylalanine loading studies. This supports the existence of some type of substance activation of the enzyme as reflected in the previously reported exponential relationship between phenylalanine concentration and phenylalanine hydroxylase activity in vitro. The use of continuous simultaneous infusions of tracer amounts of stable isotope-labeled phenylalanine and tyrosine provides a direct means for studying physiological regulation of phenylalanine hydroxylase activity in vivo.

  14. An Overview of Phenylalanine and Tyrosine Kinetics in Humans12

    PubMed Central

    Matthews, Dwight E.

    2008-01-01

    The initial use of a tracer of phenylalanine was by Moss and Schoenheimer in rats in 1940 to determine that phenylalanine was hydroxylated to tyrosine, defining for the first time the primacy of this pathway. Phenylalanine and tyrosine kinetics were not measured in humans until the 1970–80s. The first application was for determination of the degree of blockage of phenylalanine hydroxylation in patients with hyperphenylalanemia and phenylketonuria, but this approach was expanded to determination of phenylalanine hydroxylation in normal subjects. Far more uses have been demonstrated for measuring rates of phenylalanine disposal and tyrosine production in relatively normal subjects than in patients with in-born errors of metabolism. Key to use of tracers to determine phenylalanine and tyrosine metabolic rates has been development of appropriate tracer models. Most applications have used relatively simple models ignoring the intracellular hydroxylation rate component. Because the liver is the primary site of hydroxylation in the body, the intracellular enrichment at the site of hydroxylation can be assessed from the tracer enrichments at isotopic steady state in rapid-turnover plasma proteins, such as Apo-B, made and secreted by the liver. Although there are potential problems with use of deuterated tracers of phenylalanine, suitable tracers are available and have been demonstrated for general measurement of phenylalanine and tyrosine kinetics in humans. PMID:17513423

  15. meta-Tyrosine in Festuca rubra ssp. commutata (Chewings fescue) is synthesized by hydroxylation of phenylalanine.

    PubMed

    Huang, Tengfang; Rehak, Ludmila; Jander, Georg

    2012-03-01

    m-Tyrosine is a non-protein amino acid that is structurally similar to the common protein amino acids p-tyrosine and phenylalanine. Copious amounts of m-tyrosine can be found in root exudates of the fine fescue cultivar, Festuca rubra L. ssp. commutata (Chewings fescue). The phytotoxicity of m-tyrosine may contribute to the allelopathic potential of F. rubra. m-Tyrosine in Euphorbia myrsinites (donkey-tail spurge), was previously shown to be synthesized via transamination of m-hydroxyphenylpyruvate. Here we show that m-tyrosine biosynthesis in F. rubra occurs through direct hydroxylation of phenylalanine in the root tips, perhaps through the activity of a cytochrome P450 enzyme. Hence, E. myrsinites and F. rubra, the only two plant species known to produce m-tyrosine, use distinct biosynthetic pathways that likely arose independently in evolutionary history.

  16. Chiral bio-nanocomposites based on thermally stable poly(amide-imide) having phenylalanine linkages and reactive organoclay containing tyrosine amino acid.

    PubMed

    Mallakpour, Shadpour; Dinari, Mohammad

    2013-03-01

    Montmorillonite clay modified with the bio-active trifunctional L-tyrosine amino acid salt was used as a reactive organoclay (OC) for the preparation of poly(amide-imide) (PAI)/OC hybrid films. One of the functional groups of the L-tyrosine as the swelling agent formed an ionic bond with the negatively charged silicates, whereas the remaining functional groups were available for further reaction with polymer matrix. The soluble PAI with amine end groups including phenylalanine amino acid was synthesised under green condition using molten tetra-butylammonium bromide by direct polymerization reaction of chiral diacid and 2-(3,5-diaminophenyl)benzimidazole. PAI/OC bio-nanocomposites films containing different contents of OC were prepared via solution intercalation method through blending of OC with the PAI solution. X-ray diffraction and transmission electron microscopy revealed that the dispersion of silicate layers in the PAI created an exfoliated structure as a result of using the trifunctional groups of the swelling agent. The structure and thermal behavior of the synthesised materials were characterized by a range of methods, including X-ray diffraction, Fourier transform infrared spectroscopy, (1)H-NMR, electron microscopy, elemental and thermogravimetric analysis techniques. Thermogravimetric analysis results indicated that the addition of OC into the PAI matrix was increased in the thermal decomposition temperatures of the resulted bio-nanocomposites.

  17. Rhodotorula glutinis Phenylalanine/Tyrosine Ammonia Lyase Enzyme Catalyzed Synthesis of the Methyl Ester of para-Hydroxycinnamic Acid and its Potential Antibacterial Activity.

    PubMed

    MacDonald, Marybeth C; Arivalagan, Pugazhendhi; Barre, Douglas E; MacInnis, Judith A; D'Cunha, Godwin B

    2016-01-01

    Biotransformation of L-tyrosine methyl ester (L-TM) to the methyl ester of para- hydroxycinnamic acid (p-HCAM) using Rhodotorula glutinis yeast phenylalanine/tyrosine ammonia lyase (PTAL; EC 4.3.1.26) enzyme was successfully demonstrated for the first time; progress of the reaction was followed by spectrophotometric determination at 315 nm. The following conditions were optimized for maximal formation of p-HCAM: pH (8.5), temperature (37°C), speed of agitation (50 rpm), enzyme concentration (0.080 μM), and substrate concentration (0.50 mM). Under these conditions, the yield of the reaction was ∼15% in 1 h incubation period and ∼63% after an overnight (∼18 h) incubation period. The product (p-HCAM) of the reaction of PTAL with L-TM was confirmed using Nuclear Magnetic Resonance spectroscopy (NMR). Fourier Transform Infra-Red spectroscopy (FTIR) was carried out to rule out potential hydrolysis of p-HCAM during overnight incubation. Potential antibacterial activity of p-HCAM was tested against several strains of Gram-positive and Gram-negative bacteria. This study describes a synthetically useful transformation, and could have future clinical and industrial applications.

  18. Rhodotorula glutinis Phenylalanine/Tyrosine Ammonia Lyase Enzyme Catalyzed Synthesis of the Methyl Ester of para-Hydroxycinnamic Acid and its Potential Antibacterial Activity

    PubMed Central

    MacDonald, Marybeth C.; Arivalagan, Pugazhendhi; Barre, Douglas E.; MacInnis, Judith A.; D’Cunha, Godwin B.

    2016-01-01

    Biotransformation of L-tyrosine methyl ester (L-TM) to the methyl ester of para- hydroxycinnamic acid (p-HCAM) using Rhodotorula glutinis yeast phenylalanine/tyrosine ammonia lyase (PTAL; EC 4.3.1.26) enzyme was successfully demonstrated for the first time; progress of the reaction was followed by spectrophotometric determination at 315 nm. The following conditions were optimized for maximal formation of p-HCAM: pH (8.5), temperature (37°C), speed of agitation (50 rpm), enzyme concentration (0.080 μM), and substrate concentration (0.50 mM). Under these conditions, the yield of the reaction was ∼15% in 1 h incubation period and ∼63% after an overnight (∼18 h) incubation period. The product (p-HCAM) of the reaction of PTAL with L-TM was confirmed using Nuclear Magnetic Resonance spectroscopy (NMR). Fourier Transform Infra-Red spectroscopy (FTIR) was carried out to rule out potential hydrolysis of p-HCAM during overnight incubation. Potential antibacterial activity of p-HCAM was tested against several strains of Gram-positive and Gram-negative bacteria. This study describes a synthetically useful transformation, and could have future clinical and industrial applications. PMID:27014206

  19. Deregulated tyrosine-phenylalanine metabolism in pulmonary tuberculosis patients.

    PubMed

    Das, Mrinal Kumar; Bishwal, Subasa Chandra; Das, Aleena; Dabral, Deepti; Badireddy, Vinod Kumar; Pandit, Bhaswati; Varghese, George M; Nanda, Ranjan Kumar

    2015-04-01

    Metabolic profiling of biofluids from tuberculosis (TB) patients would help us in understanding the disease pathophysiology and may also be useful for the development of novel diagnostics and host-directed therapy. In this pilot study we have compared the urine metabolic profiles of two groups of subjects having similar TB symptoms and categorized as active TB (ATB, n = 21) and non-TB (NTB, n = 21) based on GeneXpert test results. Silylation, gas chromatography mass spectrometry, and standard chemometric methods were employed to identify the important molecules and deregulated metabolic pathways. Eleven active TB patients were followed up on longitudinally for comparative urine metabolic profiling with healthy controls (n = 11). A set of 42 features qualified to have a variable importance parameter score of > 1.5 of a partial least-squares discriminate analysis model and fold change of > 1.5 at p value < 0.05 between ATB and NTB. Using these variables, a receiver operating characteristics curve was plotted and the area under the curve was calculated to be 0.85 (95% CI: 0.72-0.96). Several of these variables that represent norepinephrine, gentisic acid, 4-hydroxybenzoic acid, hydroquinone, and 4-hydroxyhippuric acid are part of the tyrosine-phenylalanine metabolic pathway. In the longitudinal study we observed a treatment-dependent trend in the urine metabolome of follow-up samples, and subjects declared as clinically cured showed similar metabolic profile as those of asymptomatic healthy subjects. The deregulated tyrosine-phenylalanine axis reveals a potential target for diagnostics and intervention in TB. PMID:25693719

  20. Deregulated tyrosine-phenylalanine metabolism in pulmonary tuberculosis patients.

    PubMed

    Das, Mrinal Kumar; Bishwal, Subasa Chandra; Das, Aleena; Dabral, Deepti; Badireddy, Vinod Kumar; Pandit, Bhaswati; Varghese, George M; Nanda, Ranjan Kumar

    2015-04-01

    Metabolic profiling of biofluids from tuberculosis (TB) patients would help us in understanding the disease pathophysiology and may also be useful for the development of novel diagnostics and host-directed therapy. In this pilot study we have compared the urine metabolic profiles of two groups of subjects having similar TB symptoms and categorized as active TB (ATB, n = 21) and non-TB (NTB, n = 21) based on GeneXpert test results. Silylation, gas chromatography mass spectrometry, and standard chemometric methods were employed to identify the important molecules and deregulated metabolic pathways. Eleven active TB patients were followed up on longitudinally for comparative urine metabolic profiling with healthy controls (n = 11). A set of 42 features qualified to have a variable importance parameter score of > 1.5 of a partial least-squares discriminate analysis model and fold change of > 1.5 at p value < 0.05 between ATB and NTB. Using these variables, a receiver operating characteristics curve was plotted and the area under the curve was calculated to be 0.85 (95% CI: 0.72-0.96). Several of these variables that represent norepinephrine, gentisic acid, 4-hydroxybenzoic acid, hydroquinone, and 4-hydroxyhippuric acid are part of the tyrosine-phenylalanine metabolic pathway. In the longitudinal study we observed a treatment-dependent trend in the urine metabolome of follow-up samples, and subjects declared as clinically cured showed similar metabolic profile as those of asymptomatic healthy subjects. The deregulated tyrosine-phenylalanine axis reveals a potential target for diagnostics and intervention in TB.

  1. Soft X-ray induced decomposition of phenylalanine and tyrosine: Acomparative study

    SciTech Connect

    Zubavichus, Y.; Zharnikov, M.; Shaporenko, A.; Fuchs, O.; Weinhart, L.; Heske, C.; Umbach, E.; Denlinger, J.D.; Grunze, M.

    2003-11-19

    The pristine state and soft X-ray induced decomposition of two aromatic amino acids, viz. phenylalanine and tyrosine, have been studied by means of XPS and NEXAFS. The spectroscopic data on the radiation decomposition have been supplemented by a mass-spectral analysis of desorbed species in the residual gas. Despite very similar chemical structures, the two amino acids show a drastically different behavior towards ionizing radiation: phenylalanine degrades very quickly whereas tyrosine shows a prominent stability against radiation damage. Reasons for this difference are discussed in relation to radical-mediated reactions responsible for the decomposition.

  2. First hyperpolarizability of the natural aromatic amino acids tryptophan, tyrosine, and phenylalanine and the tripeptide lysine-tryptophan-lysine determined by hyper-Rayleigh scattering.

    PubMed

    Duboisset, J; Matar, G; Russier-Antoine, I; Benichou, E; Bachelier, G; Jonin, Ch; Ficheux, D; Besson, F; Brevet, P F

    2010-11-01

    We report the first hyperpolarizability of tryptophan (Trp) and tyrosine (Tyr) and an upper limit for that of phenylalanine (Phe), three natural aromatic amino acids. The measurements were performed with hyper-Rayleigh scattering in an aqueous Tris buffer solution at a pH of 8.5 and 150 mM salt concentration with a fundamental wavelength of 780 nm. A value of (4.7 ± 0.7) × 10(-30) esu is found for Trp and (4.1 ± 0.7) × 10(-30) esu for Tyr whereas the upper limit of 1.4 × 10(-30) esu is found for that of Phe due to its limited solubility. The influence of the presence of lysine (Lys) in close vicinity of Trp is investigated with a measurement of the first hyperpolarizabilty of Trp in an excess of Lys and compared to the first hyperpolarizability obtained for the tripeptide Lys-Trp-Lys. The clear decrease of the values measured in these two cases indicates that the first hyperpolarizabilty of Trp is very sensitive to its local environment.

  3. No change in spontaneous behavior of rats after acute oral doses of aspartame, phenylalanine, and tyrosine.

    PubMed

    Mullenix, P J; Tassinari, M S; Schunior, A; Kernan, W J

    1991-04-01

    Spontaneous behavior subsequent to acute oral administration of high doses of aspartame, phenylalanine, or tyrosine was analyzed using a computer pattern recognition system. Sprague-Dawley male rats (250-300 g) were dosed orally with aspartame (500 or 1000 mg/kg), phenylalanine (281 or 562 mg/kg), or tyrosine (309 or 618 mg/kg), and their behavior was analyzed 1 hr after dosing. The computer pattern recognition system recorded and classified 13 different behavioral acts performed by the animals during the first 15-min exploration of a novel environment. Three measures that provide independent information concerning motor output from the central nervous system were taken: the number of behavioral initiations, total time, and time structure. These results were compared with the effects induced by d-amphetamine. Plasma concentrations of phenylalanine and tyrosine were determined from blood samples taken immediately after behavioral examination. Data analysis revealed that these doses of aspartame, phenylalanine, and tyrosine did not induce any significant changes in spontaneous behavior. Unlike low doses of amphetamine and despite high plasma concentrations of phenylalanine and tyrosine, no behavioral alteration was detected by the computer pattern recognition system. Absence of behavioral changes in this study using an objective analysis of behavior raises questions concerning the relationship between amino acid precursor loading and purported anecdotal changes in behavior following aspartame administration.

  4. Study of Interaction Between Tryptophan, Tyrosine, and Phenylalanine Separately with Silver Nanoparticles by Fluorescence Quenching Method

    NASA Astrophysics Data System (ADS)

    Roy, S.; Das, T. K.

    2015-09-01

    Using the spectroscopic method, the individual interaction of the three biochemically important amino acids, which are constituents of protein, namely, tryptophan, tyrosine, and phenylalanine with biologically synthesized silver nanoparticles has been investigated. The obtained UV-Vis spectra show the formation of ground-state complexes between tryptophan, tyrosine, and phenylalanine with silver nanoparticles. Silver nanoparticles possess the ability to quench the intrinsic fluorescence of the aforesaid amino acids by a dynamic quenching process. The binding constant, number of binding sites, and corresponding thermodynamic parameters (Δ H, Δ S, and Δ G) based on the interaction system were calculated for 293, 303, and 313 K. In the case of tryptophan and phenylalanine, with increase in temperature, the binding constant K was found to decrease; conversely, it was found to increase with increase in temperature in the case of tyrosine. The thermodynamic results revealed that the binding process was spontaneous; hydrogen bonding and van der Waals interaction were the predominant forces responsible for the complex stabilization in the case of tryptophan and phenylalanine, respectively, whereas in the case of tyrosine, hydrophobic interaction was the sole force conferring stability. Moreover, the Förster non-radiation energy transfer theory has been applied to calculate the average binding distance among the above amino acids and silver nanoparticles. The results show a binding distance of <7 nm, which ensures that energy transfer does occur between the said amino acids and silver nanoparticles.

  5. Production of tyrosine through phenylalanine hydroxylation bypasses the intrinsic feedback inhibition in Escherichia coli.

    PubMed

    Huang, Jin; Lin, Yuheng; Yuan, Qipeng; Yan, Yajun

    2015-04-01

    Tyrosine is a proteinogenic aromatic amino acid that is often used as a supplement of food and animal feed, as well as a (bio-)synthetic precursor to various pharmaceutically or industrially important molecules. Extensive metabolic engineering efforts have been made towards the efficient and cost-effective microbial production of tyrosine. Conventional strategies usually focus on eliminating intrinsic feedback inhibition and redirecting carbon flux into the shikimate pathway. In this study, we found that continuous conversion of phenylalanine into tyrosine by the action of tetrahydromonapterin (MH4)-utilizing phenylalanine 4-hydroxylase (P4H) can bypass the feedback inhibition in Escherichia coli, leading to tyrosine accumulation in the cultures. First, expression of the P4H from Xanthomonas campestris in combination with an MH4 recycling system in wild-type E. coli allowed the strain to accumulate tyrosine at 262 mg/L. On this basis, enhanced expression of the key enzymes associated with the shikimate pathway and the MH4 biosynthetic pathway resulted in the elevation of tyrosine production up to 401 mg/L in shake flasks. This work demonstrated a novel approach to tyrosine production and verified the possibility to alleviate feedback inhibition by creating a phenylalanine sink.

  6. Response to pentagastrin after acute phenylalanine and tyrosine depletion in healthy men: a pilot study.

    PubMed Central

    Coupland, N; Zedkova, L; Sanghera, G; Leyton, M; Le Mellédo, J M

    2001-01-01

    OBJECTIVE: To assess the effects of the acute depletion of the catecholamine precursors phenylalanine and tyrosine on mood and pentagastrin-induced anxiety. DESIGN: Randomized, double-blind controlled multiple crossover study. SETTING: University department of psychiatry. PARTICIPANTS: 6 healthy male volunteers. INTERVENTIONS: 3 treatments were compared: pretreatment with a nutritionally balanced amino acid mixture, followed 5 hours later by a bolus injection of normal saline placebo; pretreatment with a balanced amino acid mixture, followed by a bolus injection of pentagastrin (0.6 microgram/kg); and pretreatment with an amino acid mixture without the catecholamine precursors phenylalanine or tyrosine, followed by pentagastrin (0.6 microgram/kg). OUTCOME MEASURES: Scores on the panic symptom scale, a visual analogue scale for anxiety, the Borg scale of respiratory exertion and the Profile of Mood States Elation-Depression Scale. RESULTS: Pentagastrin produced the expected increases in anxiety symptoms, but there was no significant or discernible influence of acute phenylalanine and tyrosine depletion on anxiety or mood. CONCLUSIONS: These pilot data do not support further study using the same design in healthy men. Under these study conditions, phenylalanine and tyrosine depletion may have larger effects on dopamine than noradrenaline. Alternative protocols to assess the role of catecholamines in mood and anxiety are proposed. PMID:11394194

  7. An alternative pathway contributes to phenylalanine biosynthesis in plants via a cytosolic tyrosine:phenylpyruvate aminotransferase.

    PubMed

    Yoo, Heejin; Widhalm, Joshua R; Qian, Yichun; Maeda, Hiroshi; Cooper, Bruce R; Jannasch, Amber S; Gonda, Itay; Lewinsohn, Efraim; Rhodes, David; Dudareva, Natalia

    2013-01-01

    Phenylalanine is a vital component of proteins in all living organisms, and in plants is a precursor for thousands of additional metabolites. Animals are incapable of synthesizing phenylalanine and must primarily obtain it directly or indirectly from plants. Although plants can synthesize phenylalanine in plastids through arogenate, the contribution of an alternative pathway via phenylpyruvate, as occurs in most microbes, has not been demonstrated. Here we show that plants also utilize a microbial-like phenylpyruvate pathway to produce phenylalanine, and flux through this route is increased when the entry point to the arogenate pathway is limiting. Unexpectedly, we find the plant phenylpyruvate pathway utilizes a cytosolic aminotransferase that links the coordinated catabolism of tyrosine to serve as the amino donor, thus interconnecting the extra-plastidial metabolism of these amino acids. This discovery uncovers another level of complexity in the plant aromatic amino acid regulatory network, unveiling new targets for metabolic engineering.

  8. Evidence that phenylalanine may not provide the full needs for aromatic amino acids in children.

    PubMed

    Hsu, Jean W C; Ball, Ronald O; Pencharz, Paul B

    2007-03-01

    Phenylalanine is nutritionally classified as an indispensable amino acid and can be converted to tyrosine by phenylalanine hydroxylation. The initial goal of the present study was to determine the aromatic amino acid (phenylalanine plus tyrosine) requirements in healthy children fed a diet without tyrosine by using the indicator amino acid oxidation (IAAO) method using lysine as the indicator amino acid. Healthy school-age children (n = 5) were fed in random order a diet with eight graded intakes of phenylalanine without tyrosine. The requirement was determined by the rate of recovery of CO2 from L-[1-C]lysine oxidation (FCO2). Phenylalanine (total aromatic amino acid) requirement, in the absence of tyrosine, for children was determined to be 28 mg/kg/d, which was only 64% of the adult requirement, which is biologically absurd. A possible reason for the lower estimate of phenylalanine requirement could be lower phenylalanine hydroxylation rate in children, which is supported by the finding of lower urinary tyrosine/phenylalanine ratios in children compared with adults. In conclusion, this study indicates that phenylalanine may not provide the total needs for aromatic amino acids in children fed an amino acid-based diet without tyrosine.

  9. Amino acid-functionalized multi-walled carbon nanotubes for improving compatibility with chiral poly(amide-ester-imide) containing L-phenylalanine and L-tyrosine linkages

    NASA Astrophysics Data System (ADS)

    Abdolmaleki, Amir; Mallakpour, Shadpour; Borandeh, Sedigheh

    2013-12-01

    Amino acid functionalized multi-walled carbon nanotubes (f-MWCNTs)/poly(amide-ester-imide) (PAEI) composites were fabricated by solution mixing method. Proper functionalization and mixing strategy of MWCNTs provides the best opportunity for better distribution and bonding of nanoparticles to the polymer matrix. MWCNTs have been chemically modified with L-phenylalanine to improve their compatibility with L-phenylalanine based PAEI. Field emission scanning electron microscopy micrographs of composite revealed that f-MWCNTs made a good interaction with polymer chains by wrapping the polymer around them, and transmission electron microscopy results confirmed well dispersion with nano size of f-MWCNTs in the polymer matrix. In addition, thermal analysis showed good enhancement in thermal properties of composites compared to pure polymer. Thermal stability of the composites containing f-MWCNTs was enhanced due to their good dispersion and improved interfacial interaction between the amino acid based PAEI matrix and f-MWCNTs.

  10. Adaptation of phenylalanine and tyrosine catabolic pathway to hibernation in bats.

    PubMed

    Pan, Yi-Hsuan; Zhang, Yijian; Cui, Jie; Liu, Yang; McAllan, Bronwyn M; Liao, Chen-Chung; Zhang, Shuyi

    2013-01-01

    Some mammals hibernate in response to harsh environments. Although hibernating mammals may metabolize proteins, the nitrogen metabolic pathways commonly activated during hibernation are not fully characterized. In contrast to the hypothesis of amino acid preservation, we found evidence of amino acid metabolism as three of five key enzymes, including phenylalanine hydroxylase (PAH), homogentisate 1,2-dioxygenase (HGD), fumarylacetoacetase (FAH), involved in phenylalanine and tyrosine catabolism were co-upregulated during hibernation in two distantly related species of bats, Myotis ricketti and Rhinolophus ferrumequinum. In addition, the levels of phenylalanine in the livers of these bats were significantly decreased during hibernation. Because phenylalanine and tyrosine are both glucogenic and ketogenic, these results indicate the role of this catabolic pathway in energy supply. Since any deficiency in the catabolism of these two amino acids can cause accumulations of toxic metabolites, these results also suggest the detoxification role of these enzymes during hibernation. A higher selective constraint on PAH, HPD, and HGD in hibernators than in non-hibernators was observed, and hibernators had more conserved amino acid residues in each of these enzymes than non-hibernators. These conserved amino acid residues are mostly located in positions critical for the structure and activity of the enzymes. Taken together, results of this work provide novel insights in nitrogen metabolism and removal of harmful metabolites during bat hibernation.

  11. Adaptation of Phenylalanine and Tyrosine Catabolic Pathway to Hibernation in Bats

    PubMed Central

    Cui, Jie; Liu, Yang; McAllan, Bronwyn M.; Liao, Chen-Chung; Zhang, Shuyi

    2013-01-01

    Some mammals hibernate in response to harsh environments. Although hibernating mammals may metabolize proteins, the nitrogen metabolic pathways commonly activated during hibernation are not fully characterized. In contrast to the hypothesis of amino acid preservation, we found evidence of amino acid metabolism as three of five key enzymes, including phenylalanine hydroxylase (PAH), homogentisate 1,2-dioxygenase (HGD), fumarylacetoacetase (FAH), involved in phenylalanine and tyrosine catabolism were co-upregulated during hibernation in two distantly related species of bats, Myotis ricketti and Rhinolophus ferrumequinum. In addition, the levels of phenylalanine in the livers of these bats were significantly decreased during hibernation. Because phenylalanine and tyrosine are both glucogenic and ketogenic, these results indicate the role of this catabolic pathway in energy supply. Since any deficiency in the catabolism of these two amino acids can cause accumulations of toxic metabolites, these results also suggest the detoxification role of these enzymes during hibernation. A higher selective constraint on PAH, HPD, and HGD in hibernators than in non-hibernators was observed, and hibernators had more conserved amino acid residues in each of these enzymes than non-hibernators. These conserved amino acid residues are mostly located in positions critical for the structure and activity of the enzymes. Taken together, results of this work provide novel insights in nitrogen metabolism and removal of harmful metabolites during bat hibernation. PMID:23620802

  12. LAT-1 activity of meta-substituted phenylalanine and tyrosine analogs.

    PubMed

    Augustyn, Evan; Finke, Karissa; Zur, Arik A; Hansen, Logan; Heeren, Nathan; Chien, Huan-Chieh; Lin, Lawrence; Giacomini, Kathleen M; Colas, Claire; Schlessinger, Avner; Thomas, Allen A

    2016-06-01

    The transporter protein Large-neutral Amino Acid Transporter 1 (LAT-1, SLC7A5) is responsible for transporting amino acids such as tyrosine and phenylalanine as well as thyroid hormones, and it has been exploited as a drug delivery mechanism. Recently its role in cancer has become increasingly appreciated, as it has been found to be up-regulated in many different tumor types, and its expression levels have been correlated with prognosis. Substitution at the meta position of aromatic amino acids has been reported to increase affinity for LAT-1; however, the SAR for this position has not previously been explored. Guided by newly refined computational models of the binding site, we hypothesized that groups capable of filling a hydrophobic pocket would increase binding to LAT-1, resulting in improved substrates relative to parent amino acid. Tyrosine and phenylalanine analogs substituted at the meta position with halogens, alkyl and aryl groups were synthesized and tested in cis-inhibition and trans-stimulation cell assays to determine activity. Contrary to our initial hypothesis we found that lipophilicity was correlated with diminished substrate activity and increased inhibition of the transporter. The synthesis and SAR of meta-substituted phenylalanine and tyrosine analogs is described.

  13. LAT-1 activity of meta-substituted phenylalanine and tyrosine analogs.

    PubMed

    Augustyn, Evan; Finke, Karissa; Zur, Arik A; Hansen, Logan; Heeren, Nathan; Chien, Huan-Chieh; Lin, Lawrence; Giacomini, Kathleen M; Colas, Claire; Schlessinger, Avner; Thomas, Allen A

    2016-06-01

    The transporter protein Large-neutral Amino Acid Transporter 1 (LAT-1, SLC7A5) is responsible for transporting amino acids such as tyrosine and phenylalanine as well as thyroid hormones, and it has been exploited as a drug delivery mechanism. Recently its role in cancer has become increasingly appreciated, as it has been found to be up-regulated in many different tumor types, and its expression levels have been correlated with prognosis. Substitution at the meta position of aromatic amino acids has been reported to increase affinity for LAT-1; however, the SAR for this position has not previously been explored. Guided by newly refined computational models of the binding site, we hypothesized that groups capable of filling a hydrophobic pocket would increase binding to LAT-1, resulting in improved substrates relative to parent amino acid. Tyrosine and phenylalanine analogs substituted at the meta position with halogens, alkyl and aryl groups were synthesized and tested in cis-inhibition and trans-stimulation cell assays to determine activity. Contrary to our initial hypothesis we found that lipophilicity was correlated with diminished substrate activity and increased inhibition of the transporter. The synthesis and SAR of meta-substituted phenylalanine and tyrosine analogs is described. PMID:27106710

  14. An isotope dilution model for partitioning phenylalanine and tyrosine uptake by the mammary gland of lactating dairy cows.

    PubMed

    Crompton, L A; France, J; Reynolds, C K; Mills, J A N; Hanigan, M D; Ellis, J L; Bannink, A; Bequette, B J; Dijkstra, J

    2014-10-21

    An isotope dilution model for partitioning phenylalanine and tyrosine uptake by the mammary gland of the lactating dairy cow is constructed and solved in the steady state. The model contains four intracellular and four extracellular pools and conservation of mass principles are applied to generate the fundamental equations describing the behaviour of the system. The experimental measurements required for model solution are milk secretion and plasma flow rate across the gland in combination with phenylalanine and tyrosine concentrations and plateau isotopic enrichments in arterial and venous plasma and free and protein bound milk during a constant infusion of [1-(13)C]phenylalanine and [2,3,5,6-(2)H]tyrosine tracer. If assumptions are made, model solution enables determination of steady state flows for phenylalanine and tyrosine inflow to the gland, outflow from it and bypass, and flows representing the synthesis and degradation of constitutive protein and phenylalanine hydroxylation. The model is effective in providing information about the fates of phenylalanine and tyrosine in the mammary gland and could be used as part of a more complex system describing amino acid metabolism in the whole ruminant.

  15. α-Ketoacids as precursors for phenylalanine and tyrosine labelling in cell-based protein overexpression.

    PubMed

    Lichtenecker, Roman J; Weinhäupl, Katharina; Schmid, Walther; Konrat, Robert

    2013-12-01

    (13)C-α-ketoacid metabolic precursors of phenylalanine and tyrosine effectively enter the metabolism of a protein overexpressing E. coli strain to label Phe- and Tyr-residues devoid of any cross-labelling. The methodology gives access to highly selective labelling patterns as valuable tools in protein NMR spectroscopy without the need of (15)N-chiral amino acid synthesis using organic chemistry.

  16. Structural and optical properties of phenylalanine and tyrosine thin films prepared by pulsed laser deposition

    NASA Astrophysics Data System (ADS)

    Hernandez-Perez, M. A.; Garapon, C.; Champeaux, C.; Orlianges, J. C.

    2007-04-01

    Thin films of the amino-acids phenylalanine (Phe) and tyrosine (Tyr) were prepared by PLD with a KrF laser at fluences of some hundreds mJ/cm2. Conservation of the chemical structure and a metastable modification of the molecular interactions are evidenced by IR spectroscopy. The evolution of the refractive indices with fluence was correlated with the structure determined by X ray diffraction. Phe plume expansion imaging was achieved.

  17. Dopamine and food reward: effects of acute tyrosine/phenylalanine depletion on appetite.

    PubMed

    Hardman, Charlotte A; Herbert, Vanessa M B; Brunstrom, Jeffrey M; Munafò, Marcus R; Rogers, Peter J

    2012-03-20

    It has been suggested that obese individuals over-eat in order to compensate for deficits in the dopaminergic reward system. The current study used acute tyrosine/phenylalanine depletion (ATPD) to investigate the effect of reduced dopamine function on appetite and the reward value of food in healthy volunteers. The compensatory-eating hypothesis would predict an increase in the reward value and consumption of food following depletion by this method. In a double-blind, counterbalanced, crossover study, 17 male participants (mean age=29.2 (SEM=2.7) years; mean body mass index=24.4 (SEM=0.6) kg/m(2)) were administered with a tyrosine/phenylalanine-free mixture (TYR/PHE-free; depletion condition) and a balanced amino acid mixture (BAL; control). Plasma amino acid levels were measured at baseline and peak depletion (300 min). Appetite, willingness to pay for food, liking, desired portion size and ad libitum food intake were also assessed. The TYR/PHE-free mixture was associated with significant decreases in tyrosine, phenylalanine, and the ratio of tyrosine+phenylalanine to the other large neutral amino acids (all p<.001). There were no effects on our measures of willingness to pay for food or liking. However, in the TYR/PHE-free condition, participants reported significantly lower levels of hunger following a fixed-test meal relative to the BAL condition. In conclusion, we found no evidence for compensatory eating following ATPD. Our results also provide support for the role of dopamine in motivational components of eating.

  18. Phenylalanine and tyrosine methyl ester intramolecular interactions and conformational analysis by (1)H NMR and infrared spectroscopies and theoretical calculations.

    PubMed

    Cormanich, Rodrigo A; Ducati, Lucas C; Tormena, Cláudio F; Rittner, Roberto

    2014-04-01

    Amino acid conformational analysis in solution are scarce, since these compounds present a bipolar zwitterionic structure ((+)H3NCHRCOO(-)) in these media. Also, intramolecular hydrogen bonds have been classified as the sole interactions governing amino acid conformational behavior in the literature. In the present work we propose phenylalanine and tyrosine methyl ester conformational studies in different solvents by (1)H NMR and infrared spectroscopies and theoretical calculations. Both experimental and theoretical results are in agreement and suggest that the conformational behavior of the phenylalanine and tyrosine methyl esters are similar and are dictated by the interplay between steric and hyperconjugative interactions.

  19. Affinity chromatography and inhibition of chorismate mutase-prephenate dehydrogenase by derivatives of phenylalanine and tyrosine.

    PubMed Central

    Smith, G D; Roberts, D V; Daday, A

    1977-01-01

    Several derivatives of phenylalanine and tyrosine were prepared and tested for inhibition of chorismate mutase-prephenate dehydrogenase (EC 1.3.1.12) from Escherichia coli K12 (strain JP 232). The best inhibitors were N-toluene-p-sulphonyl-L-phenylalanine, N-benzenesulphonyl-L-phenylalanine and N-benzloxycarbonyl-L-phenylalanine. Consequently two compounds, N-toluene-sulphonyl-L-p-aminophenylalanine and N-p-aminobenzenesulphonyl-L-phenylalanine, were synthesized for coupling to CNBr-activated Sepharose-4B. The N-toluene-p-sulphonyl-L-p-aminophenylalanine-Sepharose-4B conjugate was shown to bind the enzyme very strongly at pH 7.5. The enzyme was not eluted by various eluents, including 1 M-NaCl, but could be quantitatively recovered by washing with buffer of pH9. Elution was more effective in the presence of 10 mM-1-adamantaneacetic acid, a competitive inhibitor of the enzyme. This affinity-chromatography procedure results in a high degree of purification of the enzyme and can be used to prepare the enzyme in a one-step procedure from the bacterial crude extract. Such a procedure may therefore prove useful in studying this enzyme in a state that closely resembles that in vivo. PMID:889568

  20. Excited State Dynamics of Protonated Phenylalanine and Tyrosine: Photo-Induced Reactions Following Electronic Excitation.

    PubMed

    Féraud, Géraldine; Broquier, Michel; Dedonder, Claude; Jouvet, Christophe; Grégoire, Gilles; Soorkia, Satchin

    2015-06-11

    The electronic spectroscopy and the electronic excited state properties of cold protonated phenylalanine and protonated tyrosine have been revisited on a large spectral domain and interpreted by comparison with ab initio calculations. The protonated species are stored in a cryogenically cooled Paul trap, maintained at ∼10 K, and the parent and all the photofragment ions are mass-analyzed in a time-of-flight mass spectrometer, which allows detecting the ionic species with an improved mass resolution compared to what is routinely achieved with a quadrupole mass spectrometer. These new results emphasize the competition around the band origin between two proton transfer reactions from the ammonium group toward either the aromatic chromophore or the carboxylic acid group. These reactions are initiated by the coupling of the locally excited ππ* state with higher charge transfer states, the positions and coupling of which depend on the conformation of the protonated molecules. Each of these reaction processes gives rise to specific fragmentation channels that supports the conformer selectivity observed in the photofragmentation spectra of protonated tyrosine and phenylalanine.

  1. Interferon-alpha therapy in patients with hepatitis C virus infection increases plasma phenylalanine and the phenylalanine to tyrosine ratio.

    PubMed

    Zoller, Heinz; Schloegl, Anna; Schroecksnadel, Sebastian; Vogel, Wolfgang; Fuchs, Dietmar

    2012-05-01

    Higher blood levels of the essential amino acid phenylalanine (Phe) together with impaired conversion of Phe to tyrosine (Tyr) have been observed in patients suffering from inflammatory conditions. Data suggest that inflammatory responses may interfere with Phe metabolism. This study aimed to investigate whether treatment with cytokine interferon-α (IFN-α) influences Phe concentrations and the Phe to Tyr ratios (Phe/Tyr) measured by HPLC. Twenty-five patients (9 females, 16 males, aged mean ± SD: 44.5 ± 11.0 years) with hepatitis C virus (HCV) infection were examined before and after 1 month of effective antiviral therapy with pegylated IFN-α and weight-based ribavirin. Results were compared to HCV-RNA titers and concentrations of neopterin. IFN-α treatment was associated with a drop of HCV load (from median 6.3 to 3.2 log10 copies/μL; P<0.001) and an increase of neopterin concentrations (from median 4.83 to 12.1 nM; P=0.001) which confirms effectiveness of therapy. Before therapy, median Phe concentration were 123.9 μM, Tyr was 98.8 μM, and Phe/Tyr was 1.23 μmol/μmol, and under therapy median Phe concentrations increased to 132.6 μM and Phe/Tyr to 1.33 (both P<0.05; paired rank test), Tyr levels remained unchanged. The increase of Phe concentrations and of Phe/Tyr in HCV infected individuals is caused by IFN-α therapy. Data indicate that activity of enzyme phenylalanine 4-hydroxylase becomes impaired. Future studies should show whether side effects of IFN-α treatment such as mood changes and depression will be associated with the alterations of Phe metabolism.

  2. New peptide architectures through C–H activation stapling between tryptophan–phenylalanine/tyrosine residues

    PubMed Central

    Mendive-Tapia, Lorena; Preciado, Sara; García, Jesús; Ramón, Rosario; Kielland, Nicola; Albericio, Fernando; Lavilla, Rodolfo

    2015-01-01

    Natural peptides show high degrees of specificity in their biological action. However, their therapeutical profile is severely limited by their conformational freedom and metabolic instability. Stapled peptides constitute a solution to these problems and access to these structures lies on a limited number of reactions involving the use of non-natural amino acids. Here, we describe a synthetic strategy for the preparation of unique constrained peptides featuring a covalent bond between tryptophan and phenylalanine or tyrosine residues. The preparation of such peptides is achieved in solution and on solid phase directly from the corresponding sequences having an iodo-aryl amino acid through an intramolecular palladium-catalysed C–H activation process. Moreover, complex topologies arise from the internal stapling of cyclopeptides and double intramolecular arylations within a linear peptide. Finally, as a proof of principle, we report the application to this new stapling method to relevant biologically active compounds. PMID:25994485

  3. Evaluation of an adsorbent based on agricultural waste (corn cobs) for removal of tyrosine and phenylalanine from aqueous solutions.

    PubMed

    Alves, Cibele C O; Franca, Adriana S; Oliveira, Leandro S

    2013-01-01

    Adsorption of phenolic amino acids, such as phenylalanine and tyrosine, is quite relevant for the production of protein hydrolysates used as dietary formulations for patients suffering from congenital disorders of amino acid metabolism, such as phenylketonuria. In this study, an adsorbent prepared from corn cobs was evaluated for the removal of tyrosine (Tyr) from both a single component solution and a binary aqueous solution with phenylalanine (Phe). The adsorption behavior of tyrosine was similar to that of phenylalanine in single component solutions, however, with a much lower adsorption capacity (14 mg g(-1) for Tyr compared to 109 mg g(-1) for Phe). Tyr adsorption kinetics was satisfactorily described by a pseudosecond-order model as it was for Phe. In adsorption equilibrium studies for binary mixtures, the presence of Tyr in Phe solutions favored Phe faster adsorption whereas the opposite behavior was observed for the presence of Phe in Tyr solutions. Such results indicate that, in binary systems, Phe will be adsorbed preferably to Tyr, and this is a welcome feature when employing the prepared adsorbent for the removal of Phe from protein hydrolysates to be used in dietary formulations for phenylketonuria treatment.

  4. Photostability of amino acids: photodissociation dynamics of phenylalanine chromophores.

    PubMed

    Tseng, Chien-Ming; Lin, Ming-Fu; Yang, Yi Lin; Ho, Yu Chieh; Ni, Chi-Kung; Chang, Jia-Lin

    2010-05-21

    The theoretical prediction of H atom elimination on the excited state of phenol, imidazole and indole, the respective chromophores for the amino acids tyrosine, histidine and tryptophan, and the confirmation of theoretical prediction by experimental observations have a great impact on the explanation of photostability of amino acids upon irradiation with UV photons. On the other hand, no theoretical prediction of the excited state photodissociation dynamics has been made on the other aromatic amino acid, phenylalanine. In this work, photodissociation dynamics for various phenylalanine chromophores, including, phenylethylamine, N-methyl-phenylethylamine, and N-acetyl phenylalanine methyl ester was investigated in a molecular beam at 248 and 193 nm using multimass ion imaging techniques. The major dissociation channel for these compounds is the C-C bond cleavage. However, the photofragment translational energy distribution of phenylethylamine contains two components. The slow component corresponds to the dissociation on the ground state surface after internal conversion, and the fast component represents the dissociation from an excited state with a large exit barrier. The competition between the dissociation on the ground state and on the excited state changes as the size of chromophores increases. Internal conversion to the ground state prior to dissociation becomes the major nonradiative process for large chromophores. This study reveals the size-dependent photostability for these amino acid chromophores.

  5. Effect of water coordination on competition between π and non-π cation binding sites in aromatic amino acids: L-phenylalanine, L-tyrosine, and L-tryptophan Li+, Na +, and K+ complexes.

    PubMed

    Remko, Milan; Šoralová, Stanislava

    2012-04-01

    Quantum chemistry methods have been applied to charged complexes of the alkali metals Li(+), Na(+), and K(+) with the aromatic amino acids (AAAs) phenylalanine (Phe), tyrosine (Tyr), and tryptophan (Trp). The geometries of 72 different complexes (Phe·M, Tyr·M, Trp·M, M is Li(+), Na(+), or K(+)) were completely optimized at the B3LYP/6-311+G(d,p) level of density functional theory. The solvent effect on the geometry and stability of individual complexes was studied by making use of a microsolvation model. The interaction enthalpies, entropies, and Gibbs energies of nine different complexes of the systems Phe·M, Tyr·M, and Trp·M (M is Li(+), Na(+), or K(+)) were also determined at the B3LYP density functional level of theory. The calculated Gibbs binding energies of the M(+)-AAA complexes follow the order Phe < Tyr < Trp for all three metal cations studied. Among the three AAAs studied, the indole ring of Trp is the best π donor for alkali metal cations. Our calculations demonstrated the existence of strong cation-π interactions between the alkali metals and the aromatic side chains of the three AAAs. These AAAs comprise about 8% of all known protein sequences. Thus, besides the potential for hydrogen-bond interaction, aromatic residues of Phe, Tyr, and Trp show great potential for π-donor interactions. The existence of cation-π interaction in proteins has also been demonstrated experimentally. However, more complex experimental studies of metal cation-π interaction in diverse biological systems will no doubt lead to more exact validation of these investigations.

  6. Production of tyrosine and other aromatic compounds from phenylalanine by rumen microorganisms.

    PubMed

    Khan, R I; Onodera, R; Amin, M R; Mohammed, N

    1999-01-01

    Rumen contents from three fistulated Japanese native goats fed Lucerne hay cubes (Medicago sativa) and concentrate mixture were collected to prepare the suspensions of mixed rumen bacteria (B), mixed protozoa (P) and a combination of the two (BP). Microbial suspensions were anaerobically incubated at 39 degrees C for 12 h with or without 1 mM of L-phenylalanine (Phe). Phe, tyrosine (Tyr) and other related compounds in both supernatant and microbial hydrolysates of the incubations were analyzed by HPLC. Tyr can be produced from Phe not only by rumen bacteria but also by rumen protozoa. The production of Tyr during 12 h incubation in B (183.6 mumol/g MN) was 4.3 times higher than that in P. One of the intermediate products between Phe and Tyr seems to be p-hydroxyphenylacetic acid. The rate of the net degradation of Phe incubation in B (76.0 mumol/g MN/h) was 2.4 times higher than in P. In the case of all rumen microorganisms, degraded Phe was mainly (> 53%) converted into phenylacetic acid. The production of benzoic acid was higher in P than in B suspensions. Small amount of phenylpyruvic acid was produced from Phe by both rumen bacteria and protozoa, but phenylpropionic acid and phenyllactic acid were produced only by rumen bacteria.

  7. Identification of β-phenylalanine as a non-protein amino acid in cultivated rice, Oryza sativa.

    PubMed

    Yokoo, Takayuki; Takata, Ryo; Yan, Jian; Matsumoto, Fuka; Teraishi, Masayoshi; Okumoto, Yutaka; Jander, Georg; Mori, Naoki

    2015-01-01

    Non-protein amino acids, often analogs of the standard 20 protein amino acids, have been discovered in many plant species. Recent research with cultivated rice (Oryza sativa) identified (3R)-β-tyrosine, as well as a tyrosine amino mutase that synthesizes (3R)-β-tyrosine from the protein amino acid (2S)-α-tyrosine. Gas chromatography-mass spectrometry (GC-MS) assays and comparison to an authentic standard showed that β-phenylalanine is also a relatively abundant non-protein amino acid in rice leaves and that its biosynthesis occurs independently from that of β-tyrosine.

  8. Identification of β-phenylalanine as a non-protein amino acid in cultivated rice, Oryza sativa

    PubMed Central

    Yokoo, Takayuki; Takata, Ryo; Yan, Jian; Matsumoto, Fuka; Teraishi, Masayoshi; Okumoto, Yutaka; Jander, Georg; Mori, Naoki

    2015-01-01

    Non-protein amino acids, often analogs of the standard 20 protein amino acids, have been discovered in many plant species. Recent research with cultivated rice (Oryza sativa) identified (3R)-β-tyrosine, as well as a tyrosine amino mutase that synthesizes (3R)-β-tyrosine from the protein amino acid (2S)-α-tyrosine. Gas chromatography-mass spectrometry (GC-MS) assays and comparison to an authentic standard showed that β-phenylalanine is also a relatively abundant non-protein amino acid in rice leaves and that its biosynthesis occurs independently from that of β-tyrosine. PMID:27066169

  9. Syntheses of halogen derivatives of L-tryptophan, L-tyrosine and L-phenylalanine labeled with hydrogen isotopes.

    PubMed

    Pająk, Małgorzata; Pałka, Katarzyna; Winnicka, Elżbieta; Kańska, Marianna

    2016-01-01

    Halogenated, labeled with tritium and doubly with deuterium and tritium, derivatives of L-tryptophan, i.e. 5'-bromo-[2-(3)H]-, 5'-bromo-[2-(2)H/(3)H]-, 5'-fluoro-[2-(3)H]-5'-fluoro-[2-(2)H/(3)H]-, 6'-fluoro-[2-(3)H]-, 6'-fluoro-[2-(2)H/(3)H]-L-tryptophan, as well as, L-tyrosine, i.e. 3'-fluoro-[2-(3)H]-, 3'-fluoro-[2-(2)H/(3)H]-, 3'-chloro-[2-(3)H]-, and 3'-chloro-[2-(2)H/(3)H]-L-tyrosine, and also L-phenylalanine, i.e. 2'-fluoro-[(3S)-(3)H]-, 2'-fluoro-[(3S)-(2)H/(3) H]-, 2'-chloro-[(3S)-(3)H]-, 2'-chloro-[(3S)-(2)H/(3)H]-, 4'-chloro-[(3S)-(3)H]-, and 4'-chloro-[(3S)-(2)H/(3)H]-L-phenylalanine were synthesized using enzymatic methods. Isotopomers of L-tryptophan were synthesized by coupling of halogenated indoles with S-methyl-L-cysteine carried out in deuteriated or tritiated incubation media. Labeled halogenated derivatives of L-tyrosine were obtained by the enzymatically supported exchange between halogenated L-tyrosine and isotopic water. Labeled halogenated isotopologues of L-Phe were synthesized by the enzymatic addition of ammonia to halogenated cinnamic acid. As a source of hydrogen tritiated water (HTO) and heavy water (D2O) with addition of HTO were used.

  10. Rapid quantitative method for the detection of phenylalanine and tyrosine in human plasma using pillar array columns and gradient elution.

    PubMed

    Song, Yanting; Takatsuki, Katsuya; Sekiguchi, Tetsushi; Funatsu, Takashi; Shoji, Shuichi; Tsunoda, Makoto

    2016-07-01

    This study reports a fast and quantitative determination method for phenylalanine (Phe) and tyrosine (Tyr) in human plasma using on-chip pressure-driven liquid chromatography. A pillar array column with low-dispersion turns and a gradient elution system was used. The separation of fluorescent derivatives of Phe, Tyr, and other hydrophobic amino acids was successfully performed within 140 s. Under the optimized conditions, Phe and Tyr in human plasma were quantified. The developed method is promising for rapid diagnosis in the clinical field.

  11. Bacteriorhodopsin mutants containing single tyrosine to phenylalanine substitutions are all active in proton translocation.

    PubMed Central

    Mogi, T; Stern, L J; Hackett, N R; Khorana, H G

    1987-01-01

    To study the possible role of the tyrosine residues in proton translocation by bacteriorhodopsin, we have replaced these residues individually by phenylalanine. The required codon changes were introduced in the bacterioopsin gene by replacement of appropriate restriction fragments by synthetic counterparts containing the desired nucleotide changes. The denatured opsin polypeptides obtained by expression of the mutant genes in Escherichia coli were purified and treated with a mixture of detergents, phospholipids, and retinal in a previously established renaturation procedure. All of the mutant proteins folded to regenerate bacteriorhodopsin-like chromophores. Three mutants with tyrosine to phenylalanine substitutions at positions 57, 83, and 185 regenerated the chromophore more slowly than the wild-type protein, and two of these mutants, Phe-57 and -83, showed slightly blue-shifted chromophores. When reconstituted into liposomes all of the mutant proteins with single Tyr----Phe substitutions pumped protons at rates and levels comparable to those of the wild-type bacteriorhodopsin. We conclude that single substitutions of tyrosine by phenylalanine do not affect folding, retinal binding, or light-dependent proton pumping in bacteriorhodopsin. PMID:3039495

  12. Polyuridylic Acid-directed Phenylalanine Incorporation in Minicell Extracts

    PubMed Central

    Fralick, J. A.; Fisher, W. D.; Adler, H. I.

    1969-01-01

    Cell-free extracts of miniature Escherichia coli cells deficient in deoxyribonucleic acid (DNA) and DNA-dependent ribonucleic acid polymerase have been shown to be capable of polyuridylic acid-directed [14C]phenylalanine incorporation. PMID:4897117

  13. Thermal broadening of Lb band of ``trehalose coated'' tyrosine and phenylalanine

    NASA Astrophysics Data System (ADS)

    Carrotta, Rita; Sanfratello, Vincenzo; Leone, Maurizio; Cordone, Lorenzo

    2000-04-01

    We studied the thermal broadening of Lb band of tyrosine and phenylalanine embedded in a trehalose matrix. Aim of this work is to obtain information on the effects of "trehalose coating" on the coupling of the electronic transition to low frequency modes in the surrounding of the chromophore. The results obtained for the two molecular complexes put in evidence that O-H groups are involved in blocking these structures within the solid trehalose matrix and shed light on the role played by hydrogen bonds on the interactions that keep "trehalose coated" proteins rigid and solid-like.

  14. Dietary tyrosine/phenylalanine depletion effects on behavioral and brain signatures of human motivational processing.

    PubMed

    Bjork, James M; Grant, Steven J; Chen, Gang; Hommer, Daniel W

    2014-02-01

    Dopamine (DA) neurotransmission is critical for motivational processing. We assessed whether disruption of DA synthesis in healthy controls using an amino-acid beverage devoid of catecholamine precursors (tyrosine-phenylalanine depletion (TPD)) would blunt recruitment of the nucleus accumbens (NAcc) by rewards. Sixteen controls ingested each of a tyr/phe-depleting beverage (DEP) or a tyr/phe-balanced (BAL) control beverage in two laboratory visits. Five hours after consumption of each drink, subjects underwent functional magnetic resonance imaging while they viewed anticipatory cues to respond to a target to either win money or avoid losing money. TPD did not exert main effects on mood or on task behavior, but affected brain activation. In right NAcc, TPD blunted activation by anticipation of high rewards. In left NAcc, recruitment anticipating high rewards was modulated by individual differences in mood change across the DEP drink day, where subjects whose mood worsened following TPD (relative to within-day mood change under BAL conditions) also showed lower activation under DEP conditions relative to BAL conditions. Exploratory analysis indicated that TPD qualitatively blunted the voxel-wise spatial extent of suprathreshold activation by reward anticipation. Finally, loss outcomes activated anterior insula under DEP conditions but not under BAL conditions. These data indicate that: (1) dietary depletion of catacholamine precursors will blunt dopaminergic mesolimbic activity, and (2) in controls, synthetic pathways of this neurocircuitry maintain sufficient buffering capacity to resist an effect on motivated behavior. Additional studies are needed to determine if clinical populations would show similar resistance to behavioral effects of TPD.

  15. Dietary tyrosine/phenylalanine depletion effects on behavioral and brain signatures of human motivational processing.

    PubMed

    Bjork, James M; Grant, Steven J; Chen, Gang; Hommer, Daniel W

    2014-02-01

    Dopamine (DA) neurotransmission is critical for motivational processing. We assessed whether disruption of DA synthesis in healthy controls using an amino-acid beverage devoid of catecholamine precursors (tyrosine-phenylalanine depletion (TPD)) would blunt recruitment of the nucleus accumbens (NAcc) by rewards. Sixteen controls ingested each of a tyr/phe-depleting beverage (DEP) or a tyr/phe-balanced (BAL) control beverage in two laboratory visits. Five hours after consumption of each drink, subjects underwent functional magnetic resonance imaging while they viewed anticipatory cues to respond to a target to either win money or avoid losing money. TPD did not exert main effects on mood or on task behavior, but affected brain activation. In right NAcc, TPD blunted activation by anticipation of high rewards. In left NAcc, recruitment anticipating high rewards was modulated by individual differences in mood change across the DEP drink day, where subjects whose mood worsened following TPD (relative to within-day mood change under BAL conditions) also showed lower activation under DEP conditions relative to BAL conditions. Exploratory analysis indicated that TPD qualitatively blunted the voxel-wise spatial extent of suprathreshold activation by reward anticipation. Finally, loss outcomes activated anterior insula under DEP conditions but not under BAL conditions. These data indicate that: (1) dietary depletion of catacholamine precursors will blunt dopaminergic mesolimbic activity, and (2) in controls, synthetic pathways of this neurocircuitry maintain sufficient buffering capacity to resist an effect on motivated behavior. Additional studies are needed to determine if clinical populations would show similar resistance to behavioral effects of TPD. PMID:23995581

  16. No Tryptophan, Tyrosine and Phenylalanine Abnormalities in Children with Attention-Deficit/Hyperactivity Disorder

    PubMed Central

    Bergwerff, Catharina Elisabeth; Luman, Marjolein; Blom, Henk J.; Oosterlaan, Jaap

    2016-01-01

    Background The aim of the current study was to explore the role of aromatic amino acids (AAAs) in blood in relation to attention-deficit/hyperactivity disorder (ADHD). Given their impact on the synthesis of serotonin and dopamine, decreased concentrations of the AAAs tryptophan, tyrosine and phenylalanine in blood may contribute to the expression of ADHD symptoms. Decreased AAA blood concentrations, in turn, may be related to lowered dietary protein intake or to abnormal AAA catabolism, as evidenced by increased urinary AAA concentrations. Methods Eighty-three children with ADHD (75% males) and 72 typically developing (TD) children (51% males), aged 6 to 13 years, participated in the study. AAA concentrations were assessed in blood spots and an 18-hour urinary sample. A nutritional diary was filled out by parents to calculate dietary protein intake. Parent and teacher questionnaires assessed symptoms of ADHD, oppositional defiant disorder, conduct disorder, and autism spectrum disorder. Results Children with ADHD showed normal AAA concentrations in blood spots and urine, as well as normal protein intake compared to controls. No associations between AAA concentrations and symptoms of ADHD or comorbid psychiatric disorders were found. Conclusions This study is the first to explore AAA metabolism in children with ADHD using a well-defined and relatively large sample. We found that AAA deficiencies are not related to ADHD. The results do not support treatment with AAA supplements in children with ADHD. Future studies regarding the cause of serotonin and dopamine alterations in ADHD should focus on other explanations, such as effects of altered transport of AAAs. PMID:26938936

  17. SERS study of transformation of phenylalanine to tyrosine under particle irradiation

    NASA Astrophysics Data System (ADS)

    Zhang, Jingjing; Huang, Qing; Yao, Guohua; Ke, Zhigang; Zhang, Hong; Lu, Yilin

    2014-08-01

    Surface enhanced Raman scattering or spectroscopy (SERS) is a very powerful analytical tool which has been widely applied in many scientific research and application fields. It is therefore also very intriguing for us to introduce SERS technique in the radiobiological research, where in many cases only a very few of biomolecules are subjected to changes which however can lead to significant biological effects. The radiation induced biochemical reactions are normally very sophisticated with different substances produced in the system, so currently it is still a big challenge for SERS to analyze such a mixture system which contains multiple analytes. In this context, this work aimed to establish and consolidate the feasibility of SERS as an effective tool in radiation chemistry, and this purpose, we employed SERS as a sensitive probe to a known process, namely, the oxidation of phenylalanine (Phe) under particle irradiation, where the energetic particles were obtained from either plasma discharge or electron-beam. During the irradiation, three types of tyrosine (Tyr), namely, p-Tyr, m-Tyr and o-Tyr were produced, and all these tyrosine isomers together with Phe could be identified and measured based on the SERS spectral analysis of the corresponding enhanced characteristic signals, namely, 1002 cm-1 for Phe, 1161 cm-1 for p-Tyr, 990 cm-1 for m-Tyr, and 970 cm-1 for o-Tyr, respectively. The estimation of the quantities of different tyrosine isomers were also given and verified by conventional method such as high performance liquid chromatography (HPLC). As for comparison of different ways of particle irradiation, our results also indicated that electron-beam irradiation was more efficient for converting Phe into Tyr than plasma discharge treatment, confirming the role of hydroxyl radicals in the Phe-Tyr conformation. Therefore, our work has not only demonstrated that SERS can be successfully applied in the radiobiological study, but also given insights into the

  18. Effect of tyrosine on the potentiation by aspartame and phenylalanine of metrazol-induced convulsions in rats.

    PubMed

    Guiso, G; Diomede, L; Romano, M; Caccia, S; Sarati, S; Salmona, M

    1991-12-01

    Male rats were treated by oral intubation with tyrosine (Tyr), at doses of 0.5 and 1.0 g/kg body weight, alone or together with 1 g aspartame (APM)/kg body weight, or an equivalent dose of phenylalanine (Phe; 0.5 g/kg body weight); the effects on seizures induced by an effective dose of metrazol (ED50) were observed. Tyr (0.5 g/kg body weight) had a protective effect against the Phe-potentiation of metrazol-induced clonic-tonic convulsions. At the same dose Tyr had no effect on the seizure-promoting activity of APM, but at 1 g/kg it reduced the proconvulsant potential of the sweetener. Analysis of the brain and plasma amino acid concentrations indicated that the Tyr to Phe ratio tended to be enhanced in Tyr-Phe treated rats compared with those treated with Phe alone. This ratio remained essentially constant in the brain of APM-treated rats, compared with those treated with APM plus 1 g Tyr/kg body weight, whereas an increase in this ratio in the plasma was observed. These results confirm that Tyr antagonizes the proconvulsant effect of Phe and APM and they further suggest that no simple relationship exists between the relative brain concentrations of the two amino acids and the response to metrazol convulsions.

  19. Enhancement of L-phenylalanine production by engineered Escherichia coli using phased exponential L-tyrosine feeding combined with nitrogen source optimization.

    PubMed

    Yuan, Peipei; Cao, Weijia; Wang, Zhen; Chen, Kequan; Li, Yan; Ouyang, Pingkai

    2015-07-01

    Nitrogen source optimization combined with phased exponential L-tyrosine feeding was employed to enhance L-phenylalanine production by a tyrosine-auxotroph strain, Escherichia coli YP1617. The absence of (NH4)2SO4, the use of corn steep powder and yeast extract as composite organic nitrogen source were more suitable for cell growth and L-phenylalanine production. Moreover, the optimal initial L-tyrosine level was 0.3 g L(-1) and exponential L-tyrosine feeding slightly improved L-phenylalanine production. Nerveless, L-phenylalanine production was greatly enhanced by a strategy of phased exponential L-tyrosine feeding, where exponential feeding was started at the set specific growth rate of 0.08, 0.05, and 0.02 h(-1) after 12, 32, and 52 h, respectively. Compared with exponential L-tyrosine feeding at the set specific growth rate of 0.08 h(-1), the developed strategy obtained a 15.33% increase in L-phenylalanine production (L-phenylalanine of 56.20 g L(-1)) and a 45.28% decrease in L-tyrosine supplementation.

  20. Serum Phenylalanine, Tyrosine, and their Ratio in Acute Ischemic Stroke: on the Trail of a Biomarker?

    PubMed

    Ormstad, Heidi; Verkerk, Robert; Sandvik, Leiv

    2016-01-01

    Fast diagnosis and appropriate treatment are of utmost importance to improving the outcome in patients with acute ischemic stroke (AIS). A rapid and sensitive blood test for ischemic stroke is required. The aim of this study was to examine the usefulness of phenylalanine (PHE) and tyrosine (TYR) as diagnostic biomarkers in AIS. Serum levels of PHE and TYR, measured using HPLC, and their ratio (PHE/TYR) were compared between 45 patients with AIS and 40 healthy control subjects. The relationship between PHE/TYR and the serum levels of several cytokines were also examined. PHE/TYR was significantly higher in AIS patients than in healthy controls (1.75 vs 1.24, p < 0.001). A receiver operating characteristic (ROC) curve analysis of PHE/TYR in AIS patients relative to healthy controls revealed promising sensitivity and specificity, which at an optimal cutoff of 1.45 were 76 and 85 %, respectively. PHE/TYR was positively correlated with interleukin (IL)-1β (r = 0.37, p = 0.011) and IL-6 (r = 0.33, p = 0.025). This study shows that PHE/TYR is highly elevated in the acute phase of AIS, and that this elevation is coupled to the inflammatory response. The ROC analysis documents the possible value of PHE/TYR as a biomarker for AIS and demonstrates its clinical potential as a blood-based test for AIS.

  1. [Difluro(phosphono)methyl]phenylalanine-containing peptide inhibitors of protein tyrosine phosphatases.

    PubMed Central

    Desmarais, S; Friesen, R W; Zamboni, R; Ramachandran, C

    1999-01-01

    Peptides containing the non-hydrolysable phosphotyrosine analogue 4-[difluro(phosphono)methyl]phenylalanine [Phe(CF2P)] were synthesized and tested as inhibitors of the protein tyrosine phosphatases (PTPs) PTP1B, CD45, PTPbeta, LAR and SHP-1. We have identified peptides containing two adjacent Phe(CF2P) residues as potent inhibitors of PTPs. The tripeptide having the sequence Glu-Phe(CF2P)-Phe(CF2P) is a potent and selective inhibitor of PTP1B. This peptide inhibits PTP1B with an IC50 of 40 nM, which is at least 100-fold lower than with other PTPs. A second tripeptide, Pro-Phe(CF2P)-Phe(CF2P), is most potent against PTPbeta, with an IC50 of 200 nM, and inhibits PTP1B with an IC50 of 300 nM. These data suggest that it is possible to develop selective, active-site-directed, reversible, potent inhibitors of PTPs. PMID:9882618

  2. The effect of acute tyrosine phenylalanine depletion on emotion-based decision-making in healthy adults.

    PubMed

    Vrshek-Schallhorn, Suzanne; Wahlstrom, Dustin; White, Tonya; Luciana, Monica

    2013-04-01

    Despite interest in dopamine's role in emotion-based decision-making, few reports of the effects of dopamine manipulations are available in this area in humans. This study investigates dopamine's role in emotion-based decision-making through a common measure of this construct, the Iowa Gambling Task (IGT), using Acute Tyrosine Phenylalanine Depletion (ATPD). In a between-subjects design, 40 healthy adults were randomized to receive either an ATPD beverage or a balanced amino acid beverage (a control) prior to completing the IGT, as well as pre- and post-manipulation blood draws for the neurohormone prolactin. Together with conventional IGT performance metrics, choice selections and response latencies were examined separately for good and bad choices before and after several key punishment events. Changes in response latencies were also used to predict total task performance. Prolactin levels increased significantly in the ATPD group but not in the control group. However, no significant group differences in performance metrics were detected, nor were there sex differences in outcome measures. However, the balanced group's bad deck latencies speeded up across the task, while the ATPD group's latencies remained adaptively hesitant. Additionally, modulation of latencies to the bad decks predicted total score for the ATPD group only. One interpretation is that ATPD subtly attenuated reward salience and altered the approach by which individuals achieved successful performance, without resulting in frank group differences in task performance.

  3. Phenylalanine supplementation improves the phenylalanine profile in tyrosinaemia.

    PubMed

    Wilson, C J; Van Wyk, K G; Leonard, J V; Clayton, P T

    2000-11-01

    Tyrosinaemia types I and II are caused by enzyme deficiencies in the tyrosine catabolism pathway. Successful treatment is possible with the novel enzyme inhibitor NTBC in tyrosinaemia type I and with dietary tyrosine and phenylalanine restriction in both conditions. This is achieved with a low natural protein intake and a supplementary amino acid formula that is phenylalanine- and tyrosine-free. Patients on this regimen had been noted, periodically, to have very low plasma phenylalanine concentrations (<20 micromol/L). The tyrosine and phenylalanine profiles in six patients were measured. Five of the six patients had very low concentrations of phenylalanine during the later half of the day. The response to phenylalanine supplementation was assessed and supplementing the diet with phenylalanine 30-40 mg/kg per day resulted in normal concentrations throughout the day. Possible complications of hypophenylalaninaemia and potential preventive treatment strategies are discussed. Further studies are needed to investigate the longer-term clinical and biochemical consequences of phenylalanine supplementation.

  4. Dopamine and pain sensitivity: neither sulpiride nor acute phenylalanine and tyrosine depletion have effects on thermal pain sensations in healthy volunteers.

    PubMed

    Becker, Susanne; Ceko, Marta; Louis-Foster, Mytsumi; Elfassy, Nathaniel M; Leyton, Marco; Shir, Yoram; Schweinhardt, Petra

    2013-01-01

    Based on animal studies and some indirect clinical evidence, dopamine has been suggested to have anti-nociceptive effects. Here, we investigated directly the effects of increased and decreased availability of extracellular dopamine on pain perception in healthy volunteers. In Study 1, participants ingested, in separate sessions, a placebo and a low dose of the centrally acting D2-receptor antagonist sulpiride, intended to increase synaptic dopamine via predominant pre-synaptic blockade. No effects were seen on thermal pain thresholds, tolerance, or temporal summation. Study 2 used the acute phenylalanine and tyrosine depletion (APTD) method to transiently decrease dopamine availability. In one session participants ingested a mixture that depletes the dopamine amino acid precursors, phenylalanine and tyrosine. In the other session they ingested a nutritionally balanced control mixture. APTD led to a small mood-lowering response following aversive thermal stimulation, but had no effects on the perception of cold, warm, or pain stimuli. In both studies the experimental manipulation of dopaminergic neurotransmission was successful as indicated by manipulation checks. The results contradict proposals that dopamine has direct anti-nociceptive effects in acute experimental pain. Based on dopamine's well-known role in reward processing, we hypothesize that also in the context of pain, dopamine acts on stimulus salience and might play a role in the initiation of avoidance behavior rather than having direct antinociceptive effects in acute experimental pain.

  5. Effect of amino acids on the formation of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) in creatinine/phenylalanine and creatinine/phenylalanine/4-oxo-2-nonenal reaction mixtures.

    PubMed

    Zamora, Rosario; Alcón, Esmeralda; Hidalgo, Francisco J

    2013-12-15

    2-Amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) formation in mixtures of creatinine, phenylalanine, amino acids and 4-oxo-2-nonenal was studied, to analyse the role of amino acids on the generation of this heterocyclic aromatic amine. When oxidised lipid was absent, cysteine, serine, aspartic acid, threonine, asparagine, tryptophan, tyrosine, proline, and methionine increased significantly (p < 0.05) the amount of PhIP formed in comparison to the control. When lipid was present, only the addition of methionine, glycine, and serine increased significantly (p < 0.05) the amount of PhIP produced, while histidine, cysteine, lysine, tryptophan, tyrosine, and alanine reduced significantly (p < 0.05) PhIP. These results may be a consequence of the different competitive reactions that occur. Thus, in the absence of lipids, thermal decomposition of the amino acids produced reactive carbonyls that converted phenylalanine into phenylacetaldehyde as a key step in the formation of PhIP. When oxidised lipid was present, amino acids competed with phenylalanine for the lipid, and amino acid degradation products were formed, among which alpha-keto acids seemed to play a role in these reactions. These results suggest that PhIP can be produced by several alternative reaction pathways from all major food components, including amino acids and lipids, in addition to carbohydrates. PMID:23993611

  6. Effect of amino acids on the formation of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) in creatinine/phenylalanine and creatinine/phenylalanine/4-oxo-2-nonenal reaction mixtures.

    PubMed

    Zamora, Rosario; Alcón, Esmeralda; Hidalgo, Francisco J

    2013-12-15

    2-Amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) formation in mixtures of creatinine, phenylalanine, amino acids and 4-oxo-2-nonenal was studied, to analyse the role of amino acids on the generation of this heterocyclic aromatic amine. When oxidised lipid was absent, cysteine, serine, aspartic acid, threonine, asparagine, tryptophan, tyrosine, proline, and methionine increased significantly (p < 0.05) the amount of PhIP formed in comparison to the control. When lipid was present, only the addition of methionine, glycine, and serine increased significantly (p < 0.05) the amount of PhIP produced, while histidine, cysteine, lysine, tryptophan, tyrosine, and alanine reduced significantly (p < 0.05) PhIP. These results may be a consequence of the different competitive reactions that occur. Thus, in the absence of lipids, thermal decomposition of the amino acids produced reactive carbonyls that converted phenylalanine into phenylacetaldehyde as a key step in the formation of PhIP. When oxidised lipid was present, amino acids competed with phenylalanine for the lipid, and amino acid degradation products were formed, among which alpha-keto acids seemed to play a role in these reactions. These results suggest that PhIP can be produced by several alternative reaction pathways from all major food components, including amino acids and lipids, in addition to carbohydrates.

  7. Molecular Characterization of a Recombinant Zea mays Phenylalanine Ammonia-Lyase (ZmPAL2) and Its Application in trans-Cinnamic Acid Production from L-Phenylalanine.

    PubMed

    Zang, Ying; Jiang, Ting; Cong, Ying; Zheng, Zhaojuan; Ouyang, Jia

    2015-06-01

    Phenylalanine ammonia-lyase (PAL) is one of the most extensively studied enzymes with its crucial role in secondary phenylpropanoid metabolism of plants. Recently, its demand has been increased for aromatic chemical production, but its applications in trans-cinnamic acid production were not much explored. In the present study, a putative PAL gene from Zea mays designated as ZmPAL2 was expressed and characterized in Escherichia coli BL21 (DE3). The recombinant ZmPAL2 exhibited a high PAL activity (7.14 U/mg) and a weak tyrosine ammonia-lyase activity. The optimal temperature of ZmPAL2 was 55 °C, and the thermal stability results showed that about 50 % of enzyme activity remained after a treatment at 60 °C for 6 h. The recombinant ZmPAL2 is a good candidate for the production of trans-cinnamic acid. The vitro conversion indicated that the recombinant ZmPAL2 could effectively catalyze the L-phenylalanine to trans-cinnamic acid, and the trans-cinnamic acid concentration can reach up to 5 g/l.

  8. Synthesis of aromatic (13)C/(2)H-α-ketoacid precursors to be used in selective phenylalanine and tyrosine protein labelling.

    PubMed

    Lichtenecker, R J

    2014-10-14

    Recent progress in protein NMR spectroscopy revealed aromatic residues to be valuable information sources for performing structure and motion analysis of high molecular weight proteins. However, the applied NMR experiments require tailored isotope labelling patterns in order to regulate spin-relaxation pathways and optimize magnetization transfer. We introduced a methodology to use α-ketoacids as metabolic amino acid precursors in cell-based overexpression of phenylalanine and/or tyrosine labelled proteins in a recent publication, which we have now developed further by providing synthetic routes to access the corresponding side-chain labelled precursors. The target compounds allow for selective introduction of (13)C-(1)H spin systems in a highly deuterated chemical environment and feature alternating (12)C-(13)C-(12)C ring-patterns. The resulting isotope distribution is especially suited to render straightforward (13)C spin relaxation experiments possible, which provide insight into the dynamic properties of the corresponding labelled proteins.

  9. Differential Laser-Induced Perturbation Spectroscopy for Analysis of Mixtures of the Fluorophores l-Phenylalanine, l-Tyrosine and l-Tryptophan Using a Fluorescence Probe.

    PubMed

    Oztekin, Erman K; Hahn, David W

    2016-09-01

    Quantitative detection of common endogenous fluorophores is accomplished using differential laser-induced perturbation spectroscopy (DLIPS) with a 193-nm UV fluorescence probe and various UV perturbation wavelengths. In this study, DLIPS is explored as an alternative to traditional fluorescence spectroscopy alone, with a goal of exploring natural fluorophores pursuant to biological samples and tissue analysis. To this end, aromatic amino acids, namely, l-phenylalanine, l-tyrosine and l-tryptophan are mixed with differing mass ratios and then classified with various DLIPS schemes. Classification with a traditional fluorescence probe is used as a benchmark. The results show a 20% improvement in classification performance of the DLIPS method over the traditional fluorescence method using partial least squares (PLS) analysis. Additional multivariate analyses are explored, and the relevant photochemistry is elucidated in the context of perturbation wavelengths. We conclude that DLIPS is a promising biosensing approach with potential for in vivo analysis given the current findings with fluorophores relevant to biological tissues.

  10. Cinnamic acid production using Streptomyces lividans expressing phenylalanine ammonia lyase.

    PubMed

    Noda, Shuhei; Miyazaki, Takaya; Miyoshi, Takanori; Miyake, Michiru; Okai, Naoko; Tanaka, Tsutomu; Ogino, Chiaki; Kondo, Akihiko

    2011-05-01

    Cinnamic acid production was demonstrated using Streptomyces as a host. A gene encoding phenylalanine ammonia lyase (PAL) from Streptomyces maritimus was introduced into Streptomyces lividans, and its expression was confirmed by Western blot analysis. After 4 days cultivation using glucose as carbon source, the maximal level of cinnamic acid reached 210 mg/L. When glycerol (30 g/L) was used as carbon source, the maximal level of produced cinnamic acid reached 450 mg/L. In addition, using raw starch, xylose or xylan as carbon source, the maximal level of cinnamic acid reached 460, 300, and 130 mg/L, respectively. We demonstrated that S. lividans has great potential to produce cinnamic acid as well as other aromatic compounds.

  11. Phenylalanine and tyrosine levels are rate-limiting factors in production of health promoting metabolites in Vitis vinifera cv. Gamay Red cell suspension.

    PubMed

    Manela, Neta; Oliva, Moran; Ovadia, Rinat; Sikron-Persi, Noga; Ayenew, Biruk; Fait, Aaron; Galili, Gad; Perl, Avichai; Weiss, David; Oren-Shamir, Michal

    2015-01-01

    Environmental stresses such as high light intensity and temperature cause induction of the shikimate pathway, aromatic amino acids (AAA) pathways, and of pathways downstream from AAAs. The induction leads to production of specialized metabolites that protect the cells from oxidative damage. The regulation of the diverse AAA derived pathways is still not well understood. To gain insight on that regulation, we increased AAA production in red grape Vitis vinifera cv. Gamay Red cell suspension, without inducing external stress on the cells, and characterized the metabolic effect of this induction. Increased AAA production was achieved by expressing a feedback-insensitive bacterial form of 3-deoxy- D-arabino-heptulosonate 7-phosphate synthase enzyme (AroG (*)) of the shikimate pathway under a constitutive promoter. The presence of AroG(*) protein led to elevated levels of primary metabolites in the shikimate and AAA pathways including phenylalanine and tyrosine, and to a dramatic increase in phenylpropanoids. The AroG (*) transformed lines accumulated up to 20 and 150 fold higher levels of resveratrol and dihydroquercetin, respectively. Quercetin, formed from dihydroquercetin, and resveratrol, are health promoting metabolites that are induced due to environmental stresses. Testing the expression level of key genes along the stilbenoids, benzenoids, and phenylpropanoid pathways showed that transcription was not affected by AroG (*). This suggests that concentrations of AAAs, and of phenylalanine in particular, are rate-limiting in production of these metabolites. In contrast, increased phenylalanine production did not lead to elevated concentrations of anthocyanins, even though they are also phenylpropanoid metabolites. This suggests a control mechanism of this pathway that is independent of AAA concentration. Interestingly, total anthocyanin concentrations were slightly lower in AroG(*) cells, and the relative frequencies of the different anthocyanins changed as well.

  12. Phenylalanine and tyrosine levels are rate-limiting factors in production of health promoting metabolites in Vitis vinifera cv. Gamay Red cell suspension

    PubMed Central

    Manela, Neta; Oliva, Moran; Ovadia, Rinat; Sikron-Persi, Noga; Ayenew, Biruk; Fait, Aaron; Galili, Gad; Perl, Avichai; Weiss, David; Oren-Shamir, Michal

    2015-01-01

    Environmental stresses such as high light intensity and temperature cause induction of the shikimate pathway, aromatic amino acids (AAA) pathways, and of pathways downstream from AAAs. The induction leads to production of specialized metabolites that protect the cells from oxidative damage. The regulation of the diverse AAA derived pathways is still not well understood. To gain insight on that regulation, we increased AAA production in red grape Vitis vinifera cv. Gamay Red cell suspension, without inducing external stress on the cells, and characterized the metabolic effect of this induction. Increased AAA production was achieved by expressing a feedback-insensitive bacterial form of 3-deoxy- D-arabino-heptulosonate 7-phosphate synthase enzyme (AroG*) of the shikimate pathway under a constitutive promoter. The presence of AroG* protein led to elevated levels of primary metabolites in the shikimate and AAA pathways including phenylalanine and tyrosine, and to a dramatic increase in phenylpropanoids. The AroG* transformed lines accumulated up to 20 and 150 fold higher levels of resveratrol and dihydroquercetin, respectively. Quercetin, formed from dihydroquercetin, and resveratrol, are health promoting metabolites that are induced due to environmental stresses. Testing the expression level of key genes along the stilbenoids, benzenoids, and phenylpropanoid pathways showed that transcription was not affected by AroG*. This suggests that concentrations of AAAs, and of phenylalanine in particular, are rate-limiting in production of these metabolites. In contrast, increased phenylalanine production did not lead to elevated concentrations of anthocyanins, even though they are also phenylpropanoid metabolites. This suggests a control mechanism of this pathway that is independent of AAA concentration. Interestingly, total anthocyanin concentrations were slightly lower in AroG* cells, and the relative frequencies of the different anthocyanins changed as well. PMID:26236327

  13. Caffeic acid production enhancement by engineering a phenylalanine over-producing Escherichia coli strain.

    PubMed

    Huang, Qin; Lin, Yuheng; Yan, Yajun

    2013-12-01

    Caffeic acid is a plant-specific phenylpropanoic acid with multiple health-improving effects reported, and its therapeutic derivatives have also been studied throughout the last decade. To meet its market need and achieve high-level production, microbial production of caffeic acid approaches have been developed in metabolically engineered Escherichia coli. In our previous work, we have established the first artificial pathway that realized de novo production of caffeic acid using E. coli endogenous 4-hydroxyphenylacetate 3-hydroxylase (4HP3H). In this work, we exploited the catalytic potential of 4HPA3H in the whole-cell bioconversion study and produced 3.82 g/L (461.12 mg/L/OD) caffeic acid from p-coumaric acid, a direct precursor. We further engineered a phenylalanine over-producer into a tyrosine over-producer and then introduced the artificial pathway. After adjusting the expression strategy and optimizing the inoculants timing, de novo production of caffeic acid reached 766.68 mg/L. Both results from the direct precursor and simple carbon sources represent the highest titers of caffeic acid from microbial production so far.

  14. Rotational Spectra of Phenylalanine, Tirosine and Tryptophan

    NASA Astrophysics Data System (ADS)

    Mata, S.; Perez, C.; Sanz, M. E.; Blanco, S.; López, J. C.; Alonso, J. L.

    2009-06-01

    The rotational spectra of the aromatic natural amino acids phenylalanine, tyrosine and tryptophan have been investigated by Laser Ablation Molecular Beam Fourier transform Microwave Spectroscopy LA-MB-FTMW. The spectra of two rotamers of phenylalanine have been detected in the supersonic expansion. Both forms are stabilized by a chain of intramolecular hydrogen bonds O-H\\cdotsN-H\\cdots{π}, being the carboxylic group incis configuration. One conformer of tyrosine, which only differs from phenylalanine in a -OH group inpara position, has been also characterized. Preliminary results on the rotational spectrum of tryptophan are presented.

  15. Discovery of novel glitazones incorporated with phenylalanine and tyrosine: synthesis, antidiabetic activity and structure-activity relationships.

    PubMed

    Prashantha Kumar, B R; Baig, Nasir R; Sudhir, Sai; Kar, Koyal; Kiranmai, M; Pankaj, M; Joghee, Nanjan M

    2012-12-01

    We report a series of new glitazones incorporated with phenylalanine and tyrosine. All the compounds were tested for their in vitro glucose uptake activity using rat-hemidiaphragm, both in presence and absence of insulin. Six of the most active compounds from the in vitro screening were taken forward for their in vivo triglyceride and glucose lowering activity against dexamethazone induced hyperlipidemia and insulin resistance in Wistar rats. The liver samples of rats that received the most active compounds, 23 and 24, in the in vivo studies, were subjected to histopathological examination to assess their short term hepatotoxicity. The investigations on the in vitro glucose uptake, in vivo triglyceride and glucose lowering activity are described here along with the quantitative structure-activity relationships.

  16. Replacement of tyrosine D with phenylalanine affects the normal proton transfer pathways for the reduction of P680+ in oxygen-evolving photosystem II particles from Chlamydomonas.

    PubMed

    Jeans, C; Schilstra, M J; Ray, N; Husain, S; Minagawa, J; Nugent, J H A; Klug, D R

    2002-12-31

    We have probed the electrostatics of P680(+) reduction in oxygenic photosynthesis using histidine-tagged and histidine-tagged Y(D)-less Photosystem II cores. We make two main observations: (i) that His-tagged Chlamydomonas cores show kinetics which are essentially identical to those of Photosystem II enriched thylakoid membranes from spinach; (ii) that the microsecond kinetics, previously shown to be proton/hydrogen transfer limited [Schilstra et al. (1998) Biochemistry 37, 3974-3981], are significantly different in Y(D)-less Chlamydomonas particles when compared with both the His-tagged Chlamydomonas particles and the spinach membranes. The oscillatory nature of the kinetics in both Chlamydomonas samples is normal, indicating that S-state cycling is unaffected by either the histidine-tagging or the replacement of tyrosine D with phenylalanine. We propose that the effects on the proton-coupled electron transfers of P680(+) reduction in the absence of Y(D) are likely to be due to pK shifts of residues in a hydrogen-bonded network of amino acids in the vicinity of Y(Z). Tyrosine D is 35 A from Y(Z) and yet has a significant influence on proton-coupled electron transfer events in the vicinity of Y(Z). This finding emphasizes the delicacy of the proton balance that Photosystem II has to achieve during the water splitting process. PMID:12501204

  17. Effect of dietary aspartame on plasma concentrations of phenylalanine and tyrosine in normal and homozygous phenylketonuric patients.

    PubMed

    Mackey, S A; Berlin, C M

    1992-07-01

    Six normal subjects each ingested a single 12-oz can of a diet cola (Diet Coke) providing 184 mg aspartame (APM), of which 104 mg is phenylalanine (Phe), and, on another occasion, a single 12-oz can of regular cola (Coke Classic). Neither cola significantly affected plasma concentrations of Phe or tyrosine over the three-hour postingestion study period. Each of five homozygous phenylketonuric (PKU) subjects (ages 11, 16, 17, 21, and 23 years) ingested a single 12-oz can of the same diet cola. In these five subjects (three with classic PKU and two with hyperphenylalinemia), the increase in plasma Phe concentrations varied from 0.26 mg/dL to 1.77 mg/dL two or three hours after ingestion (baseline levels, 5.04 to 17.2 mg/dL). Tyrosine concentrations did not differ significantly from baseline levels. The data indicate that ingestion of dietary Phe, as supplied in a single can of diet cola, is readily handled in both normal and PKU subjects. The small increases in plasma Phe concentrations in the homozygous PKU patients are not considered clinically significant.

  18. Identification of a plastidial phenylalanine exporter that influences flux distribution through the phenylalanine biosynthetic network.

    PubMed

    Widhalm, Joshua R; Gutensohn, Michael; Yoo, Heejin; Adebesin, Funmilayo; Qian, Yichun; Guo, Longyun; Jaini, Rohit; Lynch, Joseph H; McCoy, Rachel M; Shreve, Jacob T; Thimmapuram, Jyothi; Rhodes, David; Morgan, John A; Dudareva, Natalia

    2015-01-01

    In addition to proteins, L-phenylalanine is a versatile precursor for thousands of plant metabolites. Production of phenylalanine-derived compounds is a complex multi-compartmental process using phenylalanine synthesized predominantly in plastids as precursor. The transporter(s) exporting phenylalanine from plastids, however, remains unknown. Here, a gene encoding a Petunia hybrida plastidial cationic amino-acid transporter (PhpCAT) functioning in plastidial phenylalanine export is identified based on homology to an Escherichia coli phenylalanine transporter and co-expression with phenylalanine metabolic genes. Radiolabel transport assays show that PhpCAT exports all three aromatic amino acids. PhpCAT downregulation and overexpression result in decreased and increased levels, respectively, of phenylalanine-derived volatiles, as well as phenylalanine, tyrosine and their biosynthetic intermediates. Metabolic flux analysis reveals that flux through the plastidial phenylalanine biosynthetic pathway is reduced in PhpCAT RNAi lines, suggesting that the rate of phenylalanine export from plastids contributes to regulating flux through the aromatic amino-acid network. PMID:26356302

  19. Identification of a plastidial phenylalanine exporter that influences flux distribution through the phenylalanine biosynthetic network.

    PubMed

    Widhalm, Joshua R; Gutensohn, Michael; Yoo, Heejin; Adebesin, Funmilayo; Qian, Yichun; Guo, Longyun; Jaini, Rohit; Lynch, Joseph H; McCoy, Rachel M; Shreve, Jacob T; Thimmapuram, Jyothi; Rhodes, David; Morgan, John A; Dudareva, Natalia

    2015-09-10

    In addition to proteins, L-phenylalanine is a versatile precursor for thousands of plant metabolites. Production of phenylalanine-derived compounds is a complex multi-compartmental process using phenylalanine synthesized predominantly in plastids as precursor. The transporter(s) exporting phenylalanine from plastids, however, remains unknown. Here, a gene encoding a Petunia hybrida plastidial cationic amino-acid transporter (PhpCAT) functioning in plastidial phenylalanine export is identified based on homology to an Escherichia coli phenylalanine transporter and co-expression with phenylalanine metabolic genes. Radiolabel transport assays show that PhpCAT exports all three aromatic amino acids. PhpCAT downregulation and overexpression result in decreased and increased levels, respectively, of phenylalanine-derived volatiles, as well as phenylalanine, tyrosine and their biosynthetic intermediates. Metabolic flux analysis reveals that flux through the plastidial phenylalanine biosynthetic pathway is reduced in PhpCAT RNAi lines, suggesting that the rate of phenylalanine export from plastids contributes to regulating flux through the aromatic amino-acid network.

  20. Identification of a plastidial phenylalanine exporter that influences flux distribution through the phenylalanine biosynthetic network

    PubMed Central

    Widhalm, Joshua R.; Gutensohn, Michael; Yoo, Heejin; Adebesin, Funmilayo; Qian, Yichun; Guo, Longyun; Jaini, Rohit; Lynch, Joseph H.; McCoy, Rachel M.; Shreve, Jacob T.; Thimmapuram, Jyothi; Rhodes, David; Morgan, John A.; Dudareva, Natalia

    2015-01-01

    In addition to proteins, L-phenylalanine is a versatile precursor for thousands of plant metabolites. Production of phenylalanine-derived compounds is a complex multi-compartmental process using phenylalanine synthesized predominantly in plastids as precursor. The transporter(s) exporting phenylalanine from plastids, however, remains unknown. Here, a gene encoding a Petunia hybrida plastidial cationic amino-acid transporter (PhpCAT) functioning in plastidial phenylalanine export is identified based on homology to an Escherichia coli phenylalanine transporter and co-expression with phenylalanine metabolic genes. Radiolabel transport assays show that PhpCAT exports all three aromatic amino acids. PhpCAT downregulation and overexpression result in decreased and increased levels, respectively, of phenylalanine-derived volatiles, as well as phenylalanine, tyrosine and their biosynthetic intermediates. Metabolic flux analysis reveals that flux through the plastidial phenylalanine biosynthetic pathway is reduced in PhpCAT RNAi lines, suggesting that the rate of phenylalanine export from plastids contributes to regulating flux through the aromatic amino-acid network. PMID:26356302

  1. Light-switched inhibitors of protein tyrosine phosphatase PTP1B based on phosphonocarbonyl phenylalanine as photoactive phosphotyrosine mimetic.

    PubMed

    Wagner, Stefan; Schütz, Anja; Rademann, Jörg

    2015-06-15

    Phosphopeptide mimetics containing the 4-phosphonocarbonyl phenylalanine (pcF) as a photo-active phosphotyrosine isoster are developed as potent, light-switchable inhibitors of the protein tyrosine phosphatase PTP1B. The photo-active inhibitors 6-10 are derived from phosphopeptide substrates and are prepared from the suitably protected pcF building block 12 by Fmoc-based solid phase peptide synthesis. All pcF-containing peptides are moderate inhibitors of PTP1B with KI values between 10 and 50μM. Irradiation of the inhibitors at 365nm in the presence of the protein PTP1B amplify the inhibitory activity of pcF-peptides up to 120-fold, switching the KI values of the best inhibitors to the sub-micromolar range. Photo-activation of the inhibitors results in the formation of triplet intermediates of the benzoylphosphonate moiety, which deactivate PTP1B following an oxidative radical mechanism. Deactivation of PTP1B proceeds without covalent crosslinking of the protein target with the photo-switched inhibitors and can be reverted by subsequent addition of reducing agent dithiothreitol (DTT).

  2. Effect of feeding garlic leaf on microbial nitrogen supply, kinetics of plasma phenylalanine, tyrosine and protein synthesis in sheep.

    PubMed

    Kamruzzaman, Md; Liang, Xi; Sekiguchi, Natsumi; Sano, Hiroaki

    2014-05-01

    The objective of the present study was to assess the feeding effects of garlic leaf on microbial N supply (MNS), turnover rates of plasma phenylalanine (PheTR) and tyrosine (TyrTR) and whole body protein synthesis (WBPS) in sheep. The sheep were fed either mixed hay (Hay-diet, as control) or hay plus garlic leaf diet (GL-diet, at a ratio of 9:1) in a crossover design each for a 21 day period. The isotope dilution method using [(2) H5 ]Phe and [(2) H2 ]Tyr was performed on the 21st day of each dietary treatment. Nitrogen intake remained similar between the diets and N absorption and N digestibility were higher (P<0.05) in the GL-diet than Hay-diet. Total purine derivatives excretion and MNS were greater (P<0.05) in the GL-diet than the Hay-diet. Plasma PheTR tended to be higher (P=0.06) during GL feeding and TyrTR did not differ between the diets. Further, WBPS tended to be greater (P=0.05) for the GL-diet compared with the Hay-diet. Hence, the present results suggest that garlic leaf may have positive effects on N metabolism by influencing MNS in sheep and could be used as a potential ruminant feed in the future.

  3. Transfer RNA gene numbers may not be completely responsible for the codon usage bias in asparagine, isoleucine, phenylalanine, and tyrosine in the high expression genes in bacteria.

    PubMed

    Satapathy, Siddhartha Sankar; Dutta, Malay; Buragohain, Alak Kumar; Ray, Suvendra Kumar

    2012-08-01

    It is generally believed that the effect of translational selection on codon usage bias is related to the number of transfer RNA genes in bacteria, which is more with respect to the high expression genes than the whole genome. Keeping this in the background, we analyzed codon usage bias with respect to asparagine, isoleucine, phenylalanine, and tyrosine amino acids. Analysis was done in seventeen bacteria with the available gene expression data and information about the tRNA gene number. In most of the bacteria, it was observed that codon usage bias and tRNA gene number were not in agreement, which was unexpected. We extended the study further to 199 bacteria, limiting to the codon usage bias in the two highly expressed genes rpoB and rpoC which encode the RNA polymerase subunits β and β', respectively. In concordance with the result in the high expression genes, codon usage bias in rpoB and rpoC genes was also found to not be in agreement with tRNA gene number in many of these bacteria. Our study indicates that tRNA gene numbers may not be the sole determining factor for translational selection of codon usage bias in bacterial genomes.

  4. Formation of self-aggregated structures of different types in water of chiral polymerizable amphiphiles from L-tyrosine and L-phenylalanine.

    PubMed

    Angayarkanny, S; Vijay, R; Baskar, Geetha; Mandal, A B

    2012-06-26

    Sodium salts of maleamic acid derivatives from lauryl esters of L-tyrosine (MTNa) and L-phenylalanine (MPNa) were synthesized and characterized. The aggregated structures of MTNa and MPNa in water were investigated, employing several independent methods. MPNa showed secondary aggregated structures in contrast to MTNa at concentrations of >1 × 10(-3) M. The results from dynamic light scattering, transmittance, conductivity, and viscosity measurements suggested the formation of aggregated structures of different types in MTNa and MPNa solutions. The measured fluorescence anisotropy (r) at 0.180 of the fluoroprobe, 1,6-diphenyl-1,3,5-hexatriene (DPH), and the d spacing of 38 Å from small-angle X-ray diffraction (SAXD) experiments confirmed the bilayer structures in MPNa. Scanning electron microscope (SEM) images provided the morphological features. The emulsion produced using MPNa solution was more stable. The confocal fluorescence microscopy image of the emulsion from MPNa confirmed the entrapment of water-soluble dye, rhodamine. The models of MTNa and MPNa molecules and the aggregated structures are presented.

  5. Quantitative analysis of phenylalanine, tyrosine, tryptophan and kynurenine in rat model for tauopathies by ultra-high performance liquid chromatography with fluorescence and mass spectrometry detection.

    PubMed

    Galba, Jaroslav; Michalicova, Alena; Parrak, Vojtech; Novak, Michal; Kovac, Andrej

    2016-01-01

    We developed and validated a simple and sensitive ultra-high performance liquid chromatography (UHPLC) method for the analysis of phenylalanine (Phe), tyrosine (Tyr), tryptophan (Trp) and kynurenine (Kyn) in rat plasma. Analytes were separated on Acquity UPLC HSS T3 column (2.1 mm×50 mm, 1.8 μm particle size) using a 4 min ammonium acetate (pH 5) gradient and detected by fluorescence and positive ESI mass spectrometry. Sample preparation involved dilution of plasma, deproteinization by trichloroacetic acid and centrifugation. The procedure was validated in compliance with the FDA guideline. The limits of quantification (LOQ) were 0.3 μM for Kyn and from 1.5 to 3 μM for Phe, Tyr, Trp. The method showed excellent linearity with regression coefficients higher than 0.99. The accuracy was within the range of 86-108%. The inter-day precision (n=5 days), expressed as % RSD, was in the range 1-13%. The benefit of using UHPLC is a short analysis period and thus, a very good sample throughput. Using this method, we analyzed plasma samples and detected significant changes of Kyn and Phe in transgenic rat model for tauopathies.

  6. Amino acid supplementation does not alter whole-body phenylalanine kinetics in Arabian geldings.

    PubMed

    Urschel, Kristine L; Geor, Raymond J; Hanigan, Mark D; Harris, Pat A

    2012-03-01

    Stable isotope infusion methods have not been extensively used in horses to study protein metabolism. The objectives were to develop infusion and sampling methodologies for [1-(13)C] phenylalanine and apply these methods to determine whether the addition of supplemental amino acids to a control diet affected whole-body phenylalanine kinetics in mature horses. Arabian geldings were studied using a 6-h primed (9 μmol/kg), constant (6 μmol · kg(-1) · h(-1)) i.v. infusion of L-[1-(13)C] phenylalanine, with blood and breath sampled every 30 min, to measure whole-body phenylalanine kinetics in response to receiving the control diet (n = 12) or the control diet supplemented with equimolar amounts of glutamate (+Glu; 55 mg · kg(-1) · d(-1); n = 5), leucine (+Leu; 49 mg · kg(-1) · d(-1); n = 5), lysine (+Lys; 55 mg · kg(-1) · d(-1); n = 5), or phenylalanine (+Phe; 62 mg · kg(-1) · d(-1); n = 6). The plasma concentrations of the supplemented amino acid in horses receiving the +Leu, +Lys, and +Phe diets were 58, 53, and 36% greater, respectively, than for the control treatment (P < 0.05). Isotopic plateau was attained in blood [1-(13)C] phenylalanine and breath (13)CO(2) enrichments by 60 and 270 min, respectively. Phenylalanine flux (+20%) and oxidation (+110%) were greater (P < 0.05) in horses receiving the +Phe treatment than in those fed the control diet. There was no effect of treatment diet on nonoxidative phenylalanine disposal or phenylalanine release from protein breakdown. The developed methods are a valuable way to study protein metabolism and assess dietary amino acid adequacy in horses and will provide a useful tool for studying amino acid requirements in the future. PMID:22259192

  7. Amino acid supplementation does not alter whole-body phenylalanine kinetics in Arabian geldings.

    PubMed

    Urschel, Kristine L; Geor, Raymond J; Hanigan, Mark D; Harris, Pat A

    2012-03-01

    Stable isotope infusion methods have not been extensively used in horses to study protein metabolism. The objectives were to develop infusion and sampling methodologies for [1-(13)C] phenylalanine and apply these methods to determine whether the addition of supplemental amino acids to a control diet affected whole-body phenylalanine kinetics in mature horses. Arabian geldings were studied using a 6-h primed (9 μmol/kg), constant (6 μmol · kg(-1) · h(-1)) i.v. infusion of L-[1-(13)C] phenylalanine, with blood and breath sampled every 30 min, to measure whole-body phenylalanine kinetics in response to receiving the control diet (n = 12) or the control diet supplemented with equimolar amounts of glutamate (+Glu; 55 mg · kg(-1) · d(-1); n = 5), leucine (+Leu; 49 mg · kg(-1) · d(-1); n = 5), lysine (+Lys; 55 mg · kg(-1) · d(-1); n = 5), or phenylalanine (+Phe; 62 mg · kg(-1) · d(-1); n = 6). The plasma concentrations of the supplemented amino acid in horses receiving the +Leu, +Lys, and +Phe diets were 58, 53, and 36% greater, respectively, than for the control treatment (P < 0.05). Isotopic plateau was attained in blood [1-(13)C] phenylalanine and breath (13)CO(2) enrichments by 60 and 270 min, respectively. Phenylalanine flux (+20%) and oxidation (+110%) were greater (P < 0.05) in horses receiving the +Phe treatment than in those fed the control diet. There was no effect of treatment diet on nonoxidative phenylalanine disposal or phenylalanine release from protein breakdown. The developed methods are a valuable way to study protein metabolism and assess dietary amino acid adequacy in horses and will provide a useful tool for studying amino acid requirements in the future.

  8. Comparison of fluorescence reagents for simultaneous determination of hydroxylated phenylalanine and nitrated tyrosine by high-performance liquid chromatography with fluorescence detection.

    PubMed

    Iwasaki, Yusuke; Mochizuki, Keisuke; Nakano, Yuki; Maruya, Natsumi; Goto, Masato; Maruyama, Yosuke; Ito, Rie; Saito, Koichi; Nakazawa, Hiroyuki

    2012-01-01

    Reactive oxygen species (ROS) and reactive nitrogen species (RNS) are well-known and important contributors to oxidative and nitrosative stress in several diseases. Hydroxylated phenylalanine and nitrated tyrosine products appear to be particularly susceptible targets of oxidative and nitrosative stress. We compared fluorescence reagents for their potential use in the analysis of hydroxylated phenylalanine and nitrated tyrosine products with a high-sensitivity and high-specificity HPLC-UV-FL technique. The analytes were extracted from serum via solid-phase extraction on Waters Oasis MCX cartridges. Chromatographic separation was achieved on an ODS column (Capcell Pak MG II; 150 × 2.0 mm) using a gradient mobile phase consisting of 20 mm sodium phosphate buffer (adjusted to pH 3.0) and acetonitrile. The method quantification limit for 4-nitrophenylalanine, m-tyrosine, and 3-nitrotyrosine was 0.1 μm. The relative standard deviation of the precision and accuracy was acceptable at the spiked concentration of 0.1 μm for 4-nitrophenylalanine, m-tyrosine and 3-nitrotyrosine. The method could be used for the in vitro analysis of serum samples.

  9. Phenylalanine ammonia lyase catalyzed synthesis of amino acids by an MIO-cofactor independent pathway.

    PubMed

    Lovelock, Sarah L; Lloyd, Richard C; Turner, Nicholas J

    2014-04-25

    Phenylalanine ammonia lyases (PALs) belong to a family of 4-methylideneimidazole-5-one (MIO) cofactor dependent enzymes which are responsible for the conversion of L-phenylalanine into trans-cinnamic acid in eukaryotic and prokaryotic organisms. Under conditions of high ammonia concentration, this deamination reaction is reversible and hence there is considerable interest in the development of PALs as biocatalysts for the enantioselective synthesis of non-natural amino acids. Herein the discovery of a previously unobserved competing MIO-independent reaction pathway, which proceeds in a non-stereoselective manner and results in the generation of both L- and D-phenylalanine derivatives, is described. The mechanism of the MIO-independent pathway is explored through isotopic-labeling studies and mutagenesis of key active-site residues. The results obtained are consistent with amino acid deamination occurring by a stepwise E1 cB elimination mechanism.

  10. Racemization of aspartic acid and phenylalanine in the sweetener aspartame at 100 degrees C.

    PubMed Central

    Boehm, M F; Bada, J L

    1984-01-01

    The racemization half-lives (i.e., the time required to reach a D/L = 0.33) at pH 6.8 for aspartic acid and phenylalanine in the sweetener aspartame (L-aspartyl-L-phenylalanine methyl ester) were determined to be 13 and 23 hours, respectively, at 100 degrees C. Racemization at this pH does not occur in aspartame but rather in its diketopiperazine decomposition product. Our results indicate that the use of aspartame to sweeten neutral pH foods and beverages that are then heated at elevated temperature could generate D-aspartic acid and D-phenylalanine. The nutritive consequences of these D-amino acids in the human diet are not well established, and thus aspartame should probably not be used as a sweetener when the exposure of neutral pH foods and beverages to elevated temperatures is required. At pH 4, a typical pH of most foods and beverages that might be sweetened with aspartame, the half-lives are 47 hours for aspartic acid and 1200 hours for phenylalanine at 100 degrees C. Racemization at pH 4 takes place in aspartame itself. Although the racemization rates at pH 4 are slow and no appreciable racemization of aspartic acid and phenylalanine should occur during the normal use of aspartame, some food and beverage components could conceivably act as catalysts. Additional studies are required to evaluate whether the use of aspartame as a sugar substitute might not in turn result in an increased human consumption of D-aspartic acid and D-phenylalanine. PMID:6591191

  11. Racemization of aspartic acid and phenylalanine in the sweetener aspartame at 100 degrees C.

    PubMed

    Boehm, M F; Bada, J L

    1984-08-01

    The racemization half-lives (i.e., the time required to reach a D/L = 0.33) at pH 6.8 for aspartic acid and phenylalanine in the sweetener aspartame (L-aspartyl-L-phenylalanine methyl ester) were determined to be 13 and 23 hours, respectively, at 100 degrees C. Racemization at this pH does not occur in aspartame but rather in its diketopiperazine decomposition product. Our results indicate that the use of aspartame to sweeten neutral pH foods and beverages that are then heated at elevated temperature could generate D-aspartic acid and D-phenylalanine. The nutritive consequences of these D-amino acids in the human diet are not well established, and thus aspartame should probably not be used as a sweetener when the exposure of neutral pH foods and beverages to elevated temperatures is required. At pH 4, a typical pH of most foods and beverages that might be sweetened with aspartame, the half-lives are 47 hours for aspartic acid and 1200 hours for phenylalanine at 100 degrees C. Racemization at pH 4 takes place in aspartame itself. Although the racemization rates at pH 4 are slow and no appreciable racemization of aspartic acid and phenylalanine should occur during the normal use of aspartame, some food and beverage components could conceivably act as catalysts. Additional studies are required to evaluate whether the use of aspartame as a sugar substitute might not in turn result in an increased human consumption of D-aspartic acid and D-phenylalanine.

  12. Phenylketonuria: protein content and amino acids profile of dishes for phenylketonuric patients. The relevance of phenylalanine.

    PubMed

    Pimentel, Filipa B; Alves, Rita C; Costa, Anabela S G; Torres, Duarte; Almeida, Manuela F; Oliveira, M Beatriz P P

    2014-04-15

    Phenylketonuria is an inborn error of metabolism, involving, in most cases, a deficient activity of phenylalanine hydroxylase. Neonatal diagnosis and a prompt special diet (low phenylalanine and natural-protein restricted diets) are essential to the treatment. The lack of data concerning phenylalanine contents of processed foodstuffs is an additional limitation for an already very restrictive diet. Our goals were to quantify protein (Kjeldahl method) and amino acid (18) content (HPLC/fluorescence) in 16 dishes specifically conceived for phenylketonuric patients, and compare the most relevant results with those of several international food composition databases. As might be expected, all the meals contained low protein levels (0.67-3.15 g/100 g) with the highest ones occurring in boiled rice and potatoes. These foods also contained the highest amounts of phenylalanine (158.51 and 62.65 mg/100 g, respectively). In contrast to the other amino acids, it was possible to predict phenylalanine content based on protein alone. Slight deviations were observed when comparing results with the different food composition databases.

  13. Inhibition of intestinal absorption of phenylalanine by phenylalaninol.

    PubMed

    Shimomura, K; Fukushima, T; Danno, T; Matsumoto, K; Miyoshi, M

    1975-08-01

    Plasma phenylalanine and tyrosine levels in rats which had been orally administered L-phenylalaninol and L-phenylalanine were determined. Since these amino acid levels in rats administered L-phenylalanine solution containing L-phenylalaninol were significantly lower than those in rats administered L-phenylalanine alone. L-phenylalaninol appears to inhibit the intestinal absorption of L-phenylalanine. This effect was more potent than that of cycloleucine. L-phenylalaninol inhibited the phenylalanine transport of everted sacs. The Km value of L-phenylalanine was 3.44 X 10(-3) M and the Ki value of L-phenylalaninol was 7.69 M 10(-3) M from Lineweaver-Burk plots. From these two curves, it appeared that L-phenylalaninol may competitively inhibit the intestinal transport of L-phenylalanine. The effects of L-phenylalanine, L-phenylalaninol and cycloleucine on the urinary excretions of Na+ and K+ in rats were also examined. Potassium excretion which increased on oral administration of L-phenylalanine, was suppressed by the administration of L-phenylalaninol but not administration of cycloleucine. L-phenylalaninol alone enhanced Na+ excretion in urine. These results confirmed that L-phenylalaninol shows inhibitory effects as potent as those of cycloleucine on the intestinal absorption of L-phenylalanine. PMID:1228171

  14. Performance of piglets in response to the standardized ileal digestible phenylalanine and tyrosine supply in low-protein diets.

    PubMed

    Gloaguen, M; Le Floc'h, N; Primot, Y; Corrent, E; van Milgen, J

    2014-09-01

    Reducing the CP level of the diet allows for a reduction in N excretion without limiting performance as long as the amino acid (AA) requirements are covered. The availability of crystalline AA has permitted for a considerable reduction in the CP level of diets, practically used in pig nutrition. The adoption of low CP diets and the extent to which the CP content can be reduced further depends on the knowledge of the minimum levels of indispensable AA that maximize growth. The standardized ileal digestible (SID) Phe : Lys and Tyr : Lys requirements and the possibility to substitute Tyr by Phe have never been studied in piglets. The objectives of this study were to estimate these requirements in 10 to 20 kg pig as well as to determine the extent to which Phe can be used to cover the Tyr requirement. In three dose-response studies, six pigs within each of 14 blocks were assigned to six low CP diets (14.5% CP) sub-limiting in Lys at 1.00% SID. In experiment 1, the SID Phe : Lys requirement estimate was assessed by supplementing a Phe-deficient diet with different levels of l-Phe to attain 33%, 39%, 46%, 52%, 58%, and 65% SID Phe : Lys. Because Phe can be used for Tyr synthesis, the diets provided a sufficient Tyr supply. A similar approach was used in experiment 2 with six levels of l-Tyr supplementation to attain 21%, 27%, 33%, 39%, 45% and 52% SID Tyr : Lys. Phenylalanine was supplied at a level sufficient to sustain maximum growth (estimated in experiment 1). The SID Phe : Lys and SID Tyr : Lys requirements for maximizing daily gain were 54% and 40% using a curvilinear-plateau model, respectively. A 10% deficiency in Phe and Tyr reduced daily gain by 3.0% and 0.7%, respectively. In experiment 3, the effect of the equimolar substitution of dietary SID Tyr by Phe to obtain 50%, 57%, and 64% SID Phe : (Phe+Tyr) was studied at two limiting levels of Phe+Tyr. From 57% to 64% SID Phe : (Phe+Tyr), performance was slightly reduced. In conclusion, it is recommended not to

  15. NON-PHYSIOLOGICAL AMINO ACID (NPAA) THERAPY TARGETING BRAIN PHENYLALANINE REDUCTION: PILOT STUDIES IN PAHENU2 MICE

    PubMed Central

    Vogel, Kara R.; Arning, Erland; Wasek, Brandi L.; Bottiglieri, Teodoro; Gibson, K. Michael

    2012-01-01

    Transport of large neutral amino acids (LNAA) across the blood brain barrier (BBB) is facilitated by the L-type amino acid transporter, LAT1. Peripheral accumulation of one LNAA (e.g., phenylalanine (phe) in PKU) is predicted to increase uptake of the offending amino acid to the detriment of others, resulting in disruption of brain amino acid homeostasis. We hypothesized that selected non-physiological amino acids (NPAAs) such as DL-norleucine (NL), 2-aminonorbornane (NB; 2-aminobicyclo-(2,1,1)-heptane-2-carboxylic acid), 2-aminoisobutyrate (AIB), and N-methyl-aminoisobutyrate (MAIB), acting as competitive inhibitors of various brain amino acid transporters, could reduce brain phe in Pahenu2 mice, a relevant murine model of PKU. Oral feeding of 5% NL, 5% AIB, 0.5% NB and 3% MAIB reduced brain phe by 56% (p<0.01), −1% (p=NS), 27% (p<0.05) and 14% (p<0.01), respectively, compared to untreated subjects. Significant effects on other LNAAs (tyrosine, methionine, branched chain amino acids) were also observed, however, with MAIB displaying the mildest effects. Of interest, MAIB represents an inhibitor of the system A (alanine) transporter that primarily traffics small amino acids and not LNAAs. Our studies represent the first in vivo use of these NPAAs in Pahenu2 mice, and provide proof-of-principle for their further preclinical development, with the long-term objective of identifying NPAA combinations and concentrations that selectively restrict brain phe transport while minimally impacting other LNAAs and downstream intermediates. PMID:22976763

  16. Non-physiological amino acid (NPAA) therapy targeting brain phenylalanine reduction: pilot studies in PAHENU2 mice.

    PubMed

    Vogel, Kara R; Arning, Erland; Wasek, Brandi L; Bottiglieri, Teodoro; Gibson, K Michael

    2013-05-01

    Transport of large neutral amino acids (LNAA) across the blood brain barrier (BBB) is facilitated by the L-type amino acid transporter, LAT1. Peripheral accumulation of one LNAA (e.g., phenylalanine (phe) in PKU) is predicted to increase uptake of the offending amino acid to the detriment of others, resulting in disruption of brain amino acid homeostasis. We hypothesized that selected non-physiological amino acids (NPAAs) such as DL-norleucine (NL), 2-aminonorbornane (NB; 2-aminobicyclo-(2,1,1)-heptane-2-carboxylic acid), 2-aminoisobutyrate (AIB), and N-methyl-aminoisobutyrate (MAIB), acting as competitive inhibitors of various brain amino acid transporters, could reduce brain phe in Pah (enu2) mice, a relevant murine model of PKU. Oral feeding of 5 % NL, 5 % AIB, 0.5 % NB and 3 % MAIB reduced brain phe by 56 % (p < 0.01), -1 % (p = NS), 27 % (p < 0.05) and 14 % (p < 0.01), respectively, compared to untreated subjects. Significant effects on other LNAAs (tyrosine, methionine, branched chain amino acids) were also observed, however, with MAIB displaying the mildest effects. Of interest, MAIB represents an inhibitor of the system A (alanine) transporter that primarily traffics small amino acids and not LNAAs. Our studies represent the first in vivo use of these NPAAs in Pah (enu2) mice, and provide proof-of-principle for their further preclinical development, with the long-term objective of identifying NPAA combinations and concentrations that selectively restrict brain phe transport while minimally impacting other LNAAs and downstream intermediates.

  17. Between peptides and bile acids: self-assembly of phenylalanine substituted cholic acids.

    PubMed

    Travaglini, Leana; D'Annibale, Andrea; di Gregorio, Maria Chiara; Schillén, Karin; Olsson, Ulf; Sennato, Simona; Pavel, Nicolae V; Galantini, Luciano

    2013-08-01

    Biocompatible molecules that undergo self-assembly are of high importance in biological and medical applications of nanoscience. Peptides and bile acids are among the most investigated due to their ability to self-organize into many different, often stimuli-sensitive, supramolecular structures. With the aim of preparing molecules mixing the aggregation properties of bile acid and amino acid-based molecules, we report on the synthesis and self-association behavior of two diastereomers obtained by substituting a hydroxyl group of cholic acid with a l-phenylalanine residue. The obtained molecules are amphoteric, and we demonstrate that they show a pH-dependent self-assembly. Both molecules aggregate in globular micelles at high pH, whereas they form tubular superstructures under acid conditions. Unusual narrow nanotubes with outer and inner cross-section diameters of about 6 and 3 nm are formed by the derivatives. The diasteroisomer with α orientation of the substituent forms in addition a wider tubule (17 nm cross-section diameter). The ability to pack in supramolecular tubules is explained in terms of a wedge-shaped bola-form structure of the derivatives. Parallel or antiparallel face-to-face dimers are hypothesized as fundamental building blocks for the formation of the narrow and wide nanotubes, respectively.

  18. Activation of phenylalanine hydroxylase by phenylalanine does not require binding in the active site.

    PubMed

    Roberts, Kenneth M; Khan, Crystal A; Hinck, Cynthia S; Fitzpatrick, Paul F

    2014-12-16

    Phenylalanine hydroxylase (PheH), a liver enzyme that catalyzes the hydroxylation of excess phenylalanine in the diet to tyrosine, is activated by phenylalanine. The lack of activity at low levels of phenylalanine has been attributed to the N-terminus of the protein's regulatory domain acting as an inhibitory peptide by blocking substrate access to the active site. The location of the site at which phenylalanine binds to activate the enzyme is unknown, and both the active site in the catalytic domain and a separate site in the N-terminal regulatory domain have been proposed. Binding of catecholamines to the active-site iron was used to probe the accessibility of the active site. Removal of the regulatory domain increases the rate constants for association of several catecholamines with the wild-type enzyme by ∼2-fold. Binding of phenylalanine in the active site is effectively abolished by mutating the active-site residue Arg270 to lysine. The k(cat)/K(phe) value is down 10⁴ for the mutant enzyme, and the K(m) value for phenylalanine for the mutant enzyme is >0.5 M. Incubation of the R270K enzyme with phenylalanine also results in a 2-fold increase in the rate constants for catecholamine binding. The change in the tryptophan fluorescence emission spectrum seen in the wild-type enzyme upon activation by phenylalanine is also seen with the R270K mutant enzyme in the presence of phenylalanine. Both results establish that activation of PheH by phenylalanine does not require binding of the amino acid in the active site. This is consistent with a separate allosteric site, likely in the regulatory domain.

  19. Phosphorylation of cystic fibrosis transmembrane conductance regulator (CFTR) serine-511 by the combined action of tyrosine kinases and CK2: the implication of tyrosine-512 and phenylalanine-508.

    PubMed

    Cesaro, Luca; Marin, Oriano; Venerando, Andrea; Donella-Deana, Arianna; Pinna, Lorenzo A

    2013-12-01

    The cystic fibrosis transmembrane conductance regulator (CFTR) harbors, close to Phe-508, whose deletion is the commonest cause of cystic fibrosis, a conserved potential CK2 phospho-acceptor site (Ser511), which however is not susceptible to phosphorylation by CK2. To shed light on this apparent paradox, a series of systematically substituted peptides encompassing Ser511 were assayed for their ability to be phosphorylated. The main outcomes of our study are the following: (a) Tyr512 plays a prominent role as a negative determinant as its replacement by Ala restores Ser511 phosphorylation by CK2; (b) an even more pronounced phosphorylation of Ser511 is promoted if Tyr512 is replaced by phospho-tyrosine instead of alanine; (c) Tyr512 and, to a lesser extent, Tyr515 are readily phosphorylated by Lyn, a protein tyrosine kinase of the Src family, in a manner which is enhanced by the concomitant Phe508 deletion. Collectively taken, our data, in conjunction with the notion that Tyr515 is phosphorylated in vivo, disclose the possibility that CFTR Ser511 can be phosphorylated by the combined action of tyrosine kinases and CK2 and disclose a new mechanism of hierarchical phosphorylation where the role of the priming kinase is that of removing negative determinant(s).

  20. The Amino Acid Specificity for Activation of Phenylalanine Hydroxylase Matches the Specificity for Stabilization of Regulatory Domain Dimers

    PubMed Central

    2016-01-01

    Liver phenylalanine hydroxylase is allosterically activated by phenylalanine. The structural changes that accompany activation have not been identified, but recent studies of the effects of phenylalanine on the isolated regulatory domain of the enzyme support a model in which phenylalanine binding promotes regulatory domain dimerization. Such a model predicts that compounds that stabilize the regulatory domain dimer will also activate the enzyme. Nuclear magnetic resonance spectroscopy and analytical ultracentrifugation were used to determine the ability of different amino acids and phenylalanine analogues to stabilize the regulatory domain dimer. The abilities of these compounds to activate the enzyme were analyzed by measuring their effects on the fluorescence change that accompanies activation and on the activity directly. At concentrations of 10–50 mM, d-phenylalanine, l-methionine, l-norleucine, and (S)-2-amino-3-phenyl-1-propanol were able to activate the enzyme to the same extent as 1 mM l-phenylalanine. Lower levels of activation were seen with l-4-aminophenylalanine, l-leucine, l-isoleucine, and 3-phenylpropionate. The ability of these compounds to stabilize the regulatory domain dimer agreed with their ability to activate the enzyme. These results support a model in which allosteric activation of phenylalanine hydroxylase is linked to dimerization of regulatory domains. PMID:26252467

  1. The Amino Acid Specificity for Activation of Phenylalanine Hydroxylase Matches the Specificity for Stabilization of Regulatory Domain Dimers.

    PubMed

    Zhang, Shengnan; Hinck, Andrew P; Fitzpatrick, Paul F

    2015-08-25

    Liver phenylalanine hydroxylase is allosterically activated by phenylalanine. The structural changes that accompany activation have not been identified, but recent studies of the effects of phenylalanine on the isolated regulatory domain of the enzyme support a model in which phenylalanine binding promotes regulatory domain dimerization. Such a model predicts that compounds that stabilize the regulatory domain dimer will also activate the enzyme. Nuclear magnetic resonance spectroscopy and analytical ultracentrifugation were used to determine the ability of different amino acids and phenylalanine analogues to stabilize the regulatory domain dimer. The abilities of these compounds to activate the enzyme were analyzed by measuring their effects on the fluorescence change that accompanies activation and on the activity directly. At concentrations of 10-50 mM, d-phenylalanine, l-methionine, l-norleucine, and (S)-2-amino-3-phenyl-1-propanol were able to activate the enzyme to the same extent as 1 mM l-phenylalanine. Lower levels of activation were seen with l-4-aminophenylalanine, l-leucine, l-isoleucine, and 3-phenylpropionate. The ability of these compounds to stabilize the regulatory domain dimer agreed with their ability to activate the enzyme. These results support a model in which allosteric activation of phenylalanine hydroxylase is linked to dimerization of regulatory domains.

  2. Enzymatic synthesis of enantiopure alpha- and beta-amino acids by phenylalanine aminomutase-catalysed amination of cinnamic acid derivatives.

    PubMed

    Wu, Bian; Szymanski, Wiktor; Wietzes, Piet; de Wildeman, Stefaan; Poelarends, Gerrit J; Feringa, Ben L; Janssen, Dick B

    2009-01-26

    The phenylalanine aminomutase (PAM) from Taxus chinensis catalyses the conversion of alpha-phenylalanine to beta-phenylalanine, an important step in the biosynthesis of the N-benzoyl phenylisoserinoyl side-chain of the anticancer drug taxol. Mechanistic studies on PAM have suggested that (E)-cinnamic acid is an intermediate in the mutase reaction and that it can be released from the enzyme's active site. Here we describe a novel synthetic strategy that is based on the finding that ring-substituted (E)-cinnamic acids can serve as a substrate in PAM-catalysed ammonia addition reactions for the biocatalytic production of several important beta-amino acids. The enzyme has a broad substrate range and a high enantioselectivity with cinnamic acid derivatives; this allows the synthesis of several non-natural aromatic alpha- and beta-amino acids in excellent enantiomeric excess (ee >99 %). The internal 5-methylene-3,5-dihydroimidazol-4-one (MIO) cofactor is essential for the PAM-catalysed amination reactions. The regioselectivity of amination reactions was influenced by the nature of the ring substituent.

  3. Structures of D-amino-acid amidase complexed with L-phenylalanine and with L-phenylalanine amide: insight into the D-stereospecificity of D-amino-acid amidase from Ochrobactrum anthropi SV3.

    PubMed

    Okazaki, Seiji; Suzuki, Atsuo; Mizushima, Tsunehiro; Komeda, Hidenobu; Asano, Yasuhisa; Yamane, Takashi

    2008-03-01

    The crystal structures of D-amino-acid amidase (DAA) from Ochrobactrum anthropi SV3 in complex with L-phenylalanine and with L-phenylalanine amide were determined at 2.3 and 2.2 A resolution, respectively. Comparison of the L-phenylalanine amide complex with the D-phenylalanine complex reveals that the D-stereospecificity of DAA might be achieved as a consequence of three structural factors: (i) the hydrophobic cavity in the region in which the hydrophobic side chain of the substrate is held, (ii) the spatial arrangement of Gln310 O and Glu114 O epsilon2 that fixes the amino N atom of the substrate and (iii) the existence of two cavities that keep the carboxyl/amide group of the substrate near or apart from Ser60 O gamma.

  4. Thermodynamic characteristics of the interaction between nicotinic acid and phenylalanine in an aqueous buffer solution at 298 K

    NASA Astrophysics Data System (ADS)

    Badelin, V. G.; Tyunina, E. Yu.; Mezhevoi, I. N.; Tarasova, G. N.

    2013-08-01

    The interaction between L-phenylalanine and nicotinic acid is studied by solution calorimetry in an aqueous buffer solution (pH 7.35) at different ratios of the reagents. Experimental data on the enthalpy of dissolution of amino acid in the buffer solution of nicotinic acid at 298.15 K are calculated. The values of thermodynamic parameters for the complexation of L-phenylalanine with nicotinic acid are calculated. It is shown that the formation of a 1: 2 molecular complex is stabilized by the entropy factor due to the dominant role of the dehydration effect of initial reagents.

  5. Roles of the tyrosine isomers meta-tyrosine and ortho-tyrosine in oxidative stress.

    PubMed

    Ipson, Brett R; Fisher, Alfred L

    2016-05-01

    The damage to cellular components by reactive oxygen species, termed oxidative stress, both increases with age and likely contributes to age-related diseases including Alzheimer's disease, atherosclerosis, diabetes, and cataract formation. In the setting of oxidative stress, hydroxyl radicals can oxidize the benzyl ring of the amino acid phenylalanine, which then produces the abnormal tyrosine isomers meta-tyrosine or ortho-tyrosine. While elevations in m-tyrosine and o-tyrosine concentrations have been used as a biological marker of oxidative stress, there is emerging evidence from bacterial, plant, and mammalian studies demonstrating that these isomers, particularly m-tyrosine, directly produce adverse effects to cells and tissues. These new findings suggest that the abnormal tyrosine isomers could in fact represent mediators of the effects of oxidative stress. Consequently the accumulation of m- and o-tyrosine may disrupt cellular homeostasis and contribute to disease pathogenesis, and as result, effective defenses against oxidative stress can encompass not only the elimination of reactive oxygen species but also the metabolism and ultimately the removal of the abnormal tyrosine isomers from the cellular amino acid pool. Future research in this area is needed to clarify the biologic mechanisms by which the tyrosine isomers damage cells and disrupt the function of tissues and organs and to identify the metabolic pathways involved in removing the accumulated isomers after exposure to oxidative stress.

  6. Self-assembled phenylalanine-α,β-dehydrophenylalanine nanotubes for sustained intravitreal delivery of a multi-targeted tyrosine kinase inhibitor.

    PubMed

    Panda, Jiban J; Yandrapu, Sarath; Kadam, Rajendra S; Chauhan, Virander S; Kompella, Uday B

    2013-12-28

    Current standard of care for sustained back of the eye drug delivery is surgical placement or injection of large, slow release implants using a relatively large 22 gauge needle. We designed novel dipeptide (phenylalanine-α,β-dehydrophenylalanine; Phe-∆Phe) based nanotubes with a diameter of ~15-30 nm and a length of ~1500 nm that could be injected with a 33 gauge needle for sustained intravitreal delivery of pazopanib, a multi-targeted tyrosine kinase inhibitor. The drug could be loaded during nanotube assembly or post-loaded after nanotube formation, with the former being more efficient at 25% w/w pazopanib loading and ~55% loading efficiency. Plain and peptide loaded nanotube were non-cytotoxic to retinal pigment epithelial cells even at a concentration of 200 μg/ml. Following intravitreal injection of fluorescently labeled nanotubes using a 33 gauge needle in a rat model, the nanotube persistence and drug delivery were monitored using noninvasive fluorophotometry, electron microscopy and mass spectrometry analysis. Nanotubes persisted in the vitreous humor during the 15 days study and pazopanib levels in the vitreous humor, retina, and choroid-RPE at the end of the study were 4.5, 5, and 2.5-folds higher, respectively, compared to the plain drug. Thus, Phe-∆Phe nanotubes allow intravitreal injections with a small gauge needle and sustain drug delivery.

  7. Reduced phenylalanine ammonia-lyase and tyrosine ammonia-lyase activities and lignin synthesis in wheat grown under low pressure sodium lamps

    NASA Technical Reports Server (NTRS)

    Guerra, D.; Anderson, A. J.; Salisbury, F. B.

    1985-01-01

    Wheat (Triticum aestivum L. cv Fremont) grown in hydroponic culture under 24-hour continuous irradiation at 560 to 580 micromoles per square meter per second from either metalhalide (MH), high pressure sodium (HPS), or low pressure sodium (LPS) lamps reached maturity in 70 days. Grain yields were similar under all three lamps, although LPS-grown plants lodged at maturity. Phenylalanine ammonia-lyase (PAL) and a tyrosine ammonia lyase (TAL) with lesser activity were detected in all extracts of leaf, inflorescence, and stem. Ammonia-lyase activities increased with age of the plant, and plants grown under the LPS lamp displayed PAL and TAL activities lower than wheat cultured under MH and HPS radiation. Greenhouse solar-grown wheat had the highest PAL and TAL activities. Lignin content of LPS-grown wheat was also significantly reduced from that of plants grown under MH or HPS lamps or in the greenhouse, showing a correlation with the reduced PAL and TAL activities. Ratios of far red-absorbing phytochrome to total phytochrome were similar for all three lamps, but the data do not yet warrant a conclusion about specific wavelengths missing from the LPS lamps that might have induced PAL and TAL activities in plants under the other lamps.

  8. Non-ionic surfactant modified ligand exchange chromatography using copper (II) complex of N,N-dimethyl-L-phenylalanine as the chiral additive for enantioselective amino acids separation.

    PubMed

    Dimitrova, Pepa; Bart, Hans-Jörg

    2010-03-17

    The influence of non-ionic surfactants on the selectivity and retention in the ligand exchange chromatography for the enantioselective separation of racemic mixtures of the amino acids dl-methionine, dl-leucine, dl-valine and dl-tyrosine applying chiral mobile phases was investigated, whereas five different surfactants were tested as modifiers. The experiments were carried out using a commercially available non-chiral RP-C8 column and the copper (II) complex of N,N-dimethyl-l-phenylalanine as the chiral additive. Varying the surfactant concentrations the retention factors and the selectivity could be controlled and in general no negative influence on the separation (due to surfactant adsorption on the non-chiral stationary phase) occurred. Changing the temperature the van't Hoff plots were obtained and the thermodynamic parameters calculated. Temperature had influence on the selectivity for each surfactant and lowered the retention times as expected.

  9. A study of the antibacterial activity of L-phenylalanine and L-tyrosine esters in relation to their CMCs and their interactions with 1,2-dipalmitoyl-sn-glycero-3-phosphocholine, DPPC as model membrane.

    PubMed

    Joondan, Nausheen; Jhaumeer-Laulloo, Sabina; Caumul, Prakashanand

    2014-01-01

    Cationic amino acid-based surfactants are known to interact with the lipid bilayer of cell membranes resulting in depolarization, lysis and cell death through a disruption of the membrane topology. A range of cationic surfactant analogues derived from L-Phenylalanine (C1-C20) and L-Tyrosine (C8-C14) esters have been synthesized and screened for their antibacterial activity. The esters were more active against gram positive than gram negative bacteria. The activity increased with increasing chain length, exhibiting a cut-off effect at C12 for gram positive and C8/C10 for gram negative bacteria. The cut-off effect for gram negative bacteria was observed at a lower alkyl chain length. The CMC was correlated with the MIC, inferring that micellar activity contribute to the cut-off effect in antibacterial activity. The interaction of the cationic surfactants with the phospholipid vesicles (1,2-dipalmitoyl-sn-glycero-3-phosphocholine, DPPC) in the presence of 1-anilino-8-naphthalene sulfonate (ANS) and 1,6-diphenyl-1,3,5-hexatriene (DPH) as fluorescence probes showed that an increase in ionic interaction causes an increase in antibacterial activity. Increase in hydrophobic interaction increases the antibacterial activity only to a certain chain length, attributing to the cut-off effect. Therefore, both electrostatic and hydrophobic interactions, involving the polar and nonpolar moieties are of paramount importance for the bactericidal properties.

  10. Phenylalanine sensitive K562-D cells for the analysis of the biochemical impact of excess amino acid

    PubMed Central

    Sanayama, Yoshitami; Matsumoto, Akio; Shimojo, Naoki; Kohno, Yoichi; Nakaya, Haruaki

    2014-01-01

    Although it is recognized that the abnormal accumulation of amino acid is a cause of the symptoms in metabolic disease such as phenylketonuria (PKU), the relationship between disease severity and serum amino acid levels is not well understood due to the lack of experimental model. Here, we present a novel in vitro cellular model using K562-D cells that proliferate slowly in the presence of excessive amount of phenylalanine within the clinically observed range, but not phenylpyruvate. The increased expression of the L-type amino acid transporter (LAT2) and its adapter protein 4F2 heavy chain appeared to be responsible for the higher sensitivity to phenylalanine in K562-D cells. Supplementation with valine over phenylalanine effectively restored cell proliferation, although other amino acids did not improve K562-D cell proliferation over phenylalanine. Biochemical analysis revealed mammalian target of rapamycin complex (mTORC) as a terminal target of phenylalanine in K562-D cell proliferation, and supplementation of valine restored mTORC1 activity. Our results show that K562-D cell can be a potent tool for the investigation of PKU at the molecular level and to explore new therapeutic approaches to the disease. PMID:25373594

  11. Phenylalanine sensitive K562-D cells for the analysis of the biochemical impact of excess amino acid.

    PubMed

    Sanayama, Yoshitami; Matsumoto, Akio; Shimojo, Naoki; Kohno, Yoichi; Nakaya, Haruaki

    2014-11-06

    Although it is recognized that the abnormal accumulation of amino acid is a cause of the symptoms in metabolic disease such as phenylketonuria (PKU), the relationship between disease severity and serum amino acid levels is not well understood due to the lack of experimental model. Here, we present a novel in vitro cellular model using K562-D cells that proliferate slowly in the presence of excessive amount of phenylalanine within the clinically observed range, but not phenylpyruvate. The increased expression of the L-type amino acid transporter (LAT2) and its adapter protein 4F2 heavy chain appeared to be responsible for the higher sensitivity to phenylalanine in K562-D cells. Supplementation with valine over phenylalanine effectively restored cell proliferation, although other amino acids did not improve K562-D cell proliferation over phenylalanine. Biochemical analysis revealed mammalian target of rapamycin complex (mTORC) as a terminal target of phenylalanine in K562-D cell proliferation, and supplementation of valine restored mTORC1 activity. Our results show that K562-D cell can be a potent tool for the investigation of PKU at the molecular level and to explore new therapeutic approaches to the disease.

  12. Eliminating tyrosine sequence variants in CHO cell lines producing recombinant monoclonal antibodies.

    PubMed

    Feeney, Lauren; Carvalhal, Veronica; Yu, X Christopher; Chan, Betty; Michels, David A; Wang, Yajun Jennifer; Shen, Amy; Ressl, Jan; Dusel, Brendon; Laird, Michael W

    2013-04-01

    Amino acid sequence variants are defined as unintended amino acid sequence changes that contribute to product variation with potential impact to product safety, immunogenicity, and efficacy. Therefore, it is important to understand the propensity for sequence variant (SV) formation during the production of recombinant proteins for therapeutic use. During the development of clinical therapeutic products, several monoclonal antibodies (mAbs) produced from Chinese Hamster Ovary (CHO) cells exhibited SVs at low levels (≤3%) in multiple locations throughout the mAbs. In these examples, the cell culture process depleted tyrosine, and the tyrosine residues in the recombinant mAbs were replaced with phenylalanine or histidine. In this work, it is demonstrated that tyrosine supplementation eliminated the tyrosine SVs, while early tyrosine starvation significantly increased the SV level in all mAbs tested. Additionally, it was determined that phenylalanine is the amino acid preferentially misincorporated in the absence of tyrosine over histidine, with no other amino acid misincorporated in the absence of tyrosine, phenylalanine, and histidine. The data support that the tyrosine SVs are due to mistranslation and not DNA mutation, most likely due to tRNA(Tyr) mischarging due to the structural similarities between tyrosine and phenylalanine.

  13. Conformational preferences of 1-amino-2-phenylcyclohexanecarboxylic acid, a phenylalanine cyclohexane analogue.

    PubMed

    Alemán, Carlos; Jiménez, Ana I; Cativiela, Carlos; Nussinov, Ruth; Casanovas, Jordi

    2009-10-16

    The intrinsic conformational preferences of the restricted phenylalanine analogue generated by including the alpha and beta carbon atoms into a cyclohexane ring (1-amino-2-phenylcyclohexanecarboxylic acid, c(6)Phe) have been determined using quantum mechanical calculations. Specifically, the conformational profile of the N-acetyl-N'-methylamide derivative of the c(6)Phe stereoisomers exhibiting either a cis or a trans relative orientation between the amino and phenyl substituents has been analyzed in different environments (gas phase, chloroform, and aqueous solutions). Calculations were performed using B3LYP, MP2, and HF methods combined with the 6-31+G(d,p) and 6-311++G(d,p) basis sets, and a self-consistent reaction-field (SCRF) method was applied to analyze the influence of the solvent. The amino acids investigated can be viewed as constrained phenylalanine analogues with a rigidly oriented aromatic side chain that may interact with the peptide backbone not only sterically but also electronically through the aromatic pi orbitals. Their conformational propensities have been found to be strongly influenced by the specific orientation of the aromatic substituent in each stereoisomer and the conformation adopted by the cyclohexane ring, as well as by the environment.

  14. Phenylalanine induces Burkholderia cenocepacia phenylacetic acid catabolism through degradation to phenylacetyl-CoA in synthetic cystic fibrosis sputum medium.

    PubMed

    Yudistira, Harry; McClarty, Leigh; Bloodworth, Ruhi A M; Hammond, Sydney A; Butcher, Haley; Mark, Brian L; Cardona, Silvia T

    2011-09-01

    Synthetic cystic fibrosis sputum medium (SCFM) is rich in amino acids and supports robust growth of Burkholderia cenocepacia, a member of the Burkholderia cepacia complex (Bcc). Previous work demonstrated that B. cenocepacia phenylacetic acid (PA) catabolic genes are up-regulated during growth in SCFM and are required for full virulence in a Caenorhabditis elegans host model. In this work, we investigated the role of phenylalanine, one of the aromatic amino acids present in SCFM, as an inducer of the PA catabolic pathway. Phenylalanine degradation intermediates were used as sole carbon sources for growth and gene reporter experiments. In addition to phenylalanine and PA, phenylethylamine, phenylpyruvate, and 2-phenylacetamide were usable as sole carbon sources by wild type B. cenocepacia K56-2, but not by a PA catabolism-defective mutant. EMSA analysis showed that the binding of PaaR, the negative regulator protein of B. cenocepacia PA catabolism, to PA regulatory DNA could only be relieved by phenylacetyl-Coenzyme A (PA-CoA), but not by any of the putative phenylalanine degradation intermediates. Taken together, our results show that in B. cenocepacia, phenylalanine is catabolized to PA and induces PA catabolism through PA activation to PA-CoA. Thus, PaaR shares the same inducer with PaaX, the regulator of PA catabolism in Escherichia coli, despite belonging to a different protein family.

  15. D- and L-[123I]-2-I-phenylalanine show a long tumour retention compared with D- and L-[123I]-2-I-tyrosine in R1M rhabdomyosarcoma tumour-bearing Wag/Rij rats.

    PubMed

    Bauwens, Matthias; Lahoutte, Tony; Kersemans, Ken; Caveliers, Vicky; Bossuyt, Axel; Mertens, John

    2007-07-01

    The aim of this study was the comparison of the tumour uptake and the long-term retention of [(123)I]-2-I-L-phenylalanine and [(123)I]-2-I-D-phenylalanine with those of [(123)I]-2-I-L-tyrosine and [(123)I]-2-I-D-tyrosine in R1M rhabdomyosarcoma tumour-bearing rats. The biodistribution of the radioactivity as a function of time in R1M tumour-bearing rats was measured by planar gamma camera imaging (dynamic and static). If dissection was applied, the activity in the tumours and tissues of interest was measured by gamma counting. [(123)I]-2-iodo-L-phenylalanine, [(123)I]-2-iodo-D-phenylalaine, [(123)I]-2-I-L-tyrosine showed a considerable tumour uptake reaching a maximum between 10 and 30 min. At 30 min p.i. the differential uptake ratio values of this uptake were, respectively, 2.1, 2.3, 2.5 and 1.7. The activity in the tumour was shown to be related to a tumour cell uptake and not to an increased blood pool activity. All the tracers showed a clearance from the blood to the bladder without renal retention. At longer times both L- and D- [(123)I]-2-I-tyrosine were cleared for a large part from the tumours and the body. [(123)I]-2-I-L-Phe and [(123)I]-2-I-D-Phe showed a considerable and equal retention in the tumours: as compared with 0.5 h, 91% at 24 h and 80% at 48 h. This was related to the longer retention of activity in the blood pool noticed for these compounds (81% at 24 h and 65% at 48 h). The tumour-to-background ratio increased with 25% at those longer times. At short times all the tracers were taken up to a considerable extent in the tumours. In the R1M-bearing Wag/Rij rat model only [(123)I]-2-I-L-phenylalanine and [(123)I]-2-I-D-phenylalanine showed an especially high retention at long times without any significant difference between the enantiomers. PMID:17683118

  16. Inhibition of the Hematopoietic Protein Tyrosine Phosphatase by Phenoxyacetic Acids.

    PubMed

    Bobkova, Ekaterina V; Liu, Wallace H; Colayco, Sharon; Rascon, Justin; Vasile, Stefan; Gasior, Carlton; Critton, David A; Chan, Xochella; Dahl, Russell; Su, Ying; Sergienko, Eduard; Chung, Thomas D Y; Mustelin, Tomas; Page, Rebecca; Tautz, Lutz

    2011-02-01

    Protein tyrosine phosphatases (PTPs) have only recently become the focus of attention in the search for novel drug targets despite the fact that they play vital roles in numerous cellular processes and are implicated in many human diseases. The hematopoietic protein tyrosine phosphatase (HePTP) is often found dysregulated in preleukemic myelodysplastic syndrome (MDS), as well as in acute myelogenous leukemia (AML). Physiological substrates of HePTP include the mitogen-activated protein kinases (MAPKs) ERK1/2 and p38. Specific modulators of HePTP catalytic activity will be useful for elucidating mechanisms of MAPK regulation in hematopietic cells, and may also provide treatments for hematopoietic malignancies such as AML. Here we report the discovery of phenoxyacetic acids as inhibitors of HePTP. Structure-activity relationship (SAR) analysis and in silico docking studies reveal the molecular basis of HePTP inhibition by these compounds. We also show that these compounds are able to penetrate cell membranes and inhibit HePTP in human T lymphocytes.

  17. Effect of aspartame and protein, administered in phenylalanine-equivalent doses, on plasma neutral amino acids, aspartate, insulin and glucose in man.

    PubMed

    Møller, S E

    1991-05-01

    Six human males each received 0.56 g phenylalanine (Phe) in the form of 1.0 g aspartame or 12.2 g bovine albumin in 200 ml water or water alone. Venous blood samples collected before consumption and during the following 4 hr were assayed for plasma levels of large, neutral amino acids (LNAA), aspartate, insulin and glucose. The area under the curve for plasma Phe was 40% greater, although not significant, after aspartame compared with albumin intake. The indicated increased clearance rate of plasma Phe after albumin may be caused by the significant increase of insulin, on which aspartame had no effect. There was a significant main effect of aspartame for plasma tyrosine but not for tryptophan, valine, isoleucine or leucine. Plasma aspartate was significantly increased at 0.25 hr after the aspartame intake. The percentage Phe/LNAA decreased slightly in response to albumin but increased 55% after aspartame and remained significantly increased for 2 hr. Tyrosine/LNAA increased and tryptophan/LNAA decreased modestly after aspartame intake. The study showed that the intake of aspartame in a not unrealistically high dose produced a marked and persistent increase of the availability of Phe to the brain, which was not observed after protein intake. The study indicated, furthermore, that Phe was cleared faster from the plasma after consumption of protein compared with aspartame.

  18. Phenylalanine-containing cyclic dipeptides--the lowest molecular weight hydrogelators based on unmodified proteinogenic amino acids.

    PubMed

    Kleinsmann, Alexander J; Nachtsheim, Boris J

    2013-09-14

    Cyclic dipeptides (diketopiperazines - DKPs) that are based on the proteinogenic amino acid phenylalanine in combination with serine, cysteine, glutamate, histidine and lysine are described as simple and remarkable low molecular weight hydrogelators. Blends of selected DKPs show remarkable pH-dependent properties and can be applied as easy to tune materials in drug delivery.

  19. Phenylketonuria: brain phenylalanine concentrations relate inversely to cerebral protein synthesis.

    PubMed

    de Groot, Martijn J; Sijens, Paul E; Reijngoud, Dirk-Jan; Paans, Anne M; van Spronsen, Francjan J

    2015-02-01

    In phenylketonuria, elevated plasma phenylalanine concentrations may disturb blood-to-brain large neutral amino acid (LNAA) transport and cerebral protein synthesis (CPS). We investigated the associations between these processes, using data obtained by positron emission tomography with l-[1-(11)C]-tyrosine ((11)C-Tyr) as a tracer. Blood-to-brain transport of non-Phe LNAAs was modeled by the rate constant for (11)C-Tyr transport from arterial plasma to brain tissue (K1), while CPS was modeled by the rate constant for (11)C-Tyr incorporation into cerebral protein (k3). Brain phenylalanine concentrations were measured by magnetic resonance spectroscopy in three volumes of interest (VOIs): supraventricular brain tissue (VOI 1), ventricular brain tissue (VOI 2), and fluid-containing ventricular voxels (VOI 3). The associations between k3 and each predictor variable were analyzed by multiple linear regression. The rate constant k3 was inversely associated with brain phenylalanine concentrations in VOIs 2 and 3 (adjusted R(2)=0.826, F=19.936, P=0.021). Since brain phenylalanine concentrations in these VOIs highly correlated with each other, the specific associations of each predictor with k3 could not be determined. The associations between k3 and plasma phenylalanine concentration, K1, and brain phenylalanine concentrations in VOI 1 were nonsignificant. In conclusion, our study shows an inverse association between k3 and increased brain phenylalanine concentrations.

  20. Phenylalanine kinetics differ between formula-fed and human milk-fed preterm infants.

    PubMed

    Darling, Pauline B; Dunn, Michael; Gilani, G Sarwar; Ball, Ronald O; Pencharz, Paul B

    2004-10-01

    Infants fed casein-dominant formulas have higher plasma phenylalanine and tyrosine concentrations than those fed mother's milk. Conversely, elevated plasma threonine concentrations are observed in infants fed whey-dominant formulas. We recently showed that formula-fed preterm infants have a lower capacity to degrade threonine than do preterm infants fed mother's milk. We hypothesized that these same infants (n = 18) would differ in their catabolism of phenylalanine in response to phenylalanine loads provided by formulas with increasing casein content of formulas (whey:casein 60:40, 40:60, and 20:80) compared with preterm infants fed mother's milk. Plasma phenylalanine concentrations significantly rose (49, 46, 79 micromol . L(-1) for whey:casein 60:40, 40:60, and 20:80, respectively, pooled SD 8, P < 0.05); and plasma phenylalanine concentrations in infants fed mother's milk were low (40 +/- 4 micromol . L(-1)). Using [1-(13)C]phenylalanine tracer and (13)CO(2) production in breath we found that although there was a significant positive relation between phenylalanine oxidation and phenylalanine intake in formula-fed infants (r(2) = 0.43, P = 0.03), these infants were not able to increase their oxidation of phenylalanine enough to prevent a significant rise in plasma phenylalanine when fed the 20:80 formula. Compared to infants fed mother's milk, formula-fed infants had significantly lower phenylalanine oxidation (39.1 vs. 30.7% of phenylalanine intake, respectively, P < 0.05). We conclude that one of the mechanisms for the differences in plasma amino acid concentration between formula-fed and mother's milk-fed preterm infants may be in vivo down-regulated catabolism of 2 important essential amino acids (phenylalanine in addition to threonine) in formula-fed preterm infants.

  1. Food products made with glycomacropeptide, a low-phenylalanine whey protein, provide a new alternative to amino Acid-based medical foods for nutrition management of phenylketonuria.

    PubMed

    van Calcar, Sandra C; Ney, Denise M

    2012-08-01

    Phenylketonuria (PKU), an inborn error in phenylalanine metabolism, requires lifelong nutrition management with a low-phenylalanine diet, which includes a phenylalanine-free amino acid-based medical formula to provide the majority of an individual's protein needs. Compliance with this diet is often difficult for older children, adolescents, and adults with PKU. The whey protein glycomacropeptide (GMP) is ideally suited for the PKU diet because it is naturally low in phenylalanine. Nutritionally complete, acceptable medical foods and beverages can be made with GMP to increase the variety of protein sources for the PKU diet. As an intact protein, GMP improves protein use and increases satiety compared with amino acids. Thus, GMP provides a new, more physiologic source of low-phenylalanine dietary protein for people with PKU.

  2. Bioavailability of phenylalanine and aspartate from aspartame (20 mg/kg) in capsules and solution.

    PubMed

    Burns, T S; Stargel, W W; Hurwitz, A

    1990-11-01

    Aspartame (L-aspartyl-L-phenylalanine methyl ester) was given in capsules or solution to compare the bioavailability of its constituent amino acids, aspartate and phenylalanine. Twenty healthy subjects received a single 20 mg/kg dose of aspartame in capsules or solution in a randomized, crossover design. Plasma amino acid concentrations and the phenylalanine to large neutral amino acid ratios (Phe/LNAA) were determined. Plasma aspartate concentrations did not increase with either treatment. For plasma phenylalanine following capsule ingestion, there was a smaller peak plasma concentration (Cmax; 103.3 v 126.6 mumol/L), a longer time to peak concentration (tmax; 108.6 v 36.6 minutes), but no significant difference in the area under the plasma concentration-time curve (AUC) (7,656 v 7,200 mumol.min/L) when compared with solution ingestion. The maximum plasma Phe/LNAA ratio was smaller (0.16 v 0.19) with capsules. The changes for plasma tyrosine were similar to those seen with phenylalanine. There were no significant differences in the plasma concentrations of the other LNAAs between capsule and solution ingestion. Thus, given the small effect on phenylalanine Cmax and Phe/LNAA and no effect on the extent of absorption of phenylalanine, aspartame ingested in capsules at doses up to 20 mg/kg is a suitable dosage form for blinded clinical studies, provided that the slower rate of absorption of phenylalanine from capsules is taken into account.

  3. The global gene expression response of Escherichia coli to L-phenylalanine.

    PubMed

    Polen, T; Krämer, M; Bongaerts, J; Wubbolts, M; Wendisch, V F

    2005-02-01

    We investigated the global gene expression changes of Escherichia coli due to the presence of different concentrations of phenylalanine or shikimate in the growth medium. The response to 0.5 g l(-1) phenylalanine primarily reflected a perturbed aromatic amino acid metabolism, in particular due to TyrR-mediated regulation. The addition of 5g l(-1) phenylalanine reduced the growth rate by half and elicited a great number of likely indirect effects on genes regulated in response to changed pH, nitrogen or carbon availability. Consistent with the observed gene expression changes, supplementation with shikimate, tyrosine and tryptophan relieved growth inhibition by phenylalanine. In contrast to the wild-type, a tyrR disruption strain showed increased expression of pckA and of tktB in the presence of phenylalanine, but its growth was not affected by phenylalanine at the concentrations tested. The absence of growth inhibition by phenylalanine suggested that at high phenylalanine concentrations TyrR-defective strains might perform better in phenylalanine production. PMID:15639085

  4. Vacuum-Ultraviolet Circular Dichroism Spectra of Escherichia coli Dihydrofolate Reductase and Its Mutants: Contributions of Phenylalanine and Tyrosine Side Chains and Exciton Coupling of Two Tryptophan Side Chains.

    PubMed

    Ohmae, Eiji; Tanaka, Suguru; Miyashita, Yurina; Katayanagi, Katsuo; Matsuo, Koichi

    2015-10-15

    Vacuum-ultraviolet (VUV) circular dichroism (CD) spectroscopy has recently been used for secondary structure analysis of proteins; however, the contribution of aromatic side chains to protein VUV CD spectra is unresolved. In this report, VUV CD spectra of 10 Escherichia coli dihydrofolate reductase (DHFR) mutants, in which each phenylalanine or tyrosine residue was mutated to leucine, were measured down to 175 nm at 25 °C and pH 8.0 to elucidate the contributions of these aromatic side chains to the high-energy transitions of peptide bonds. The VUV CD spectra of these mutants were different from the spectrum of the wild-type protein, indicating that the contribution of the phenylalanine and tyrosine side chains of DHFR extends to the VUV region. Furthermore, the VUV CD spectrum and the folate- or NADP(+)-induced spectral change of F103L mutant DHFR indicated a modification and regeneration of exciton coupling between the Trp47 and Trp74 side chains, respectively, suggesting that exciton coupling may also contribute to the CD spectrum of DHFR in the VUV region. These results should be useful for theoretically characterizing the contribution of aromatic side chains to protein CD spectra, leading to the improvement of protein secondary-structure analysis by VUV CD spectroscopy.

  5. Intravenous infusion of L-isomers of phenylalanine and tryptophan stimulate gastric acid secretion at physiologic plasma concentrations in normal subjects and after parietal cell vagotomy.

    PubMed Central

    McArthur, K E; Isenberg, J I; Hogan, D L; Dreier, S J

    1983-01-01

    To determine whether intravenous infusion of individual amino acids stimulated gastric acid secretion in man, graded doses of phenylalanine, tryptophan, glycine, alanine, histidine, and NaCl control were infused on separate days in nine healthy subjects. Intravenous infusion of phenylalanine and tryptophan significantly stimulated gastric acid secretion to 50 and 52%, respectively, of the acid secretory response to intragastric peptone. Intravenous alanine and histidine were without effect, whereas glycine produced a slight response. Serum gastrin concentrations did not significantly change during intravenous amino acid infusion, except in response to 0.1 M phenylalanine. However, the increase in serum gastrin occurred 2 h after acid secretion had significantly increased in response to the 0.025 M phenylalanine infusion. Plasma amino acid concentrations were measured during intravenous amino acid infusion and in response to a steak meal in five of the subjects. At a time when acid secretion was significantly increased during intravenous infusion of phenylalanine and tryptophan, plasma amino acids were similar to, or less than, that observed after the steak meal, suggesting that circulating levels of these three amino acids have a physiologic effect on gastric secretion in man. Intravenous infusion of a combination of graded doses of phenylalanine plus a continuous infusion of 0.01 M tryptophan shifted the dose-response curve to the left and resulted in a significantly greater response than to either amino acid alone. In five subjects with parietal cell vagotomy, intravenous phenylalanine and tryptophan stimulated acid secretion, whereas histidine was without effect, similar to normal subjects. These studies indicate that intravenous infusion of small amounts of phenylalanine (0.025 M, 3.1 mmol/h) and tryptophan (0.01 M, 1.25 mmol/h) stimulated gastric acid secretion at plasma concentrations similar to those observed after a steak meal, suggesting a physiologic role

  6. Terreic Acid, a Quinone Epoxide Inhibitor of Bruton's Tyrosine Kinase

    NASA Astrophysics Data System (ADS)

    Kawakami, Yuko; Hartman, Stephen E.; Kinoshita, Eiji; Suzuki, Hidefumi; Kitaura, Jiro; Yao, Libo; Inagaki, Naoki; Franco, Alessandra; Hata, Daisuke; Maeda-Yamamoto, Mari; Fukamachi, Hiromi; Nagai, Hiroichi; Kawakami, Toshiaki

    1999-03-01

    Bruton's tyrosine kinase (Btk) plays pivotal roles in mast cell activation as well as in B cell development. Btk mutations lead to severe impairments in proinflammatory cytokine production induced by cross-linking of high-affinity IgE receptor on mast cells. By using an in vitro assay to measure the activity that blocks the interaction between protein kinase C and the pleckstrin homology domain of Btk, terreic acid (TA) was identified and characterized in this study. This quinone epoxide specifically inhibited the enzymatic activity of Btk in mast cells and cell-free assays. TA faithfully recapitulated the phenotypic defects of btk mutant mast cells in high-affinity IgE receptor-stimulated wild-type mast cells without affecting the enzymatic activities and expressions of many other signaling molecules, including those of protein kinase C. Therefore, this study confirmed the important roles of Btk in mast cell functions and showed the usefulness of TA in probing into the functions of Btk in mast cells and other immune cell systems. Another insight obtained from this study is that the screening method used to identify TA is a useful approach to finding more efficacious Btk inhibitors.

  7. Mechanism of fluorescence quenching of tyrosine derivatives by amide group

    NASA Astrophysics Data System (ADS)

    Wiczk, Wiesław; Rzeska, Alicja; Łukomska, Joanna; Stachowiak, Krystyna; Karolczak, Jerzy; Malicka, Joanna; Łankiewicz, Leszek

    2001-06-01

    The difference between fluorescence lifetimes of the following amino acids: phenylalanine (Phe), tyrosine (Tyr), ( O-methyl)tyrosine (Tyr(Me)), (3-hydroxy)tyrosine (Dopa), (3,4-dimethoxy)phenylalanine (Dopa(Me) 2) and their amides was used to testify the mechanism of fluorescence quenching of aromatic amino acids by the amide group. On the basis of the Marcus theory of photoinduced electron transfer parabolic relationships between ln kET and ionization potentials reduced by energy of excitation ( IP-E ∗0,0) for the above-mentioned amino acids were obtained. This finding indicates the occurrence of photoinduced electron transfer from the excited chromophore group to the amide group.

  8. A conserved acidic residue in phenylalanine hydroxylase contributes to cofactor affinity and catalysis.

    PubMed

    Ronau, Judith A; Paul, Lake N; Fuchs, Julian E; Liedl, Klaus R; Abu-Omar, Mahdi M; Das, Chittaranjan

    2014-11-01

    The catalytic domains of aromatic amino acid hydroxylases (AAAHs) contain a non-heme iron coordinated to a 2-His-1-carboxylate facial triad and two water molecules. Asp139 from Chromobacterium violaceum PAH (cPAH) resides within the second coordination sphere and contributes key hydrogen bonds with three active site waters that mediate its interaction with an oxidized form of the cofactor, 7,8-dihydro-l-biopterin, in crystal structures. To determine the catalytic role of this residue, various point mutants were prepared and characterized. Our isothermal titration calorimetry (ITC) analysis of iron binding implies that polarity at position 139 is not the sole criterion for metal affinity, as binding studies with D139E suggest that the size of the amino acid side chain also appears to be important. High-resolution crystal structures of the mutants reveal that Asp139 may not be essential for holding the bridging water molecules together, because many of these waters are retained even in the Ala mutant. However, interactions via the bridging waters contribute to cofactor binding at the active site, interactions for which charge of the residue is important, as the D139N mutant shows a 5-fold decrease in its affinity for pterin as revealed by ITC (compared to a 16-fold loss of affinity in the case of the Ala mutant). The Asn and Ala mutants show a much more pronounced defect in their kcat values, with nearly 16- and 100-fold changes relative to that of the wild type, respectively, indicating a substantial role of this residue in stabilization of the transition state by aligning the cofactor in a productive orientation, most likely through direct binding with the cofactor, supported by data from molecular dynamics simulations of the complexes. Our results indicate that the intervening water structure between the cofactor and the acidic residue masks direct interaction between the two, possibly to prevent uncoupled hydroxylation of the cofactor before the arrival of

  9. Aspartame and sucrose produce a similar increase in the plasma phenylalanine to large neutral amino acid ratio in healthy subjects.

    PubMed

    Burns, T S; Stargel, W W; Tschanz, C; Kotsonis, F N; Hurwitz, A

    1991-01-01

    Aspartame (L-aspartyl-L-phenylalanine methyl ester) consumption has been postulated to increase brain phenylalanine levels by increasing the molar ratio of the plasma phenylalanine concentration to the sum of the plasma concentrations of the other large neutral amino acids (Phe/LNAA). Dietary manipulations with carbohydrate or protein can also produce changes in the Phe/LNAA value. To compare the effects of aspartame and carbohydrate on Phe/LNAA, beverages sweetened with aspartame, sucrose, and aspartame plus sucrose, and unsweetened beverage were ingested by 8 healthy, fasted subjects in a randomized, four-way crossover design. The beverages were sweetened with an amount of aspartame (500 mg) and/or sucrose (100 g) approximately equivalent to that used to sweeten 1 liter of soft drink. The baseline-corrected plasma Phe/LNAA values did not differ significantly following ingestion of aspartame or sucrose. Following aspartame alone, the high mean ratio increased 26% over baseline 1 h after ingestion. Following sucrose alone, the high mean ratio increased 19% at 2.5 h. Sucrose increased the Phe/LNAA value due to an insulin-mediated decrease in the plasma LNAA, while aspartame increased the ratio by increasing the plasma Phe concentration. These findings indicate that similar increases in plasma Phe/LNAA occur when healthy, fasting subjects ingest amounts of equivalent sweetness of sucrose or aspartame.

  10. Gibberellic Acid-Promoted Lignification and Phenylalanine Ammonia-lyase Activity in a Dwarf Pea (Pisum sativum) 1

    PubMed Central

    Cheng, Christina K.-C.; Marsh, H. V.

    1968-01-01

    The effects of gibberellic acid on lignification in seedlings of a dwarf and a tall cultivar of pea (Pisum sativum) grown under red or white light or in the darkness, were studied. Gibberellic acid (10−6-10−4 m) promoted stem elongation in both light and dark and increased the percentage of lignin in the stems of the light-grown dwarf pea. The gibberellin had no effect on the lignin content of the tall pea although high concentrations (10−4 m) promoted growth of the tall plants. Time course studies indicated that the enhanced lignification in the gibberellin-treated dwarf plants occurred only after a lag period of several days. It was concluded that gibberellic acid-enhanced ligmification had no direct relation to gibberellic acid-promoted growth. The activity of phenylalanine ammonia-lyase (E.C. 4.3.1.5) was higher in gibberellin-treated dwarf plants grown under white or red light than in untreated dwarf plants. Gibberellic acid had no detectable effect on the activity of this enzyme when the plants were grown in darkness, just as it had no effect on lignification under dark conditions. The data suggest that in gibberellin-deficient peas the activity of phenylalanine ammonia-lyase is one of the limiting factors in lignification. PMID:16656968

  11. Gibberellic Acid-Promoted Lignification and Phenylalanine Ammonia-lyase Activity in a Dwarf Pea (Pisum sativum).

    PubMed

    Cheng, C K; Marsh, H V

    1968-11-01

    The effects of gibberellic acid on lignification in seedlings of a dwarf and a tall cultivar of pea (Pisum sativum) grown under red or white light or in the darkness, were studied. Gibberellic acid (10(-6)-10(-4)m) promoted stem elongation in both light and dark and increased the percentage of lignin in the stems of the light-grown dwarf pea. The gibberellin had no effect on the lignin content of the tall pea although high concentrations (10(-4)m) promoted growth of the tall plants. Time course studies indicated that the enhanced lignification in the gibberellin-treated dwarf plants occurred only after a lag period of several days. It was concluded that gibberellic acid-enhanced ligmification had no direct relation to gibberellic acid-promoted growth. The activity of phenylalanine ammonia-lyase (E.C. 4.3.1.5) was higher in gibberellin-treated dwarf plants grown under white or red light than in untreated dwarf plants. Gibberellic acid had no detectable effect on the activity of this enzyme when the plants were grown in darkness, just as it had no effect on lignification under dark conditions. The data suggest that in gibberellin-deficient peas the activity of phenylalanine ammonia-lyase is one of the limiting factors in lignification. PMID:16656968

  12. Effect of aspartame-derived phenylalanine on neutral amino acid uptake in human brain: a positron emission tomography study.

    PubMed

    Koeppe, R A; Shulkin, B L; Rosenspire, K C; Shaw, L A; Betz, A L; Mangner, T; Price, J C; Agranoff, B W

    1991-05-01

    The possible effects of elevation of the plasma phenylalanine level secondary to the ingestion of aspartame on brain amino acid uptake in human subjects have been investigated by means of positron emission tomography (PET). 1-[11C]Aminocyclohexanecarboxylate [( 11C]ACHC) is a poorly metabolized synthetic amino acid that crosses the blood-brain barrier by the same carrier that transports naturally occurring large neutral amino acids. Quantitative test-retest PET studies were performed on 15 individuals. Seven received two identical baseline scans, whereas eight received a baseline scan followed by a scan performed approximately 40-45 min following ingestion of an orange-flavored beverage containing 34 mg/kg of body weight of the low-calorie sweetener aspartame, a dose equivalent to the amount in 5 L of diet soft drink consumed all at once by the study subjects, weighing an average of 76 kg. The 40-45-min interval was selected to maximize the detection of possible decreases in ACHC uptake resulting from increased competition for the carrier, because the plasma phenylalanine level is known to peak at this time. We observed an 11.5% decrease in the amino acid transport rate constant K1 and a smaller decrease in the tissue distribution volume of ACHC (6%). Under conditions of normal dietary use, aspartame is thus unlikely to cause changes in brain amino acid uptake that are measurable by PET.

  13. Expression, purification, crystallization, data collection and preliminary biochemical characterization of methicillin-resistant Staphylococcus aureus Sar2028, an aspartate/tyrosine/phenylalanine pyridoxal-5′-phosphate-dependent aminotransferase

    SciTech Connect

    Seetharamappa, Jaldappagari; Oke, Muse; Liu, Huanting; McMahon, Stephen A.; Johnson, Kenneth A.; Carter, Lester; Dorward, Mark; Zawadzki, Michal; Overton, Ian M.; Niekirk, C. A. Johannes van; Graham, Shirley; Botting, Catherine H.; Taylor, Garry L.; White, Malcolm F.; Barton, Geoffrey J.; Coote, Peter J.; Naismith, James H.

    2007-05-01

    As part of work on S. aureus, the crystallization of Sar2028, a protein that is upregulated in MRSA, is reported. Sar2028, an aspartate/tyrosine/phenylalanine pyridoxal-5′-phosphate-dependent aminotransferase with a molecular weight of 48 168 Da, was overexpressed in methicillin-resistant Staphylococcus aureus compared with a methicillin-sensitive strain. The protein was expressed in Escherichia coli, purified and crystallized. The protein crystallized in a primitive orthorhombic Laue group with unit-cell parameters a = 83.6, b = 91.3, c = 106.0 Å, α = β = γ = 90°. Analysis of the systematic absences along the three principal axes indicated the space group to be P2{sub 1}2{sub 1}2{sub 1}. A complete data set was collected to 2.5 Å resolution.

  14. 21 CFR 862.1555 - Phenylalanine test system.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Phenylalanine test system. 862.1555 Section 862....1555 Phenylalanine test system. (a) Identification. A phenylalanine test system is a device intended to measure free phenylalanine (an amino acid) in serum, plasma, and urine. Measurements of phenylalanine...

  15. 21 CFR 862.1555 - Phenylalanine test system.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Phenylalanine test system. 862.1555 Section 862....1555 Phenylalanine test system. (a) Identification. A phenylalanine test system is a device intended to measure free phenylalanine (an amino acid) in serum, plasma, and urine. Measurements of phenylalanine...

  16. 21 CFR 862.1555 - Phenylalanine test system.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Phenylalanine test system. 862.1555 Section 862....1555 Phenylalanine test system. (a) Identification. A phenylalanine test system is a device intended to measure free phenylalanine (an amino acid) in serum, plasma, and urine. Measurements of phenylalanine...

  17. 21 CFR 862.1555 - Phenylalanine test system.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Phenylalanine test system. 862.1555 Section 862....1555 Phenylalanine test system. (a) Identification. A phenylalanine test system is a device intended to measure free phenylalanine (an amino acid) in serum, plasma, and urine. Measurements of phenylalanine...

  18. 21 CFR 862.1555 - Phenylalanine test system.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Phenylalanine test system. 862.1555 Section 862....1555 Phenylalanine test system. (a) Identification. A phenylalanine test system is a device intended to measure free phenylalanine (an amino acid) in serum, plasma, and urine. Measurements of phenylalanine...

  19. Mechanistic and computational studies of the reductive half-reaction of tyrosine to phenylalanine active site variants of D-arginine dehydrogenase.

    PubMed

    Gannavaram, Swathi; Sirin, Sarah; Sherman, Woody; Gadda, Giovanni

    2014-10-21

    The flavin-mediated enzymatic oxidation of a CN bond in amino acids can occur through hydride transfer, carbanion, or polar nucleophilic mechanisms. Previous results with D-arginine dehydrogenase from Pseudomonas aeruginosa (PaDADH) using multiple deuterium kinetic isotope effects (KIEs) and computational studies established preferred binding of the substrate protonated on the α-amino group, with cleavages of the NH and CH bonds occurring in asynchronous fashion, consistent with the three possible mechanisms. The hydroxyl groups of Y53 and Y249 are ≤4 Å from the imino and carboxylate groups of the reaction product iminoarginine, suggesting participation in binding and catalysis. In this study, we have investigated the reductive half-reactions of the Y53F and Y249F variants of PaDADH using substrate and solvent deuterium KIEs, solvent viscosity and pH effects, and quantum mechanical/molecular mechanical computational approaches to gain insights into the catalytic roles of the tyrosines and evaluate whether their mutations affect the transition state for substrate oxidation. Both Y53F and Y249F enzymes oxidized D-arginine with steady-state kinetic parameters similar to those of the wild-type enzyme. Rate constants for flavin reduction (k(red)) with D-leucine, a slow substrate amenable to rapid kinetics, were 3-fold smaller than the wild-type value with similar pKa values for an unprotonated group of ∼10.0. Similar pKa values were observed for (app)Kd in the variant and wild-type enzymes. However, cleavage of the substrate NH and CH bonds in the enzyme variants occurred in synchronous fashion, as suggested by multiple deuterium KIEs on k(red). These data can be reconciled with a hydride transfer mechanism, but not with carbanion and polar nucleophilic mechanisms.

  20. Study of the effects of proline, phenylalanine, and urea foliar application to Tempranillo vineyards on grape amino acid content. Comparison with commercial nitrogen fertilisers.

    PubMed

    Garde-Cerdán, T; López, R; Portu, J; González-Arenzana, L; López-Alfaro, I; Santamaría, P

    2014-11-15

    The aim of this work was to study the influence of foliar application of different nitrogen sources on grape amino acid content. The nitrogen sources applied to Tempranillo grapevines were proline, phenylalanine, urea, and two commercial nitrogen fertilisers, both without and with amino acids in their formulations. All treatments were applied at veraison and one week later. Proline treatment did not affect the must nitrogen composition. However, phenylalanine and urea foliar application enhanced the plants' synthesis of most of the amino acids, producing similar effects. In addition, the spray of commercial nitrogen fertilisers over leaves also induced a rise in grape amino acid concentrations regardless of the presence or absence of amino acids in their formulation. The most effective treatments were phenylalanine and urea followed by nitrogen fertilisers. This finding is of oenological interest for improved must nitrogen composition, ensuring better fermentation kinetics and most likely enhancing wine quality.

  1. Modeling of facilitated transport of phenylalanine by emulsion liquid membranes with di(2-ethylhexyl)phosphoric acid as a carrier

    SciTech Connect

    Liu, X.; Liu, D.

    1998-12-01

    A mathematical model is developed in this paper to simulate the facilitated transport of phenylalanine (Phe) in emulsion liquid membrane (ELM) systems with di(2-ethylhexyl)phosphoric acid as a carrier. The model takes into account the mass transfer in both the external aqueous phase and the organic membrane phase interfacial reaction as well as membrane breakage during agitation. The model is tested by comparing theoretical predications with experimental results using Phe extraction by ELM processes. It is found that the model is valid for simulating the facilitated transport of Phe with ELM under various experimental conditions.

  2. Growth, structural, spectral, optical, and thermal studies on amino acid based new NLO single crystal: L-phenylalanine-4-nitrophenol.

    PubMed

    Prakash, M; Lydia Caroline, M; Geetha, D

    2013-05-01

    A new organic nonlinear optical single crystal, L-phenylalanine-4-nitrophenol (LPAPN) belonging to the amino acid group has been successfully grown by slow evaporation technique. The lattice parameters of the grown crystal have been determined by X-ray diffraction studies. FT-IR spectrum was recorded to identify the presence of functional group and molecular structure was confirmed by NMR spectrum. Thermal strength of the grown crystal has been studied using TG-DTA analyses. The grown crystals were found to be transparent in the entire visible region. The existence of second harmonic generation signals was observed using Nd:YAG laser with fundamental wavelength of 1064 nm.

  3. Growth, structural, spectral, optical, and thermal studies on amino acid based new NLO single crystal: L-phenylalanine-4-nitrophenol

    NASA Astrophysics Data System (ADS)

    Prakash, M.; Lydia Caroline, M.; Geetha, D.

    2013-05-01

    A new organic nonlinear optical single crystal, L-phenylalanine-4-nitrophenol (LPAPN) belonging to the amino acid group has been successfully grown by slow evaporation technique. The lattice parameters of the grown crystal have been determined by X-ray diffraction studies. FT-IR spectrum was recorded to identify the presence of functional group and molecular structure was confirmed by NMR spectrum. Thermal strength of the grown crystal has been studied using TG-DTA analyses. The grown crystals were found to be transparent in the entire visible region. The existence of second harmonic generation signals was observed using Nd:YAG laser with fundamental wavelength of 1064 nm.

  4. Comparison of Growth Performance and Whole-body Amino Acid Composition in Red Seabream (Pagrus major) Fed Free or Dipeptide Form of Phenylalanine.

    PubMed

    Kim, Sung-Sam; Rahimnejad, Samad; Song, Jin-Woo; Lee, Kyeong-Jun

    2012-08-01

    This study was conducted to evaluate the efficacy of the dipeptide form of phenylalanine as a new source of amino acid in terms of growth performance and whole-body amino acid composition in comparison to the free form for red seabream (Pagrus major). Fish (1.46±0.001 g) were fed four isonitrogenous and isocaloric experimental diets containing 0.7 or 1.4% phenylalanine either in free or dipeptide form. A feeding trial was carried out in three replicates and the fish were fed to apparent satiation for six weeks. At the end of the feeding trial, feed intake of fish was influenced by both phenylalanine form and level and significantly higher values were obtained at an inclusion level of 0.7% and by the use of dipeptide form. However, the other growth parameters did not significantly differ among treatments. Whole-body amino acid compositions revealed no significant changes in concentrations of both essential and non-essential amino acids regardless of the increase in phenylalanine levels or the use of its different forms. The finding in this study indicates that juvenile red seabream can utilize dipeptide phenylalanine as efficiently as free form without any undesirable effects on growth performance or whole-body amino acid composition.

  5. Spectroscopic analyses on interaction of o-Vanillin- D-Phenylalanine, o-Vanillin- L-Tyrosine and o-Vanillin- L-Levodopa Schiff Bases with bovine serum albumin (BSA)

    NASA Astrophysics Data System (ADS)

    Gao, Jingqun; Guo, Yuwei; Wang, Jun; Wang, Zhiqiu; Jin, Xudong; Cheng, Chunping; Li, Ying; Li, Kai

    2011-04-01

    In this work, three o-Vanillin Schiff Bases (o-VSB: o-Vanillin- D-Phenylalanine (o-VDP), o-Vanillin- L-Tyrosine (o-VLT) and o-Vanillin- L-Levodopa (o-VLL)) with alanine constituent were synthesized by direct reflux method in ethanol solution, and then were used to study the interaction to bovine serum albumin (BSA) molecules by fluorescence spectroscopy. Based on the fluorescence quenching calculation, the bimolecular quenching constant ( Kq), apparent quenching constant ( Ksv), effective binding constant ( KA) and corresponding dissociation constant ( KD) as well as binding site number ( n) were obtained. In addition, the binding distance ( r) was also calculated according to Foster's non-radioactive energy transfer theory. The results show that these three o-VSB can efficiently bind to BSA molecules, but the binding array order is o-VDP-BSA > o-VLT-BSA > o-VLL-BSA. Synchronous fluorescence spectroscopy indicates that the o-VDP is more accessibility to tryptophan (Trp) residues of BSA molecules than to tyrosine (Tyr) residues. Nevertheless, the o-VLT and o-VLL are more accessibility to Tyr residues than to Trp residues.

  6. Tyrosine can protect against oxidative stress through ferryl hemoglobin reduction.

    PubMed

    Lu, Naihao; He, Yingjie; Chen, Chao; Tian, Rong; Xiao, Qiang; Peng, Yi-Yuan

    2014-08-01

    The toxic mechanism of hemoglobin (Hb) under oxidative stress is linked to the formations of highly cytotoxic ferryl species and subsequently heme-to-protein cross-linked derivative of Hb (Hb-X). In this study, we have examined the effects of free tyrosine and its analogues (3-chlorotyrosine, phenylalanine) on the stability of ferryl hemoglobin and the formation of Hb-X. The results showed that free tyrosine (not phenylalanine, 10-500 μM) was an efficient reducing agent of ferryl species and also effective at preventing the formation of cytotoxic Hb-X. Meanwhile, the dimeric tyrosine was formed as the oxidation product of tyrosine during Hb redox reaction. Compared with free tyrosine, 3-chlorotyrosine, an oxidation product of tyrosine and a proposed biomarker for hypochlorous acid (HOCl) in vivo, exhibited stronger antioxidant properties in Hb-induced oxidative stress, which was consistent with its more efficient ability in the reduction of ferryl species. These results showed that the presence of tyrosine and its derivative in vivo and vitro could ameliorate oxidative damage through ferryl heme reduction. The antioxidant ability, therefore, may provide new insights into the nutritional and physiological significance of free tyrosine with redox active heme proteins-related oxidative stress.

  7. Inhibition of lactoperoxidase-catalyzed 2,2'-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS) and tyrosine oxidation by tyrosine-containing random amino acid copolymers.

    PubMed

    Clausen, Morten R; Skibsted, Leif H; Stagsted, Jan

    2008-09-24

    Oxidation of 2,2'-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) by lactoperoxidase was found to be inhibited by tyrosine-containing random amino acid copolymers but not by tyrosine. Both electrostatic effects and polymer size were found to be important by comparison of negatively and positively charged copolymers of varying lengths, with poly(Glu, Tyr)4:1 ([E 4Y 1] approximately 40) as the strongest competitive inhibitor (EC 50 approximately 20 nM). This polymer did not form dityrosine in the presence of lactoperoxidase (LPO) and peroxide. Furthermore, incubation with tert-butyl hydroperoxide, as opposed to hydrogen peroxide, resulted in a peculiar long lag phase of the reaction between the redox intermediate compound II and [E 4Y 1] approximately 40, indicating a very tight association between enzyme and inhibitor. We propose that interactions between multiple positively charged areas on the surface of LPO and the polymer are required for optimal inhibition.

  8. Effects of oral aspartame on plasma phenylalanine in humans and experimental rodents. Short note.

    PubMed

    Wurtman, R J; Maher, T J

    1987-01-01

    All aspartame does given to humans cause greater elevations in plasma (and, presumably, brain) phenylalanine than in plasma tyrosine. In contrast, doses of aspartame usually used in experiments on rodents preferentially elevate tyrosine. Since phenylalanine can inhibit brain catecholamine synthesis while tyrosine is the antidote for this effect, we determined the aspartame dose that would be needed to elevate phenylalanine more than tyrosine in rodents, using published data. In general rodents need 60 times as much aspartame, on a mg/kg basis, as humans to obtain comparable elevations in phenylalanine with respect to tyrosine.

  9. Inhibition of lymphoid tyrosine phosphatase by benzofuran salicylic acids.

    PubMed

    Vang, Torkel; Xie, Yuli; Liu, Wallace H; Vidović, Dusica; Liu, Yidong; Wu, Shuangding; Smith, Deborah H; Rinderspacher, Alison; Chung, Caty; Gong, Gangli; Mustelin, Tomas; Landry, Donald W; Rickert, Robert C; Schürer, Stephan C; Deng, Shi-Xian; Tautz, Lutz

    2011-01-27

    The lymphoid tyrosine phosphatase (Lyp, PTPN22) is a critical negative regulator of T cell antigen receptor (TCR) signaling. A single-nucleotide polymorphism (SNP) in the ptpn22 gene correlates with the incidence of various autoimmune diseases, including type 1 diabetes, rheumatoid arthritis, and systemic lupus erythematosus. Since the disease-associated allele is a more potent inhibitor of TCR signaling, specific Lyp inhibitors may become valuable in treating autoimmunity. Using a structure-based approach, we synthesized a library of 34 compounds that inhibited Lyp with IC(50) values between 0.27 and 6.2 μM. A reporter assay was employed to screen for compounds that enhanced TCR signaling in cells, and several inhibitors displayed a dose-dependent, activating effect. Subsequent probing for Lyp's direct physiological targets by immunoblot analysis confirmed the ability of the compounds to inhibit Lyp in T cells. Selectivity profiling against closely related tyrosine phosphatases and in silico docking studies with the crystal structure of Lyp yielded valuable information for the design of Lyp-specific compounds. PMID:21190368

  10. Hydration profiles of aromatic amino acids: conformations and vibrations of L-phenylalanine-(H2O)n clusters.

    PubMed

    Ebata, Takayuki; Hashimoto, Takayo; Ito, Takafumi; Inokuchi, Yoshiya; Altunsu, Fuat; Brutschy, Bernhard; Tarakeshwar, P

    2006-11-01

    IR-UV double resonance spectroscopy and ab initio calculations were employed to investigate the structures and vibrations of the aromatic amino acid, L-phenylalanine-(H(2)O)(n) clusters formed in a supersonic free jet. Our results indicate that up to three water molecules are preferentially bound to both the carbonyl oxygen and the carboxyl hydrogen of L-phenylalanine (L-Phe) in a bridged hydrogen-bonded conformation. As the number of water molecules is increased, the bridge becomes longer. Two isomers are found for L-Phe-(H(2)O)(1), and both of them form a cyclic hydrogen-bond between the carboxyl group and the water molecule. In L-Phe-(H(2)O)(2), only one isomer was identified, in which two water molecules form extended cyclic hydrogen bonds with the carboxyl group. In the calculated structure of L-Phe-(H(2)O)(3) the bridge of water molecules becomes larger and exhibits an extended hydrogen-bond to the pi-system. Finally, in isolated L-Phe, the D conformer was found to be the most stable conformer by the experiment and by the ab initio calculation.

  11. Plasma and brain kinetics of large neutral amino acids and of striatum monoamines in rats given aspartame.

    PubMed

    Romano, M; Diomede, L; Guiso, G; Caccia, S; Perego, C; Salmona, M

    1990-05-01

    Two doses (250 and 1000 mg/kg body weight) of aspartame were administered orally to male rats, and plasma and brain phenylalanine and tyrosine kinetic profiles were studied. In both plasma and brain the maximum increase in phenylalanine and tyrosine levels was reached 60 min after treatment. The changes in brain levels of phenylalanine or tyrosine 0, 60, 120 or 180 min after treatment with 1000 mg AMP/kg were directly correlated with the ratio of the plasma concentration of phenylalanine or tyrosine to the overall plasma concentration of the other large neutral amino acids. The time course of monoamine and metabolite concentrations, in the corpora striatum of the brain, was studied after an oral dose of 500 mg phenylalanine/kg. No significant modifications of monoamine levels were found at any of the times studied, up to 5 hr after dosing.

  12. Utilization of [14C]phenylalanine derived from arylphorin or free amino acid in Manduca sexta pharate adults

    NASA Technical Reports Server (NTRS)

    Wu, M.; Tischler, M. E.

    1995-01-01

    The role of arylphorin as a storage protein was studied using 14C-arylphorin. 14C-arylphorin was produced optimally by incubating one-half fat body from Manduca sexta fifth instar larvae at 22 degrees C for 24 h, in 1 ml of medium containing amino acids at 25% of their physiological concentration with [U-14C]-phenylalanine (phe) provided initially without nonlabeled phenylalanine. Nonlabeled phe was provided after 1 h at 16% of its physiological concentration. The specific activity of 14C-arylphorin produced in vitro was 30 times greater than that generated in vivo. Injection of 14C-arylphorin into pharate adults was used to study the distribution of 14C-phe derived from this protein into 14CO2 and tissues for comparison with injection of free 14C-phe during the middle (days 6 to 12 pharate adult) and late (days 12 to 17 pharate adult) stages of adult development. Appearance of 14CO2 from 14C-arylphorin as compared to 14C-phenylalanine showed a slower time course during both the middle and late stages of development, in keeping with the time needed for degradation of the protein. In accord with faster phe turnover near the end of adult development, total 14CO2 production was greater and the retention of 14C in hemolymph and fat body was less compared to the middle stage of development regardless of whether 14C-arylphorin or 14C-phe was injected. In the middle stage of development, the appearance of 14C in the cuticle and head parts was greater, whereas incorporation into abdomen and thorax was less than during the late stage of development. Since the pattern of 14C distribution from 14C-arylphorin and 14C-phe was similar, one major function of arylphorin must be as a storage protein replenishing the supply of free amino acids used for synthesis of adult tissues. These results also suggest a limited contribution of M. sexta arylphorin to formation of the cuticle subsequent to day-6 pharate adult.

  13. A comparative study of the vibrational spectra of the anticancer drug melphalan and its fundamental molecules 3-phenylpropionic acid and L-phenylalanine

    NASA Astrophysics Data System (ADS)

    Badawi, Hassan M.; Khan, Ibrahim

    2016-04-01

    The structural stability and the vibrational spectra of the anticancer drug melphalan and its parent compounds 3-phenylpropionic acid and L-phenylalanine were investigated by the DFT B3LYP/6-311G** calculations. Melphalan and its fundamental compounds were predicted to exist predominantly in non-planar structures. The vibrational frequencies of the low energy structures of melphalan, 3-phenylpropionic acid, and phenylalanine were computed at the DFT B3LYP level of theory. Complete vibrational assignments of the normal modes of melphalan, 3-phenylpropionic acid, and phenylalanine were provided by combined theoretical and experimental data of the molecules. The experimental infrared spectra of phenylalanine and melphalan show a significantly different pattern of the Cdbnd O stretching mode as compared to those of normal carboxylic acids. A comparison of the 3700-2000 cm-1 infrared spectral region of the three molecules suggests the presence of similar intermolecular H-bonding in their condensed phases. The observed infrared and Raman spectra are consistent with the presence of one predominant melphalan conformation at room temperature.

  14. Role of Tyrosine Isomers in Acute and Chronic Diseases Leading to Oxidative Stress - A Review

    PubMed Central

    Molnár, Gergő A.; Kun, Szilárd; Sélley, Eszter; Kertész, Melinda; Szélig, Lívia; Csontos, Csaba; Böddi, Katalin; Bogár, Lajos; Miseta, Attila; Wittmann, István

    2016-01-01

    Oxidative stress plays a major role in the pathogenesis of a variety of acute and chronic diseases. Measurement of the oxidative stress-related end products may be performed, e.g. that of structural isomers of the physiological para-tyrosine, namely meta- and ortho-tyrosine, that are oxidized derivatives of phenylalanine. Recent data suggest that in sepsis, serum level of meta-tyrosine increases, which peaks on the 2nd and 3rd days (p<0.05 vs. controls), and the kinetics follows the intensity of the systemic inflammation correlating with serum procalcitonin levels. In a similar study subset, urinary meta-tyrosine excretion correlated with both need of daily insulin dose and the insulin-glucose product in non-diabetic septic cases (p<0.01 for both). Using linear regression model, meta-tyrosine excretion, urinary meta-tyrosine/para-tyrosine, urinary ortho-tyrosine/para-tyrosine and urinary (meta- + ortho-tyrosine)/para-tyrosine proved to be markers of carbohydrate homeostasis. In a chronic rodent model, we tried to compensate the abnormal tyrosine isomers using para-tyrosine, the physiological amino acid. Rats were fed a standard high cholesterol-diet, and were given para-tyrosine or vehicle orally. High-cholesterol feeding lead to a significant increase in aortic wall meta-tyrosine content and a decreased vasorelaxation of the aorta to insulin and the glucagon-like peptide-1 analogue, liraglutide, that both could be prevented by administration of para-tyrosine. Concluding, these data suggest that meta- and ortho-tyrosine are potential markers of oxidative stress in acute diseases related to oxidative stress, and may also interfere with insulin action in septic humans. Competition of meta- and ortho-tyrosine by supplementation of para-tyrosine may exert a protective role in oxidative stress-related diseases. PMID:26785996

  15. Amino acid-bile acid based molecules: extremely narrow surfactant nanotubes formed by a phenylalanine-substituted cholic acid.

    PubMed

    Travaglini, Leana; D'Annibale, Andrea; Schillén, Karin; Olsson, Ulf; Sennato, Simona; Pavel, Nicolae V; Galantini, Luciano

    2012-12-21

    An amino acid-substituted bile acid forms tubular aggregates with inner and outer diameters of about 3 and 6 nm. The diameters are unusually small for surfactant self-assembled tubes. The results enhance the spectrum of applications of supramolecular tubules and open up possibilities for investigating a novel class of biological amphiphiles.

  16. Lack of behavioural effects after acute tyrosine depletion in healthy volunteers.

    PubMed

    Lythe, K E; Anderson, I M; Deakin, J F W; Elliott, R; Strickland, P L

    2005-01-01

    Acute dietary tyrosine depletion has previously been shown to reduce dopamine neurotransmission in both animals and humans. In this study, we investigated the effects of brain dopamine depletion, through acute tyrosine and phenylalanine depletion, on plasma prolactin, mood and neuropsychological function in 12 normal subjects. In a randomized, double-blind, cross-over design, subjects received two amino-acid drinks separated by a week, a nutritionally balanced mixture (Bal) and on the other occasion a tyrosine and phenylalanine deficient mixture (TP-). The plasma ratio of tyrosine and phenylalanine to the other large neutral amino acids decreased significantly on the TP- occasion (-78.7%, p < 0.0001) and there was an increase in plasma prolactin concentration relative to the balanced drink in the seven subjects for whom results were available for both occasions (p < 0.02). Acute tyrosine depletion did not alter mood as measured by visual analogue scale ratings, and measures of memory, attention and behavioural inhibition were also unaffected. Our results are consistent with acute dietary tyrosine depletion causing a reduction in brain dopamine neurotransmission but raise questions about how robust or consistent the effects are on psychological function. PMID:15671123

  17. Identification of a highly conserved valine-glycine-phenylalanine amino acid triplet required for HIV-1 Nef function

    PubMed Central

    2012-01-01

    Background The Nef protein of HIV facilitates virus replication and disease progression in infected patients. This role as pathogenesis factor depends on several genetically separable Nef functions that are mediated by interactions of highly conserved protein-protein interaction motifs with different host cell proteins. By studying the functionality of a series of nef alleles from clinical isolates, we identified a dysfunctional HIV group O Nef in which a highly conserved valine-glycine-phenylalanine (VGF) region, which links a preceding acidic cluster with the following proline-rich motif into an amphipathic surface was deleted. In this study, we aimed to study the functional importance of this VGF region. Results The dysfunctional HIV group O8 nef allele was restored to the consensus sequence, and mutants of canonical (NL4.3, NA-7, SF2) and non-canonical (B2 and C1422) HIV-1 group M nef alleles were generated in which the amino acids of the VGF region were changed into alanines (VGF→AAA) and tested for their capacity to interfere with surface receptor trafficking, signal transduction and enhancement of viral replication and infectivity. We found the VGF motif, and each individual amino acid of this motif, to be critical for downregulation of MHC-I and CXCR4. Moreover, Nef’s association with the cellular p21-activated kinase 2 (PAK2), the resulting deregulation of cofilin and inhibition of host cell actin remodeling, and targeting of Lck kinase to the trans-golgi-network (TGN) were affected as well. Of particular interest, VGF integrity was essential for Nef-mediated enhancement of HIV virion infectivity and HIV replication in peripheral blood lymphocytes. For targeting of Lck kinase to the TGN and viral infectivity, especially the phenylalanine of the triplet was essential. At the molecular level, the VGF motif was required for the physical interaction of the adjacent proline-rich motif with Hck. Conclusion Based on these findings, we propose that this highly

  18. Fungal and Plant Phenylalanine Ammonia-lyase

    PubMed Central

    Hyun, Min Woo; Yun, Yeo Hong; Kim, Jun Young

    2011-01-01

    L-Phenylalanine is one of the essential amino acids that cannot be synthesized in mammals in adequate amounts to meet the requirements for protein synthesis. Fungi and plants are able to synthesize phenylalanine via the shikimic acid pathway. L-Phenylalanine, derived from the shikimic acid pathway, is used directly for protein synthesis in plants or metabolized through the phenylpropanoid pathway. This phenylpropanoid metabolism leads to the biosynthesis of a wide array of phenylpropanoid secondary products. The first step in this metabolic sequence involves the action of phenylalanine ammonia-lyase (PAL). The discovery of PAL enzyme in fungi and the detection of 14CO2 production from 14C-ring-labeled phenylalanine and cinnamic acid demonstrated that certain fungi can degrade phenylalanine by a pathway involving an initial deamination to cinnamic acid, as happens in plants. In this review, we provide background information on PAL and a recent update on the presence of PAL genes in fungi. PMID:22783113

  19. Growth, thermal, dielectric and mechanical properties of L-phenylalanine-benzoic acid: A nonlinear optical single crystal

    NASA Astrophysics Data System (ADS)

    Tamilselvan, S.; Vimalan, M.; Vetha Potheher, I.; Rajasekar, S.; Jeyasekaran, R.; Antony Arockiaraj, M.; Madhavan, J.

    2013-10-01

    An efficient amino acid family nonlinear optical single crystal L-phenylalanine-benzoic acid (LPB) was conveniently grown by slow evaporation technique at room temperature. The crystal system and the lattice parameters were analyzed by single crystal X-ray diffraction studies. The grown crystal has excellent transmission in the entire visible region and its lower cut-off wavelength was found to be 248 nm. The SHG efficiency of the grown crystal was found to be 1.6 times higher than that of KDP crystal. The Laser damage threshold value of LPB has been found to be 6.5 GW/cm2. The sample was thermally stable up to 134 °C. Microhardness, dielectric and AC/DC conductivity measurements were made along (0 0 1) plane and reported for the first time. Microhardness studies revealed that the sample belongs to hard nature. Frequency dependent dielectric constant was measured for different temperatures and found maximum dielectric constant of 14 for 363 K. Photoconductivity studies of LPB divulged its negative photoconducting nature.

  20. Molecular aggregation in selected crystalline 1:1 complexes of hydrophobic D- and L-amino acids. IV. The L-phenylalanine series.

    PubMed

    Görbitz, Carl Henrik; Rissanen, Kari; Valkonen, Arto; Husabø, Asmund

    2009-06-01

    The amino acid L-phenylalanine has been cocrystallized with D-2-aminobutyric acid, C(9)H(11)NO(2).C(4)H(9)NO(2), D-norvaline, C(9)H(11)NO(2).C(5)H(11)NO(2), and D-methionine, C(9)H(11)NO(2).C(5)H(11)NO(2)S, with linear side chains, as well as with D-leucine, C(9)H(11)NO(2).C(6)H(13)NO(2), D-isoleucine, C(9)H(11)NO(2).C(6)H(13)NO(2), and D-allo-isoleucine, C(9)H(11)NO(2).C(6)H(13)NO(2), with branched side chains. The structures of these 1:1 complexes fall into two classes based on the observed hydrogen-bonding pattern. From a comparison with other L:D complexes involving hydrophobic amino acids and regular racemates, it is shown that the structure-directing properties of phenylalanine closely parallel those of valine and isoleucine but not those of leucine, which shares side-chain branching at C(gamma) with phenylalanine and is normally considered to be the most closely related non-aromatic amino acid.

  1. Enantioselective acylation of β-phenylalanine acid and its derivatives catalyzed by penicillin G acylase from Alcaligenes faecalis.

    PubMed

    Li, Dengchao; Ji, Lilian; Wang, Xinfeng; Wei, Dongzhi

    2013-01-01

    This study developed a simple, efficient method for producing racemic β-phenylalanine acid (BPA) and its derivatives via the enantioselective acylation catalyzed by the penicillin G acylase from Alcaligenes faecalis (Af-PGA). When the reaction was run at 25°C and pH 10 in an aqueous medium containing phenylacetamide and BPA in a molar ratio of 2:1, 8 U/mL enzyme and 0.1 M BPA, the maximum BPA conversion efficiency at 40 min only reached 36.1%, which, however, increased to 42.9% as the pH value and the molar ratio of phenylacetamide to BPA were elevated to 11 and 3:1, respectively. Under the relatively optimum reaction conditions, the maximum conversion efficiencies of BPA derivatives all reached about 50% in a relatively short reaction time (45-90 min). The enantiomeric excess value of product (ee(p)) and enantiomeric excess value of substrate (ee(s)) were all above 98% and 95%, respectively. These results suggest that the method established in this study is practical, effective, and environmentally benign and may be applied to industrial production of enantiomerically pure BPA and its derivatives. PMID:23302108

  2. Uncoupling of biliary lipid from bile acid secretion by formyl-methionyl-leucyl-phenylalanine in the rat.

    PubMed

    Mizuno, K; Hoshino, M; Hayakawa, T; Yamada, H; Nakazawa, T; Inagaki, T; Takeuchi, T; Kunimatsu, M

    1996-11-01

    A neutrophil chemotactic factor N-formyl-methionyl-leucyl-phenylalanine (fMLP), produced by Escherichia coli under conditions of intestinal inflammation, is reported to circulate enterohepatically in the presence of experimental colitis, but its effect on bile secretion is unclear. Therefore, we investigated the effect of fMLP on bile secretion in a single-pass isolated perfused rat liver system. Infusion of fMLP at different concentrations (2 micromol/L, 10 micromol/L, and 20 micromol/L) into the portal vein resulted in excretion into bile in the native form, independent of sodium taurocholate (1 micromol/min) infusion. Excretion of fMLP increased dose dependently, and approximately 12% of the infused dose was detected at each concentration. With constant infusion of sodium taurocholate (1 micromol/min), fMLP (20 micromol/L) increased bile flow but decreased phospholipid and cholesterol secretion. Bile acid secretion was not affected. Phospholipid/bile acid molar ratios decreased from 0.069 +/- 0.002 to 0.038 +/- 0.002, and cholesterol/bile acid molar ratios decreased from 0.0074 +/- 0.0009 to 0.0029 +/- 0.0008. Thus, administration of fMLP resulted in the uncoupling of biliary excretion of phospholipid and cholesterol from that of bile acids; this effect proved reversible. The increase in bile flow caused by fMLP infusion appeared to result from osmotic choleresis. When 25 mg of horseradish peroxidase, a conventional marker of transcytotic vesicle transport pathway, was infused for 1 minute as a pulse load into the portal vein after continuous infusion of taurocholate, its late peak excretion was reduced by fMLP (10 micromol/L) from 9.59 +/- 1.09 to 6.05 +/- 0.66 (ng/g liver). Gel-permeation chromatography of bile showed a specific association of fMLP with bile acids. These results suggest an uncoupling of biliary lipids from bile acids by fMLP because of inhibition of transcellular vesicle transport and interaction between fMLP and bile acid micelles in the bile

  3. Behavioral alteration of plasma phenylalanine concentration.

    PubMed

    Jevning, R; Pirkle, H C; Wilson, A F

    1977-11-01

    The concentration of 13 neutral and acidic plasma amino acids was measured before, during and after either 40 min of control relaxation or 40 min of the process known as transcendental meditation (TM). An electro-oculogram, electroencephalogram, and electromyogram were simultaneously monitored in these subjects. Increased phenylalanine concentration was noted during TM practice with no change during control relaxation; no difference between the groups of total time slept or sleep stage percent was observed. The stability of phenylalanine concentration in controls and lack of correlation of increased phenylalanine with sleep in the long-term practitioners seem to suggest a relationship of the phenylalanine increase to TM practice.

  4. Mutation of tyrosine-141 inhibits insulin-promoted tyrosine phosphorylation and increased responsiveness of the human beta 2-adrenergic receptor.

    PubMed Central

    Valiquette, M; Parent, S; Loisel, T P; Bouvier, M

    1995-01-01

    The ability of insulin to promote phosphorylation of the human beta 2-adrenergic receptor (beta 2AR) was assessed in Chinese hamster fibroblasts transfected with beta 2AR cDNA. Phosphotyrosine residues were detected in purified beta 2AR using a polyclonal anti-phosphotyrosine antibody and by phosphoamino acid analysis following metabolic labelling with inorganic 32P. Treatment of the cells with insulin induced a 2.4-fold increase in the phosphotyrosine content of the receptor. The insulin-promoted phosphorylation of the beta 2AR was accompanied by an increase in the beta-adrenergic-stimulated adenyl cyclase activity. Substitution of a phenylalanine residue for tyrosine-141 completely prevented both the increased tyrosine phosphorylation and the enhanced responsiveness of the beta 2AR promoted by insulin treatment. Mutation of three other tyrosines located in the cytoplasmic domain of the receptor, tyrosine-366, tyrosine-350 and tyrosine-354, did not abolish the insulin-promoted tyrosine phosphorylation. Taken together, these results suggest that insulin promotes phosphorylation of the beta 2AR on tyrosine-141 and that such phosphorylation leads to a supersensitization of the receptor. Images PMID:8521811

  5. Are carboxyl groups the most acidic sites in amino acids? Gas-phase acidities, photoelectron spectra, and computations on tyrosine, p-hydroxybenzoic acid, and their conjugate bases.

    PubMed

    Tian, Zhixin; Wang, Xue-Bin; Wang, Lai-Sheng; Kass, Steven R

    2009-01-28

    Deprotonation of tyrosine in the gas phase was found to occur preferentially at the phenolic site, and the conjugate base consists of a 70:30 mixture of phenoxide and carboxylate anions at equilibrium. This result was established by developing a chemical probe for differentiating these two isomers, and the presence of both ions was confirmed by photoelectron spectroscopy. Equilibrium acidity measurements on tyrosine indicated that deltaG(acid)(o) = 332.5 +/- 1.5 kcal mol(-1) and deltaH(acid)(o) = 340.7 +/- 1.5 kcal mol(-1). Photoelectron spectra yielded adiabatic electron detachment energies of 2.70 +/- 0.05 and 3.55 +/- 0.10 eV for the phenoxide and carboxylate anions, respectively. The H/D exchange behavior of deprotonated tyrosine was examined using three different alcohols (CF3CH2OD, C6H5CH2OD, and CH3CH2OD), and incorporation of up to three deuterium atoms was observed. Two pathways are proposed to account for these results, and all of the experimental findings are supplemented with B3LYP/aug-cc-pVDZ and G3B3 calculations. In addition, it was found that electrospray ionization of tyrosine from a 3:1 (v/v) CH3OH/H2O solution using a commercial source produces a deprotonated [M-H]- anion with the gas-phase equilibrium composition rather than the structure of the ion that exists in aqueous media. Electrospray ionization from acetonitrile, however, leads largely to the liquid-phase (carboxylate) structure. A control molecule, p-hydroxybenzoic acid, was found to behave in a similar manner. Thus, the electrospray conditions that are employed for the analysis of a compound can alter the isomeric composition of the resulting anion.

  6. [Phenylalanine ammonia-lyase of pigmented yeasts].

    PubMed

    Mushi, N Iu; Kupletskaia, M B; Bab'eva, I P; Egorov, N S

    1980-01-01

    116 pigmented yeast cultures were tested for the presence of L-phenylalanine-ammonia lyase transforming L-phenylalanine into trans-cinnamic acid. The enzyme was found in 54 strains. Most of these strains belonged to the genera Rhodotorula and Sporobolomyces. Toluene, along with acetone, was successfully used to increase cellular permeability of the yeast cultures while determining the activity of phenylalanine-ammonia lyase.

  7. Identification of the LIV-I/LS system as the third phenylalanine transporter in Escherichia coli K-12.

    PubMed

    Koyanagi, Takashi; Katayama, Takane; Suzuki, Hideyuki; Kumagai, Hidehiko

    2004-01-01

    In Escherichia coli, the active transport of phenylalanine is considered to be performed by two different systems, AroP and PheP. However, a low level of accumulation of phenylalanine was observed in an aromatic amino acid transporter-deficient E. coli strain (DeltaaroP DeltapheP Deltamtr Deltatna DeltatyrP). The uptake of phenylalanine by this strain was significantly inhibited in the presence of branched-chain amino acids. Genetic analysis and transport studies revealed that the LIV-I/LS system, which is a branched-chain amino acid transporter consisting of two periplasmic binding proteins, the LIV-binding protein (LIV-I system) and LS-binding protein (LS system), and membrane components, LivHMGF, is involved in phenylalanine accumulation in E. coli cells. The K(m) values for phenylalanine in the LIV-I and LS systems were determined to be 19 and 30 micro M, respectively. Competitive inhibition of phenylalanine uptake by isoleucine, leucine, and valine was observed for the LIV-I system and, surprisingly, also for the LS system, which has been assumed to be leucine specific on the basis of the results of binding studies with the purified LS-binding protein. We found that the LS system is capable of transporting isoleucine and valine with affinity comparable to that for leucine and that the LIV-I system is able to transport tyrosine with affinity lower than that seen with other substrates. The physiological importance of the LIV-I/LS system for phenylalanine accumulation was revealed in the growth of phenylalanine-auxotrophic E. coli strains under various conditions.

  8. In vitro effects of cinnamic acid derivatives on protein tyrosine phosphatase 1B.

    PubMed

    Adisakwattana, Sirichai; Pongsuwan, Jirawan; Wungcharoen, Chompunut; Yibchok-anun, Sirintorn

    2013-10-01

    Protein Tyrosine Phosphatase 1B (PTP1B) is a major negative regulator of insulin signaling pathways. Finding selective PTP1B inhibitors from natural sources has been widely recognized as a potential drug target for the treatment of diabetes mellitus and obesity. In the present study, we evaluated the inhibitory activity of cinnamic acid derivatives against PTP1B in vitro. Among 14 cinnamic acid derivatives and related compounds, the most potent inhibitor PTP1Bs were o-hydroxycinnamic acid and p-hydroxycinnamic acid, which had IC50 values of 137.67 ± 13.37 and 181.60 ± 9.34 µM, respectively. The kinetics analysis revealed that PTP1B was inhibited by o-hydroxycinnamic acid and p-hydroxycinnamic acid in a non-competitive manner. o-Hydroxycinnamic acid (25 μM) and p-hydroxycinnamic acid (25 μM), in combination with sodium orthovanadate (0.0125 μM), demonstrated a synergistic effect to inhibit PTP1B activity. In conclusion, the findings provide a new insight into naturally occurring PTP1B inhibitors that could be useful for treatment of diabetes and obesity.

  9. Phenylalanine metabolism regulates reproduction and parasite melanization in the malaria mosquito.

    PubMed

    Fuchs, Silke; Behrends, Volker; Bundy, Jacob G; Crisanti, Andrea; Nolan, Tony

    2014-01-01

    The blood meal of the female malaria mosquito is a pre-requisite to egg production and also represents the transmission route for the malaria parasite. The proper and rapid assimilation of proteins and nutrients in the blood meal creates a significant metabolic challenge for the mosquito. To better understand this process we generated a global profile of metabolite changes in response to blood meal of Anopheles gambiae, using Gas Chromatography-Mass Spectrometry (GC-MS). To disrupt a key pathway of amino acid metabolism we silenced the gene phenylalanine hydroxylase (PAH) involved in the conversion of the amino acid phenylalanine into tyrosine. We observed increased levels of phenylalanine and the potentially toxic metabolites phenylpyruvate and phenyllactate as well as a reduction in the amount of tyrosine available for melanin synthesis. This in turn resulted in a significant impairment of the melanotic encapsulation response against the rodent malaria parasite Plasmodium berghei. Furthermore silencing of PAH resulted in a significant impairment of mosquito fertility associated with reduction of laid eggs, retarded vitellogenesis and impaired melanisation of the chorion. Carbidopa, an inhibitor of the downstream enzyme DOPA decarboxylase that coverts DOPA into dopamine, produced similar effects on egg melanization and hatching rate suggesting that egg chorion maturation is mainly regulated via dopamine. This study sheds new light on the role of amino acid metabolism in regulating reproduction and immunity.

  10. Genetic Incorporation of the Unnatural Amino Acid p-Acetyl Phenylalanine into Proteins for Site-Directed Spin Labeling

    PubMed Central

    Evans, Eric G.B.; Millhauser, Glenn L.

    2016-01-01

    Site-directed spin labeling (SDSL) is a powerful tool for the characterization of protein structure and dynamics; however, its application in many systems is hampered by the reliance on unique and benign cysteine substitutions for the site-specific attachment of the spin label. An elegant solution to this problem involves the use of genetically encoded unnatural amino acids (UAAs) containing reactive functional groups that are chemically orthogonal to those of the 20 amino acids found naturally in proteins. These unique functional groups can then be selectively reacted with an appropriately functionalized spin probe. In this chapter, we detail the genetic incorporation of the ketone-bearing amino acid p-acetyl phenylalanine (pAcPhe) into recombinant proteins expressed in E. coli. Incorporation of pAcPhe is followed by chemoselective reaction of the ketone side chain with a hydroxylamine-functionalized nitroxide to afford the spin-labeled side chain “K1,” and we present two protocols for successful K1 labeling of proteins bearing site-specific pAcPhe. We outline the basic requirements for pAcPhe incorporation and labeling, with an emphasis on practical aspects that must be considered by the researcher if high yields of UAA incorporation and efficient labeling reactions are to be achieved. To this end, we highlight recent advances that have led to increased yields of pAcPhe incorporation, and discuss the use of aniline-based catalysts allowing for facile conjugation of the hydroxylamine spin label under mild reaction conditions. To illustrate the utility of K1 labeling in proteins where traditional cysteine-based SDSL methods are problematic, we site-specifically K1 label the cellular prion protein at two positions in the C-terminal domain and determine the interspin distance using double electron–electron resonance EPR. Recent advances in UAA incorporation and ketone-based bioconjugation, in combination with the commercial availability of all requisite

  11. Genetic Incorporation of the Unnatural Amino Acid p-Acetyl Phenylalanine into Proteins for Site-Directed Spin Labeling.

    PubMed

    Evans, Eric G B; Millhauser, Glenn L

    2015-01-01

    Site-directed spin labeling (SDSL) is a powerful tool for the characterization of protein structure and dynamics; however, its application in many systems is hampered by the reliance on unique and benign cysteine substitutions for the site-specific attachment of the spin label. An elegant solution to this problem involves the use of genetically encoded unnatural amino acids (UAAs) containing reactive functional groups that are chemically orthogonal to those of the 20 amino acids found naturally in proteins. These unique functional groups can then be selectively reacted with an appropriately functionalized spin probe. In this chapter, we detail the genetic incorporation of the ketone-bearing amino acid p-acetyl phenylalanine (pAcPhe) into recombinant proteins expressed in E. coli. Incorporation of pAcPhe is followed by chemoselective reaction of the ketone side chain with a hydroxylamine-functionalized nitroxide to afford the spin-labeled side chain "K1," and we present two protocols for successful K1 labeling of proteins bearing site-specific pAcPhe. We outline the basic requirements for pAcPhe incorporation and labeling, with an emphasis on practical aspects that must be considered by the researcher if high yields of UAA incorporation and efficient labeling reactions are to be achieved. To this end, we highlight recent advances that have led to increased yields of pAcPhe incorporation, and discuss the use of aniline-based catalysts allowing for facile conjugation of the hydroxylamine spin label under mild reaction conditions. To illustrate the utility of K1 labeling in proteins where traditional cysteine-based SDSL methods are problematic, we site-specifically K1 label the cellular prion protein at two positions in the C-terminal domain and determine the interspin distance using double electron-electron resonance EPR. Recent advances in UAA incorporation and ketone-based bioconjugation, in combination with the commercial availability of all requisite reagents

  12. Genetic Incorporation of the Unnatural Amino Acid p-Acetyl Phenylalanine into Proteins for Site-Directed Spin Labeling.

    PubMed

    Evans, Eric G B; Millhauser, Glenn L

    2015-01-01

    Site-directed spin labeling (SDSL) is a powerful tool for the characterization of protein structure and dynamics; however, its application in many systems is hampered by the reliance on unique and benign cysteine substitutions for the site-specific attachment of the spin label. An elegant solution to this problem involves the use of genetically encoded unnatural amino acids (UAAs) containing reactive functional groups that are chemically orthogonal to those of the 20 amino acids found naturally in proteins. These unique functional groups can then be selectively reacted with an appropriately functionalized spin probe. In this chapter, we detail the genetic incorporation of the ketone-bearing amino acid p-acetyl phenylalanine (pAcPhe) into recombinant proteins expressed in E. coli. Incorporation of pAcPhe is followed by chemoselective reaction of the ketone side chain with a hydroxylamine-functionalized nitroxide to afford the spin-labeled side chain "K1," and we present two protocols for successful K1 labeling of proteins bearing site-specific pAcPhe. We outline the basic requirements for pAcPhe incorporation and labeling, with an emphasis on practical aspects that must be considered by the researcher if high yields of UAA incorporation and efficient labeling reactions are to be achieved. To this end, we highlight recent advances that have led to increased yields of pAcPhe incorporation, and discuss the use of aniline-based catalysts allowing for facile conjugation of the hydroxylamine spin label under mild reaction conditions. To illustrate the utility of K1 labeling in proteins where traditional cysteine-based SDSL methods are problematic, we site-specifically K1 label the cellular prion protein at two positions in the C-terminal domain and determine the interspin distance using double electron-electron resonance EPR. Recent advances in UAA incorporation and ketone-based bioconjugation, in combination with the commercial availability of all requisite reagents

  13. Shikimate and Phenylalanine Biosynthesis in the Green Lineage

    PubMed Central

    Tohge, Takayuki; Watanabe, Mutsumi; Hoefgen, Rainer; Fernie, Alisdair R.

    2013-01-01

    The shikimate pathway provides carbon skeletons for the aromatic amino acids l-tryptophan, l-phenylalanine, and l-tyrosine. It is a high flux bearing pathway and it has been estimated that greater than 30% of all fixed carbon is directed through this pathway. These combined pathways have been subjected to considerable research attention due to the fact that mammals are unable to synthesize these amino acids and the fact that one of the enzymes of the shikimate pathway is a very effective herbicide target. However, in addition to these characteristics these pathways additionally provide important precursors for a wide range of important secondary metabolites including chlorogenic acid, alkaloids, glucosinolates, auxin, tannins, suberin, lignin and lignan, tocopherols, and betalains. Here we review the shikimate pathway of the green lineage and compare and contrast its evolution and ubiquity with that of the more specialized phenylpropanoid metabolism which this essential pathway fuels. PMID:23543266

  14. Siglec-5 (CD170) can mediate inhibitory signaling in the absence of immunoreceptor tyrosine-based inhibitory motif phosphorylation.

    PubMed

    Avril, Tony; Freeman, Sylvie D; Attrill, Helen; Clarke, Rosemary G; Crocker, Paul R

    2005-05-20

    Siglec-5 (CD170) is a member of the recently described human CD33-related siglec subgroup of sialic acid binding Ig-like lectins and is expressed on myeloid cells of the hemopoietic system. Similar to other CD33-related siglecs, Siglec-5 contains two tyrosine-based motifs in its cytoplasmic tail implicated in signaling functions. To investigate the role of these motifs in Siglec-5-dependent signaling, we used transfected rat basophil leukemia cells as a model system. Tyrosine phosphorylation of Siglec-5 led to recruitment of the tyrosine phosphatases SHP-1 and SHP-2, as seen in both pull-down assays and microscopy. Siglec-5 could efficiently inhibit FcepsilonRI-mediated calcium fluxing and serotonin release after co-cross-linking. Surprisingly, a double tyrosine to alanine mutant of Siglec-5 could still mediate strong inhibition of serotonin release in the absence of detectable tyrosine phosphorylation, whereas a double tyrosine to phenylalanine mutant lost all inhibitory activity. In comparison, suppression of Siglec-5-dependent adhesion to red blood cells was reversed by either tyrosine to alanine or tyrosine to phenylalanine mutations of the membrane proximal tyrosine-based motif. Using an in vitro phosphatase assay with synthetic and recombinant forms of the cytoplasmic tail, it was shown that a double alanine mutant of Siglec-5 had weak, but significant SHP-1 activating properties similar to those of wild type, non-phosphorylated cytoplasmic tail, whereas a double phenylalanine mutant was inactive. These findings establish that Siglec-5 can be classified as an inhibitory receptor with the potential to mediate SHP-1 and/or SHP-2-dependent signaling in the absence of tyrosine phosphorylation. PMID:15769739

  15. Disturbed Amino Acid Metabolism in HIV: Association with Neuropsychiatric Symptoms

    PubMed Central

    Gostner, Johanna M.; Becker, Kathrin; Kurz, Katharina; Fuchs, Dietmar

    2015-01-01

    Blood levels of the amino acid phenylalanine, as well as of the tryptophan breakdown product kynurenine, are found to be elevated in human immunodeficiency virus type 1 (HIV-1)-infected patients. Both essential amino acids, tryptophan and phenylalanine, are important precursor molecules for neurotransmitter biosynthesis. Thus, dysregulated amino acid metabolism may be related to disease-associated neuropsychiatric symptoms, such as development of depression, fatigue, and cognitive impairment. Increased phenylalanine/tyrosine and kynurenine/tryptophan ratios are associated with immune activation in patients with HIV-1 infection and decrease upon effective antiretroviral therapy. Recent large-scale metabolic studies have confirmed the crucial involvement of tryptophan and phenylalanine metabolism in HIV-associated disease. Herein, we summarize the current status of the role of tryptophan and phenylalanine metabolism in HIV disease and discuss how inflammatory stress-associated dysregulation of amino acid metabolism may be part of the pathophysiology of common HIV-associated neuropsychiatric conditions. PMID:26236243

  16. From scratch to value: engineering Escherichia coli wild type cells to the production of L-phenylalanine and other fine chemicals derived from chorismate.

    PubMed

    Sprenger, Georg A

    2007-06-01

    Recombinant strains of Escherichia coli K-12 for the production of the three aromatic amino acids (L-phenylalanine, L-tryptophan, L-tyrosine) have been constructed. The largest demand is for L-phenylalanine (L-Phe), as it can be used as a building block for the low-calorie sweetener, aspartame. Besides L-Phe, an increasing number of shikimic acid pathway intermediates can be produced from appropriate E. coli mutants with blocks in this pathway. The last common intermediate, chorismate, in E. coli not only serves for production of aromatic amino acids but can also be used for high-titer production of non-aromatic compounds, e.g., cyclohexadiene-transdiols. In an approach to diversity-oriented metabolic engineering (metabolic grafting), platform strains with increased flux through the general aromatic pathway were created by suitable gene deletions, additions, or rearrangements. Examples for rational strain constructions for L-phenylalanine and chorismate derivatives are given with emphasis on genetic engineering. As a result, L-phenylalanine producers are available, which were derived through several defined steps from E. coli K-12 wild type. These mutant strains showed L-phenylalanine titers of up to 38 g/l of L-phenylalanine (and up to 45.5 g/l using in situ product recovery). Likewise, two cyclohexadiene-transdiols could be recovered.

  17. Expression Analysis of Phenylalanine Ammonia Lyase Gene and Rosmarinic Acid Production in Salvia officinalis and Salvia virgata Shoots Under Salicylic Acid Elicitation.

    PubMed

    Ejtahed, Roghayeh Sadat; Radjabian, Tayebeh; Hoseini Tafreshi, Sayed Ali

    2015-08-01

    Partial fragments of phenylalanine ammonia lyase (PAL) genes were cloned and characterized from Salvia officinalis (SoPAL) and Salvia virgata (SvPAL). Different concentrations (250 and 500 μM) of exogenous salicylic acid (SA) were used when correlation between PAL expression and rosmarinic acid (RA) accumulation was compared. The results showed that the deduced cDNA sequences of the partial genes had high similarities with those of known PAL gene from other plant species. Semi-quantitative reverse transcription PCR (RT-PCR) analysis revealed that exogenous application of SA led to up-regulating of the PAL expression. Further analysis showed that in S. virgata, at higher concentration of SA, higher accumulation of RA was achieved, while in S. officinalis, the higher RA accumulation was observed at lower concentration of SA. It was concluded that there was no positive correlation between the intensity of PAL transcription and the RA accumulation in the studied species. Therefore, despite of the increase in transcription rate of the PAL at the higher concentration of SA, the lower amounts of RA were accumulated in the case of S. officinalis. Consequently, the hypothesis that PAL is the rate-determining step in RA biosynthesis is not always valid and probably some other unknown factors participate in the synthesis of phenolics.

  18. Heterologous expression and characterization of tyrosine decarboxylase from Enterococcus faecalis R612Z1 and Enterococcus faecium R615Z1.

    PubMed

    Liu, Fang; Xu, Wenjuan; Du, Lihui; Wang, Daoying; Zhu, Yongzhi; Geng, Zhiming; Zhang, Muhan; Xu, Weimin

    2014-04-01

    Tyrosine decarboxylase (TDC) is responsible for tyramine production and can catalyze phenylalanine to produce β-phenylethylamine. Enterococcus strains are a group of bacteria predominantly producing tyramine and β-phenylethylamine in water-boiled salted duck. In this study, the heterologous expression and characterization of two TDCs from Enterococcus faecalis R612Z1 (612TDC) and Enterococcus faecium R615Z1 (615TDC) were studied. The recombinant putative proteins of 612TDC and 615TDC were heterologously expressed in Escherichia coli. 612TDC is a 620-amino-acid protein with a molecular mass of 70.0 kDa, whereas 615TDC is a 625-amino-acid protein with a molecular mass of 70.3 kDa. Both 612TDC and 615TDC showed an optimum temperature of 25 °C for the tyrosine and phenylalanine substrates. However, 612TDC revealed maximal activity at pH 5.5, whereas 615TDC revealed maximal activity at pH 6.0. Kinetic studies showed that 612TDC and 615TDC exhibited higher specificity for tyrosine than for phenylalanine. The catalysis abilities of both 612TDC and 615TDC for phenylalanine were restrained significantly with the increase in NaCl concentration, but this was not the case for tyrosine. This study revealed that the enzyme properties of the purified recombinant 612TDC and 615TDC were similar, although their amino acid sequences had 84% identity. PMID:24680070

  19. 4-Quinolone-3-carboxylic acids as cell-permeable inhibitors of protein tyrosine phosphatase 1B.

    PubMed

    Zhi, Ying; Gao, Li-Xin; Jin, Yi; Tang, Chun-Lan; Li, Jing-Ya; Li, Jia; Long, Ya-Qiu

    2014-07-15

    Protein tyrosine phosphatase 1B is a negative regulator in the insulin and leptin signaling pathways, and has emerged as an attractive target for the treatment of type 2 diabetes and obesity. However, the essential pharmacophore of charged phosphotyrosine or its mimetic confer low selectivity and poor cell permeability. Starting from our previously reported aryl diketoacid-based PTP1B inhibitors, a drug-like scaffold of 4-quinolone-3-carboxylic acid was introduced for the first time as a novel surrogate of phosphotyrosine. An optimal combination of hydrophobic groups installed at C-6, N-1 and C-3 positions of the quinolone motif afforded potent PTP1B inhibitors with low micromolar IC50 values. These 4-quinolone-3-carboxylate based PTP1B inhibitors displayed a 2-10 fold selectivity over a panel of PTP's. Furthermore, the bidentate inhibitors of 4-quinolone-3-carboxylic acids conjugated with aryl diketoacid or salicylic acid were cell permeable and enhanced insulin signaling in CHO/hIR cells. The kinetic studies and molecular modeling suggest that the 4-quinolone-3-carboxylates act as competitive inhibitors by binding to the PTP1B active site in the WPD loop closed conformation. Taken together, our study shows that the 4-quinolone-3-carboxylic acid derivatives exhibit improved pharmacological properties over previously described PTB1B inhibitors and warrant further preclinical studies.

  20. Interferon signaling is dependent on specific tyrosines located within the intracellular domain of IFNAR2c. Expression of IFNAR2c tyrosine mutants in U5A cells.

    PubMed

    Wagner, T Charis; Velichko, Sharlene; Vogel, David; Rani, M R Sandhya; Leung, Stewart; Ransohoff, Richard M; Stark, George R; Perez, H Daniel; Croze, Ed

    2002-01-11

    Type I interferons (IFNs) are cytokines that play a central role in mediating antiviral, antiproliferative, and immunomodulatory activities in virtually all cells. These activities are entirely dependent on the interaction of IFNs with their particular cell surface receptor. In this report, we identify two specific tyrosine residues located within the cytoplasmic domain of IFNAR2c that are obligatory for IFN-dependent signaling. Various IFNAR2c tyrosine mutants were expressed in a human lung fibroscarcoma cell line lacking IFNAR2c (U5A). Stable clones expressing these mutants were analyzed for their ability to induce STAT1 and STAT2 activation, ISGF3 transcriptional complex formation, gene expression, and cell growth regulation in response to stimulation with type I IFNs. The replacement of all seven cytoplasmic tyrosine residues of IFNAR2c with phenylalanine resulted in a receptor unable to respond to IFN stimulation. Substitution of single tyrosines at amino acid residue 269, 316, 318, 337, or 512 with phenylalanine had no effect on IFN-dependent signaling, suggesting that no single tyrosine is essential for IFN receptor-mediated signaling. In addition, IFNAR2c retaining five proximal tyrosines residues (269, 306, 316, 318, and 337) or either two distal tyrosine residues (411 or 512) continued to be responsive to IFN stimulation. Surprisingly, the presence of only a single tyrosine at either position 337 or 512 was sufficient to restore a complete IFN response. These results indicate that IFN-dependent signaling proceeds through the redundant usage of two tyrosine residues in the cytoplasmic domain of IFNAR2c.

  1. [Change in the content of salicylic acid and activities of phenylalanine ammonia-lyase and catalase in wheat seedling roots under the influence of Azospirilium lectins].

    PubMed

    Alen'kina, S A; Trutneva, K A; Nikitina, V E

    2013-01-01

    The time course of changes in the endogenous content of salicylic acid, the ratio between the acid's free and bound forms, and changes in the activities of phenylalanine ammonia-lyase and catalase in wheat seedling roots under the effect of lectins of two strains of the associative nitrogen-fixing bacterium Azospirillum (A. brasilense Sp7 and its mutant defective in lectin activity, A. brasilense Sp7.2.3) is investigated. Differences in plant response to the action of the lectins from these two strains are established. On the basis of the obtained data, a model is proposed for lectin-assisted induction of resistance, according to which the lectin effect on the roots of seedlings results in the accumulation of free salicylic acid, which inhibits catalase activity, ultimately leading to accumulation of hydrogen peroxide and formation of induced resistance. PMID:25518563

  2. [Change in the content of salicylic acid and activities of phenylalanine ammonia-lyase and catalase in wheat seedling roots under the influence of Azospirilium lectins].

    PubMed

    Alen'kina, S A; Trutneva, K A; Nikitina, V E

    2013-01-01

    The time course of changes in the endogenous content of salicylic acid, the ratio between the acid's free and bound forms, and changes in the activities of phenylalanine ammonia-lyase and catalase in wheat seedling roots under the effect of lectins of two strains of the associative nitrogen-fixing bacterium Azospirillum (A. brasilense Sp7 and its mutant defective in lectin activity, A. brasilense Sp7.2.3) is investigated. Differences in plant response to the action of the lectins from these two strains are established. On the basis of the obtained data, a model is proposed for lectin-assisted induction of resistance, according to which the lectin effect on the roots of seedlings results in the accumulation of free salicylic acid, which inhibits catalase activity, ultimately leading to accumulation of hydrogen peroxide and formation of induced resistance.

  3. Substrates and enzyme activities related to biotransformation of resveratrol from phenylalanine by Alternaria sp. MG1.

    PubMed

    Zhang, Jinhua; Shi, Junling; Liu, Yanlin

    2013-12-01

    To identify the substrates and enzymes related to resveratrol biosynthesis in Alternaria sp. MG1, different substrates were used to produce resveratrol, and their influence on resveratrol production was analyzed using high performance liquid chromatography (HPLC). Formation of resveratrol and related intermediates was identified using mass spectrum. During the biotransformation, activities of related enzymes, including phenylalanine ammonia-lyase (PAL), trans-cinnamate 4-hydroxylase (C4H), and 4-coumarate-CoA ligase (4CL), were analyzed and tracked. The reaction system contained 100 mL 0.2 mol/L phosphate buffer (pH 6.5), 120 g/L Alternaria sp. MG1 cells, 0.1 g/L MgSO₄, and 0.2 g/L CaSO₄ and different substrates according to the experimental design. The biotransformation was carried out for 21 h at 28 °C and 120 rpm. Resveratrol formation was identified when phenylalanine, tyrosine, cinnamic acid, and p-coumaric acid were separately used as the only substrate. Accumulation of cinnamic acid, p-coumaric acid, and resveratrol and the activities of PAL, C4H, and 4CL were identified and changed in different trends during transformation with phenylalanine as the only substrate. The addition of carbohydrates and the increase of phenylalanine concentration promoted resveratrol production and yielded the highest value (4.57 μg/L) when 2 g/L glucose, 1 g/L cyclodextrin, and phenylalanine (4.7 mmol/L) were used simultaneously.

  4. Molecular cloning and promoter analysis of the specific salicylic acid biosynthetic pathway gene phenylalanine ammonia-lyase (AaPAL1) from Artemisia annua.

    PubMed

    Zhang, Ying; Fu, Xueqing; Hao, Xiaolong; Zhang, Lida; Wang, Luyao; Qian, Hongmei; Zhao, Jingya

    2016-07-01

    Phenylalanine ammonia-lyase (PAL) is the key enzyme in the biosynthetic pathway of salicylic acid (SA). In this study, a full-length cDNA of PAL gene (named as AaPAL1) was cloned from Artemisia annua. The gene contains an open reading frame of 2,151 bps encoding 716 amino acids. Comparative and bioinformatics analysis revealed that the polypeptide protein of AaPAL1 was highly homologous to PALs from other plant species. Southern blot analysis revealed that it belonged to a gene family with three members. Quantitative RT-PCR analysis of various tissues of A. annua showed that AaPAL1 transcript levels were highest in the young leaves. A 1160-bp promoter region was also isolated resulting in identification of distinct cis-regulatory elements including W-box, TGACG-motif, and TC-rich repeats. Quantitative RT-PCR indicated that AaPAL1 was upregulated by salinity, drought, wounding, and SA stresses, which were corroborated positively with the identified cis-elements within the promoter region. AaPAL1 was successfully expressed in Escherichia. coli and the enzyme activity of the purified AaPAL1 was approximately 287.2 U/mg. These results substantiated the involvement of AaPAL1 in the phenylalanine pathway. PMID:26040426

  5. Production of phenylpyruvic acid from L-phenylalanine using an L-amino acid deaminase from Proteus mirabilis: comparison of enzymatic and whole-cell biotransformation approaches.

    PubMed

    Hou, Ying; Hossain, Gazi Sakir; Li, Jianghua; Shin, Hyun-Dong; Liu, Long; Du, Guocheng

    2015-10-01

    Phenylpyruvic acid (PPA) is an important organic acid that has a wide range of applications. In this study, the membrane-bound L-amino acid deaminase (L-AAD) gene from Proteus mirabilis KCTC 2566 was expressed in Escherichia coli BL21(DE3) and then the L-AAD was purified. After that, we used the purified enzyme and the recombinant E. coli whole-cell biocatalyst to produce PPA via a one-step biotransformation from L-phenylalanine. L-AAD was solubilized from the membrane and purified 52-fold with an overall yield of 13 %, which corresponded to a specific activity of 0.94 ± 0.01 μmol PPA min(-1)·mg(-1). Then, the biotransformation conditions for the pure enzyme and the whole-cell biocatalyst were optimized. The maximal production was 2.6 ± 0.1 g·L(-1) (specific activity of 1.02 ± 0.02 μmol PPA min(-1)·mg(-1) protein, 86.7 ± 5 % mass conversion rate, and 1.04 g·L(-1)·h(-1) productivity) and 3.3 ± 0.2 g L(-1) (specific activity of 0.013 ± 0.003 μmol PPA min(-1)·mg(-1) protein, 82.5 ± 4 % mass conversion rate, and 0.55 g·L(-1)·h(-1) productivity) for the pure enzyme and whole-cell biocatalyst, respectively. Comparative studies of the enzymatic and whole-cell biotransformation were performed in terms of specific activity, production, conversion, productivity, stability, need of external cofactors, and recycling. We have developed two eco-friendly and efficient approaches for PPA production. The strategy described herein may aid the biotransformational synthesis of other α-keto acids from their corresponding amino acids.

  6. Conformational Structure of Tyrosine, Tyrosyl-Glycine, and Tyrosyl-Glycyl-Glycine by Double Resonance Spectroscopy

    NASA Technical Reports Server (NTRS)

    Abo-Riziq, Ali; Grace, Louis; Crews, Bridgit; Callahan, Michael P,; van Mourik, Tanja; de Vries, Mattanjah S,

    2011-01-01

    We investigated the variation in conformation for the amino acid tyrosine (Y), alone and in the small peptides tyrosine-glycine (YC) and tyrosine-glycine-glycine (YGG), in the gas phase by using UV-UV and IR-UV double resonance spectroscopy and density functional theory calculations. For tyrosine we found seven different conformations, for YG we found four different conformations, and for YGG we found three different conformations. As the peptides get larger, we observe fewer stable conformers, despite the increasing complexity and number of degrees of freedom. We find structural trends similar to those in phenylalanine-glycine glycine (FGG) and tryptophan-glycine-glycine (WGG)j however) the effect of dispersive forces in FGG for stabilizing a folded structure is replaced by that of hydrogen bonding in YGG.

  7. Kinetic mechanism of phenylalanine hydroxylase: intrinsic binding and rate constants from single-turnover experiments.

    PubMed

    Roberts, Kenneth M; Pavon, Jorge Alex; Fitzpatrick, Paul F

    2013-02-12

    Phenylalanine hydroxylase (PheH) catalyzes the key step in the catabolism of dietary phenylalanine, its hydroxylation to tyrosine using tetrahydrobiopterin (BH(4)) and O(2). A complete kinetic mechanism for PheH was determined by global analysis of single-turnover data in the reaction of PheHΔ117, a truncated form of the enzyme lacking the N-terminal regulatory domain. Formation of the productive PheHΔ117-BH(4)-phenylalanine complex begins with the rapid binding of BH(4) (K(d) = 65 μM). Subsequent addition of phenylalanine to the binary complex to form the productive ternary complex (K(d) = 130 μM) is approximately 10-fold slower. Both substrates can also bind to the free enzyme to form inhibitory binary complexes. O(2) rapidly binds to the productive ternary complex; this is followed by formation of an unidentified intermediate, which can be detected as a decrease in absorbance at 340 nm, with a rate constant of 140 s(-1). Formation of the 4a-hydroxypterin and Fe(IV)O intermediates is 10-fold slower and is followed by the rapid hydroxylation of the amino acid. Product release is the rate-determining step and largely determines k(cat). Similar reactions using 6-methyltetrahydropterin indicate a preference for the physiological pterin during hydroxylation.

  8. Simultaneous and selective decarboxylation of L-serine and deamination of L-phenylalanine in an amino acid mixture--a means of separating amino acids for synthesizing biobased chemicals.

    PubMed

    Teng, Yinglai; Scott, Elinor L; Witte-van Dijk, Susan C M; Sanders, Johan P M

    2016-01-25

    Amino acids (AAs) obtained from the hydrolysis of biomass-derived proteins are interesting feedstocks for the chemical industry. They can be prepared from the byproduct of biofuel production and agricultural wastes. They are rich in functionalities needed in petrochemicals, providing the opportunity to save energy, reagents, and process steps. However, their separation is required before they can be applied for further applications. Electrodialysis (ED) is a promising separation method, but its efficiency needs to be improved when separating AAs with similar isoelectric points. Thus, specific conversions are required to form product with different charges. Here we studied the enzymatic conversions which can be used as a means to aid the ED separation of neutral AAs. A model mixture containing L-serine, L-phenylalanine and L-methionine was used. The reactions of L-serine decarboxylase and L-phenylalanine ammonia-lyase were employed to specifically convert serine and phenylalanine into ethanolamine and trans-cinnamic acid. At the isoelectric point of methionine (pH 5.74), the charge of ethanolamine and trans-cinnamic acid are +1 and -1, therefore facilitating potential separation into three different streams by electrodialysis. Here the enzyme kinetics, specificity, inhibition and the operational stabilities were studied, showing that both enzymes can be applied simultaneously to aid the ED separation of neutral AAs. PMID:25976628

  9. Simultaneous and selective decarboxylation of L-serine and deamination of L-phenylalanine in an amino acid mixture--a means of separating amino acids for synthesizing biobased chemicals.

    PubMed

    Teng, Yinglai; Scott, Elinor L; Witte-van Dijk, Susan C M; Sanders, Johan P M

    2016-01-25

    Amino acids (AAs) obtained from the hydrolysis of biomass-derived proteins are interesting feedstocks for the chemical industry. They can be prepared from the byproduct of biofuel production and agricultural wastes. They are rich in functionalities needed in petrochemicals, providing the opportunity to save energy, reagents, and process steps. However, their separation is required before they can be applied for further applications. Electrodialysis (ED) is a promising separation method, but its efficiency needs to be improved when separating AAs with similar isoelectric points. Thus, specific conversions are required to form product with different charges. Here we studied the enzymatic conversions which can be used as a means to aid the ED separation of neutral AAs. A model mixture containing L-serine, L-phenylalanine and L-methionine was used. The reactions of L-serine decarboxylase and L-phenylalanine ammonia-lyase were employed to specifically convert serine and phenylalanine into ethanolamine and trans-cinnamic acid. At the isoelectric point of methionine (pH 5.74), the charge of ethanolamine and trans-cinnamic acid are +1 and -1, therefore facilitating potential separation into three different streams by electrodialysis. Here the enzyme kinetics, specificity, inhibition and the operational stabilities were studied, showing that both enzymes can be applied simultaneously to aid the ED separation of neutral AAs.

  10. Dual role for the tyrosine decarboxylation pathway in Enterococcus faecium E17: response to an acid challenge and generation of a proton motive force.

    PubMed

    Pereira, C I; Matos, D; San Romão, M V; Crespo, M T Barreto

    2009-01-01

    In this work we investigated the role of the tyrosine decarboxylation pathway in the response of Enterococcus faecium E17 cells to an acid challenge. It was found that 91% of the cells were able to remain viable in the presence of tyrosine when they were incubated for 3 h in a complex medium at pH 2.5. This effect was shown to be related to the tyrosine decarboxylation pathway. Therefore, the role of tyrosine decarboxylation in pH homeostasis was studied. The membrane potential and pH gradient, the parameters that compose the proton motive force (PMF), were measured at different pHs (pH 4.5 to 7). We obtained evidence showing that the tyrosine decarboxylation pathway generates a PMF composed of a pH gradient formed due to proton consumption in the decarboxylation reaction and by a membrane potential which results from electrogenic transport of tyrosine in exchange for the corresponding biogenic amine tyramine. The properties of the tyrosine transporter were also studied in this work by using whole cells and right-side-out vesicles. The results showed that the transporter catalyzes homologous tyrosine/tyrosine antiport, as well as electrogenic heterologous tyrosine-tyramine exchange. The tyrosine transporter had properties of a typical precursor-product exchanger operating in a proton motive decarboxylation pathway. Therefore, the tyrosine decarboxylation pathway contributes to an acid response mechanism in E. faecium E17. This decarboxylation pathway gives the strain a competitive advantage in nutrient-depleted conditions, as well as in harsh acidic environments, and a better chance of survival, which contributes to higher cell counts in food fermentation products. PMID:19011061

  11. Stress enhances the gene expression and enzyme activity of phenylalanine ammonia-lyase and the endogenous content of salicylic acid to induce flowering in pharbitis.

    PubMed

    Wada, Kaede C; Mizuuchi, Kaori; Koshio, Aya; Kaneko, Kentaro; Mitsui, Toshiaki; Takeno, Kiyotoshi

    2014-07-01

    The involvement of salicylic acid (SA) in the regulation of stress-induced flowering in the short-day plant pharbitis (also called Japanese morning glory) Ipomoea nil (formerly Pharbitis nil) was studied. Pharbitis cv. Violet was induced to flower when grown in 1/100-strength mineral nutrient solution under non-inductive long-day conditions. All fully expanded true leaves were removed from seedlings, leaving only the cotyledons, and flowering was induced under poor-nutrition stress conditions. This indicates that cotyledons can play a role in the regulation of poor-nutrition stress-induced flowering. The expression of the pharbitis homolog of PHENYLALANINE AMMONIA-LYASE, the enzyme activity of phenylalanine ammonia-lyase (PAL; E.C. 4.3.1.5) and the content of SA in the cotyledons were all up-regulated by the stress treatment. The Violet was also induced to flower by low-temperature stress, DNA demethylation and short-day treatment. Low-temperature stress enhanced PAL activity, whereas non-stress factors such as DNA demethylation and short-day treatment decreased the activity. The PAL enzyme activity was also examined in another cultivar, Tendan, obtaining similar results to Violet. The exogenously applied SA did not induce flowering under non-stress conditions but did promote flowering under weak stress conditions in both cultivars. These results suggest that stress-induced flowering in pharbitis is induced, at least partly, by SA, and the synthesis of SA is promoted by PAL.

  12. Domain Movements upon Activation of Phenylalanine Hydroxylase Characterized by Crystallography and Chromatography-Coupled Small-Angle X-ray Scattering.

    PubMed

    Meisburger, Steve P; Taylor, Alexander B; Khan, Crystal A; Zhang, Shengnan; Fitzpatrick, Paul F; Ando, Nozomi

    2016-05-25

    Mammalian phenylalanine hydroxylase (PheH) is an allosteric enzyme that catalyzes the first step in the catabolism of the amino acid phenylalanine. Following allosteric activation by high phenylalanine levels, the enzyme catalyzes the pterin-dependent conversion of phenylalanine to tyrosine. Inability to control elevated phenylalanine levels in the blood leads to increased risk of mental disabilities commonly associated with the inherited metabolic disorder, phenylketonuria. Although extensively studied, structural changes associated with allosteric activation in mammalian PheH have been elusive. Here, we examine the complex allosteric mechanisms of rat PheH using X-ray crystallography, isothermal titration calorimetry (ITC), and small-angle X-ray scattering (SAXS). We describe crystal structures of the preactivated state of the PheH tetramer depicting the regulatory domains docked against the catalytic domains and preventing substrate binding. Using SAXS, we further describe the domain movements involved in allosteric activation of PheH in solution and present the first demonstration of chromatography-coupled SAXS with Evolving Factor Analysis (EFA), a powerful method for separating scattering components in a model-independent way. Together, these results support a model for allostery in PheH in which phenylalanine stabilizes the dimerization of the regulatory domains and exposes the active site for substrate binding and other structural changes needed for activity. PMID:27145334

  13. Mechanistic, Mutational, and Structural Evaluation of a Taxus Phenylalanine Aminomutase

    SciTech Connect

    Feng, Lei; Wanninayake, Udayanga; Strom, Susan; Geiger, James; Walker, Kevin D.

    2014-10-02

    The structure of a phenylalanine aminomutase (TcPAM) from Taxus canadensis has been determined at 2.4 {angstrom} resolution. The active site of the TcPAM contains the signature 4-methylidene-1H-imidazol-5(4H)-one prosthesis, observed in all catalysts of the class I lyase-like family. This catalyst isomerizes (S)-{alpha}-phenylalanine to the (R)-{beta}-isomer by exchange of the NH{sub 2}/H pair. The stereochemistry of the TcPAM reaction product is opposite of the (S)-{beta}-tyrosine made by the mechanistically related tyrosine aminomutase (SgTAM) from Streptomyces globisporus. Since TcPAM and SgTAM share similar tertiary- and quaternary-structures and have several highly conserved aliphatic residues positioned analogously in their active sites for substrate recognition, the divergent product stereochemistries of these catalysts likely cannot be explained by differences in active site architecture. The active site of the TcPAM structure also is in complex with (E)-cinnamate; the latter functions as both a substrate and an intermediate. To account for the distinct (3R)-{beta}-amino acid stereochemistry catalyzed by TcPAM, the cinnamate skeleton must rotate the C{sub 1}-C{sub {alpha}} and C{sub ipso}-C{sub {beta}} bonds 180{sup o} in the active site prior to exchange and rebinding of the NH{sub 2}/H pair to the cinnamate, an event that is not required for the corresponding acrylate intermediate in the SgTAM reaction. Moreover, the aromatic ring of the intermediate makes only one direct hydrophobic interaction with Leu-104. A L104A mutant of TcPAM demonstrated an 1.5-fold increase in k{sub cat} and a decrease in K{sub M} values for sterically demanding 3'-methyl-{alpha}-phenylalanine and styryl-{alpha}-alanine substrates, compared to the kinetic parameters for TcPAM. These parameters did not change significantly for the mutant with 4'-methyl-{alpha}-phenylalanine compared to those for TcPAM.

  14. A root-expressed L-phenylalanine:4-hydroxyphenylpyruvate aminotransferase is required for tropane alkaloid biosynthesis in Atropa belladonna.

    PubMed

    Bedewitz, Matthew A; Góngora-Castillo, Elsa; Uebler, Joseph B; Gonzales-Vigil, Eliana; Wiegert-Rininger, Krystle E; Childs, Kevin L; Hamilton, John P; Vaillancourt, Brieanne; Yeo, Yun-Soo; Chappell, Joseph; DellaPenna, Dean; Jones, A Daniel; Buell, C Robin; Barry, Cornelius S

    2014-09-01

    The tropane alkaloids, hyoscyamine and scopolamine, are medicinal compounds that are the active components of several therapeutics. Hyoscyamine and scopolamine are synthesized in the roots of specific genera of the Solanaceae in a multistep pathway that is only partially elucidated. To facilitate greater understanding of tropane alkaloid biosynthesis, a de novo transcriptome assembly was developed for Deadly Nightshade (Atropa belladonna). Littorine is a key intermediate in hyoscyamine and scopolamine biosynthesis that is produced by the condensation of tropine and phenyllactic acid. Phenyllactic acid is derived from phenylalanine via its transamination to phenylpyruvate, and mining of the transcriptome identified a phylogenetically distinct aromatic amino acid aminotransferase (ArAT), designated Ab-ArAT4, that is coexpressed with known tropane alkaloid biosynthesis genes in the roots of A. belladonna. Silencing of Ab-ArAT4 disrupted synthesis of hyoscyamine and scopolamine through reduction of phenyllactic acid levels. Recombinant Ab-ArAT4 preferentially catalyzes the first step in phenyllactic acid synthesis, the transamination of phenylalanine to phenylpyruvate. However, rather than utilizing the typical keto-acid cosubstrates, 2-oxoglutarate, pyruvate, and oxaloacetate, Ab-ArAT4 possesses strong substrate preference and highest activity with the aromatic keto-acid, 4-hydroxyphenylpyruvate. Thus, Ab-ArAT4 operates at the interface between primary and specialized metabolism, contributing to both tropane alkaloid biosynthesis and the direct conversion of phenylalanine to tyrosine.

  15. Amino acids and immune response: a role for cysteine, glutamine, phenylalanine, tryptophan and arginine in T-cell function and cancer?

    PubMed

    Sikalidis, Angelos K

    2015-01-01

    While proteins are critical for immunity, T-cells constitute a critical component of adaptive immunity by clearing cancerous cells among other abnormal cells. However, cancer cells exhibit a potential to escape T-cell control by employing mechanisms not completely delineated. Interesting work has investigated how certain amino acids affect the proliferation rate of T-cells as well as their effectiveness in clearing tumors. The role of amino acids cysteine, glutamine, phenylalanine, tryptophan and arginine in immunomodulation and particularly regarding T-cell proliferation and activation is discussed. The redox balance is reported to affect T-cell proliferation via modulation of cysteine availability. In addition antigen presenting cells (APCs), similar to myeloid cells determine the availability of amino acids in the extracellular microenvironment affecting T-cell proliferation and activation. A better mechanistic understanding of T-cell function modulation via amino acid signaling or metabolic properties may be helpful towards optimization of adaptive immunity with implications for cancer prognosis and treatment.

  16. Brain catechol synthesis - Control by brain tyrosine concentration

    NASA Technical Reports Server (NTRS)

    Wurtman, R. J.; Larin, F.; Mostafapour, S.; Fernstrom, J. D.

    1974-01-01

    Brain catechol synthesis was estimated by measuring the rate at which brain dopa levels rose following decarboxylase inhibition. Dopa accumulation was accelerated by tyrosine administration, and decreased by treatments that lowered brain tyrosine concentrations (for example, intraperitoneal tryptophan, leucine, or parachlorophenylalanine). A low dose of phenylalanine elevated brain tyrosine without accelerating dopa synthesis. Our findings raise the possibility that nutritional and endocrine factors might influence brain catecholamine synthesis by controlling the availability of tyrosine.

  17. Direct synthesis of unprotected 4-aryl phenylalanines via the Suzuki reaction under microwave irradiation.

    PubMed

    Gong, Yong; He, Wei

    2002-10-31

    [formula: see text] 4-Aryl phenylalanines were prepared as free amino acids from the Suzuki coupling of 4-borono phenylalanine with aryl halides in high yields within 5-10 min under microwave irradiation.

  18. Interactions in L-phenylalanine/L-leucine/L-glutamic Acid/L-proline + 2 M aqueous NaCl/2 M NaNO3 systems at different temperatures

    NASA Astrophysics Data System (ADS)

    Riyazuddeen, Imran Khan; Afrin, Sadaf

    2012-12-01

    Density (ρ) and speed of sound ( u) in 2 M aqueous NaCl and 2 M NaNO3 solutions of amino acids: L-phenylalanine, L-leucine, L-glutamic acid, and L-proline have been measured for several molal concentrations of amino acids at different temperatures. The ρ and u data have been used to calculate the values of isothermal compressibility and internal pressure at different temperatures. The trends of variations of κ T and P i with an increase in molal concentration of amino acid and temperature have been discussed in terms of solute-solvent and solute-solute interactions in the systems.

  19. Poly(carbonate–amide)s Derived from Bio-Based Resources: Poly(ferulic acid-co-tyrosine)

    PubMed Central

    2015-01-01

    Ferulic acid (FA), a bio-based resource found in fruits and vegetables, was coupled with a hydroxyl-amino acid to generate a new class of monomers to afford poly(carbonate–amide)s with potential to degrade into natural products. l-Serine was first selected as the hydroxyl-amino partner for FA, from which the activated p-nitrophenyl carbonate monomer was synthesized. Unfortunately, polymerizations were unsuccessful, and the elimination product was systematically obtained. To avoid elimination, we revised our strategy and used l-tyrosine ethyl ester, which lacks an acidic proton on the α position of the ethyl ester. Four new monomers were synthesized and converted into the corresponding poly(carbonate–amide)s with specific regioselectivities. The polymers were fully characterized through thermal and spectroscopic analyses. Preliminary fluorescent studies revealed interesting photophysical properties for the monomers and their corresponding poly(carbonate–amide)s, beyond the fluorescence characteristics of l-tyrosine and FA, making these materials potentially viable for sensing and/or imaging applications, in addition to their attractiveness as engineering materials derived from renewable resources. PMID:24839309

  20. Molecular Docking Study of Catecholamines and [4-(Propan-2-yl) Phenyl]Carbamic acid with Tyrosine Hydroxylase

    PubMed Central

    Parveen, Zahida; Nawaz, Muhammad Sulaman; Shakil, Shazi; Greig, Nigel H.; Kamal, Mohammad A.

    2016-01-01

    Parkinson’s disease is a major age-related neurodegenerative disorder. As the classical disease-related motor symptoms are associated with the loss of dopamine-generating cells within the substantia nigra, tyrosine hydroxylase (TH), the rate-limiting enzyme in the synthesis of catecholamines has become an important target in the development of Parkinson’s disease drug candidates, with the focus to augment TH levels or its activity. By contrast, TH inhibitors are of relevance in the treatment of conditions associated with catecholamine over-production, as occurs in pheochromocytomas. To aid characterizing new drug candidates, a molecular docking study of catecholamines and a novel hypothetical compound [4-(propan-2-yl) phenyl]carbamic acid (PPCA) with TH is described. Docking was performed using Autodock4.2 and results were analyzed using Chimera1.5.2. All the studied ligands were found to bind within a deep narrow groove lined with polar aromatic and acidic residues within TH. Our results corroborated a ‘hexa interacting amino acids unit’ located in this deep narrow groove crucial to the interaction of PPCA and the studied catecholamines with TH, whereby the ‘His361-His336 dyad’ was found to be even more crucial to these binding interactions. PPCA displayed a binding interaction with human TH that was comparable to the original TH substrate, L-tyrosine. Hence PPCA may warrant in vitro and in vivo characterization with TH to assess its potential as a candidate therapeutic. PMID:22583429

  1. Oxidation of the aromatic amino acids tryptophan and tyrosine disrupts their anabolic effects on bone marrow mesenchymal stem cells.

    PubMed

    El Refaey, Mona; Watkins, Christopher P; Kennedy, Eileen J; Chang, Andrew; Zhong, Qing; Ding, Ke-Hong; Shi, Xing-ming; Xu, Jianrui; Bollag, Wendy B; Hill, William D; Johnson, Maribeth; Hunter, Monte; Hamrick, Mark W; Isales, Carlos M

    2015-07-15

    Age-induced bone loss is associated with greater bone resorption and decreased bone formation resulting in osteoporosis and osteoporosis-related fractures. The etiology of this age-induced bone loss is not clear but has been associated with increased generation of reactive oxygen species (ROS) from leaky mitochondria. ROS are known to oxidize/damage the surrounding proteins/amino acids/enzymes and thus impair their normal function. Among the amino acids, the aromatic amino acids are particularly prone to modification by oxidation. Since impaired osteoblastic differentiation from bone marrow mesenchymal stem cells (BMMSCs) plays a role in age-related bone loss, we wished to examine whether oxidized amino acids (in particular the aromatic amino acids) modulated BMMSC function. Using mouse BMMSCs, we examined the effects of the oxidized amino acids di-tyrosine and kynurenine on proliferation, differentiation and Mitogen-Activated Protein Kinase (MAPK) pathway. Our data demonstrate that amino acid oxides (in particular kynurenine) inhibited BMMSC proliferation, alkaline phosphatase expression and activity and the expression of osteogenic markers (Osteocalcin and Runx2). Taken together, our data are consistent with a potential pathogenic role for oxidized amino acids in age-induced bone loss.

  2. Interaction of bacterial fatty-acid-displaced regulators with DNA is interrupted by tyrosine phosphorylation in the helix-turn-helix domain

    PubMed Central

    Derouiche, Abderahmane; Bidnenko, Vladimir; Grenha, Rosa; Pigonneau, Nathalie; Ventroux, Magali; Franz-Wachtel, Mirita; Nessler, Sylvie; Noirot-Gros, Marie-Françoise; Mijakovic, Ivan

    2013-01-01

    Bacteria possess transcription regulators (of the TetR family) specifically dedicated to repressing genes for cytochrome P450, involved in oxidation of polyunsaturated fatty acids. Interaction of these repressors with operator sequences is disrupted in the presence of fatty acids, and they are therefore known as fatty-acid-displaced regulators. Here, we describe a novel mechanism of inactivating the interaction of these proteins with DNA, illustrated by the example of Bacillus subtilis regulator FatR. FatR was found to interact in a two-hybrid assay with TkmA, an activator of the protein-tyrosine kinase PtkA. We show that FatR is phosphorylated specifically at the residue tyrosine 45 in its helix-turn-helix domain by the kinase PtkA. Structural modelling reveals that the hydroxyl group of tyrosine 45 interacts with DNA, and we show that this phosphorylation reduces FatR DNA binding capacity. Point mutants mimicking phosphorylation of FatR in vivo lead to a strong derepression of the fatR operon, indicating that this regulatory mechanism works independently of derepression by polyunsaturated fatty acids. Tyrosine 45 is a highly conserved residue, and PtkA from B. subtilis can phosphorylate FatR homologues from other bacteria. This indicates that phosphorylation of tyrosine 45 may be a general mechanism of switching off bacterial fatty-acid-displaced regulators. PMID:23939619

  3. Protein tyrosine kinase regulates α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor trafficking induced by acute hypoxia in cultured brainstem neurons.

    PubMed

    Wang, H; Yu, L C; Li, Y C

    2016-01-01

    This study was performed to investigate the modulation effect of protein tyrosine kinase on postsynaptic a-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor trafficking induced by acute hypoxia in cultured brainstem neurons. The cultured neurons were exposed to 1% O2 and the expression of AMPA receptor subunit GluR2 on the cell surface was significantly increased, while total GluR2 was not markedly changed. Furthermore, the hypoxia-induced increase in GluR2 expression on the cell surface was partially blocked by the protein tyrosine kinase membrane-permeable inhibitor genistein. In contrast, both the protein tyrosine kinase agonist nerve growth factor and protein tyrosine phosphatase inhibitor vanadate promoted the hypoxia-induced increase of GluR2 expression on cell surface. Moreover, GluR2 could be phosphorylated by tyrosine under normoxia and hypoxia conditions in vitro on brainstem neurons, and tyrosine phosphorylation of GluR2 was significantly stronger under hypoxia conditions. Our results indicate that acute hypoxia induces the AMPA receptor subunit GluR2 to rapidly migrate to the cell membrane to modify the strength of the synapse. This study indicates that tyrosine phosphorylation of the receptor is an important pathway regulating the rapid migration of GluR2 in the postsynaptic domain induced by hypoxia. PMID:27525851

  4. New Method for Determining Isotopic Values of Glutamic Acid and Phenylalanine for Estimation of Precise Trophic Position in Food Web Studies

    NASA Astrophysics Data System (ADS)

    Kamath, T.; Broek, T.; McCarthy, M.

    2012-12-01

    Compound Specific Isotope Analysis of Amino Acids (CSI-AA) has emerged as a highly precise new method of determining trophic levels of both aquatic and terrestrial organisms. Multiple studies have now shown that δ15N values for glutamic acid (Glu) and phenylalanine (Phe) can be coupled to provide extremely precise estimates of trophic position in diverse food web studies. The standard gas chromatography—isotope ratio mass spectrometer (GC-IRMS) approach is presently limited to a select number of labs since necessary equipment is both expensive and not widely accessible. Furthermore, typical GC-IRMS δ15N precision (±1‰) is significantly lower than usual bulk δ15N values (±0.1‰), thus presenting a considerable setback for precise trophic level calculations. In this study, we develop a new dual-column method to purify Glu and Phe using high performance liquid chromatography (HPLC). Phe is purified using an analytical scale reverse phase column embedded with anionic ion-pairing reagents and collected using automated fraction collection. Glu is separated from the non-polar amino acids using the same column and further purified using a hydrophilic interaction liquid chromatography (HILIC) cation and anion-exchange column and collected via automated fraction collection. Isotopic analysis of the purified AAs is then conducted on an elemental analyzer—isotope ratio mass spectrometer (EA-IRMS). As a test of this method, we present and compare the trophic position of five marine organisms—cyanobacteria, deep-sea bamboo coral, juvenile and adult white sea bass, and harbor seal, calculated using Glu and Phe δ15N values produced by both GC-IRMS and our HPLC-EA-IRMS approach. The preliminary results of this study suggest that the HPLC-EA-IRMS method is a viable alternative to GC-IRMS, which should allow accurate trophic position estimates to be made by more researchers using more readily available instrumentation.

  5. Metabolic engineering of the L-phenylalanine pathway in Escherichia coli for the production of S- or R-mandelic acid

    PubMed Central

    2011-01-01

    Background Mandelic acid (MA), an important component in pharmaceutical syntheses, is currently produced exclusively via petrochemical processes. Growing concerns over the environment and fossil energy costs have inspired a quest to develop alternative routes to MA using renewable resources. Herein we report the first direct route to optically pure MA from glucose via genetic modification of the L-phenylalanine pathway in E. coli. Results The introduction of hydroxymandelate synthase (HmaS) from Amycolatopsis orientalis into E. coli led to a yield of 0.092 g/L S-MA. By combined deletion of competing pathways, further optimization of S-MA production was achieved, and the yield reached 0.74 g/L within 24 h. To produce R-MA, hydroxymandelate oxidase (Hmo) from Streptomyces coelicolor and D-mandelate dehydrogenase (DMD) from Rhodotorula graminis were co-expressed in an S-MA-producing strain, and the resulting strain was capable of producing 0.68 g/L R-MA. Finally, phenylpyruvate feeding experiments suggest that HmaS is a potential bottleneck to further improvement in yields. Conclusions We have constructed E. coli strains that successfully accomplished the production of S- and R-MA directly from glucose. Our work provides the first example of the completely fermentative production of S- and R-MA from renewable feedstock. PMID:21910908

  6. L-leucine, L-methionine, and L-phenylalanine share a Na(+)/K (+)-dependent amino acid transporter in shrimp hepatopancreas.

    PubMed

    Duka, Ada; Ahearn, Gregory A

    2013-08-01

    Hepatopancreatic brush border membrane vesicles (BBMV), made from Atlantic White shrimp (Litopenaeus setiferus), were used to characterize the transport properties of (3)H-L-leucine influx by these membrane systems and how other essential amino acids and the cations, sodium and potassium, interact with this transport system. (3)H-L-leucine uptake by BBMV was pH-sensitive and occurred against transient transmembrane concentration gradients in both Na(+)- and K(+)-containing incubation media, suggesting that either cation was capable of providing a driving force for amino acid accumulation. (3)H-L-leucine uptake in NaCl or KCl media were each three times greater in acidic pH (pH 5.5) than in alkaline pH (pH 8.5). The essential amino acid, L-methionine, at 20 mM significantly (p < 0.0001) inhibited the 2-min uptakes of 1 mM (3)H-L-leucine in both Na(+)- and K(+)-containing incubation media. The residual (3)H-L-leucine uptake in the two media were significantly greater than zero (p < 0.001), but not significantly different from each other (p > 0.05) and may represent an L-methionine- and cation-independent transport system. (3)H-L-leucine influxes in both NaCl and KCl incubation media were hyperbolic functions of [L-leucine], following the carrier-mediated Michaelis-Menten equation. In NaCl, (3)H-L-leucine influx displayed a low apparent K M (high affinity) and low apparent J max, while in KCl the transport exhibited a high apparent K M (low affinity) and high apparent J max. L-methionine or L-phenylalanine (7 and 20 mM) were competitive inhibitors of (3)H-L-leucine influxes in both NaCl and KCl media, producing a significant (p < 0.01) increase in (3)H-L-leucine influx K M, but no significant response in (3)H-L-leucine influx J max. Potassium was a competitive inhibitor of sodium co-transport with (3)H-L-leucine, significantly (p < 0.01) increasing (3)H-L-leucine influx K M in the presence of sodium, but having negligible effect on (3)H-L-leucine influx J

  7. Aromatic/heterocyclic amino acids and the simulated sunlight-ultraviolet inactivation of the Heliothis/Helicoverpa baculovirus

    SciTech Connect

    Ignoffo, C.M.; Garcia, C.

    1995-04-01

    Tryptophan, of five aromatic/heterocyclic amino acids (tyrosine, phenylalanine, proline, histidine) provided significant protection of the Heliothis baculovirus (HzSNPV) from inactivation by simulated ultraviolet (SUV). Fifty percent of SUV protection of HzSNPV with tryptophan or tyrosine was obtained at 0.03 mg/ml and 0.5 mg/ml, respectively. Rates as high as 100.0 mg/ml of phenylalanine, histidine, or proline provided <50% protection. The extent of tryptophan protection was correlated with its absorption in the sunlight UV-B spectra. 16 refs., 2 tabs.

  8. A strategy for the study of cerebral amino acid transport using iodine-123-labeled amino acid radiopharmaceutical: 3-iodo-alpha-methyl-L-tyrosine

    SciTech Connect

    Kawai, K.; Fujibayashi, Y.; Saji, H.; Yonekura, Y.; Konishi, J.; Kubodera, A.; Yokoyama, A. )

    1991-05-01

    We examined the brain accumulation of iodine-123-iodo-alpha-methyl-L-tyrosine ({sup 123}I-L-AMT) in mice and rats. I-L-AMT showed high brain accumulation in mice, and in rats; rat brain uptake index exceeded that of {sup 14}C-L-tyrosine. The brain uptake index and the brain slice studies indicated the affinity of I-L-AMT for carrier-mediated and stereoselective active transport systems, respectively; both operating across the blood-brain barrier and cell membranes of the brain. The tissue homogenate analysis revealed that most of the accumulated radioactivity belonged to intact I-L-AMT, an indication of its stability. Thus, {sup 123}I-L-AMT appears to be a useful radiopharmaceutical for the selective measurement of cerebral amino acid transport.

  9. Synthesis of D- and L-phenylalanine derivatives by phenylalanine ammonia lyases: a multienzymatic cascade process.

    PubMed

    Parmeggiani, Fabio; Lovelock, Sarah L; Weise, Nicholas J; Ahmed, Syed T; Turner, Nicholas J

    2015-04-01

    The synthesis of substituted D-phenylalanines in high yield and excellent optical purity, starting from inexpensive cinnamic acids, has been achieved with a novel one-pot approach by coupling phenylalanine ammonia lyase (PAL) amination with a chemoenzymatic deracemization (based on stereoselective oxidation and nonselective reduction). A simple high-throughput solid-phase screening method has also been developed to identify PALs with higher rates of formation of non-natural D-phenylalanines. The best variants were exploited in the chemoenzymatic cascade, thus increasing the yield and ee value of the D-configured product. Furthermore, the system was extended to the preparation of those L-phenylalanines which are obtained with a low ee value using PAL amination.

  10. Differential inductions of phenylalanine ammonia-lyase and chalcone synthase during wounding, salicylic acid treatment, and salinity stress in safflower, Carthamus tinctorius.

    PubMed

    Dehghan, Sara; Sadeghi, Mahnaz; Pöppel, Anne; Fischer, Rainer; Lakes-Harlan, Reinhard; Kavousi, Hamid Reza; Vilcinskas, Andreas; Rahnamaeian, Mohammad

    2014-01-01

    Safflower (Carthamus tinctorius L.) serves as a reference dicot for investigation of defence mechanisms in Asteraceae due to abundant secondary metabolites and high resistance/tolerance to environmental stresses. In plants, phenylpropanoid and flavonoid pathways are considered as two central defence signalling cascades in stress conditions. Here, we describe the isolation of two major genes in these pathways, CtPAL (phenylalanine ammonia-lyase) and CtCHS (chalcone synthase) in safflower along with monitoring their expression profiles in different stress circumstances. The aa (amino acid) sequence of isolated region of CtPAL possesses the maximum identity up to 96% to its orthologue in Cynara scolymus, while that of CtCHS retains the highest identity to its orthologue in Callistephus chinensis up to 96%. Experiments for gene expression profiling of CtPAL and CtCHS were performed after the treatment of seedlings with 0.1 and 1 mM SA (salicylic acid), wounding and salinity stress. The results of semi-quantitative RT-PCR revealed that both CtPAL and CtCHS genes are further responsive to higher concentration of SA with dissimilar patterns. Regarding wounding stress, CtPAL gets slightly induced upon injury at 3 hat (hours after treatment) (hat), whereas CtCHS gets greatly induced at 3 hat and levels off gradually afterward. Upon salinity stress, CtPAL displays a similar expression pattern by getting slightly induced at 3 hat, but CtCHS exhibits a biphasic expression profile with two prominent peaks at 3 and 24 hat. These results substantiate the involvement of phenylpropanoid and particularly flavonoid pathways in safflower during wounding and especially salinity stress. PMID:24865400

  11. Differential inductions of phenylalanine ammonia-lyase and chalcone synthase during wounding, salicylic acid treatment, and salinity stress in safflower, Carthamus tinctorius

    PubMed Central

    Dehghan, Sara; Sadeghi, Mahnaz; Pöppel, Anne; Fischer, Rainer; Lakes-Harlan, Reinhard; Kavousi, Hamid Reza; Vilcinskas, Andreas; Rahnamaeian, Mohammad

    2014-01-01

    Safflower (Carthamus tinctorius L.) serves as a reference dicot for investigation of defence mechanisms in Asteraceae due to abundant secondary metabolites and high resistance/tolerance to environmental stresses. In plants, phenylpropanoid and flavonoid pathways are considered as two central defence signalling cascades in stress conditions. Here, we describe the isolation of two major genes in these pathways, CtPAL (phenylalanine ammonia-lyase) and CtCHS (chalcone synthase) in safflower along with monitoring their expression profiles in different stress circumstances. The aa (amino acid) sequence of isolated region of CtPAL possesses the maximum identity up to 96% to its orthologue in Cynara scolymus, while that of CtCHS retains the highest identity to its orthologue in Callistephus chinensis up to 96%. Experiments for gene expression profiling of CtPAL and CtCHS were performed after the treatment of seedlings with 0.1 and 1 mM SA (salicylic acid), wounding and salinity stress. The results of semi-quantitative RT–PCR revealed that both CtPAL and CtCHS genes are further responsive to higher concentration of SA with dissimilar patterns. Regarding wounding stress, CtPAL gets slightly induced upon injury at 3 hat (hours after treatment) (hat), whereas CtCHS gets greatly induced at 3 hat and levels off gradually afterward. Upon salinity stress, CtPAL displays a similar expression pattern by getting slightly induced at 3 hat, but CtCHS exhibits a biphasic expression profile with two prominent peaks at 3 and 24 hat. These results substantiate the involvement of phenylpropanoid and particularly flavonoid pathways in safflower during wounding and especially salinity stress. PMID:24865400

  12. Differential inductions of phenylalanine ammonia-lyase and chalcone synthase during wounding, salicylic acid treatment, and salinity stress in safflower, Carthamus tinctorius.

    PubMed

    Dehghan, Sara; Sadeghi, Mahnaz; Pöppel, Anne; Fischer, Rainer; Lakes-Harlan, Reinhard; Kavousi, Hamid Reza; Vilcinskas, Andreas; Rahnamaeian, Mohammad

    2014-06-25

    Safflower (Carthamus tinctorius L.) serves as a reference dicot for investigation of defence mechanisms in Asteraceae due to abundant secondary metabolites and high resistance/tolerance to environmental stresses. In plants, phenylpropanoid and flavonoid pathways are considered as two central defence signalling cascades in stress conditions. Here, we describe the isolation of two major genes in these pathways, CtPAL (phenylalanine ammonia-lyase) and CtCHS (chalcone synthase) in safflower along with monitoring their expression profiles in different stress circumstances. The aa (amino acid) sequence of isolated region of CtPAL possesses the maximum identity up to 96% to its orthologue in Cynara scolymus, while that of CtCHS retains the highest identity to its orthologue in Callistephus chinensis up to 96%. Experiments for gene expression profiling of CtPAL and CtCHS were performed after the treatment of seedlings with 0.1 and 1 mM SA (salicylic acid), wounding and salinity stress. The results of semi-quantitative RT-PCR revealed that both CtPAL and CtCHS genes are further responsive to higher concentration of SA with dissimilar patterns. Regarding wounding stress, CtPAL gets slightly induced upon injury at 3 hat (hours after treatment) (hat), whereas CtCHS gets greatly induced at 3 hat and levels off gradually afterward. Upon salinity stress, CtPAL displays a similar expression pattern by getting slightly induced at 3 hat, but CtCHS exhibits a biphasic expression profile with two prominent peaks at 3 and 24 hat. These results substantiate the involvement of phenylpropanoid and particularly flavonoid pathways in safflower during wounding and especially salinity stress.

  13. GPR139, an Orphan Receptor Highly Enriched in the Habenula and Septum, Is Activated by the Essential Amino Acids L-Tryptophan and L-Phenylalanine.

    PubMed

    Liu, Changlu; Bonaventure, Pascal; Lee, Grace; Nepomuceno, Diane; Kuei, Chester; Wu, Jiejun; Li, Qingqin; Joseph, Victory; Sutton, Steven W; Eckert, William; Yao, Xiang; Yieh, Lynn; Dvorak, Curt; Carruthers, Nicholas; Coate, Heather; Yun, Sujin; Dugovic, Christine; Harrington, Anthony; Lovenberg, Timothy W

    2015-11-01

    GPR139 is an orphan G-protein-coupled receptor expressed in the central nervous system. To identify its physiologic ligand, we measured GPR139 receptor activity from recombinant cells after treatment with amino acids, orphan ligands, serum, and tissue extracts. GPR139 activity was measured using guanosine 5'-O-(3-[(35)S]thio)-triphosphate binding, calcium mobilization, and extracellular signal-regulated kinases phosphorylation assays. Amino acids L-tryptophan (L-Trp) and L-phenylalanine (L-Phe) activated GPR139, with EC50 values in the 30- to 300-μM range, consistent with the physiologic concentrations of L-Trp and L-Phe in tissues. Chromatography of rat brain, rat serum, and human serum extracts revealed two peaks of GPR139 activity, which corresponded to the elution peaks of L-Trp and L-Phe. With the purpose of identifying novel tools to study GPR139 function, a high-throughput screening campaign led to the identification of a selective small-molecule agonist [JNJ-63533054, (S)-3-chloro-N-(2-oxo-2-((1-phenylethyl)amino)ethyl) benzamide]. The tritium-labeled JNJ-63533054 bound to cell membranes expressing GPR139 and could be specifically displaced by L-Trp and L-Phe. Sequence alignment revealed that GPR139 is highly conserved across species, and RNA sequencing studies of rat and human tissues indicated its exclusive expression in the brain and pituitary gland. Immunohistochemical analysis showed specific expression of the receptor in circumventricular regions of the habenula and septum in mice. Together, these findings suggest that L-Trp and L-Phe are candidate physiologic ligands for GPR139, and we hypothesize that this receptor may act as a sensor to detect dynamic changes of L-Trp and L-Phe in the brain.

  14. Gambogic Acid Inhibits STAT3 Phosphorylation Through Activation of Protein Tyrosine Phosphatase SHP-1: Potential Role in Proliferation and Apoptosis

    PubMed Central

    Prasad, Sahdeo; Pandey, Manoj K.; Yadav, Vivek R.; Aggarwal, Bharat B.

    2011-01-01

    The transcription factor, signal transducer and activator of transcription 3 (STAT3), is associated with proliferation, survival, and metastasis of cancer cells. We investigated whether gambogic acid (GA), a xanthone derived from the resin of traditional Chinese medicine, Gamboge hanburyi (mangosteen), can regulate the STAT3 pathway, leading to suppression of growth and sensitization of cancer cells. We found that GA induced apoptosis in human multiple myeloma cells that correlated with the inhibition of both constitutive and inducible STAT3 activation. STAT3 phosphorylation at both tyrosine residue 705 and serine residue 727 was inhibited by GA. STAT3 suppression was mediated through the inhibition of activation of the protein tyrosine kinases Janus-activated kinase (JAK) 1, and JAK2. Treatment with the protein tyrosine phosphatase (PTP) inhibitor pervanadate reversed the GA-induced down-regulation of STAT3, suggesting the involvement of a PTP. We also found that GA induced the expression of the PTP SHP-1. Deletion of the SHP-1 gene by small interfering RNA suppressed the ability of GA to inhibit STAT3 activation and to induce apoptosis, suggesting the critical role of SHP-1 in its action. Moreover, GA down-regulated the expression of STAT3-regulated antiapoptotic (Bcl-2, Bcl-xL, and Mcl-1), proliferative (cyclin D1), and angiogenic (VEGF) proteins, and this correlated with suppression of proliferation and induction of apoptosis. Overall, these results suggest that GA blocks STAT3 activation, leading to suppression of tumor cell proliferation and induction of apoptosis. PMID:21490133

  15. Synthesis and carbonic anhydrase inhibitory properties of amino acid - coumarin/quinolinone conjugates incorporating glycine, alanine and phenylalanine moieties.

    PubMed

    Küçükbay, F Zehra; Küçükbay, Hasan; Tanc, Muhammet; Supuran, Claudiu T

    2016-12-01

    N-Protected amino acids (Gly, Ala and Phe) were reacted with amino substituted coumarin and quinolinone derivatives, leading to the corresponding N-protected amino acid-coumarin/quinolinone conjugates. The carbonic anhydrase (CA, EC 4.2.1.1) inhibitory activity of the new compounds was assessed against various human (h) isoforms, such as hCA I, hCA II, hCA IV and hCA XII. The quinolinone conjugates were inactive as enzyme inhibitors, whereas the coumarins were ineffective hCA I/II inhibitors (KIs > 50 μM) but were submicromolar hCA IV and XII inhibitors, with inhibition constants ranging between 92 nM and 1.19 μM for hCA IV, and between 0.11 and 0.79 μM for hCA XII. These coumarin derivatives, as many others reported earlier, thus show an interesting selective inhibitory profile for the membrane-bound over the cytosolic CA isoforms.

  16. Highly Active and Specific Tyrosine Ammonia-Lyases from Diverse Origins Enable Enhanced Production of Aromatic Compounds in Bacteria and Saccharomyces cerevisiae

    PubMed Central

    Stahlhut, Steen Gustav; Li, Mingji; Gaspar, Paula; Siedler, Solvej; Förster, Jochen; Maury, Jérôme; Borodina, Irina

    2015-01-01

    Phenylalanine and tyrosine ammonia-lyases form cinnamic acid and p-coumaric acid, which are precursors of a wide range of aromatic compounds of biotechnological interest. Lack of highly active and specific tyrosine ammonia-lyases has previously been a limitation in metabolic engineering approaches. We therefore identified 22 sequences in silico using synteny information and aiming for sequence divergence. We performed a comparative in vivo study, expressing the genes intracellularly in bacteria and yeast. When produced heterologously, some enzymes resulted in significantly higher production of p-coumaric acid in several different industrially important production organisms. Three novel enzymes were found to have activity exclusively for phenylalanine, including an enzyme from the low-GC Gram-positive bacterium Brevibacillus laterosporus, a bacterial-type enzyme from the amoeba Dictyostelium discoideum, and a phenylalanine ammonia-lyase from the moss Physcomitrella patens (producing 230 μM cinnamic acid per unit of optical density at 600 nm [OD600]) in the medium using Escherichia coli as the heterologous host). Novel tyrosine ammonia-lyases having higher reported substrate specificity than previously characterized enzymes were also identified. Enzymes from Herpetosiphon aurantiacus and Flavobacterium johnsoniae resulted in high production of p-coumaric acid in Escherichia coli (producing 440 μM p-coumaric acid OD600 unit−1 in the medium) and in Lactococcus lactis. The enzymes were also efficient in Saccharomyces cerevisiae, where p-coumaric acid accumulation was improved 5-fold over that in strains expressing previously characterized tyrosine ammonia-lyases. PMID:25911487

  17. Phenylalanine and aspartame fail to alter feeding behavior, mood and arousal in men.

    PubMed

    Ryan-Harshman, M; Leiter, L A; Anderson, G H

    1987-01-01

    Two experiments were designed to investigate the neurobehavioral effects of phenylalanine (PHE; 0.84, 2.52, 5.04, and 10.08 g) and aspartame (APM; 5.04 and 10.08 g) on energy and macronutrient selection and on subjective feelings of hunger, mood and arousal in normal weight adult males. Neither phenylalanine nor aspartame altered mean energy intakes or macronutrient selection at a lunch begun 60 to 105 min after the amino acids were consumed. During this time, increased (p less than 0.05) visual analog scale (VAS) scores for emptiness, rumbling, weakness, degree of hunger and urge to eat were found in both experiments, but no treatment effects or interactions were seen for any variable in either experiment. Plasma PHE levels and ratios to other large neutral amino acids (NAA) rose significantly (p less than 0.05) after all treatments except 0.84 g PHE; plasma tyrosine (TYR) levels increased (p less than 0.05) only when greater than or equal to 2.52 g PHE was given. TYR/NAA ratios were higher (p less than 0.05) after 2.52 and 5.04 g PHE, and 10.08 g APM. No relationships were found between food intake and plasma amino acid levels. We conclude that, in normal weight men, PHE and APM, in doses up to 10 g, do not affect short-term energy and macronutrient intakes, or subjective feelings of hunger, mood and arousal.

  18. In vivo amino acid transport of subacute and chronic cerebral infarction evaluated by 12-18F-phenylalanine

    SciTech Connect

    Shimosegawa, E.; Miura, S.; Murakami, M.

    1994-05-01

    On the basis of previous validation of kinetic two-compartment model and the determination of normal values of three parameters (k{sub 1}:influx rate constant, k{sub 2}:outflux rate constant, Vd:distribution volume), PET measurements of in vivo amino acid transport from blood to brain using L-(2-18F)-fluorophenylalanine ({sup 18}F-Phe) were undergone in the patients with cerebral infarction. The purposes of this study are to evaluate the alteration of amino acid transport in subacute and chronic stage of cerebral infarction and to compare with cerebral blood flow (CBF) and oxygen metabolism. Dynamic {sup 18}F-Phe PET studies for 50 minutes were performed in 7 patients with cerebral infarction. The input function was obtained by 27 points of arterial sampling. In all patients, measurements of CBF, cerebral blood volume (CBV), cerebral metabolic rate of oxygen (CMRO{sub 2}), and oxygen extraction fraction (OEF) were made on the same day of {sup 18}F-Phe PET measurement. Each patient was studied twice, within 2 weeks of the onset and 3 months later. Weighted integration technique with table look-up method was applied for the reconstruction of parametric images of {sup 18}F-Phe and ROI analysis of k{sub 1}, k{sub 2}, and Vd. In subacute stage, significant reduction of k{sub 2} value in infarct area was observed when compared to that in periinfarct area (p<0.05) and in normal cortices (p<0.001). k{sub 1} value in this stage showed only slightly decrease in infarct area, therefore, Vd value in infarct area increased significantly compared to normal cortices (p<0.001). In chronic stage, both k{sub 1} and k{sub 2} values in infarct area were significantly lower than that in normal cortices (p<0.001), and corresponding Vd value reduced to normal level. Correlativity between kinetic parameters of {sup 18}F-Phe and CBF or oxygen metabolism was not observed both in subacute and chronic stage of infarction.

  19. Molecular cloning and characterization of tyrosine aminotransferase and hydroxyphenylpyruvate reductase, and rosmarinic acid accumulation in Scutellaria baicalensis.

    PubMed

    Kim, Yeon Bok; Uddina, Md Romij; Kim, YeJi; Park, Chun Geon; Park, Sang Un

    2014-09-01

    Rosmarinic acid (a-O-caffeoyl-3,4-dihydroxyphenylacetic acid, RA) is a caffeoyl ester widely distributed in plants. cDNA clones encoding tyrosine aminotransferase (TAT1 and 2) and hydroxyphenylpyruvate reductase (HPPR) have been isolated from Scutellaria baicalensis. The open reading frames (ORFs) of SbTAT1 and 2 were 1230 and 1272 bp long and encoded 409 and 423 amino acid residues, respectively. HPPR corresponded to a 942-bp ORF and 313 amino acid residues of translated protein. To study the molecular mechanisms of TAT and HPPR and investigate RA accumulation in S. baicalensis, we examined the transcript levels of TAT isoforms and HPPR with quantitative real-time PCR and analyzed the RA content in different organs by using high-performance liquid chromatography. The transcript levels of SbTATI SbTAT2, and SbHPPR in the flowers were higher than those in other organs. RA was also highly accumulated in the flowers and with a trace amount in the roots. No RA was detected in the leaves and stems of S. baicalensis. The amount of accumulated RA in the flowers was 28.7 times higher than that in the roots. Our results will be helpful in elucidating the mechanisms of RA biosynthesis in S. baicalensis. PMID:25918800

  20. The catalytic role of aspartic acid-92 in a human dual-specific protein-tyrosine-phosphatase.

    PubMed

    Denu, J M; Zhou, G; Guo, Y; Dixon, J E

    1995-03-14

    The mechanism of catalysis for the human dual-specific (vaccinia H1-related) protein-tyrosine-phosphatase was investigated. The pH dependence of the kcat value is bell-shaped when p-nitrophenyl phosphate was employed as a model substrate. The kcat/Km pH profile rises with a slope of 2 and decreases with a slope of -1, indicating that two groups must be unprotonated and one group must be protonated for activity. An amino acid residue with an apparent pKa value of 5.5 +/- 0.2 must be unprotonated and a residue with a pKa value of 5.7 must be unprotonated for activity. The pKa value of the catalytic cysteine-124 (C124) was 5.6 +/- 0.1. The aspartic acid-92-asparagine (D92N) mutant enzyme was 100-fold less active than the native enzyme and exhibited the loss of the basic limb in the pH profiles, suggesting that in the native enzyme D92 must be protonated for activity. The D92 residue is conserved throughout the entire family of dual-specific phosphatases. Mutants glutamic acid-6-glutamine, glutamic acid-32-glutamine, aspartic acid-14-asparagine, and aspartic acid-110-asparagine had less than a 2-fold effect on the kinetic parameters when compared to native enzyme. Based upon the lack of a "burst" in rapid reaction kinetics, formation of the intermediate is rate-limiting with both native and D92N mutant enzymes. In agreement with rate-limiting formation of the intermediate, the pKa value of 5.5 for the group which must be unprotonated for activity was assigned to C124.(ABSTRACT TRUNCATED AT 250 WORDS)

  1. Wound-Inducible Biosynthesis of Phytoalexin Hydroxycinnamic Acid Amides of Tyramine in Tryptophan and Tyrosine Decarboxylase Transgenic Tobacco Lines1

    PubMed Central

    Guillet, Gabriel; De Luca, Vincenzo

    2005-01-01

    The wound-activated biosynthesis of phytoalexin hydroxycinnamic acid amides of tyramine was compared in untransformed and transgenic tobacco (Nicotiana tabacum) lines that express tryptophan decarboxylase (TDC), tyrosine decarboxylase (TYDC), or both activities. Transgenic in vitro-grown tobacco lines expressing TDC activity accumulated high levels of tryptamine but not hydroxycinnamic amides of tryptamine. In contrast, transgenic tobacco lines expressing TYDC accumulated tyramine as well as p-coumaroyltyramine and feruloyltyramine. The MeOH-soluble and cell wall fractions showed higher concentrations of wound-inducible p-coumaroyltyramine and feruloyltyramine, especially at and around wound sites, in TYDC and TDC ×TYDC tobacco lines compared to wild-type or TDC lines. All the enzymes involved in the biosynthesis of hydroxycinnamic acid amides of tyramine were found to be similarly wound inducible in all tobacco genotypes investigated. These results provide experimental evidence that, under some circumstances, TYDC activity can exert a rate-limiting control over the carbon flux allocated to the biosynthesis of hydroxycinnamic acid amides of tyramine. PMID:15665252

  2. Systems engineering of tyrosine 195, tyrosine 260, and glutamine 265 in cyclodextrin glycosyltransferase from Paenibacillus macerans to enhance maltodextrin specificity for 2-O-(D)-glucopyranosyl-(L)-ascorbic acid synthesis.

    PubMed

    Han, Ruizhi; Liu, Long; Shin, Hyun-Dong; Chen, Rachel R; Li, Jianghua; Du, Guocheng; Chen, Jian

    2013-01-01

    In this work, the site saturation mutagenesis of tyrosine 195, tyrosine 260 and glutamine 265 in the cyclodextrin glycosyltransferase (CGTase) from Paenibacillus macerans was conducted to improve the specificity of CGTase for maltodextrin, which can be used as a cheap and easily soluble glycosyl donor for the synthesis of 2-O-d-glucopyranosyl-l-ascorbic acid (AA-2G). Specifically, the site-saturation mutagenesis of three sites-tyrosine 195, tyrosine 260, and glutamine 265-was performed, and it was found that the resulting mutants (containing the mutations Y195S [tyrosine → serine], Y260R [tyrosine → arginine], and Q265K [glutamine → lysine]) produced higher AA-2G yields than the wild type and the other mutant CGTases when maltodextrin was used as the glycosyl donor. Furthermore, double and triple mutations were introduced, and four mutants (containing Y195S/Y260R, Y195S/Q265K, Y260R/Q265K, and Y260R/Q265K/Y195S) were obtained and evaluated for the capacity to produce AA-2G. The Y260R/Q265K/Y195S triple mutant produced the highest titer of AA-2G at 1.92 g/liter, which was 60% higher than that (1.20 g/liter) produced by the wild-type CGTase. The kinetics analysis of AA-2G synthesis by the mutant CGTases confirmed the enhanced maltodextrin specificity, and it was also found that compared with the wild-type CGTase, all seven mutants had lower cyclization activities and higher hydrolysis and disproportionation activities. Finally, the mechanism responsible for the enhanced substrate specificity was explored by structure modeling, which indicated that the enhancement of maltodextrin specificity may be related to the changes of hydrogen bonding interactions between the side chain of residue at the three positions (195, 260, and 265) and the substrate sugars. This work adds to our understanding of the synthesis of AA-2G and makes the Y260R/Q265K/Y195S mutant a good starting point for further development by protein engineering.

  3. New tyrosinase inhibitory decapeptide: Molecular insights into the role of tyrosine residues.

    PubMed

    Ochiai, Akihito; Tanaka, Seiya; Imai, Yuta; Yoshida, Hisashi; Kanaoka, Takumi; Tanaka, Takaaki; Taniguchi, Masayuki

    2016-06-01

    Tyrosinase, a rate-limiting enzyme in melanin biosynthesis, catalyzes the hydroxylation of l-tyrosine to 3,4-dihydroxy-l-phenylalanine (l-dopa) (monophenolase reaction) and the subsequent oxidation of l-dopa to l-dopaquinone (diphenolase reaction). Thus, tyrosinase inhibitors have been proposed as skin-lightening agents; however, many of the existing inhibitors cannot be widely used in the cosmetic industry due to their high cytotoxicity and instability. On the other hand, some tyrosinase inhibitory peptides have been reported as safe. In this study, we found that the peptide TH10, which has a similar sequence to the characterized inhibitory peptide P4, strongly inhibits the monophenolase reaction with a half-maximal inhibitory concentration of 102 μM. Seven of the ten amino acid residues in TH10 were identical to P4; however, TH10 possesses one N-terminal tyrosine, whereas P4 contains three tyrosine residues located at its N-terminus, center, and C-terminus. Subsequent analysis using sequence-shuffled variants indicated that the tyrosine residues located at the N-terminus and center of P4 have little to no contribution to its inhibitory activity. Furthermore, docking simulation analysis of these peptides with mushroom tyrosinase demonstrated that the active tyrosine residue was positioned close to copper ions, suggesting that TH10 and P4 bind to tyrosinase as a substrate analogue. PMID:26589783

  4. Laboratory simulation of ultraviolet irradiation from the Sun on amino acids. III. irradiation of glycine-tyrosine

    NASA Astrophysics Data System (ADS)

    Scappini, F.; Capobianco, M. L.; Casadei, F.; Zamboni, R.

    2009-04-01

    The effects of near ultraviolet (UV) radiation on water solutions of tyrosine and glycine-tyrosine are investigated using a broadband xenon lamp in the region 200-800 nm. These experiments form a contribution in the laboratory simulation of the solar irradiation on the building blocks of life with regard to the origin of life. Results are presented showing the photodecomposition of tyrosine and glycine-tyrosine, at different concentrations, against UV doses. The analysis of the irradiated solutions is carried out by spectroscopic and analytical techniques. The findings of our laboratory simulations are used to constrain the early stages of the life emerging process.

  5. Plasma concentrations and pharmacokinetics of phenylalanine in rats and mice administered aspartame.

    PubMed

    Hjelle, J J; Dudley, R E; Marietta, M P; Sanders, P G; Dickie, B C; Brisson, J; Kotsonis, F N

    1992-01-01

    Aspartame (L-aspartyl-L-phenylalanine methyl ester) is an esterified, dipeptide sweetener that is rapidly and completely metabolized in the gastrointestinal tract to phenylalanine, aspartic acid and methanol. The pharmacokinetics of phenylalanine (PHE) and tyrosine (TYR) were examined following the administration of oral doses of aspartame (APM) to fasted male Sprague-Dawley rats (0, 50, 100, 200, 500 and 1,000 mg/kg) and CD-1 mice (0, 100, 200, 500, 1,000 and 2,000 mg/kg). Peak plasma PHE/large neutral amino acid (LNAA) ratios were calculated. Maximal plasma PHE and TYR concentrations were observed within 1 h after dosing and returned to baseline within 4-8 h in both species regardless of the dose of APM. Mean PHE Cmaxs ranged from 73.6 to 1,161 nmol/ml in the rat, and from 78.6 to 1,967 nmol/ml in the mouse. TYR Cmaxs ranged from 91.6 to 502 nmol/ml and from 89.2 to 792 nmol/ml in the rat and mouse, respectively. AUCs and Cmaxs were linear with dose in both species. Peak plasma PHE/LNAA ratios ranged from 0.112 to 1.117 in rats and from 0.121 to 1.769 in mice. Comparison of these ratios with those observed previously in humans indicates that rodents require a 2-6 times higher dose of APM than humans to produce similar increases in plasma PHE/LNAA ratios.

  6. Treatment of hyperactive children with D-phenylalanine.

    PubMed

    Zametkin, A J; Karoum, F; Rapoport, J L

    1987-06-01

    Eleven hyperactive boys were treated for 2 weeks with D-phenylalanine (20 mg/kg per day) and for 2 weeks with placebo in a double-blind crossover study. Tests included parent and teacher behavior ratings, cognitive measures, and blood and urine measures of norepinephrine, amino acids, and trace amines. No significant improvement or deterioration in behavior and no side effects were noted, and only serum phenylalanine was increased by the active treatment phase. This provides reassurance about the toxicity of aspartame, a food additive that contains phenylalanine, but argues against precursor loading treatment of hyperactivity.

  7. A modern view of phenylalanine ammonia lyase.

    PubMed

    MacDonald, M Jason; D'Cunha, Godwin B

    2007-06-01

    Phenylalanine ammonia lyase (PAL; E.C.4.3.1.5), which catalyses the biotransformation of L-phenylalanine to trans-cinnamic acid and ammonia, was first described in 1961 by Koukol and Conn. Since its discovery, much knowledge has been gathered with reference to the enzyme's catabolic role in microorganisms and its importance in the phenyl propanoid pathway of plants. The 3-dimensional structure of the enzyme has been characterized using X-ray crystallography. This has led to a greater understanding of the mechanism of PAL-catalyzed reactions, including the discovery of a recently described cofactor, 3,5-dihydro-5-methyldiene-4H-imidazol-4-one. In the past 3 decades, PAL has gained considerable significance in several clinical, industrial, and biotechnological applications. The reversal of the normal physiological reaction can be effectively employed in the production of optically pure L-phenylalanine, which is a precursor of the noncalorific sweetener aspartame (L-phenylalanyl-L-aspartyl methyl ester). The enzyme's natural ability to break down L-phenylalanine makes PAL a reliable treatment for the genetic condition phenylketonuria. In this mini-review, we discuss prominent details relating to the physiological role of PAL, the mechanism of catalysis, methods of determination and purification, enzyme kinetics, and enzyme activity in nonaqueous media. Two topics of current study on PAL, molecular biology and crystal structure, are also discussed. PMID:17612622

  8. A modern view of phenylalanine ammonia lyase.

    PubMed

    MacDonald, M Jason; D'Cunha, Godwin B

    2007-06-01

    Phenylalanine ammonia lyase (PAL; E.C.4.3.1.5), which catalyses the biotransformation of L-phenylalanine to trans-cinnamic acid and ammonia, was first described in 1961 by Koukol and Conn. Since its discovery, much knowledge has been gathered with reference to the enzyme's catabolic role in microorganisms and its importance in the phenyl propanoid pathway of plants. The 3-dimensional structure of the enzyme has been characterized using X-ray crystallography. This has led to a greater understanding of the mechanism of PAL-catalyzed reactions, including the discovery of a recently described cofactor, 3,5-dihydro-5-methyldiene-4H-imidazol-4-one. In the past 3 decades, PAL has gained considerable significance in several clinical, industrial, and biotechnological applications. The reversal of the normal physiological reaction can be effectively employed in the production of optically pure L-phenylalanine, which is a precursor of the noncalorific sweetener aspartame (L-phenylalanyl-L-aspartyl methyl ester). The enzyme's natural ability to break down L-phenylalanine makes PAL a reliable treatment for the genetic condition phenylketonuria. In this mini-review, we discuss prominent details relating to the physiological role of PAL, the mechanism of catalysis, methods of determination and purification, enzyme kinetics, and enzyme activity in nonaqueous media. Two topics of current study on PAL, molecular biology and crystal structure, are also discussed.

  9. Protein tyrosine phosphatases regulate arachidonic acid release, StAR induction and steroidogenesis acting on a hormone-dependent arachidonic acid-preferring acyl-CoA synthetase.

    PubMed

    Cano, Florencia; Poderoso, Cecilia; Cornejo Maciel, Fabiana; Castilla, Rocío; Maloberti, Paula; Castillo, Fernanda; Neuman, Isabel; Paz, Cristina; Podestá, Ernesto J

    2006-06-01

    The activation of the rate-limiting step in steroid biosynthesis, that is the transport of cholesterol into the mitochondria, is dependent on PKA-mediated events triggered by hormones like ACTH and LH. Two of such events are the protein tyrosine dephosphorylation mediated by protein tyrosine phosphatases (PTPs) and the release of arachidonic acid (AA) mediated by two enzymes, ACS4 (acyl-CoA synthetase 4) and Acot2 (mitochondrial thioesterase). ACTH and LH regulate the activity of PTPs and Acot2 and promote the induction of ACS4. Here we analyzed the involvement of PTPs on the expression of ACS4. We found that two PTP inhibitors, acting through different mechanisms, are both able to abrogate the hormonal effect on ACS4 induction. PTP inhibitors also reduce the effect of cAMP on steroidogenesis and on the level of StAR protein, which facilitates the access of cholesterol into the mitochondria. Moreover, our results indicate that exogenous AA is able to overcome the inhibition produced by PTP inhibitors on StAR protein level and steroidogenesis. Then, here we describe a link between PTP activity and AA release, since ACS4 induction is under the control of PTP activity, being a key event for AA release, StAR induction and steroidogenesis.

  10. Microbial Protein-tyrosine Kinases*

    PubMed Central

    Chao, Joseph D.; Wong, Dennis; Av-Gay, Yossef

    2014-01-01

    Microbial ester kinases identified in the past 3 decades came as a surprise, as protein phosphorylation on Ser, Thr, and Tyr amino acids was thought to be unique to eukaryotes. Current analysis of available microbial genomes reveals that “eukaryote-like” protein kinases are prevalent in prokaryotes and can converge in the same signaling pathway with the classical microbial “two-component” systems. Most microbial tyrosine kinases lack the “eukaryotic” Hanks domain signature and are designated tyrosine kinases based upon their biochemical activity. These include the tyrosine kinases termed bacterial tyrosine kinases (BY-kinases), which are responsible for the majority of known bacterial tyrosine phosphorylation events. Although termed generally as bacterial tyrosine kinases, BY-kinases can be considered as one family belonging to the superfamily of prokaryotic protein-tyrosine kinases in bacteria. Other members of this superfamily include atypical “odd” tyrosine kinases with diverse mechanisms of protein phosphorylation and the “eukaryote-like” Hanks-type tyrosine kinases. Here, we discuss the distribution, phylogeny, and function of the various prokaryotic protein-tyrosine kinases, focusing on the recently discovered Mycobacterium tuberculosis PtkA and its relationship with other members of this diverse family of proteins. PMID:24554699

  11. Serum albumin and α-1 acid glycoprotein impede the killing of Schistosoma mansoni by the tyrosine kinase inhibitor Imatinib

    PubMed Central

    Beckmann, Svenja; Long, Thavy; Scheld, Christina; Geyer, Rudolf; Caffrey, Conor R.; Grevelding, Christoph G.

    2014-01-01

    In the search for new drugs and drug targets to treat the flatworm disease schistosomiasis, protein kinases (PKs) have come under particular scrutiny because of their essential roles in developmental and physiological processes in schistosome parasites. In this context the application of the anti-cancer Abl tyrosine kinase (TK) inhibitor Imatinib (Gleevec/Glivec; STI-571) to adult Schistosoma mansoni in vitro has indicated negative effects on diverse physiological processes including survival. Motivated by these in vitro findings, we performed in vivo experiments in rodent models of S. mansoni infection. Unexpectedly, Imatinib had no effect on worm burden or egg-production. We found that the blood components serum albumin (SA) and alpha-1 acid glycoprotein (AGP or orosomucoid) negated Imatinib’s deleterious effects on adult S. mansoni and schistosomula (post-infective larvae) in vitro. This negative effect was partially reversed by erythromycin. AGP synthesis can increase as a consequence of inflammatory processes or infection; in addition upon infection AGP levels are 6–8 times higher in mice compared to humans. Therefore, mice and probably other rodents are poor infection models for measuring the effects of Imatinib in vivo. Accordingly, we suggest the routine evaluation of the ability of AGP and SA to block in vitro anti-schistosomal effects of small molecules like Imatinib prior to laborious and expensive animal experiments. PMID:25516839

  12. Salicylic Acid Based Small Molecule Inhibitor for the Oncogenic Src Homology-2 Domain Containing Protein Tyrosine Phosphatase-2 (SHP2)

    SciTech Connect

    Zhang, Xian; He, Yantao; Liu, Sijiu; Yu, Zhihong; Jiang, Zhong-Xing; Yang, Zhenyun; Dong, Yuanshu; Nabinger, Sarah C.; Wu, Li; Gunawan, Andrea M.; Wang, Lina; Chan, Rebecca J.; Zhang, Zhong-Yin

    2010-08-13

    The Src homology-2 domain containing protein tyrosine phosphatase-2 (SHP2) plays a pivotal role in growth factor and cytokine signaling. Gain-of-function SHP2 mutations are associated with Noonan syndrome, various kinds of leukemias, and solid tumors. Thus, there is considerable interest in SHP2 as a potential target for anticancer and antileukemia therapy. We report a salicylic acid based combinatorial library approach aimed at binding both active site and unique nearby subpockets for enhanced affinity and selectivity. Screening of the library led to the identification of a SHP2 inhibitor II-B08 (compound 9) with highly efficacious cellular activity. Compound 9 blocks growth factor stimulated ERK1/2 activation and hematopoietic progenitor proliferation, providing supporting evidence that chemical inhibition of SHP2 may be therapeutically useful for anticancer and antileukemia treatment. X-ray crystallographic analysis of the structure of SHP2 in complex with 9 reveals molecular determinants that can be exploited for the acquisition of more potent and selective SHP2 inhibitors.

  13. Au-Pd/reduced graphene oxide composite as a new sensing layer for electrochemical determination of ascorbic acid, acetaminophen and tyrosine.

    PubMed

    Tadayon, Fariba; Vahed, Saba; Bagheri, Hasan

    2016-11-01

    An Au-Pd/reduced graphene oxide composite was employed as a novel electrode material for the sensitive and simultaneous determination of ascorbic acid, acetaminophen and tyrosine. The electrochemical response characteristics of the modified electrode toward the analytes were investigated by differential pulse voltammetry and cyclic voltammetry. The responses of the electrochemical sensor for the target analytes were found to be improved significantly in comparison with those obtained using a conventional carbon paste electrode (CPE) and reduced graphene oxide/CPE. The experimental conditions for simultaneous determination of these species have been established. Ternary mixtures of analytes can be determined in the ranges of 0.03-9.50μM. Under optimal conditions, the limits of detection were 15.7, 7.6 and 11.1nM for ascorbic acid, acetaminophen, and tyrosine, respectively. The method was applied successfully to determine the analytes in urine, serum and pharmaceutical samples simultaneously. PMID:27524083

  14. Total branched-chain amino acids requirement in patients with maple syrup urine disease by use of indicator amino acid oxidation with L-[1-13C]phenylalanine.

    PubMed

    Riazi, Roya; Rafii, Mahroukh; Clarke, Joe T R; Wykes, Linda J; Ball, Ronald O; Pencharz, Paul B

    2004-07-01

    Maple syrup urine disease (MSUD) is an autosomal recessive disorder caused by defects in the mitochondrial multienzyme complex branched-chain alpha-keto acid dehydrogenase (BCKD; EC 1.2.4.4), responsible for the oxidative decarboxylation of the branched-chain ketoacids (BCKA) derived from the branched-chain amino acids (BCAA) leucine, valine, and isoleucine. Deficiency of the enzyme results in increased concentrations of the BCAA and BCKA in body cells and fluids. The treatment of the disease is aimed at keeping the concentration of BCAA below the toxic concentrations, primarily by dietary restriction of BCAA intake. The objective of this study was to determine the total BCAA requirements of patients with classical MSUD caused by marked deficiency of BCKD by use of the indicator amino acid oxidation (IAAO) technique. Five MSUD patients from the MSUD clinic of The Hospital for Sick Children participated in the study. Each was randomly assigned to different intakes of BCAA mixture (0, 20, 30, 50, 60, 70, 90, 110, and 130 mg.kg(-1).day(-1)), in which the relative proportion of BCAA was the same as that in egg protein. Total BCAA requirement was determined by measuring the oxidation of l-[1-(13)C]phenylalanine to (13)CO(2). The mean total BCAA requirement was estimated using a two-phase linear regression crossover analysis, which showed that the mean total BCAA requirement was 45 mg.kg(-1).day(-1), with the safe level of intake (upper 95% confidence interval) at 62 mg.kg(-1).day(-1). This is the first time BCAA requirements in patients with MSUD have been determined directly.

  15. Two-step energy transfer enables use of phenylalanine in action-EET for distance constraint determination in gaseous biomolecules.

    PubMed

    Hendricks, Nathan G; Julian, Ryan R

    2015-08-18

    Two-step energy transfer is potentially useful for exploring macromolecular structure, but it has not been observed previously in the gas-phase. Single step excitation energy transfer (EET) has been recently documented for tyrosine and tryptophan containing peptides, but not for phenylalanine. Herein, we report sequential energy transfer from phenylalanine to tyrosine to a disulfide, resulting in homolytic cleavage of a sulfur-sulfur bond. Interestingly, energy transfer from phenylalanine is only observed in the presence of tyrosine and only occurs within certain distance constraints. Isolated, electronically excited phenylalanine is known to have an extremely long lifetime in the gas phase, potentially suggesting quicker relaxation occurs via energy transfer to tyrosine. Alternatively, the direct overlap of states between phenylalanine and disulfide bonds is predicted to be poor, in which case tyrosine would serve to bridge the gap. In either case, the distance constraints imposed by this two-step EET are shown to be useful for evaluation and determination of gaseous biomolecular structure.

  16. Induction of L-phenylalanine ammonia-lyase during utilization of phenylalanine as a carbon or nitrogen source in Rhodotorula glutinis

    SciTech Connect

    Marusich, W.C.; Jensen, R.A.; Zamir, L.O.

    1981-06-01

    Rhodotorula glutinis is a convenient source of L-phenylalanine ammonia-lyase, an enzyme that is useful as a biochemical reagent in the assay of L-phenylalanine. There have been previous descriptions of induced lyase production in complex medium where induction occurs late in exponential growth, suggesting a role in secondary metabolism such as is the case in higher plants. A higher specific activity of L-phenylalanine ammonia-lyase (sixfold higher than in complex medium) can be obtained during midexponential growth in a defined medium containing L-phenylalanine as the sole source of carbon. L-phenylalanine will also induce lyase synthesis during exponential growth in minimal medium in which L-phenylalanine is the sole source of nitrogen. The appearance of lyase in complex medium supplemented with L-phenylalanine is probably triggered fortuitously by exhaustion late in growth of a prime source of nitrogen. In this study, R. glutinis appeared to express a single lyase enzyme, regardless of whether induction was nitrogen signaled or carbon signaled. Thin-layer chromatographic analysis of ether extracts prepared fom cultures induced with doubly labeled (U-/sup 14/C; ring-4-/sup 3/H) L-phenylalanine provided evidence of a catabolic sequence containing cinnamic acid, benzoic acid, and 4-hydroxybenzoic acid as degradative intermediates. 3,4-Dihydroxybenzoic acid was not identified as a catabolic intermediate.

  17. Studies on a novel approach for the separation of L-phenylalanine amino acid from fermentation broth using a newly developed charged ultrafiltration membrane. I. Single solute system

    SciTech Connect

    Agarwal, A.K.; Huang, R.Y.M. )

    1994-01-01

    A newly developed thin-film composite (TFC) ultrafiltration membrane made of sulfonated poly(phenylene oxide) (SPPO) was used to establish the feasibility of separating L-phenylalanine from the fermentation broth containing a number of dissolved inorganic and organic solutes as an alternative approach to the currently used complex and uneconomical conventional ion-exchange schemes. It was found that the rejection of inorganic salts in a single component system was highly dependent on the feed solution concentration and varied inversely with it. The pH of the feed solution was found to have a strong effect on the rejection of L-phenylalanine, changing it from - 10 to 90%. This rejection behavior was identical for the two TFC-SPPO membrane samples which had molecular weight cutoff ratings of 10,000 and 20,000, respectively, although the permeate flux of the latter sample was almost twice that of the former sample. It was found that glucose molecules were not rejected by the membrane. 11 refs., 18 figs., 2 tab.

  18. Phenylalanine Ammonia-Lyase-Catalyzed Deamination of an Acyclic Amino Acid: Enzyme Mechanistic Studies Aided by a Novel Microreactor Filled with Magnetic Nanoparticles.

    PubMed

    Weiser, Diána; Bencze, László Csaba; Bánóczi, Gergely; Ender, Ferenc; Kiss, Róbert; Kókai, Eszter; Szilágyi, András; Vértessy, Beáta G; Farkas, Ödön; Paizs, Csaba; Poppe, László

    2015-11-01

    Phenylalanine ammonia-lyase (PAL), found in many organisms, catalyzes the deamination of l-phenylalanine (Phe) to (E)-cinnamate by the aid of its MIO prosthetic group. By using PAL immobilized on magnetic nanoparticles and fixed in a microfluidic reactor with an in-line UV detector, we demonstrated that PAL can catalyze ammonia elimination from the acyclic propargylglycine (PG) to yield (E)-pent-2-ene-4-ynoate. This highlights new opportunities to extend MIO enzymes towards acyclic substrates. As PG is acyclic, its deamination cannot involve a Friedel-Crafts-type attack at an aromatic ring. The reversibility of the PAL reaction, demonstrated by the ammonia addition to (E)-pent-2-ene-4-ynoate yielding enantiopure l-PG, contradicts the proposed highly exothermic single-step mechanism. Computations with the QM/MM models of the N-MIO intermediates from L-PG and L-Phe in PAL show similar arrangements within the active site, thus supporting a mechanism via the N-MIO intermediate.

  19. Tyrosine Aminotransferase: Biochemical and Structural Properties and Molecular Dynamics Simulations

    SciTech Connect

    P Mehere; Q Han; J Lemkul; C Vavricka; H Robinson; D Bevan; J Li

    2011-12-31

    Tyrosine aminotransferase (TAT) catalyzes the transamination of tyrosine and other aromatic amino acids. The enzyme is thought to play a role in tyrosinemia type II, hepatitis and hepatic carcinoma recovery. The objective of this study is to investigate its biochemical and structural characteristics and substrate specificity in order to provide insight regarding its involvement in these diseases. Mouse TAT (mTAT) was cloned from a mouse cDNA library, and its recombinant protein was produced using Escherichia coli cells and purified using various chromatographic techniques. The recombinant mTAT is able to catalyze the transamination of tyrosine using {alpha}-ketoglutaric acid as an amino group acceptor at neutral pH. The enzyme also can use glutamate and phenylalanine as amino group donors and p-hydroxy-phenylpyruvate, phenylpyruvate and alpha-ketocaproic acid as amino group acceptors. Through macromolecular crystallography we have determined the mTAT crystal structure at 2.9 {angstrom} resolution. The crystal structure revealed the interaction between the pyridoxal-5'-phosphate cofactor and the enzyme, as well as the formation of a disulphide bond. The detection of disulphide bond provides some rational explanation regarding previously observed TAT inactivation under oxidative conditions and reactivation of the inactive TAT in the presence of a reducing agent. Molecular dynamics simulations using the crystal structures of Trypanosoma cruzi TAT and human TAT provided further insight regarding the substrate-enzyme interactions and substrate specificity. The biochemical and structural properties of TAT and the binding of its cofactor and the substrate may help in elucidation of the mechanism of TAT inhibition and activation.

  20. Tyrosine aminotransferase: biochemical and structural properties and molecular dynamics simulations

    SciTech Connect

    Mehere, P.; Robinson, H.; Han, Q.; Lemkul, J. A.; Vavricka, C. J.; Bevan, D. R.; Li, J.

    2010-11-01

    Tyrosine aminotransferase (TAT) catalyzes the transamination of tyrosine and other aromatic amino acids. The enzyme is thought to play a role in tyrosinemia type II, hepatitis and hepatic carcinoma recovery. The objective of this study is to investigate its biochemical and structural characteristics and substrate specificity in order to provide insight regarding its involvement in these diseases. Mouse TAT (mTAT) was cloned from a mouse cDNA library, and its recombinant protein was produced using Escherichia coli cells and purified using various chromatographic techniques. The recombinant mTAT is able to catalyze the transamination of tyrosine using {alpha}-ketoglutaric acid as an amino group acceptor at neutral pH. The enzyme also can use glutamate and phenylalanine as amino group donors and p-hydroxy-phenylpyruvate, phenylpyruvate and alpha-ketocaproic acid as amino group acceptors. Through macromolecular crystallography we have determined the mTAT crystal structure at 2.9 {angstrom} resolution. The crystal structure revealed the interaction between the pyridoxal-5'-phosphate cofactor and the enzyme, as well as the formation of a disulphide bond. The detection of disulphide bond provides some rational explanation regarding previously observed TAT inactivation under oxidative conditions and reactivation of the inactive TAT in the presence of a reducing agent. Molecular dynamics simulations using the crystal structures of Trypanosoma cruzi TAT and human TAT provided further insight regarding the substrate-enzyme interactions and substrate specificity. The biochemical and structural properties of TAT and the binding of its cofactor and the substrate may help in elucidation of the mechanism of TAT inhibition and activation.

  1. Tyrosine aminotransferase: biochemical and structural properties and molecular dynamics simulations.

    PubMed

    Mehere, Prajwalini; Han, Qian; Lemkul, Justin A; Vavricka, Christopher J; Robinson, Howard; Bevan, David R; Li, Jianyong

    2010-11-01

    Tyrosine aminotransferase (TAT) catalyzes the transamination of tyrosine and other aromatic amino acids. The enzyme is thought to play a role in tyrosinemia type II, hepatitis and hepatic carcinoma recovery. The objective of this study is to investigate its biochemical and structural characteristics and substrate specificity in order to provide insight regarding its involvement in these diseases. Mouse TAT (mTAT) was cloned from a mouse cDNA library, and its recombinant protein was produced using Escherichia coli cells and purified using various chromatographic techniques. The recombinant mTAT is able to catalyze the transamination of tyrosine using α-ketoglutaric acid as an amino group acceptor at neutral pH. The enzyme also can use glutamate and phenylalanine as amino group donors and p-hydroxy-phenylpyruvate, phenylpyruvate and alpha-ketocaproic acid as amino group acceptors. Through macromolecular crystallography we have determined the mTAT crystal structure at 2.9 Å resolution. The crystal structure revealed the interaction between the pyridoxal-5'-phosphate cofactor and the enzyme, as well as the formation of a disulphide bond. The detection of disulphide bond provides some rational explanation regarding previously observed TAT inactivation under oxidative conditions and reactivation of the inactive TAT in the presence of a reducing agent. Molecular dynamics simulations using the crystal structures of Trypanosoma cruzi TAT and human TAT provided further insight regarding the substrate-enzyme interactions and substrate specificity. The biochemical and structural properties of TAT and the binding of its cofactor and the substrate may help in elucidation of the mechanism of TAT inhibition and activation.

  2. Boswellic Acid Blocks STAT3 Signaling, Proliferation, and Survival of Multiple Myeloma via the Protein Tyrosine Phosphatase SHP-1

    PubMed Central

    Kunnumakkara, Ajaikumar B.; Nair, Asha S.; Sung, Bokyung; Pandey, Manoj K.; Aggarwal, Bharat B.

    2009-01-01

    Activation of signal transducers and activators of transcription (STAT)-3 factors has been linked with survival, proliferation, chemoresistance and angiogenesis of tumor cells, including human multiple myeloma (MM). Thus agents that can suppress STAT3 activation have potential as cancer therapeutics. In our search for such agents, we identified acetyl-11-keto-β-boswellic acid (AKBA), originally isolated from Boswellia serrata. Our results show that AKBA inhibited constitutive STAT3 activation in human MM cells. AKBA suppressed IL-6-induced STAT3 activation, and the inhibition was reversible. The phosphorylation of both Jak 2 and Src, constituents of the STAT3 pathway, was inhibited by AKBA. Interestingly, treatment of cells with pervanadate suppressed AKBA’s effect to inhibit the phosphorylation of STAT3, thus suggesting the involvement of a protein tyrosine phosphatase. We found that AKBA induced Src homology region 2 domain-containing phosphatase 1 (SHP-1), which may account for its role in dephosphorylation of STAT3. Moreover, deletion of SHP-1 gene by SiRNA abolished the ability of AKBA to inhibit STAT3 activation. The inhibition of STAT3 activation by AKBA led to the suppression of gene products involved in proliferation (cyclin D1), survival (Bcl-2, Bcl-xL and Mcl-1), and angiogenesis (VEGF). This affect correlated with the inhibition of proliferation and apoptosis in MM cells. Consistent with these results, overexpression of constitutive active STAT3 significantly reduced the AKBA induced apoptosis. Overall, our results suggest that AKBA is a novel inhibitor of STAT3 activation and has potential in the treatment of cancer. PMID:19147543

  3. Acute effects of aspartame on large neutral amino acids and monoamines in rat brain.

    PubMed

    Fernstrom, J D; Fernstrom, M H; Gillis, M A

    1983-04-01

    The dipeptide aspartame (APM; aspartylphenylalanine methylester), an artificial sweetener, was studied in vivo for its ability to influence brain levels of the large neutral amino acids and the rates of hydroxylation of the aromatic amino acids. The administration by gavage of APM (200 mg/kg) caused large increments in blood and brain levels of phenylalanine and tyrosine by 60 minutes. Brain tryptophan level was occasionally reduced significantly, but the brain levels of the branched-chain amino acids were always unaffected. Smaller doses (50, 100 mg/kg) also raised blood and brain tyrosine and phenylalanine, but did not reduce brain tryptophan levels. At the highest dose (200 mg/kg), APM gavage caused an insignificant increase in dopa accumulation (after NSD-1015), and a modest reduction in 5-hydroxytryptophan accumulation. No changes in the brain levels of serotonin, 5-hydroxyindoleacetic acid, dopamine, dihydroxyphenylacetic acid, homovanillic acid, or norepinephrine were produced by APM administration (200 mg/kg). These results thus indicate that APM, even when administered in amounts that cause large increments in brain tyrosine and phenylalanine, produce minimal effects on the rates of formation of monoamine transmitters.

  4. Biomolecular interactions of emerging two-dimensional materials with aromatic amino acids

    NASA Astrophysics Data System (ADS)

    Mallineni, Sai Sunil Kumar; Karakaya, Mehmet; Podila, Ramakrishna; Rao, Apparao

    The present work experimentally investigates the interaction of aromatic amino acids, viz., tyrosine, tryptophan, and phenylalanine with novel two-dimensional (2D) materials including graphene (G), graphene oxide (GO), and boron nitride (BN). Photoluminescence, micro-Raman spectroscopy and cyclic voltammetry were employed to investigate the nature of interactions and possible charge transfer between 2D materials and amino acids. Consistent with previous theoretical studies, graphene and BN were observed to interact with amino acids through π- π interactions. Furthermore, we found that GO exhibits strong interactions with tryptophan and tyrosine as compared to graphene and BN, which we attribute to the formation of H-bonds between tryptophan and GO as shown theoretically in Ref. 2. On the other hand, phenylalanine did not exhibit much difference in interactions with G, GO, and BN. Clemson Nanomaterials Center, Clemson University, Clemson, SC, USA.

  5. 21 CFR 201.21 - Declaration of presence of phenylalanine as a component of aspartame in over-the-counter and...

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 4 2010-04-01 2010-04-01 false Declaration of presence of phenylalanine as a...: GENERAL LABELING General Labeling Provisions § 201.21 Declaration of presence of phenylalanine as a... methylester of a dipeptide composed of two amino acids, phenylalanine and aspartic acid. When these two...

  6. 21 CFR 201.21 - Declaration of presence of phenylalanine as a component of aspartame in over-the-counter and...

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 4 2012-04-01 2012-04-01 false Declaration of presence of phenylalanine as a...: GENERAL LABELING General Labeling Provisions § 201.21 Declaration of presence of phenylalanine as a... methylester of a dipeptide composed of two amino acids, phenylalanine and aspartic acid. When these two...

  7. 21 CFR 201.21 - Declaration of presence of phenylalanine as a component of aspartame in over-the-counter and...

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 4 2011-04-01 2011-04-01 false Declaration of presence of phenylalanine as a...: GENERAL LABELING General Labeling Provisions § 201.21 Declaration of presence of phenylalanine as a... methylester of a dipeptide composed of two amino acids, phenylalanine and aspartic acid. When these two...

  8. 21 CFR 201.21 - Declaration of presence of phenylalanine as a component of aspartame in over-the-counter and...

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 4 2013-04-01 2013-04-01 false Declaration of presence of phenylalanine as a...: GENERAL LABELING General Labeling Provisions § 201.21 Declaration of presence of phenylalanine as a... methylester of a dipeptide composed of two amino acids, phenylalanine and aspartic acid. When these two...

  9. 21 CFR 201.21 - Declaration of presence of phenylalanine as a component of aspartame in over-the-counter and...

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 4 2014-04-01 2014-04-01 false Declaration of presence of phenylalanine as a...: GENERAL LABELING General Labeling Provisions § 201.21 Declaration of presence of phenylalanine as a... methylester of a dipeptide composed of two amino acids, phenylalanine and aspartic acid. When these two...

  10. Amino acids

    MedlinePlus

    ... amino acids are: histidine, isoleucine, leucine, lysine, methionine, phenylalanine, threonine, tryptophan , and valine. Nonessential amino acids "Nonessential" means that our bodies produce an amino ...

  11. Elimination of amino acids in acute renal failure.

    PubMed

    Druml, W; Bürger, U; Kleinberger, G; Lenz, K; Laggner, A

    1986-01-01

    Plasma amino acid concentrations and the elimination of parenterally administered amino acids were investigated in 12 patients with nonhypercatabolic acute renal failure. A distinctive plasma amino acid pattern could be observed: plasma concentrations of phenylalanine and methionine were increased, those of valine and leucine decreased. Of the nonessential amino acids, cystine, taurine und tyrosine had elevated but none of them reduced plasma concentrations. The elimination of amino acids was evaluated in a monocompartment model after bolus injection of an amino acid solution containing essential and nonessential amino acids. Pharmacokinetic parameters of 17 amino acids were calculated. The mean elimination half-time was raised by 25%. The elimination half-time of phenylalanine, methionine, glutamic acid, proline and ornithine was increased. Histidine was the only amino acid with--however insignificantly--accelerated elimination from the intravascular compartment. The total clearance rate and total transfer rate was not altered (107 and 97% of normal, respectively). The clearance of threonine, lysine, serine, glycine and histidine was increased, of valine, phenylalanine, glutamic acid and to a minor degree of methionine was decreased. The transfer rate of methionine, lysine, glycine was elevated, of valine, aspartic acid, glutamic acid and ornithine reduced. The demonstration of these pronounced alterations of amino acid elimination in acute renal failure may have major consequences in parenteral amino acid therapy.

  12. Ammonia lyases and aminomutases as biocatalysts for the synthesis of α-amino and β-amino acids.

    PubMed

    Turner, Nicholas J

    2011-04-01

    Ammonia lyases catalyse the reversible addition of ammonia to cinnamic acid (1: R=H) and p-hydroxycinnamic (1: R=OH) to generate L-phenylalanine (2: R=H) and L-tyrosine (2: R=OH) respectively (Figure 1a). Both phenylalanine ammonia lyase (PAL) and tyrosine ammonia lyase (TAL) are widely distributed in plants, fungi and prokaryotes. Recently there has been interest in the use of these enzymes for the synthesis of a broader range of L-arylalanines. Aminomutases catalyse a related reaction, namely the interconversion of α-amino acids to β-amino acids (Figure 1b). In the case of L-phenylalanine, this reaction is catalysed by phenylalanine aminomutase (PAM) and proceeds stereospecifically via the intermediate cinnamic acid to generate β-Phe 3. Ammonia lyases and aminomutases are related in sequence and structure and share the same active site cofactor 4-methylideneimidazole-5-one (MIO). There is currently interest in the possibility of using these biocatalysts to prepare a wide range of enantiomerically pure l-configured α-amino and β-amino acids. Recent reviews have focused on the mechanism of these MIO containing enzymes. The aim of this review is to review recent progress in the application of ammonia lyase and aminomutase enzymes to prepare enantiomerically pure α-amino and β-amino acids.

  13. The internalization signal in the cytoplasmic tail of lysosomal acid phosphatase consists of the hexapeptide PGYRHV.

    PubMed Central

    Lehmann, L E; Eberle, W; Krull, S; Prill, V; Schmidt, B; Sander, C; von Figura, K; Peters, C

    1992-01-01

    Lysosomal acid phosphatase (LAP) is rapidly internalized from the cell surface due to a tyrosine-containing internalization signal in its 19 amino acid cytoplasmic tail. Measuring the internalization of a series of LAP cytoplasmic tail truncation and substitution mutants revealed that the N-terminal 12 amino acids of the cytoplasmic tail are sufficient for rapid endocytosis and that the hexapeptide 411-PGYRHV-416 is the tyrosine-containing internalization signal. Truncation and substitution mutants of amino acid residues following Val416 can prevent internalization even though these residues do not belong to the internalization signal. It was shown recently that part of the LAP cytoplasmic tail peptide corresponding to 410-PPGY-413 forms a well-ordered beta turn structure in solution. Two-dimensional NMR spectroscopy of two modified LAP tail peptides, in which the single tyrosine was substituted either by phenylalanine or by alanine, revealed that the tendency to form a beta turn is reduced by 25% in the phenylalanine-containing peptide and by approximately 50% in the alanine-containing mutant peptide. Our results suggest, that in the short cytoplasmic tail of LAP tyrosine is required for stabilization of the right turn and that the aromatic ring system of the tyrosine residue is a contact point to the putative cytoplasmic receptor. Images PMID:1425575

  14. Calcium alginate bead immobilization of cells containing tyrosine ammonia lyase activity for use in the production of p-hydroxycinnamic acid.

    PubMed

    Trotman, Robert J; Camp, Carl E; Ben-Bassat, Arie; DiCosimo, Robert; Huang, Lixuan; Crum, Grace A; Sariaslani, F Sima; Haynie, Sharon L

    2007-01-01

    An Escherichia coli catalyst with tyrosine ammonia lyase activity (TAL) has been stabilized for repeated use in batch conversions of high tyrosine solids to p-hydroxycinnamic acid (pHCA). The TAL biocatalyst was stabilized by controlling the reaction pH to 9.8 +/- 0.1 and immobilizing the cells within a calcium alginate matrix that was cross-linked with glutaraldehyde and polyethyleneimine (GA/PEI). We found a GA range where the bead-encapsulated TAL was not inactivated, and the resulting cross-linking provided the beads with the mechanical stability necessary for repeated use in consecutive batch reactions with catalyst recycle. The GA/PEI calcium alginate TAL catalyst was used in 41 1-L batch reactions where 50 g L(-1) tyrosine was converted to 39 +/- 4 g L(-1) pHCA in each batch. The practical usefulness and ease of this process was demonstrated by scaling up the TAL bead immobilization and using the immobilized TAL catalyst in four 125-L bioconversion reactions to produce over 12 kg of purified pHCA.

  15. Effects of aspartame and glucose administration on brain and plasma levels of large neutral amino acids and brain 5-hydroxyindoles.

    PubMed

    Yokogoshi, H; Roberts, C H; Caballero, B; Wurtman, R J

    1984-07-01

    Administration of the artificial sweetener aspartame (L-aspartylphenylalanylmethyl ester; 200 mg/kg) by gavage to rats caused large increments in brain and plasma levels of phenylalanine and its product tyrosine. Glucose administration (3 g/kg, by gavage, a dose sufficient to cause insulin-mediated reductions in plasma levels of the large neutral amino acids leucine, isoleucine, and valine) also elevated brain phenylalanine and tyrosine, and enhanced the increments caused by the aspartame, nearly doubling the rise in brain phenylalanine. Each animal's brain phenylalanine or tyrosine levels were highly correlated (r = 0.97 and 0.99, respectively) with its plasma phenylalanine or tyrosine ratios, affirming that aspartame's effects on the brain amino acids result from the changes it produces in plasma composition. As described previously, glucose consumption increased brain tryptophan levels, and consequently, brain levels of the 5-hydroxyindoles serotonin and 5-hydroxyindoleacetic acid. Aspartame alone had no effect on these compounds but completely blocked the changes in 5-hydroxyindoles caused by glucose. Each animal's brain level of tryptophan (r = 0.89) and 5-hydroxyindoles (r = 0.74) was also significantly correlated with its plasma tryptophan ratio, affirming that the effects of glucose or aspartame on these brain constituents also result from the changes they produce in plasma composition. The aspartame-glucose combination also reduced brain levels of leucine, isoleucine, and valine to a significantly greater extent than aspartame or glucose alone. These observations indicate that high aspartame doses can generate major neurochemical changes in rats, especially when consumed along with carbohydrate-containing foods. However, they should not in any way be interpreted as demonstrating that aspartame significantly affects the human brain.

  16. Regulation and function of syk tyrosine kinase in mast cell signaling and beyond.

    PubMed

    de Castro, Rodrigo Orlandini

    2011-01-01

    The protein tyrosine kinase Syk plays a critical role in FcεRI signaling in mast cells. Binding of Syk to phosphorylated immunoreceptor tyrosine-based activation motifs (p-ITAM) of the receptor subunits results in conformational changes and tyrosine phosphorylation at multiple sites that leads to activation of Syk. The phosphorylated tyrosines throughout the molecule play an important role in the regulation of Syk-mediated signaling. Reconstitution of receptor-mediated signaling in Syk(-/-) cells by wild-type Syk or mutants which have substitution of these tyrosines with phenylalanine together with in vitro assays has been useful strategies to understand the regulation and function of Syk.

  17. Progesterone receptor isoforms differentially regulate the expression of tryptophan and tyrosine hydroxylase and glutamic acid decarboxylase in the rat hypothalamus.

    PubMed

    González-Flores, Oscar; Gómora-Arrati, Porfirio; García-Juárez, Marcos; Miranda-Martínez, Alfredo; Armengual-Villegas, Alejandra; Camacho-Arroyo, Ignacio; Guerra-Araiza, Christian

    2011-10-01

    Progesterone exerts a variety of actions in the brain through the interaction with its receptors (PR) which have two isoforms with different function and regulation: PR-A and PR-B. Progesterone may modulate neurotransmission by regulating the expression of neurotransmitters synthesizing enzymes or their receptors in several brain regions. The role of PR isoforms in this modulation is unknown. We explored the role of PR isoforms in the regulation of tryptophan (TPH) and tyrosine (TH) hydroxylase, and glutamic acid decarboxylase (GAD) expression in the hypothalamus of ovariectomized rats. Two weeks after ovariectomy, animals were subcutaneously injected with 5 μg of estradiol benzoate (EB), and 40 h later, progesterone (P) was intracerebroventricularly (ICV) injected. Each animal received two ICV injections of 1 μg/μl (4 nmol) of PR-B and total PR (PR-A+PR-B) sense or antisense (As) oligonucleotides (ODNs). First injection was made immediately before sc EB injection, and 24h later animals received the second one. Twenty-four hours after P administration, rats were euthanized and brains removed to measure the expression of PR-A and PR-B, TPH, TH and GAD by Western blot. We observed that sense ODNs modified neither PR isoforms nor enzymes expression in the hypothalamus, whereas PR A+B antisense (PR A+B As) clearly decreased the expression of both PR isoforms in this region. ICV administration of PR-B As only decreased PR-B isoform expression with no significant effects on PR-A expression. A differential protein expression of TPH, TH and GAD was observed after PR isoforms antisense administration. PR-B As administration decreased the expression of TPH (65% with respect to control). In contrast, PR A+B As and PR-B As administration increased (51.6% and 34.4%, respectively) TH expression. The administration of PR A+B As and PR-B As diminished GAD expression (33.4% and 41.6%, respectively). Our findings indicate that PR isoforms play a differential role in the

  18. Phenylalanine hydroxylase (PAH) from the lower eukaryote Leishmania major

    PubMed Central

    Lye, Lon-Fye; Kang, Song Ok; Nosanchuk, Joshua D.; Casadevall, Arturo; Beverley, Stephen M.

    2010-01-01

    Aromatic amino acid hydroxylases (AAAH) typically use tetrahydrobiopterin (H4B) as the cofactor. The protozoan parasite Leishmania major requires biopterin for growth and expresses strong salvage and regeneration systems to maintain H4B levels. Here we explored the consequences of genetic manipulation of the sole L. major phenylalanine hydroxylase (PAH) to explore whether it could account for the Leishmania H4B requirement. L. major PAH resembles AAAHs of other organisms, bearing eukaryotic-type domain organization, and conservation of key catalytic residues including those implicated in pteridine binding. A pah− null mutant and an episomal complemented overexpressing derivative (pah−/+PAH) were readily obtained, and metabolic labeling studies established that PAH was required to hydroxylate Phe to Tyr. Neither WT nor overexpressing lines were able to hydroxylate radiolabeled tyrosine or tryptophan, nor to synthesize catecholamines. WT but not pah− parasites showed reactivity with an antibody to melanin when grown with L-3,4-dihydroxyphenylalanine (L-DOPA), although the reactive product is unlikely to be melanin sensu strictu. WT was auxotrophic for Phe, Trp and Tyr, suggesting that PAH activity was insufficient to meet normal Tyr requirements. However, pah− showed an increased sensitivity to Tyr deprivation, while the pah−/+PAH overexpressor showed increased survival and could be adapted to grow well without added Tyr. pah− showed no alterations in H4B-dependent differentiation, as established by in vitro metacyclogenesis, or survival in mouse or macrophage infections. Thus Leishmania PAH may mitigate but not alleviate Tyr auxotrophy, but plays no essential role in the steps of the parasite infectious cycle. These findings suggest PAH is unlikely to explain the Leishmania requirement for biopterin. PMID:20887755

  19. Phenylalanine transfer across the isolated perfused human placenta: an experimental and modeling investigation

    PubMed Central

    Lofthouse, E. M.; Perazzolo, S.; Brooks, S.; Crocker, I. P.; Glazier, J. D.; Johnstone, E. D.; Panitchob, N.; Sibley, C. P.; Widdows, K. L.; Sengers, B. G.

    2015-01-01

    Membrane transporters are considered essential for placental amino acid transfer, but the contribution of other factors, such as blood flow and metabolism, is poorly defined. In this study we combine experimental and modeling approaches to understand the determinants of [14C]phenylalanine transfer across the isolated perfused human placenta. Transfer of [14C]phenylalanine across the isolated perfused human placenta was determined at different maternal and fetal flow rates. Maternal flow rate was set at 10, 14, and 18 ml/min for 1 h each. At each maternal flow rate, fetal flow rates were set at 3, 6, and 9 ml/min for 20 min each. Appearance of [14C]phenylalanine was measured in the maternal and fetal venous exudates. Computational modeling of phenylalanine transfer was undertaken to allow comparison of the experimental data with predicted phenylalanine uptake and transfer under different initial assumptions. Placental uptake (mol/min) of [14C]phenylalanine increased with maternal, but not fetal, flow. Delivery (mol/min) of [14C]phenylalanine to the fetal circulation was not associated with fetal or maternal flow. The absence of a relationship between placental phenylalanine uptake and net flux of phenylalanine to the fetal circulation suggests that factors other than flow or transporter-mediated uptake are important determinants of phenylalanine transfer. These observations could be explained by tight regulation of free amino acid levels within the placenta or properties of the facilitated transporters mediating phenylalanine transport. We suggest that amino acid metabolism, primarily incorporation into protein, is controlling free amino acid levels and, thus, placental transfer. PMID:26676251

  20. Protein tyrosine nitration

    PubMed Central

    Chaki, Mounira; Leterrier, Marina; Barroso, Juan B

    2009-01-01

    Nitric oxide metabolism in plant cells has a relative short history. Nitration is a chemical process which consists of introducing a nitro group (-NO2) into a chemical compound. in biological systems, this process has been found in different molecules such as proteins, lipids and nucleic acids that can affect its function. This mini-review offers an overview of this process with special emphasis on protein tyrosine nitration in plants and its involvement in the process of nitrosative stress. PMID:19826215

  1. THE STRUCTURE OF THE ULTRAVIOLET ABSORPTION SPECTRA OF CERTAIN PROTEINS AND AMINO ACIDS

    PubMed Central

    Coulter, Calvin B.; Stone, Florence M.; Kabat, Elvin A.

    1936-01-01

    1. The absorption spectra of a number of proteins in the region 2500 to 3000 A. have been found to comprise from six to nine narrow bands. In consequence of variation in the relative intensity of these bands from protein to protein, the absorption curve has a characteristic configuration for each protein. 2. These bands correspond closely in position with the narrow bands which appear in the absorption spectra of tryptophan, tyrosin, and phenylalanine. Tryptophan and tyrosin each present three bands, phenylalanine shows nine. 3. The bands in the proteins are accordingly attributed to these amino acids. In the proteins the bands are displaced from the positions which they occupy in the uncombined amino acids, in most instances, by 10 to 35 A. toward longer wavelengths. 4. The absorption spectrum of Pneumococcus Type I antibody resembles that of normal pseudoglobulin but shows characteristic differences. PMID:19872958

  2. OGP functionalized phenylalanine-based poly(ester urea) for enhancing osteoinductive potential of human mesenchymal stem cells.

    PubMed

    Policastro, Gina M; Lin, Fei; Smith Callahan, Laura A; Esterle, Andrew; Graham, Matthew; Sloan Stakleff, Kimberly; Becker, Matthew L

    2015-04-13

    Amino acid-based poly(ester urea)s (PEU) are high modulus, resorbable polymers with many potential uses, including the surgical repair of bone defects. In vitro and in vivo studies have previously shown that phenylalanine-based PEUs have nontoxic hydrolytic byproducts and tunable degradation times. Phenylalanine PEUs (poly(1-PHE-6)) have been further modified by tethering osteogenic growth peptide (OGP) to tyrosine-based monomer subunits. These OGP-tethered PEUs have been fabricated into porous scaffolds and cultured in vitro to examine their effect on differentiation of human mesenchymal stem cells (hMSCs) toward the osteogenic lineage. The influence of tethered OGP on the hMSC proliferation and differentiation profile was measured using immunohistochemistry, biochemistry, and quantitative real time polymerase chain reaction (qRT-PCR). In vitro data indicated an enhanced expression of BSP by 130-160% for hMSCs on OGP-tethered scaffolds compared to controls. By 4 weeks, there was a significant drop (60-85% decrease) in BSP expression on OGP-functionalized scaffolds, which is characteristic of osteogenic differentiation. ALP and OSC expression was significantly enhanced for OGP-functionalized scaffolds by week 4, with values reaching 145% and 300% greater, respectively, compared to nonfunctionalized controls. In vivo subcutaneous implantation of poly(1-PHE-6) scaffolds revealed significant tissue-scaffold integration, as well as the promotion of both osteogenesis and angiogenesis.

  3. Liquid-liquid distribution of aromatic α-amino acids in multicomponent systems

    NASA Astrophysics Data System (ADS)

    Korenman, Ya. I.; Mokshina, N. Ya.; Pakhomova, O. A.

    2010-02-01

    Distribution coefficients and recovery factors of phenylalanine, tyrosine, and tryptophan are measured in extraction systems with butanol, pentanol, acetone, and ethyl acetate, their binary and ternary mixtures, and water-soluble polymers. Extraction conditions—extractant composition, salting-out agents, and pH—are optimized. Efficient systems providing maximum quantitative characteristics of the process of liquid-liquid distribution of aromatic α-amino acids are proposed.

  4. Aspergillus oryzae pathways that convert phenylalanine into the flavor volatile 2-phenylethanol.

    PubMed

    Masuo, Shunsuke; Osada, Lisa; Zhou, Shengmin; Fujita, Tomoya; Takaya, Naoki

    2015-04-01

    The filamentous fungus Aspergillus oryzae RIB40 produced 2-phenylethanol (PE) when cultured in minimum medium containing l-phenylalanine as a sole source of nitrogen. The fungus accumulated less PE in the absence of l-phenylalanine, indicating that it converted l-phenylalanine to PE. The PE production associated with fungal glucose consumption was repressed by exogenous ammonium, indicating that nitrogen-metabolite repression controls the pathway that produces PE. We identified the A. oryzae ppdA gene that is expressed at high levels in the presence of exogenous l-phenylalanine and its encoded protein was an active phenylpyruvate decarboxylase. The fungal genome encodes predicted aminotransferases of phenylalanine and PE dehydrogenases, which, together with PpdA, are likely to constitute an Erlich pathway similar to that in Saccharomyces cerevisiae that produces PE. We also identified an A. oryzae aromatic amino acid decarboxylase (AadA) that converted l-phenylalanine to phenylethylamine (PEA), and phenylalanine-inducible PEA oxidase activity in fungal cell extracts, and found that both constitute an alternative pathway through which PEA generates PE. Incubating fungal cultures with l-[(2)H8] phenylalanine to distinguish PE produced by these pathways, indicated that the fungus produced PE by both pathways, but to a greater extent by the Erlich pathway. Gene disruption of ppdA and aadA showed that both pathways participate in the fungal conversion of l-phenylalanine to PE.

  5. Glycoproteomic Analysis of Prostate Cancer Tissues by SWATH Mass Spectrometry Discovers N-acylethanolamine Acid Amidase and Protein Tyrosine Kinase 7 as Signatures for Tumor Aggressiveness*

    PubMed Central

    Liu, Yansheng; Chen, Jing; Sethi, Atul; Li, Qing K.; Chen, Lijun; Collins, Ben; Gillet, Ludovic C. J.; Wollscheid, Bernd; Zhang, Hui; Aebersold, Ruedi

    2014-01-01

    The identification of biomarkers indicating the level of aggressiveness of prostate cancer (PCa) will address the urgent clinical need to minimize the general overtreatment of patients with non-aggressive PCa, who account for the majority of PCa cases. Here, we isolated formerly N-linked glycopeptides from normal prostate (n = 10) and from non-aggressive (n = 24), aggressive (n = 16), and metastatic (n = 25) PCa tumor tissues and analyzed the samples using SWATH mass spectrometry, an emerging data-independent acquisition method that generates a single file containing fragment ion spectra of all ionized species of a sample. The resulting datasets were searched using a targeted data analysis strategy in which an a priori spectral reference library representing known N-glycosites of the human proteome was used to identify groups of signals in the SWATH mass spectrometry data. On average we identified 1430 N-glycosites from each sample. Out of those, 220 glycoproteins showed significant quantitative changes associated with diverse biological processes involved in PCa aggressiveness and metastasis and indicated functional relationships. Two glycoproteins, N-acylethanolamine acid amidase and protein tyrosine kinase 7, that were significantly associated with aggressive PCa in the initial sample cohort were further validated in an independent set of patient tissues using tissue microarray analysis. The results suggest that N-acylethanolamine acid amidase and protein tyrosine kinase 7 may be used as potential tissue biomarkers to avoid overtreatment of non-aggressive PCa. PMID:24741114

  6. Transformation of phenol into phenylalanine by a methanogenic consortium

    SciTech Connect

    Lepine, F.; Milot, S.; Beaudet, R.; Villemur, R.

    1996-03-01

    Phenol is a widely used chemical found in many wastewaters of industrial origin. The degradation of phenol by methanogenic bacterial consortia has been reported by many investigators. To better characterise the metabolism of this consortium, a new metabolic pathway of benzoic acid, an intermediary in the degradation of phenol, is reported. This study describes the transformations of benzoic acid into 3-phenylpropionic acid and phenylalanine. 25 refs., 5 figs.

  7. Synthesis, uptake mechanism characterization and biological evaluation of 18F labeled fluoroalkyl phenylalanine analogs as potential PET imaging agents

    PubMed Central

    Wang, Limin; Qu, Wenchao; Lieberman, Brian P.; Plössl, Karl; Kung, Hank F.

    2010-01-01

    Introduction Amino acids based tracers represent a promising class of tumor metabolic imaging agents with successful clinical applications. Two new phenylalanine derivatives, p-(2-[18F]fluoroethyl)-L-phenylalanine (FEP, [18F]2) and p-(3-[18F]fluoropropyl)-L-phenylalanine (FPP, [18F]3) were synthesized and evaluated in comparison to clinically utilized O-(2-[18F]fluoroethyl)-L-tyrosine (FET, [18F]1). Methods FEP ([18F]2) and FPP ([18F]3) were successfully synthesized by a rapid and efficient two-step nucleophilic fluorination of tosylate precursors and deprotection reaction. In vitro cell uptake studies were carried out in 9L glioma cells. In vivo studies, 9L tumor xenografts were implanted in Fisher 344 rats. Results FEP ([18F]2) and FPP ([18F]3) could be efficiently labeled within 90 min with good enantiomeric purity (>95%), good yield (11–37%) and high specific activity (21–69 GBq/μmol). Cell uptake studies showed FEP had higher uptake than FPP as well as reference ligand FET ([18F]1). Uptake mechanism studies suggested that FEP is a selective substrate for system L and prefers its subtype LAT1. In vivo biodistribution studies demonstrated FEP had specific accumulation in tumor cells and tumor to background ratio reached 1.45 at 60 min. Small animal PET imaging studies showed FEP was comparable to FET for imaging rats bearing 9L tumor model. FEP had high uptake in 9L tumor compared to surrounding tissue and was quickly excreted through urinary tract. Conclusion Biological evaluations indicate that FEP ([18F]2) is a potential useful tracer for tumor imaging with PET. PMID:21220129

  8. Effect of methyl jasmonate application to grapevine leaves on grape amino acid content.

    PubMed

    Garde-Cerdán, Teresa; Portu, Javier; López, Rosa; Santamaría, Pilar

    2016-07-15

    Over the last few years, considerable attention has been paid to the application of elicitors to vineyard. However, research about the effect of elicitors on grape amino acid content is scarce. Therefore, the aim of this study was to evaluate the influence of foliar application of methyl jasmonate on must amino acid content. Results revealed that total amino acid content was not modified by the application of methyl jasmonate. However, the individual content of certain amino acids was increased as consequence of methyl jasmonate foliar application, i.e., histidine, serine, tryptophan, phenylalanine, tyrosine, asparagine, methionine, and lysine. Among them, phenylalanine content was considerably increased; this amino acid is precursor of phenolic and aromatic compounds. In conclusion, foliar application of methyl jasmonate improved must nitrogen composition. This finding suggests that methyl jasmonate treatment might be conducive to obtain wines of higher quality since must amino acid composition could affect the wine volatile composition and the fermentation kinetics.

  9. Effect of methyl jasmonate application to grapevine leaves on grape amino acid content.

    PubMed

    Garde-Cerdán, Teresa; Portu, Javier; López, Rosa; Santamaría, Pilar

    2016-07-15

    Over the last few years, considerable attention has been paid to the application of elicitors to vineyard. However, research about the effect of elicitors on grape amino acid content is scarce. Therefore, the aim of this study was to evaluate the influence of foliar application of methyl jasmonate on must amino acid content. Results revealed that total amino acid content was not modified by the application of methyl jasmonate. However, the individual content of certain amino acids was increased as consequence of methyl jasmonate foliar application, i.e., histidine, serine, tryptophan, phenylalanine, tyrosine, asparagine, methionine, and lysine. Among them, phenylalanine content was considerably increased; this amino acid is precursor of phenolic and aromatic compounds. In conclusion, foliar application of methyl jasmonate improved must nitrogen composition. This finding suggests that methyl jasmonate treatment might be conducive to obtain wines of higher quality since must amino acid composition could affect the wine volatile composition and the fermentation kinetics. PMID:26948648

  10. Neutral penta- and hexacoordinate silicon(IV) complexes containing two bidentate ligands derived from the alpha-amino acids (S)-alanine, (S)-phenylalanine, and (S)-tert-leucine.

    PubMed

    Cota, Smaranda; Beyer, Matthias; Bertermann, Rüdiger; Burschka, Christian; Götz, Kathrin; Kaupp, Martin; Tacke, Reinhold

    2010-06-11

    The neutral hexacoordinate silicon(IV) complex 6 (SiO(2)N(4) skeleton) and the neutral pentacoordinate silicon(IV) complexes 7-11 (SiO(2)N(2)C skeletons) were synthesized from Si(NCO)(4) and RSi(NCO)(3) (R = Me, Ph), respectively. The compounds were structurally characterized by solid-state NMR spectroscopy (6-11), solution NMR spectroscopy (6 and 10), and single-crystal X-ray diffraction (8 and 11 were studied as the solvates 8 x CH(3)CN and 11 x C(5)H(12) x 0.5 CH(3)CN, respectively). The silicon(IV) complexes 6 (octahedral Si-coordination polyhedron) and 7-11 (trigonal-bipyramidal Si-coordination polyhedra) each contain two bidentate ligands derived from an alpha-amino acid: (S)-alanine, (S)-phenylalanine, or (S)-tert-leucine. The deprotonated amino acids act as monoanionic (6) or as mono- and dianionic ligands (7-11). The experimental investigations were complemented by computational studies of the stereoisomers of 6 and 7.

  11. 4-Vinylphenol biosynthesis from cellulose as the sole carbon source using phenolic acid decarboxylase- and tyrosine ammonia lyase-expressing Streptomyces lividans.

    PubMed

    Noda, Shuhei; Kawai, Yoshifumi; Tanaka, Tsutomu; Kondo, Akihiko

    2015-03-01

    Streptomyces lividans was adopted as a host strain for 4-vinylphenol (4VPh) production directly from cellulose. In order to obtain novel phenolic acid decarboxylase (PAD) expressed in S. lividans, PADs distributed among Streptomyces species were screened. Three novel PADs, derived from Streptomycessviceus, Streptomyceshygroscopicus, and Streptomycescattleya, were successfully obtained and expressed in S. lividans. S. sviceus PAD (SsPAD) could convert p-hydroxycinnamic acid (pHCA) to 4VPh more efficiently than the others both in vitro and in vivo. For 4VPh production directly from cellulose, l-tyrosine ammonia lyase derived from Rhodobacter sphaeroides and SsPAD were introduced into endoglucanase-secreting S. lividans, and the 4VPh biosynthetic pathway was constructed therein. The created transformants successfully produced 4VPh directly from cellulose.

  12. 21 CFR 582.5590 - Phenylalanine.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 6 2011-04-01 2011-04-01 false Phenylalanine. 582.5590 Section 582.5590 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS... 1 § 582.5590 Phenylalanine. (a) Product. Phenylalanine (L- and DL-forms). (b) Conditions of...

  13. 21 CFR 582.5590 - Phenylalanine.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 6 2012-04-01 2012-04-01 false Phenylalanine. 582.5590 Section 582.5590 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS... 1 § 582.5590 Phenylalanine. (a) Product. Phenylalanine (L- and DL-forms). (b) Conditions of...

  14. 21 CFR 582.5590 - Phenylalanine.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 6 2014-04-01 2014-04-01 false Phenylalanine. 582.5590 Section 582.5590 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS... 1 § 582.5590 Phenylalanine. (a) Product. Phenylalanine (L- and DL-forms). (b) Conditions of...

  15. 21 CFR 582.5590 - Phenylalanine.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Phenylalanine. 582.5590 Section 582.5590 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS... 1 § 582.5590 Phenylalanine. (a) Product. Phenylalanine (L- and DL-forms). (b) Conditions of...

  16. 21 CFR 582.5590 - Phenylalanine.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 6 2013-04-01 2013-04-01 false Phenylalanine. 582.5590 Section 582.5590 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS... 1 § 582.5590 Phenylalanine. (a) Product. Phenylalanine (L- and DL-forms). (b) Conditions of...

  17. Fluorescent biphenyl derivatives of phenylalanine suitable for protein modification.

    PubMed

    Chen, Shengxi; Fahmi, Nour Eddine; Bhattacharya, Chandrabali; Wang, Lin; Jin, Yuguang; Benkovic, Stephen J; Hecht, Sidney M

    2013-11-26

    In a recent study, we demonstrated that structurally compact fluorophores incorporated into the side chains of amino acids could be introduced into dihydrofolate reductase from Escherichia coli (ecDHFR) with minimal disruption of protein structure or function, even when the site of incorporation was within a folded region of the protein. The modified proteins could be employed for FRET measurements, providing sensitive monitors of changes in protein conformation. The very favorable results achieved in that study encouraged us to prepare additional fluorescent amino acids of potential utility for studying protein dynamics. Presently, we describe the synthesis and photophysical characterization of four positional isomers of biphenyl-phenylalanine, all of which were found to exhibit potentially useful fluorescent properties. All four phenylalanine derivatives were used to activate suppressor tRNA transcripts and incorporated into multiple positions of ecDHFR. All phenylalanine derivatives were incorporated with good efficiency into position 16 of ecDHFR and afforded modified proteins that consumed NADPH at rates up to about twice the rate measured for wild type. This phenomenon has been noted on a number of occasions previously and shown to be due to an increase in the off-rate of tetrahydrofolate from the enzyme, altering a step that is normally rate limiting. When introduced into sterically accessible position 49, the four phenylalanine derivatives afforded DHFRs having catalytic function comparable to wild type. The four phenylalanine derivatives were also introduced into position 115 of ecDHFR, which is known to be a folded region of the protein less tolerant of structural alteration. As anticipated, significant differences were noted in the catalytic efficiencies of the derived proteins. The ability of two of the sizable biphenyl-phenylalanine derivatives to be accommodated at position 115 with minimal perturbation of DHFR function is attributed to rotational

  18. Ultraviolet resonance Raman and absorption difference spectroscopy of myoglobins: titration behavior of individual tyrosine residues.

    PubMed

    Asher, S A; Larkin, P J; Teraoka, J

    1991-06-18

    The UV resonance Raman spectra of horse and sperm whale myoglobin excited at 240 nm show bands between 600 and 1700 cm-1 which derive from tyrosyl and tryptophyl residues. No significant contribution from phenylalanine and peptide backbone vibrations occurs at this excitation wavelength. We examine the pH dependence of the UV resonance Raman and UV absorption difference spectra of these myoglobins to correlate the local protein environment of the tyrosyl residues as given by the protein crystal structure to their pKa values, molar absorptivities, and Raman cross sections. Some of our pKa values for the tyrosinate residues of horse Mb differ from those of previous studies. We show that the lambda max values, the molar absorptivities, and the Raman cross sections are sensitive to the local environment of the tyrosinate residues in the protein. We relate differences in the tyrosyl absorption spectra to differences in Raman cross sections. In addition, we discuss the importance to the Raman cross sections of the local electromagnetic field enhancement due to the dielectric environment of the tyrosinate residues in the protein. This local field should scale the Raman cross sections in a way useful as a probe of the average aromatic amino acid residue environment.

  19. Hybride magnetic nanostructure based on amino acids functionalized polypyrrole

    NASA Astrophysics Data System (ADS)

    Nan, Alexandrina; Bunge, Alexander; Turcu, Rodica

    2015-12-01

    Conducting polypyrrole is especially promising for many commercial applications because of its unique optical, electric, thermal and mechanical properties. We report the synthesis and characterization of novel pyrrole functionalized monomers and core-shell hybrid nanostructures, consisting of a conjugated polymer layer (amino acids functionalized pyrrole copolymers) and a magnetic nanoparticle core. For functionalization of the pyrrole monomer we used several amino acids: tryptophan, leucine, phenylalanine, serine and tyrosine. These amino acids were linked via different types of hydrophobic linkers to the nitrogen atom of the pyrrole monomer. The magnetic core-shell hybrid nanostructures are characterized by various methods such as FTIR spectroscopy, transmission electron microscopy (TEM) and magnetic measurements.

  20. Hybride magnetic nanostructure based on amino acids functionalized polypyrrole

    SciTech Connect

    Nan, Alexandrina Bunge, Alexander; Turcu, Rodica

    2015-12-23

    Conducting polypyrrole is especially promising for many commercial applications because of its unique optical, electric, thermal and mechanical properties. We report the synthesis and characterization of novel pyrrole functionalized monomers and core-shell hybrid nanostructures, consisting of a conjugated polymer layer (amino acids functionalized pyrrole copolymers) and a magnetic nanoparticle core. For functionalization of the pyrrole monomer we used several amino acids: tryptophan, leucine, phenylalanine, serine and tyrosine. These amino acids were linked via different types of hydrophobic linkers to the nitrogen atom of the pyrrole monomer. The magnetic core-shell hybrid nanostructures are characterized by various methods such as FTIR spectroscopy, transmission electron microscopy (TEM) and magnetic measurements.

  1. Aspartame as a source of essential phenylalanine for the growth of oral anaerobes.

    PubMed

    Wyss, C

    1993-04-15

    Phenylalanine and aspartic acid requirements were determined for 13 species of oral bacteria using the chemically defined medium OMIZ-W1. None of Actinobacillus actinomycetemcomitans, Bacteroides forsythus, Eikenella corrodens, Selenomonas sputigena, Treponema pectinovorum, T. socranskii, or Wolinella recta required either of these amino acid constituents of aspartame (L-aspartyl-L-phenylalanine methylester). Phenylalanine was essential for the growth of Capnocytophaga gingivalis, Eubacterium timidum, Fusobacterium nucleatum, Porphyromonas gingivalis, T. denticola, and T. vincentii, while aspartic acid was not required. With the exception of E. timidum, all phenylalanine-dependent strains could grow when the free amino acid was replaced by aspartame at concentrations at least 10-fold lower than those used for aspartame as an artificial sweetener.

  2. p-Hydroxyphenylacetaldehyde is the major product of L-tyrosine oxidation by activated human phagocytes. A chloride-dependent mechanism for the conversion of free amino acids into reactive aldehydes by myeloperoxidase.

    PubMed

    Hazen, S L; Hsu, F F; Heinecke, J W

    1996-01-26

    Reactive aldehydes generated during lipid peroxidation have been implicated in the pathogenesis of atherosclerosis as well as other inflammatory diseases. A potential catalyst for such reactions is myeloperoxidase, a hemeprotein secreted by activated phagocytes. We now report that activated neutrophils utilize the myeloperoxidase-H2O2-chloride system to convert L-tyrosine to p-hydroxyphenylacetaldehyde. Production of p-hydroxyphenylacetaldehyde was nearly quantitative at physiological concentrations of L-tyrosine and chloride. Aldehyde generation required myeloperoxidase, H2O2, L-tyrosine, and chloride ion; it was inhibited by the H2O2 scavenger catalase and by the heme poisons azide and cyanide. Phorbol ester- and calcium ionophore-stimulated human neutrophils likewise generated p-hydroxyphenylacetaldehyde from L-tyrosine by a pathway inhibited by azide, cyanide, and catalase. Aldehyde production accounted for 75% of H2O2 generated by optimally stimulated neutrophils at plasma concentrations of L-tyrosine and chloride. Collectively, these results indicate that activated phagocytes, under physiological conditions, utilize myeloperoxidase to execute the chloride-dependent conversion of L-tyrosine to the lipid-soluble aldehyde, p-hydroxyphenylacetaldehyde, in near quantitative yield. Moreover, like aldehydes derived from lipid peroxidation, amino acid-derived aldehydes may exert potent biological effects in vascular lesions and other sites of inflammation.

  3. Amino acids, precursors for cationic and anionic intercalation synthesis and characterization of amino acid pillared materials

    NASA Astrophysics Data System (ADS)

    Fudala, Á.; Pálinkó, I.; Kiricsi, I.

    1999-05-01

    The preparation and characterization of amino acid pillared materials are reported in this contribution. Host substances were Na-montmorillonite for cationic and hydrotalcite for anionic pillaring. Guest molecules were L-phenylalanine and L-tyrosine. The pillared materials were characterized by powder X-ray diffraction, BET measurements and FT-IR spectroscopy. Pillaring was successful: the layers propped open and the basal distances increased significantly. For hydrotalcite this increase was always significantly larger than for montmorillonite. This fact indicated that the spatial arrangement of the amino acid moieties is widely different. A model for this arrangement is given.

  4. A Possible Role of Divalent Manganese Ions in the Photoinduction of Phenylalanine Ammonia-Lyase

    PubMed Central

    Engelsma, G.

    1972-01-01

    Divalent Mn ions cause an increase in the level of phenylalanine ammonia-lyase in gherkin hypocotyls. With the exception of Mg ions, which had a small effect, no other metal ion has so far been found which could replace the Mn ion in this respect. Invertase and peroxidase were not significantly affected by the Mn treatment. The increase in phenylalanine ammonialyase activity is explained by the removal, under the influence of Mn ions, of hydroxycinnamic acids, which cause repression of phenylalanine ammonia-lyase synthesis and/or inactivation of phenylalanine ammonia-lyase. Arguments are advanced for the hypothesis that photochemical transformations of Mn complexes are involved in the photoinduction of phenylalanine ammonia-lyase in dark-grown gherkin seedlings. PMID:16658225

  5. Targeted tyrosine iodination in a multi-tyrosine vasopressin analog.

    PubMed

    Durr, Jacques A; Blankenship, Mary; Chauhan, Satendra S; Pennington, Michael W

    2007-11-01

    Iodination of the conserved 2-tyrosine (Tyr(2)) residue in the pressin and tocin rings of arginine- or lysine-vasopressin (AVP or LVP), and oxytocin, respectively, impairs binding to their respective receptors. Synthetic antagonists that have their Tyr(2) either replaced by another amino acid or irreversibly blocked by an O-methyl or O-ethyl ether, but have, instead, an iodinatable phenol moiety outside the pressin/tocin ring, are used for radiolabeling. We explored another approach to avoid iodinating Tyr(2) by capping this residue with a reversible O-acetyl group, incorporated during peptide synthesis. The O-acetyl-Tyr(2) LVP peptide, with a free iodinatable tyrosine attached to the epsilon-amine of 8-lysine, is iodinated at a neutral pH and purified by reverse-phase high-pressure liquid chromatography (HPLC) at an acidic pH, conditions under which the O-acetyl groups are stable. Deacetylation with hydroxylamine is selective, and leaves intact the disulfide bridge. The marked shortening of the HPLC retention time after deblocking produces a chemically homogeneous label, iodinated exclusively on the free tyrosine residue attached to the epsilon-amine of LVP. Hitherto, this (125)I labeled vasopressin agonist could be obtained only in low yield, via conjugation labeling with iodinated N-t-Boc-tyrosine succinimidyl ester. This fully reversible tyrosine protection strategy does not require special equipment, and retains the conserved Tyr(2), typical of vasopressin and oxytocin agonists.

  6. Tyrosine Detoxification Is an Essential Trait in the Life History of Blood-Feeding Arthropods.

    PubMed

    Sterkel, Marcos; Perdomo, Hugo D; Guizzo, Melina G; Barletta, Ana Beatriz F; Nunes, Rodrigo D; Dias, Felipe A; Sorgine, Marcos H F; Oliveira, Pedro L

    2016-08-22

    Blood-feeding arthropods are vectors of infectious diseases such as dengue, Zika, Chagas disease, and malaria [1], and vector control is essential to limiting disease spread. Because these arthropods ingest very large amounts of blood, a protein-rich meal, huge amounts of amino acids are produced during digestion. Previous work on Rhodnius prolixus, a vector of Chagas disease, showed that, among all amino acids, only tyrosine degradation enzymes were overexpressed in the midgut compared to other tissues [2]. Here we demonstrate that tyrosine detoxification is an essential trait in the life history of blood-sucking arthropods. We found that silencing Rhodnius tyrosine aminotransferase (TAT) and 4-hydroxyphenylpyruvate dioxygenase (HPPD), the first two enzymes of the phenylalanine/tyrosine degradation pathway, caused the death of insects after a blood meal. This was confirmed by using the HPPD inhibitor mesotrione, which selectively killed hematophagous arthropods but did not affect non-hematophagous insects. In addition, mosquitoes and kissing bugs died after feeding on mice that had previously received a therapeutic effective oral dose (1 mg/kg) of nitisinone, another HPPD inhibitor used in humans for the treatment of tyrosinemia type I [3]. These findings indicate that HPPD (and TAT) can be a target for the selective control of blood-sucking disease vector populations. Because HPPD inhibitors are extensively used as herbicides and in medicine, these compounds may provide an alternative less toxic to humans and more environmentally friendly than the conventional neurotoxic insecticides that are currently used, with the ability to affect only hematophagous arthropods.

  7. Tyrosine Detoxification Is an Essential Trait in the Life History of Blood-Feeding Arthropods.

    PubMed

    Sterkel, Marcos; Perdomo, Hugo D; Guizzo, Melina G; Barletta, Ana Beatriz F; Nunes, Rodrigo D; Dias, Felipe A; Sorgine, Marcos H F; Oliveira, Pedro L

    2016-08-22

    Blood-feeding arthropods are vectors of infectious diseases such as dengue, Zika, Chagas disease, and malaria [1], and vector control is essential to limiting disease spread. Because these arthropods ingest very large amounts of blood, a protein-rich meal, huge amounts of amino acids are produced during digestion. Previous work on Rhodnius prolixus, a vector of Chagas disease, showed that, among all amino acids, only tyrosine degradation enzymes were overexpressed in the midgut compared to other tissues [2]. Here we demonstrate that tyrosine detoxification is an essential trait in the life history of blood-sucking arthropods. We found that silencing Rhodnius tyrosine aminotransferase (TAT) and 4-hydroxyphenylpyruvate dioxygenase (HPPD), the first two enzymes of the phenylalanine/tyrosine degradation pathway, caused the death of insects after a blood meal. This was confirmed by using the HPPD inhibitor mesotrione, which selectively killed hematophagous arthropods but did not affect non-hematophagous insects. In addition, mosquitoes and kissing bugs died after feeding on mice that had previously received a therapeutic effective oral dose (1 mg/kg) of nitisinone, another HPPD inhibitor used in humans for the treatment of tyrosinemia type I [3]. These findings indicate that HPPD (and TAT) can be a target for the selective control of blood-sucking disease vector populations. Because HPPD inhibitors are extensively used as herbicides and in medicine, these compounds may provide an alternative less toxic to humans and more environmentally friendly than the conventional neurotoxic insecticides that are currently used, with the ability to affect only hematophagous arthropods. PMID:27476595

  8. Structural evolution of differential amino acid effector regulation in plant chorismate mutases.

    PubMed

    Westfall, Corey S; Xu, Ang; Jez, Joseph M

    2014-10-10

    Chorismate mutase converts chorismate into prephenate for aromatic amino acid biosynthesis. To understand the molecular basis of allosteric regulation in the plant chorismate mutases, we analyzed the three Arabidopsis thaliana chorismate mutase isoforms (AtCM1-3) and determined the x-ray crystal structures of AtCM1 in complex with phenylalanine and tyrosine. Functional analyses show a wider range of effector control in the Arabidopsis chorismate mutases than previously reported. AtCM1 is activated by tryptophan with phenylalanine and tyrosine acting as negative effectors; however, tryptophan, cysteine, and histidine activate AtCM3. AtCM2 is a nonallosteric form. The crystal structure of AtCM1 in complex with tyrosine and phenylalanine identifies differences in the effector sites of the allosterically regulated yeast enzyme and the other two Arabidopsis isoforms. Site-directed mutagenesis of residues in the effector site reveals key features leading to differential effector regulation in these enzymes. In AtCM1, mutations of Gly-213 abolish allosteric regulation, as observed in AtCM2. A second effector site position, Gly-149 in AtCM1 and Asp-132 in AtCM3, controls amino acid effector specificity in AtCM1 and AtCM3. Comparisons of chorismate mutases from multiple plants suggest that subtle differences in the effector site are conserved in different lineages and may lead to specialized regulation of this branch point enzyme.

  9. Influence of magnetic fields on the hydration process of amino acids: vibrational spectroscopy study of L-phenylalanine and L-glutamine.

    PubMed

    De Ninno, Antonella; Congiu Castellano, Agostina

    2014-02-01

    Attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy has been used to investigate the effect of weak electromagnetic fields on the structure of L-glutamine (L-Gln) and L-phenylalnine (L-Phe) in aqueous solution. It has been found that the exposure to a DC field or a 50 Hz AC field, for a short time induces modifications in the spectra of exposed samples in agreement with our preceding observations on glutamic acid. Furthermore, the acid-base equilibrium has been investigated by using the ratio of the intensity of the deprotonated on protonated species. In the case of L-Phe, the exposure induces a measurable shift of acid dissociation constant pKa1 out of the experimental errors, while in case of L-Gln, the effect is under the limit detectable with this method. The phenomenon of the shift of the acid-base equilibrium has been connected elsewhere to modification of the water-water hydrogen bonds in the water around both the backbone and the residue (R). Here we suggest that the magnetic field modifies the water structure around the molecules and changes the hydrophobic interactions allowing the molecules of amino acids to aggregate. The differences observed in the behavior of L-Phe and L-Gln may be related to the differences in the polarity of their residues.

  10. A Novel Phosphatidic Acid-Protein-tyrosine Phosphatase D2 Axis Is Essential for ERBB2 Signaling in Mammary Epithelial Cells*

    PubMed Central

    Ramesh, Mathangi; Krishnan, Navasona; Muthuswamy, Senthil K.; Tonks, Nicholas K.

    2015-01-01

    We used a loss-of-function screen to investigate the role of classical protein-tyrosine phosphatases (PTPs) in three-dimensional mammary epithelial cell morphogenesis and ERBB2 signaling. The study revealed a novel role for PTPD2 as a positive regulator of ERBB2 signaling. Suppression of PTPD2 attenuated the ERBB2-induced multiacinar phenotype in three-dimensional cultures specifically by inhibiting ERBB2-mediated loss of polarity and lumen filling. In contrast, overexpression of PTPD2 enhanced the ERBB2 phenotype. We also found that a lipid second messenger, phosphatidic acid, bound PTPD2 in vitro and enhanced its catalytic activity. Small molecule inhibitors of phospholipase D (PLD), an enzyme that produces phosphatidic acid in cells, also attenuated the ERBB2 phenotype. Exogenously added phosphatidic acid rescued the PLD-inhibition phenotype, but only when PTPD2 was present. These findings illustrate a novel pathway involving PTPD2 and the lipid second messenger phosphatidic acid that promotes ERBB2 function. PMID:25681440

  11. Increased levels of tyrosine hydroxylase and glutamic acid decarboxylase in locus coeruleus neurons after rapid eye movement sleep deprivation in rats.

    PubMed

    Majumdar, S; Mallick, B N

    2003-03-01

    Norepinephrine, acetylcholine and GABA levels alter during rapid eye movement (REM) sleep and its deprivation. Increased synthesis of those neurotransmitters is necessary for their sustained release. Hence, in this study, the concentrations of tyrosine hydroxylase (TH), choline acetyl transferase (ChAT) and glutamic acid decarboxylase (GAD), the enzymes responsible for their synthesis, were immunohistochemically estimated within the neurons in locus coeruleus, laterodorsal tegmentum and pedunculopontine tegmentum and medial preoptic area in REM sleep deprived and control rats. It was observed that as compared to controls, deprivation increased TH and GAD significantly in the locus coeruleus only, while in other areas, they remained unchanged. The findings help explaining the mechanism of increase in neurotransmitter levels in the brain after REM sleep deprivation and their significance has been discussed.

  12. Effects of running the Bostom Marathon on plasma concentrations of large neutral amino acids

    NASA Technical Reports Server (NTRS)

    Conlay, L. A.; Wurtman, R. J.; Lopez G-Coviella, I.; Blusztajn, J. K.; Vacanti, C. A.; Logue, M.; During, M.; Caballero, B.; Maher, T. J.; Evoniuk, G.

    1989-01-01

    Plasma large neutral amino acid concentrations were measured in thirty-seven subjects before and after completing the Boston Marathon. Concentrations of tyrosine, phenylalanine, and methionine increased, as did their 'plasma ratios' (i.e., the ratio of each amino acid's concentration to the summed plasma concentrations of the other large neutral amino acids which compete with it for brain uptake). No changes were noted in the plasma concentrations of tryptophan, leucine, isoleucine, nor valine; however, the 'plasma ratios' of valine, leucine, and isoleucine all decreased. These changes in plasma amino acid patterns may influence neurotransmitter synthesis.

  13. A critical tyrosine residue determines the uncoupling protein-like activity of the yeast mitochondrial oxaloacetate carrier.

    PubMed

    Luévano-Martínez, Luis A; Barba-Ostria, Carlos; Araiza-Olivera, Daniela; Chiquete-Félix, Natalia; Guerrero-Castillo, Sergio; Rial, Eduardo; Georgellis, Dimitris; Uribe-Carvajal, Salvador

    2012-04-01

    The mitochondrial Oac (oxaloacetate carrier) found in some fungi and plants catalyses the uptake of oxaloacetate, malonate and sulfate. Despite their sequence similarity, transport specificity varies considerably between Oacs. Indeed, whereas ScOac (Saccharomyces cerevisiae Oac) is a specific anion-proton symporter, the YlOac (Yarrowia lipolytica Oac) has the added ability to transport protons, behaving as a UCP (uncoupling protein). Significantly, we identified two amino acid changes at the matrix gate of YlOac and ScOac, tyrosine to phenylalanine and methionine to leucine. We studied the role of these amino acids by expressing both wild-type and specifically mutated Oacs in an Oac-null S. cerevisiae strain. No phenotype could be associated with the methionine to leucine substitution, whereas UCP-like activity was dependent on the presence of the tyrosine residue normally expressed in the YlOac, i.e. Tyr-ScOac mediated proton transport, whereas Phe-YlOac lost its protonophoric activity. These findings indicate that the UCP-like activity of YlOac is determined by the tyrosine residue at position 146.

  14. The Tyrosine Aminomutase TAM1 Is Required for β-Tyrosine Biosynthesis in Rice

    PubMed Central

    Yan, Jian; Aboshi, Takako; Teraishi, Masayoshi; Strickler, Susan R.; Spindel, Jennifer E.; Tung, Chih-Wei; Takata, Ryo; Matsumoto, Fuka; Maesaka, Yoshihiro; McCouch, Susan R.; Okumoto, Yutaka; Mori, Naoki; Jander, Georg

    2015-01-01

    Non-protein amino acids, often isomers of the standard 20 protein amino acids, have defense-related functions in many plant species. A targeted search for jasmonate-induced metabolites in cultivated rice (Oryza sativa) identified (R)-β-tyrosine, an isomer of the common amino acid (S)-α-tyrosine in the seeds, leaves, roots, and root exudates of the Nipponbare cultivar. Assays with 119 diverse cultivars showed a distinct presence/absence polymorphism, with β-tyrosine being most prevalent in temperate japonica cultivars. Genetic mapping identified a candidate gene on chromosome 12, which was confirmed to encode a tyrosine aminomutase (TAM1) by transient expression in Nicotiana benthamiana and in vitro enzyme assays. A point mutation in TAM1 eliminated β-tyrosine production in Nipponbare. Rice cultivars that do not produce β-tyrosine have a chromosome 12 deletion that encompasses TAM1. Although β-tyrosine accumulation was induced by the plant defense signaling molecule jasmonic acid, bioassays with hemipteran and lepidopteran herbivores showed no negative effects at physiologically relevant β-tyrosine concentrations. In contrast, root growth of Arabidopsis thaliana and other tested dicot plants was inhibited by concentrations as low as 1 μM. As β-tyrosine is exuded into hydroponic medium at higher concentrations, it may contribute to the allelopathic potential of rice. PMID:25901084

  15. Lack of effect of aspartame or of L-phenylalanine on photically induced myoclonus in the baboon, Papio papio.

    PubMed

    Meldrum, B S; Nanji, N; Cornell, R G

    1989-01-01

    The effects of large doses of L-phenylalanine and of aspartame on seizure susceptibility and severity have been assessed in baboons Papio papio from Senegal which show photosensitive epileptic responses similar to primary generalised epilepsy in man. L-Phenylalanine, 50, 150 or 450 mg/kg, or aspartame, 300 or 1000 mg/kg, were administered orally. Peak plasma L-phenylalanine concentrations of approximately 2000 mumoles/l occurred 1-4 h after the highest dose of L-phenylalanine or aspartame. The plasma L-phenylalanine to large neutral amino acid ratio increased approximately 30-fold at this time. Compared with water administration there were no changes in epileptic responses 1-5 h after either treatment. In this primate model of epilepsy acute increases in plasma phenylalanine concentration are neither pro- nor anticonvulsant.

  16. Aminoacyl-tRNA enrichment after a flood of labeled phenylalanine: insulin effect on muscle protein synthesis.

    PubMed

    Caso, Giuseppe; Ford, G Charles; Nair, K Sreekumaran; Garlick, Peter J; McNurlan, Margaret A

    2002-05-01

    Muscle protein synthesis in dogs measured by flooding with L-[(2)H(5)]phenylalanine (70 mg/kg) was significantly stimulated by infusion of insulin with amino acids. The stimulation of muscle protein synthesis was similar when calculated from the enrichment of phenylalanyl-tRNA (61 +/- 10%, P < 0.001), plasma phenylalanine (61 +/- 10%, P < 0.001), or tissue fluid phenylalanine (54 +/- 10%, P < 0.001). The time course for changes in enrichment of L-[(2)H(5)]phenylalanine throughout the flooding period was determined for plasma, tissue fluid, and phenylalanyl-tRNA in the basal state and during the infusion of insulin with amino acids. Enrichments of plasma free phenylalanine and phenylalanyl-tRNA were equalized between 20 and 45 min, although the enrichment of phenylalanyl-tRNA was lower at early time points. Rates of muscle protein synthesis obtained with the flooding method and calculated from plasma phenylalanine enrichment were comparable to those calculated from phenylalanyl-tRNA and also to those obtained previously with a continuous infusion of phenylalanine with phenylalanyl-tRNA as precursor. This study confirms that, with a bolus injection of labeled phenylalanine, the enrichment of aminoacyl-tRNA, the true precursor pool for protein synthesis, can be assessed from more readily sampled plasma phenylalanine.

  17. Poly(ferulic acid-co-tyrosine): Effect of the Regiochemistry on the Photophysical and Physical Properties en Route to Biomedical Applications

    PubMed Central

    2015-01-01

    The photophysical and mechanical properties of novel poly(carbonate-amide)s derived from two biorenewable resources, ferulic acid (FA) and l-tyrosine ethyl ester, were evaluated in detail. From these two bio-based precursors, a series of four monomers were generated (having amide and/or carbonate coupling units with remaining functionalities to allow for carbonate formation) and transformed to a series of four poly(carbonate-amide)s. The simplest monomer, which was biphenolic and was obtained in a single amidation synthetic step, displayed bright, visible fluorescence that was twice brighter than FA. Multidimensional fluorescence spectroscopy of the polymers in solution highlighted the strong influence that regioselectivity and the degree of polymerization have on their photophysical properties. The regiochemistry of the system had little effect on the wettability, surface free energy, and Young’s modulus (ca. 2.5 GPa) in the solid state. Confocal imaging of solvent-cast films of each polymer revealed microscopically flat surfaces with fluorescent emission deep into the visible region. Fortuitously, one of the two regiorandom polymers (obtainable from the biphenolic monomer in only an overall two synthetic steps from FA and l-tyrosine ethyl ester) displayed the most promising fluorescent properties both in the solid state and in solution, allowing for the possibility of translating this system as a self-reporting or imaging agent in future applications. To further evaluate the potential of this polymer as a biodegradable material, hydrolytic degradation studies at different pH values and temperatures were investigated. Additionally, the antioxidant properties of the degradation products of this polymer were compared with its biphenolic monomer and FA. PMID:25364040

  18. Erythrocytes encapsulated with phenylalanine hydroxylase exhibit improved pharmacokinetics and lowered plasma phenylalanine levels in normal mice.

    PubMed

    Yew, Nelson S; Dufour, Emmanuelle; Przybylska, Malgorzata; Putelat, Julie; Crawley, Cristin; Foster, Meta; Gentry, Sarah; Reczek, David; Kloss, Alla; Meyzaud, Aurélien; Horand, Françoise; Cheng, Seng H; Godfrin, Yann

    2013-08-01

    Enzyme replacement therapy is often hampered by the rapid clearance and degradation of the administered enzyme, limiting its efficacy and requiring frequent dosing. Encapsulation of therapeutic molecules into red blood cells (RBCs) is a clinically proven approach to improve the pharmacokinetics and efficacy of biologics and small molecule drugs. Here we evaluated the ability of RBCs encapsulated with phenylalanine hydroxylase (PAH) to metabolize phenylalanine (Phe) from the blood and confer sustained enzymatic activity in the circulation. Significant quantities of PAH were successfully encapsulated within murine RBCs (PAH-RBCs) with minimal loss of endogenous hemoglobin. While intravenously administered free PAH enzyme was rapidly eliminated from the blood within a few hours, PAH-RBCs persisted in the circulation for at least 10days. A single injection of PAH-RBCs was able to decrease Phe levels by nearly 80% in normal mice. These results demonstrate the ability of enzyme-loaded RBCs to metabolize circulating amino acids and highlight the potential to treat disorders of amino acid metabolism.

  19. Tyrosine - Effects on catecholamine release

    NASA Technical Reports Server (NTRS)

    Acworth, Ian N.; During, Matthew J.; Wurtman, Richard J.

    1988-01-01

    Tyrosine administration elevates striatal levels of dopamine metabolites in animals given treatments that accelerate nigrostriatal firing, but not in untreated rats. We examined the possibility that the amino acid might actually enhance dopamine release in untreated animals, but that the technique of measuring striatal dopamine metabolism was too insensitive to demonstrate such an effect. Dopamine release was assessed directly, using brain microdialysis of striatal extracellular fluid. Tyrosine administration (50-200 mg/kg IP) did indeed cause a dose related increase in extracellular fluid dopamine levels with minor elevations in levels of DOPAC and HVA, its major metabolites, which were not dose-related. The rise in dopamine was short-lived, suggesting that receptor-mediated feedback mechanisms responded to the increased dopamine release by diminishing neuronal firing or sensitivity to tyrosine. These observations indicate that measurement of changes in striatal DOPAC and HVA, if negative, need not rule out increases in nigrostriatal dopamine release.

  20. Mutational analysis of Agrobacterium tumefaciens virD2: tyrosine 29 is essential for endonuclease activity.

    PubMed Central

    Vogel, A M; Das, A

    1992-01-01

    Agrobacterium tumefaciens VirD2 polypeptide, in the presence of VirD1, catalyzes a site- and strand-specific nicking reaction at the T-DNA border sequences. VirD2 is found tightly attached to the 5' end of the nicked DNA. The protein-DNA complex is presumably formed via a tyrosine residue of VirD2 (F. Durrenberger, A. Crameri, B. Hohn, and Z. Koukolikova-Nicola, Proc. Natl. Acad. Sci. USA 86:9154-9158, 1989). A mutational approach was used to study whether a tyrosine residue(s) of VirD2 is required for its activity. By site-specific mutagenesis, a tyrosine (Y) residue at position 29, 68, 99, 119, 121, 160, or 195 of the octopine Ti plasmid pTiA6 VirD2 was altered to phenylalanine (F). The Y-29-F or Y-121-F mutation completely abolished nicking activity of VirD2 in vivo in Escherichia coli. Two other substitutions, Y-68-F and Y-160-F, drastically reduced VirD2 activity. A substitution at position 99, 119, or 195 had no effect on VirD2 activity. Additional mutagenesis experiments showed that at position 29, no other amino acid could substitute for tyrosine without destroying VirD2 activity. At position 121, only a tryptophan (W) residue could be substituted. This, however, yielded a mutant protein with significantly reduced VirD2 activity. The nicked DNA from strains bearing a Y-68-F, Y-99-F, Y-119-F, Y-160-F, Y-195-F, or Y-121-W mutation in VirD2 was always found to contain a tightly linked protein. Images PMID:1309520

  1. Raman and surface-enhanced Raman spectroscopy of amino acids and nucleotide bases for target bacterial vibrational mode identification

    NASA Astrophysics Data System (ADS)

    Guicheteau, Jason; Argue, Leanne; Hyre, Aaron; Jacobson, Michele; Christesen, Steven D.

    2006-05-01

    Raman and surface-enhanced Raman spectroscopy (SERS) studies of bacteria have reported a wide range of vibrational mode assignments associated with biological material. We present Raman and SER spectra of the amino acids phenylalanine, tyrosine, tryptophan, glutamine, cysteine, alanine, proline, methionine, asparagine, threonine, valine, glycine, serine, leucine, isoleucine, aspartic acid and glutamic acid and the nucleic acid bases adenosine, guanosine, thymidine, and uridine to better characterize biological vibrational mode assignments for bacterial target identification. We also report spectra of the bacteria Bacillus globigii, Pantoea agglomerans, and Yersinia rhodei along with band assignments determined from the reference spectra obtained.

  2. Plasma Amino Acid Concentrations in 108 Children Receiving a Pediatric Amino Acid Formulation as Part of Parenteral Nutrition

    PubMed Central

    Shelton, Chasity M.; Clark, Amanda J.; Storm, Michael C.; Helms, Richard A.

    2010-01-01

    BACKGROUND Plasma amino acid (PAA) levels can be largely normalized during parenteral nutrition (PN) in infants and children using a pediatric-specific amino acid (AA) formulation. However, these previous results were based on individual clinical studies of small populations of neonates and infants. OBJECTIVE We have now examined AA levels in 108 children (0–7 years of age) receiving a pediatric-specific AA formulation in PN using a single analytical methodology. METHODS Infants and children were enrolled in specific protocols and parents/caregivers gave informed consent. Patients were stable and receiving age-appropriate intakes of AA and non-protein calories. Samples were obtained between 8 and10 am, processed immediately, deproteinized, and AA concentrations (μmol/L) were determined on a Beckman 6300 analyzer. Means and SD were calculated for sub-populations stratified by age: 0–1 month (48 patients, n=139), 1–6 months (36 patients, n=124), 7–12 months (11 patients, n=41), and 1–7 years (13 patients, n=51). Z scores were calculated for each amino acid [(observed mean - normal control mean)/normal control SD]. RESULTS When compared to the neonatal reference range, nonessential AA had Z scores that ranged from −1.84 (asparagine) to +1.48 (threonine). Only plasma free cystine, free tyrosine, and phenylalanine had Z scores outside the −2.0 to +2.0 range (95% confidence limits). Plasma free cystine values were low in all groups except neonates. Free tyrosine levels were low in all groups despite the presence of N-acetyl-L-tyrosine in the pediatric AA formulation. Phenylalanine levels were elevated only in neonates. When children 1 to 7 years old were compared with an age-matched reference range, plasma free cystine values were low (Z score −2.47), as were plasma glutamine values (−3.11), but elevations were found in the dicarboxylic amino acids aspartic acid (+2.5) and glutamic acid (+4.27). Regardless of reference range used for comparison, all

  3. Chlamydia pneumoniae encodes a functional aromatic amino acid hydroxylase.

    PubMed

    Abromaitis, Stephanie; Hefty, P Scott; Stephens, Richard S

    2009-03-01

    Chlamydia pneumoniae is a community-acquired respiratory pathogen that has been associated with the development of atherosclerosis. Analysis of the C. pneumoniae genome identified a gene (Cpn1046) homologous to eukaryotic aromatic amino acid hydroxylases (AroAA-Hs). AroAA-Hs hydroxylate phenylalanine, tyrosine, and tryptophan into tyrosine, dihydroxyphenylalanine, and 5-hydroxytryptophan, respectively. Sequence analysis of Cpn1046 demonstrated that residues essential for AroAA-H enzymatic function are conserved and that a subset of Chlamydia species contain an AroAA-H homolog. The chlamydial AroAA-Hs are transcriptionally linked to a putative bacterial membrane transport protein. We determined that recombinant Cpn1046 is able to hydroxylate phenylalanine, tyrosine, and tryptophan with roughly equivalent activity for all three substrates. Cpn1046 is expressed within 24 h of infection, allowing C. pneumoniae to hydroxylate host stores of aromatic amino acids during the period of logarithmic bacterial growth. From these results we can conclude that C. pneumoniae, as well as a subset of other Chlamydia species, encode an AroAA-H that is able to use all three aromatic amino acids as substrates. The maintenance of this gene within a number of Chlamydia suggests that the enzyme may have an important role in shaping the metabolism or overall pathogenesis of these bacteria. PMID:19141112

  4. Examination of tyrosine/adenine stacking interactions in protein complexes.

    PubMed

    Copeland, Kari L; Pellock, Samuel J; Cox, James R; Cafiero, Mauricio L; Tschumper, Gregory S

    2013-11-14

    The π-stacking interactions between tyrosine amino acid side chains and adenine-bearing ligands are examined. Crystalline protein structures from the protein data bank (PDB) exhibiting face-to-face tyrosine/adenine arrangements were used to construct 20 unique 4-methylphenol/N9-methyladenine (p-cresol/9MeA) model systems. Full geometry optimization of the 20 crystal structures with the M06-2X density functional theory method identified 11 unique low-energy conformations. CCSD(T) complete basis set (CBS) limit interaction energies were estimated for all of the structures to determine the magnitude of the interaction between the two ring systems. CCSD(T) computations with double-ζ basis sets (e.g., 6-31G*(0.25) and aug-cc-pVDZ) indicate that the MP2 method overbinds by as much as 3.07 kcal mol(-1) for the crystal structures and 3.90 kcal mol(-1) for the optimized structures. In the 20 crystal structures, the estimated CCSD(T) CBS limit interaction energy ranges from -4.00 to -6.83 kcal mol(-1), with an average interaction energy of -5.47 kcal mol(-1), values remarkably similar to the corresponding data for phenylalanine/adenine stacking interactions. Geometry optimization significantly increases the interaction energies of the p-cresol/9MeA model systems. The average estimated CCSD(T) CBS limit interaction energy of the 11 optimized structures is 3.23 kcal mol(-1) larger than that for the 20 crystal structures.

  5. Polyclonal antibody to soman-tyrosine

    PubMed Central

    Li, Bin; Duysen, Ellen G.; Froment, Marie-Thérèse; Masson, Patrick; Nachon, Florian; Jiang, Wei; Schopfer, Lawrence M.; Thiele, Geoffrey M.; Klassen, Lynell W.; Cashman, John; Williams, Gareth R.; Lockridge, Oksana

    2013-01-01

    Soman forms a stable, covalent bond with tyrosine 411 of human albumin, with tyrosines 257 and 593 in human transferrin, and with tyrosine in many other proteins. The pinacolyl group of soman is retained, suggesting that pinacolyl methylphosphonate bound to tyrosine could generate specific antibodies. Tyrosine in the pentapeptide RYGRK was covalently modified with soman simply by adding soman to the peptide. The phosphonylated-peptide was linked to keyhole limpet hemocyanin, and the conjugate was injected into rabbits. The polyclonal antiserum recognized soman-labeled human albumin, soman-mouse albumin, and soman human transferrin, but not non-phosphonylated control proteins. The soman-labeled tyrosines in these proteins are surrounded by different amino acid sequences, suggesting that the polyclonal recognizes soman-tyrosine independent of the amino acid sequence. Antiserum obtained after 4 antigen injections over a period of 18 weeks was tested in a competition ELISA where it had an IC50 of 10−11 M. The limit of detection on Western blots was 0.01 μg (15 picomoles) of soman-labeled albumin. In conclusion, a high-affinity, polyclonal antibody that specifically recognizes soman adducts on tyrosine in a variety of proteins has been produced. Such an antibody could be useful for identifying secondary targets of soman toxicity. PMID:23469927

  6. Aromatic amino acids in high selectivity bismuth(III) recognition.

    PubMed

    Ghatak, Sumanta Kumar; Dey, Debarati; Sen, Souvik; Sen, Kamalika

    2013-04-21

    The three aromatic amino acids, tyrosine, tryptophan and phenylalanine, play different physiological roles in life processes. Metal ions capable of binding these amino acids may aid in the reduction of effective concentration of these amino acids in any physiological system. Here we have studied the efficacy of some heavy metals for their complexation with these three amino acids. Bismuth has been found to bind selectively with these aromatic amino acids and this was confirmed using spectrofluorimetric, spectrophotometric and cyclic voltammetric studies. The series of heavy metals has been chosen because each of these metals remains associated with the others at very low concentration levels and Bi(III) is the least toxic amongst the other elements. So, selective recognition for Bi(III) would also mean no response for the other heavy elements if contaminants are present even at low concentration levels. The affinity towards these amino acids has been found to be in the order tryptophan < phenylalanine < tyrosine. The association constants of these amino acids have been calculated using Benesi-Hildebrand equations and the corresponding free energy change has also been calculated. The values of the association constants obtained from BH equations using absorbance values corroborate with the Stern-Volmer constants obtained from fluorimetric studies. The evidence for complexation is also supported by the results of cyclic voltammetry.

  7. Discovery and Evaluation of Novel Inhibitors of Mycobacterium Protein Tyrosine Phosphatase B from the 6-Hydroxy-Benzofuran-5-Carboxylic Acid Scaffold

    PubMed Central

    He, Yantao; Xu, Jie; Yu, Zhi-hong; Gunawan, Andrea M.; Wu, Li; Wang, Lina; Zhang, Zhong-Yin

    2013-01-01

    Mycobacterium tuberculosis (Mtb) protein tyrosine phosphatase B (mPTPB) is a virulence factor secreted by the pathogen and mediates mycobacterial survival in macrophages by targeting host cell immune responses. Consequently, mPTPB represents an exciting new target to combat TB infection. We describe a medicinal chemistry-oriented approach that transforms a benzofuran salicylic acid scaffold into a highly potent (IC50 = 38 nM) and selective mPTPB inhibitor (>50 fold against a large panel of PTPs). Importantly, the inhibitor is capable of reversing the altered host immune responses induced by the bacterial phosphatase and restoring the macrophage’s full capacity to secrete IL-6 and undergo apoptosis in response to IFN-γ stimulation, validating the concept that chemical inhibition of mPTPB may be therapeutically useful for novel TB treatment. The study further demonstrates that bicyclic salicylic acid pharmacophores can be used to deliver PTP inhibitors with high potency, selectivity, and cellular efficacy. PMID:23305444

  8. VUV state-selected photoionization of thermally-desorbed biomolecules by coupling an aerosol source to an imaging photoelectron/photoion coincidence spectrometer: case of the amino acids tryptophan and phenylalanine.

    PubMed

    Gaie-Levrel, François; Garcia, Gustavo A; Schwell, Martin; Nahon, Laurent

    2011-04-21

    Gas phase studies of biological molecules provide structural and dynamical information on isolated systems. The lack of inter- or intra-molecular interactions facilitates the interpretation of the experimental results through theoretical calculations, and constitutes an informative complement to the condensed phase. However advances in the field are partially hindered by the difficulty of vaporising these systems, most of which are thermally unstable. In this work we present a newly developed aerosol mass thermodesorption setup, which has been coupled to a Velocity Map Imaging (VMI) analyzer operated in coincidence with a Wiley-McLaren Time of Flight spectrometer, using synchrotron radiation as a single photon ionization source. Although it has been previously demonstrated that thermolabile molecules such as amino acids can be produced intact by the aerosol vaporisation technique, we show how its non-trivial coupling to a VMI analyzer plus the use of electron/ion coincidences greatly improves the concept in terms of the amount of spectroscopic and dynamic information that can be extracted. In this manner, we report on the valence shell ionization of two amino acids, tryptophan and phenylalanine, for which threshold photoelectron spectra have been recorded within the first 3 eV above the first ionization energy using synchrotron radiation emitted from the DESIRS beamline located at SOLEIL in France. Their adiabatic ionization energies (IEs) have been measured at 7.40 ± 0.05 and 8.65 ± 0.02 eV, respectively, and their spectra analyzed using existing theoretical data from the literature. The IE values agree well with previously published ones, but are given here with a considerably reduced uncertainty by up to a factor of 5. The photostability of both amino acids is also described in detail, through the measurement of the state-selected fragmentation pathways via the use of threshold electron/ion coincidences (TPEPICO), with appearance energies for the different

  9. The role of L-type amino acid transporter 1 in human tumors

    PubMed Central

    Zhao, Yu; Wang, Lin; Pan, Jihong

    2015-01-01

    Summary L-type amino acid transporter 1 (LAT1) is an L-type amino acid transporter and transports large neutral amino acids such as leucine, isoleucine, valine, phenylalanine, tyrosine, tryptophan, methionine, and histidine. LAT1 was found to be highly expressed especially in human cancer tissues, and up-regulated LAT1 can lead to dysfunction in human tumor cells. These findings suggest that LAT1 plays an important role in human tumors. This review provides an overview of the current understanding of LAT1 expression and its clinical significance and function in tumors. PMID:26668776

  10. Phosphorylation of caveolin-1 on tyrosine-14 induced by ROS enhances palmitate-induced death of beta-pancreatic cells.

    PubMed

    Wehinger, Sergio; Ortiz, Rina; Díaz, María Inés; Aguirre, Adam; Valenzuela, Manuel; Llanos, Paola; Mc Master, Christopher; Leyton, Lisette; Quest, Andrew F G

    2015-05-01

    A considerable body of evidence exists implicating high levels of free saturated fatty acids in beta pancreatic cell death, although the molecular mechanisms and the signaling pathways involved have not been clearly defined. The membrane protein caveolin-1 has long been implicated in cell death, either by sensitizing to or directly inducing apoptosis and it is normally expressed in beta cells. Here, we tested whether the presence of caveolin-1 modulates free fatty acid-induced beta cell death by reexpressing this protein in MIN6 murine beta cells lacking caveolin-1. Incubation of MIN6 with palmitate, but not oleate, induced apoptotic cell death that was enhanced by the presence of caveolin-1. Moreover, palmitate induced de novo ceramide synthesis, loss of mitochondrial transmembrane potential and reactive oxygen species (ROS) formation in MIN6 cells. ROS generation promoted caveolin-1 phosphorylation on tyrosine-14 that was abrogated by the anti-oxidant N-acetylcysteine or the incubation with the Src-family kinase inhibitor, PP2 (4-amino-5-(4-chlorophenyl)-7(dimethylethyl)pyrazolo[3,4-d]pyrimidine). The expression of a non-phosphorylatable caveolin-1 tyrosine-14 to phenylalanine mutant failed to enhance palmitate-induced apoptosis while for MIN6 cells expressing the phospho-mimetic tyrosine-14 to glutamic acid mutant caveolin-1 palmitate sensitivity was comparable to that observed for MIN6 cells expressing wild type caveolin-1. Thus, caveolin-1 expression promotes palmitate-induced ROS-dependent apoptosis in MIN6 cells in a manner requiring Src family kinase mediated tyrosine-14 phosphorylation. PMID:25572853

  11. Highest Plasma Phenylalanine Levels in (Very) Premature Infants on Intravenous Feeding; A Need for Concern

    PubMed Central

    Cortés-Castell, Ernesto; Sánchez-González, Pablo; Palazón-Bru, Antonio; Bosch-Giménez, Vicente; Manero-Soler, Herminia; Juste-Ruiz, Mercedes; Rizo-Baeza, María Mercedes; Gil-Guillén, Vicente Francisco

    2015-01-01

    Objective To analyse the association in newborns between blood levels of phenylalanine and feeding method and gestational age. Study Design This observational, cross-sectional study included a sample of 11,829 infants between 2008 and 2013 in a Spanish region. Data were recorded on phenylalanine values, feeding method [breast, formula, mixed (breast plus formula), or partial or fully intravenous feeding], gestational age in weeks (<32, 32–37, ≥37), gender and days since birth at the moment of blood collection. Outcomes were [phenylalanine] and [phenylalanine] ≥95th percentile. Associations were analysed using multivariate models [linear (means difference) and logistic regression (adjusted odds ratios)]. Results Higher phenylalanine values were associated with lower gestational age (p<0.001) and with intravenous feeding (p<0.001). Conclusion The degree of prematurity and intravenous feeding influenced the plasma concentration of phenylalanine in the newborn. Caution should be taken in [phenylalanine] for newborns with intravenous feeding, monitoring them carefully. Very preterm infants given the recommended amount of amino acids should also be strictly monitored. These findings should be taken into consideration and call for adapting the amounts to the needs of the infant. PMID:26389596

  12. Role of the phenylalanine-hydroxylating system in aromatic substance degradation and lipid metabolism in the oleaginous fungus Mortierella alpina.

    PubMed

    Wang, Hongchao; Chen, Haiqin; Hao, Guangfei; Yang, Bo; Feng, Yun; Wang, Yu; Feng, Lu; Zhao, Jianxin; Song, Yuanda; Zhang, Hao; Chen, Yong Q; Wang, Lei; Chen, Wei

    2013-05-01

    Mortierella alpina is a filamentous fungus commonly found in soil that is able to produce lipids in the form of triacylglycerols that account for up to 50% of its dry weight. Analysis of the M. alpina genome suggests that there is a phenylalanine-hydroxylating system for the catabolism of phenylalanine, which has never been found in fungi before. We characterized the phenylalanine-hydroxylating system in M. alpina to explore its role in phenylalanine metabolism and its relationship to lipid biosynthesis. Significant changes were found in the profile of fatty acids in M. alpina grown on medium containing an inhibitor of the phenylalanine-hydroxylating system compared to M. alpina grown on medium without inhibitor. Genes encoding enzymes involved in the phenylalanine-hydroxylating system (phenylalanine hydroxylase [PAH], pterin-4α-carbinolamine dehydratase, and dihydropteridine reductase) were expressed heterologously in Escherichia coli, and the resulting proteins were purified to homogeneity. Their enzymatic activity was investigated by high-performance liquid chromatography (HPLC) or visible (Vis)-UV spectroscopy. Two functional PAH enzymes were observed, encoded by distinct gene copies. A novel role for tetrahydrobiopterin in fungi as a cofactor for PAH, which is similar to its function in higher life forms, is suggested. This study establishes a novel scheme for the fungal degradation of an aromatic substance (phenylalanine) and suggests that the phenylalanine-hydroxylating system is functionally significant in lipid metabolism.

  13. Photolysis of phenylalanine in the presence of oxidized carbon nanotubes.

    PubMed

    Humeres, Eduardo; de Souza, Eduardo Pinheiro; Debacher, Nito Angelo; Moreira, Regina de F P M; Lopes, Cristiane Nunes; Fernández, Ma Isabel; Santaballa, J Arturo; Canle, Moisés L; Schreiner, Wido H; Aliev, Abil E

    2015-01-01

    Photolyses at 254 nm of phenylalanine (Phe) in aqueous solutions, were carried out in the presence of oxidized carbon nanotubes modified by the reaction with SO2 (mNTO). Kinetics of the photolyses were followed by UV spectrophotometry at 220 nm, and the products were characterized by HPLC, XPS, and (13)C-SSNMR. The ratio of the initial rates of photolysis in the presence and absence of mNTO, k*/ko*, showed a systematic decrease. The photolytic decay of Phe occurs with minor formation of tyrosine. The mass of nanotubes produced an exponential attenuation of the photolytic decomposition of Phe. Total carbon analyses (TCA) showed no inorganic carbon formation after the photolyses. The first-order rate constant of photofunctionalization of mNTO by the insertion of phenylalanine onto the nanotube matrix was calculated from TCA to be kin = 30.1 min(-1). Comparison of the XPS spectra of the mNTO before and after the photolysis, using the atom inventory technique, suggests the insertion of Phe along with the extrusion of a sulfide radical anion ((•)S(-)) which undergo subsequent oxidation to SO4(2-). The obtained results show the effects of mNTO on the photolysis of Phe and provide a new method of photofunctionalization of carbon materials, modified by the intermediates of the reduction of SO2, with an organic moiety.

  14. A Tyrosine Aminomutase from Rice (Oryza sativa) Isomerizes (S)-α- to (R)-β-Tyrosine with Unique High Enantioselectivity and Retention of Configuration.

    PubMed

    Walter, Tyler; King, Zayna; Walker, Kevin D

    2016-01-12

    A recently discovered 3,5-dihydro-5-methylidene-4H-imidazol-4-one (MIO)-dependent tyrosine aminomutase (OsTAM) from rice [Yan, J., et al. (2015) Plant Cell 27, 1265] converts (S)-α-tyrosine to a mixture of (R)- and (S)-β-tyrosines, with high (94%) enantiomeric excess, which does not change with pH, like it does for two bacterial TAMs. The K(M) of 490 μM and the k(cat) of 0.005 s(-1) are similar for other TAM enzymes. OsTAM is unique and also catalyzes (R)-β- from (S)-α-phenylalanine. OsTAM principally retains the configuration at the reactive C(α) and C(β) centers during catalysis much like the phenylalanine aminomutase on the Taxol biosynthetic pathway in Taxus plants.

  15. Laser Desorption Supersonic Jet Spectroscopy of Hydrated Tyrosine

    NASA Astrophysics Data System (ADS)

    Oba, Hikari; Shimozono, Yoko; Ishiuchi, Shun-Ichi; Fujii, Masaaki; Carcabal, Pierre

    2013-06-01

    The structure of tyrosine (tyr) consists of amino-acid chain and phenol, and it has roughly two possible binding sites for water, amino-acid site and phenolic OH site. Investigating how water molecule binds to tyr will give fundamental information for hydrations of peptide and protein. Resonance enhanced multi photon ionization (REMPI) spectrum of tyr-water 1:1 cluster has already been reported by de Vries and co-workers, however, no analysis on the hydrated structures has been reported. In the REMPI spectrum, two clusters of bands are observed; one appears at ˜35600 cm^{-1} energy region which is the almost same with 0-0 transitions of tyr monomer, and another is observed at ˜300 cm^{-1} lower than the former. Based on the electronic transition energy of phenylalanine and the hydrated clusters, the former is expected to be derived from a structure that water binds to amino acid site. On the other hand, it is plausibly predicted that the latter originates from a structure that water binds to phenolic OH group, because the electronic transition of mono hydrated phenol is ˜300 cm^{-1} red-shifted from the monomer. We applied IR dip spectroscopy which can measure conformer selective IR spectra to the tyr-(H_{2}O)_{1} clusters by using laser desorption supersonic jet technique to confirm the assignments. Especially in the phenolic OH bound isomer, it was found that the intra molecular hydrogen bond within amino-acid chain, which is far from the water molecule and cannot interact directly with each other, is strengthened by the hydration. A. Abio-Riziq et al., J. Phys. Chem. A, 115, 6077 (2011). Y. Shimozono, et al., Phys. Chem. Chem. Phys., (2013) DOI: 10.1039/c3cp43573c. T. Ebata et al., Phys. Chem. Chem. Phys., 8, 4783 (2006). T. Watanabe et al., J. Chem. Phys., 105, 408 (1996).

  16. Phenylalanine hydroxylase deficiency: diagnosis and management guideline.

    PubMed

    Vockley, Jerry; Andersson, Hans C; Antshel, Kevin M; Braverman, Nancy E; Burton, Barbara K; Frazier, Dianne M; Mitchell, John; Smith, Wendy E; Thompson, Barry H; Berry, Susan A

    2014-02-01

    Phenylalanine hydroxylase deficiency, traditionally known as phenylketonuria, results in the accumulation of phenylalanine in the blood of affected individuals and was the first inborn error of metabolism to be identified through population screening. Early identification and treatment prevent the most dramatic clinical sequelae of the disorder, but new neurodevelopmental and psychological problems have emerged in individuals treated from birth. The additional unanticipated recognition of a toxic effect of elevated maternal phenylalanine on fetal development has added to a general call in the field for treatment for life. Two major conferences sponsored by the National Institutes of Health held >10 years apart reviewed the state of knowledge in the field of phenylalanine hydroxylase deficiency, but there are no generally accepted recommendations for therapy. The purpose of this guideline is to review the strength of the medical literature relative to the treatment of phenylalanine hydroxylase deficiency and to develop recommendations for diagnosis and therapy of this disorder. Evidence review from the original National Institutes of Health consensus conference and a recent update by the Agency for Healthcare Research and Quality was used to address key questions in the diagnosis and treatment of phenylalanine hydroxylase deficiency by a working group established by the American College of Medical Genetics and Genomics. The group met by phone and in person over the course of a year to review these reports, develop recommendations, and identify key gaps in our knowledge of this disorder. Above all, treatment of phenylalanine hydroxylase deficiency must be life long, with a goal of maintaining blood phenylalanine in the range of 120-360 µmol/l. Treatment has predominantly been dietary manipulation, and use of low protein and phenylalanine medical foods is likely to remain a major component of therapy for the immediate future. Pharmacotherapy for phenylalanine

  17. Effect of aspartame on plasma amino acid profiles of diabetic patients with chronic renal failure.

    PubMed

    Gupta, V; Cochran, C; Parker, T F; Long, D L; Ashby, J; Gorman, M A; Liepa, G U

    1989-06-01

    A randomized, double-blind study was conducted to determine the possible effects of aspartame on the plasma amino acid profiles of 23 diabetic patients with renal failure who were undergoing maintenance hemodialysis. Subjects were given a single dose of 10 mg aspartame/kg (approximately equivalent to 25 packets of Equal [NutraSweet Consumer Products, Inc, Chicago, IL] or the amount of phenylalanine in a 300-mL glass of milk) or a placebo in a crossover study design. Three postdialysis blood samples were drawn just before and 1 and 2 h after aspartame or placebo consumption. After aspartame consumption statistically significant increases in only two amino acids, phenylalanine and tyrosine, were noted at 1 and 2 h when compared with placebo values. The increases in phenylalanine were within the normal postprandial range for healthy subjects; no other increases in essential or nonessential amino acids, except for tyrosine, were detected. This study supports the view that aspartame is safe for diabetic subjects with chronic renal failure.

  18. New platform for controlled and sustained delivery of the EGF receptor tyrosine kinase inhibitor AG1478 using poly(lactic-co-glycolic acid) microspheres

    PubMed Central

    Robinson, Rebecca; Bertram, James P.; Reiter, Jill L.; Lavik, Erin B.

    2015-01-01

    Inhibition of the epidermal growth factor receptor (EGFR) has been shown to reduce tumor growth and metastases and promote axon regeneration in the central nervous system. Current strategies for inhibiting EGFR include the administration of reversible or irreversible small-molecule tyrosine kinase inhibitors (TKIs). However, to be effective in vivo constant and sustained delivery is required. This study explored the feasibility of encapsulating the tyrphostin 4-(3-chloroanilino)-6,7-dimethoxyquinazoline (AG1478) in poly(lactic-co-glycolic acid) (PLGA) microspheres to achieve sustained delivery of the TKI. We characterized microspheres prepared using three different emulsion methods: solid-in-oil-in-water, oil-in-water, and oil-in-water with co-solvent. Addition of a co-solvent increased the loading and release of AG1478, and significantly (P<0.001) decreased the size of the microspheres which facilitates administration of the spheres. On average, sustained delivery of AG1478 from microspheres was achieved for six months. However, the addition of a co-solvent prolonged release for over nine months (266 days). In addition, AG1478 retained its bioactivity upon delivery, and inhibited EGFR in both immortalized rat fibroblasts and in EGFR-amplified human carcinoma cells. These results demonstrate that AG1478 can be encapsulated in PLGA and retain bioactivity; thereby providing a new platform for controlled administration of EGFR TKIs. PMID:20055747

  19. The influence of the cell free solution of lactic acid bacteria on tyramine production by food borne-pathogens in tyrosine decarboxylase broth.

    PubMed

    Toy, Nurten; Özogul, Fatih; Özogul, Yesim

    2015-04-15

    The function of cell-free solutions (CFSs) of lactic acid bacteria (LAB) on tyramine and other biogenic amine production by different food borne-pathogens (FBPs) was investigated in tyrosine decarboxylase broth (TDB) using HPLC. Cell free solutions were prepared from four LAB strains. Two different concentrations which were 50% (5 ml CFS+5 ml medium/1:1) and 25% (2.5 ml CFS+7.5 ml medium/1:3) CFS and the control without CFS were prepared. Both concentration of CFS of Streptococcus thermophilus and 50% CFS of Pediococcus acidophilus inhibited tyramine production up to 98% by Salmonella paratyphi A. Tyramine production by Escherichia coli was also inhibited by 50% CFS of Lactococcus lactis subsp. lactis and 25% CFS of Leuconostoc lactis. subsp. cremoris. The inhibitor effect of 50% CFS of P. acidophilus was the highest on tyramine production (55%) by Listeria monocytogenes, following Lc. lactis subsp. lactis and Leuconostoc mesenteroides subsp. cremoris (20%) whilst 25% CFS of Leu. mes. subsp. cremoris and Lc. lactis subsp. lactis showed stimulator effects (160%). The stimulation effects of 50% CFS of S. thermophilus and Lc. lactis subsp. lactis were more than 70% by Staphylococcus aureus comparing to the control. CFS of LAB strains showed statistically inhibitor effect since lactic acid inhibited microbial growth, decreased pH quickly and reduced the formation of AMN and BAs. Consequently, in order to avoid the formation of high concentrations of biogenic amines in fermented food by bacteria, it is advisable to use CFS for food and food products.

  20. Influence of Phenylalanine on Carotenoid Aggregation

    NASA Astrophysics Data System (ADS)

    Lu, L.; Ni, X.; Luo, X.

    2015-01-01

    The carotenoids lutein and β-carotene form, in 1:1 ethanol-water mixtures H-aggregates, of different strengths. The effects of phenylalanine on these aggregates were recorded by UV-Vis absorption, steady-state fluorescence, and Raman spectra. The H-aggregate of lutein was characterized by a large 78 nm blue shift in the absorption spectra, confirming the strong coupling between hydroxyl groups of adjacent molecules. The 15 nm blue shift in the β-carotene mixture also indicates that it was assembled by weak coupling between polyenes. After adding phenylalanine, the reducing absorption strength of the aggregates of lutein and reappearance of vibrational substructure indicate that the hydroxyl and amino groups of phenylalanine may coordinate to lutein and disaggregate the H-aggregates. However, phenylalanine had no effect on aggregates of β-carotene. The Raman spectra show three bands of carotenoids whose intensities decreased with increasing phenylalanine concentration. The frequency of ν1 corresponding to the length of the conjugated region was more sensitive to the solution of lutein. This coordination of phenylalanine to lutein could increase the length of the conjugated region. In addition, phenylalanine significantly affected the excited electronic states of carotenoids, which were crucial in the energy transfer from carotenoids to chlorophyll a in vivo.

  1. Distribution and enantiomeric composition of amino acids in the Murchison meteorite

    NASA Technical Reports Server (NTRS)

    Engel, M. H.; Nagy, B.

    1982-01-01

    Studies of the amino acid contents and enantiomeric compositions of a single stone from the Murchison meteorite are reported. Water-extracted and 6M HCl-extracted samples from the meteorite interior of meteorite fragments were analyzed by gas chromatography and combined gas chromatography-chemical ionization mass spectrometry. Examination of the D/L ratios of glutamic acid, aspartic acid, proline, leucine and alanine reveals those amino acids extractable by water to be partially racemized, whereas the acid-extracted amino acids were less racemized. The amino acid composition of the stone is similar to those previously reported, including the absence of serine, threonine, tyrosine phenylalanine and methionine and the presence of unusual amino acids including such as isovaline, alpha-aminoisobutyric acid and pseudoleucine. It is concluded that the most likely mechanism accounting for the occurrence of nonracemic amino acid mixtures in the Murchison meteorite is by extraterrestrial stereoselective synthesis or decomposition reactions.

  2. Amino acids inhibit kynurenic acid formation via suppression of kynurenine uptake or kynurenic acid synthesis in rat brain in vitro.

    PubMed

    Sekine, Airi; Okamoto, Misaki; Kanatani, Yuka; Sano, Mitsue; Shibata, Katsumi; Fukuwatari, Tsutomu

    2015-01-01

    The tryptophan metabolite, kynurenic acid (KYNA), is a preferential antagonist of the α7 nicotinic acetylcholine receptor at endogenous brain concentrations. Recent studies have suggested that increase of brain KYNA levels is involved in psychiatric disorders such as schizophrenia and depression. KYNA-producing enzymes have broad substrate specificity for amino acids, and brain uptake of kynurenine (KYN), the immediate precursor of KYNA, is via large neutral amino acid transporters (LAT). In the present study, to find out amino acids with the potential to suppress KYNA production, we comprehensively investigated the effects of proteinogenic amino acids on KYNA formation and KYN uptake in rat brain in vitro. Cortical slices of rat brain were incubated for 2 h in Krebs-Ringer buffer containing a physiological concentration of KYN with individual amino acids. Ten out of 19 amino acids (specifically, leucine, isoleucine, phenylalanine, methionine, tyrosine, alanine, cysteine, glutamine, glutamate, and aspartate) significantly reduced KYNA formation at 1 mmol/L. These amino acids showed inhibitory effects in a dose-dependent manner, and partially inhibited KYNA production at physiological concentrations. Leucine, isoleucine, methionine, phenylalanine, and tyrosine, all LAT substrates, also reduced tissue KYN concentrations in a dose-dependent manner, with their inhibitory rates for KYN uptake significantly correlated with KYNA formation. These results suggest that five LAT substrates inhibit KYNA formation via blockade of KYN transport, while the other amino acids act via blockade of the KYNA synthesis reaction in brain. Amino acids can be a good tool to modulate brain function by manipulation of KYNA formation in the brain. This approach may be useful in the treatment and prevention of neurological and psychiatric diseases associated with increased KYNA levels.

  3. Pitfalls in phenylalanine loading test in the diagnosis of dopa-responsive dystonia.

    PubMed

    Opladen, Thomas; Hoffmann, Georg F; Kühn, Andrea A; Blau, Nenad

    2013-03-01

    Phenylalanine (Phe) loading test is a useful tool in the differential diagnosis of dopa-responsive dystonia due to autosomal dominant or recessive GTP cyclohydrolase I (GTPCH) deficiency or autosomal recessive sepiapterin reductase (SR) deficiency. In these patients hepatic phenylalanine hydroxylase system is compromised due to subnormal tetrahydrobiopterin (BH(4)) levels and hydroxylation of phenylalanine (Phe) to tyrosine (Tyr) is reduced with elevated Phe/Tyr ratio 1-2 h after oral Phe administration (100 mg/kg bw) administration. In healthy persons there is only a modest increase in Tyr production and blood Phe normalizes after 4 h. We report on a challenge with Phe (100 mg/kg bw) in a patient with dopa-responsive dystonia while on therapy with BH(4) and l-dopa. During Phe challenge Phe concentration remained below the normal range while a transient mild hypertyrosinemia was observed, leading to an extremely low Phe/Tyr ratio. A repeated test, after BH(4) withdrawal, reversed the findings and resulted normal. These data suggest activation of hepatic phenylalanine hydroxylase by BH(4). Thus, the Phe loading test should not be performed during substitution with BH(4).

  4. Endo[metallo] SWNT-amino acid interactions: A theoretical study

    NASA Astrophysics Data System (ADS)

    Jalbout, Abraham F.

    We propose that an atom of calcium (Ca) which is an alkaline earth metal on encapsulation inside of a metallic armchair (5,5) (SWNT) species can have stronger amino acid interactions. From our calculations of various physical parameters we depict several configurations in which such an endo[metallo] SWNT can be modified by an internally placed Ca atom. Density functional theory (DFT) calculations reveal that the most favorable interactions of the SWNT system is with tryptophan, tyrosine, and phenylalanine that can be directly correlated to the backbone geometry of the amino acid species.

  5. Arginine-responsive terbium luminescent hybrid sensors triggered by two crown ether carboxylic acids.

    PubMed

    Jiang, Lasheng; Tang, Ke; Ding, Xiaoping; Wang, Qianming; Zhou, Zhan; Xiao, Rui

    2013-12-01

    Crown ether carboxylic acids constitute main building blocks for the synthesis of terbium containing covalent cross-linked luminescent materials. Both the complexes and the hybrid nanomaterials could exhibit remarkable green emissions in pure water. More importantly, they were found to have a profound effect on the luminescence responses to arginine compared with glutamic acid, histidine, tryptophan, threonine, tyrosine and phenylalanine in aqueous environment. The present study provided the possibility of using a host-guest mechanism as a way of signal transduction based on lanthanide supramolecular hybrid materials.

  6. Plasmon resonance enhancement of nonlinear properties of amino acids

    NASA Astrophysics Data System (ADS)

    de Araujo, Renato E.; Rativa, Diego; Gomes, Anderson S. L.

    2007-02-01

    Here we analyze the influence of 9 nm (mean diameter) silver particles on the nonlinear properties of intrinsic cell molecules. A novel high sensitivity thermal managed eclipse Z-scan technique with a femtosecond laser system was used to analyze the nonlinear susceptibility of water solution of fluorescent and non-fluorescent amino acids (Tryptophan, Tyrosine, Phenylalanine, Proline and Histidine) with different concentration of silver nanoparticles. The generalized Maxwell Garnett model is used to explain the behavior of the measured nonlinear refractive index with the change of the nanoparticles concentration in the sample.

  7. Introduction of two mutations into AroG increases phenylalanine production in Escherichia coli.

    PubMed

    Ding, Rui; Liu, Lifei; Chen, Xuhui; Cui, Zhenhai; Zhang, Ao; Ren, Daming; Zhang, Lijun

    2014-10-01

    L-Phenylalanine is an important amino acid commercially, and therefore optimization of its manufacture is of interest. We constructed a range of mutant alleles of AroG, the enzyme involved in the first step of phenylalanine biosynthesis. Three single-site mutant alleles were constructed (aroG8, aroG15, and aroG29), which were then combined to generate three double-site aroG (fbr) mutant alleles (aroG8/15, aroG8/29, and aroG15/29). Enzymatic activity, feedback inhibition, and fermentation were analyzed in all of the mutants. All double-site mutants, except AroG15/29, showed higher enzymatic activity and greater resistance to feedback inhibition than their respective single-site mutants. The E. coli strain carrying the aroG8/15 allele produced a phenylalanine titer of 26.78 g/l, a 116 % improvement over the control phenylalanine overproducing strain (12.41 g/l). Our findings provide an effective method for modifying phenylalanine biosynthetic genes, which may be applied to optimize the commercial manufacture of phenylalanine.

  8. Effect of dietary phenylalanine restriction on visual attention span in mentally retarded subjects with phenylketonuria.

    PubMed

    Giffin, F D; Clarke, J T; d'Entremont, D M

    1980-05-01

    An ABA within-subject study design was used to assess the effect of dietary phenylalanine restriction on the visual attention span of three mentally retarded males (9, 15, and 21 years old) with phenylketonuria. Visual attention span was measured by recording the amount of time the subjects visually fixated pictures projected on a screen according to a standardized test protocol. After 4-6 weeks of baseline testing (A-phase), each subject was placed on a phenylalanine-restricted diet, designed to maintain plasma phenylalanine levels at 0.3-0.9 mM, for 8-14 weeks (B-phase). A return to baseline phenylalanine intake (A-phase) was achieved by surreptitiously adding sufficient L-phenylalanine to the therapeutic diets to increase plasma concentrations of the amino acid to pretreatment levels. Diet treatment (B-phase) was associated with highly significant improvements in visual attention span in two subjects (multivariate analysis of variance, P less than 0.001); the third subject, the most retarded, showed no effect. No objectively demonstrable change in behavior was documented in any subject. However, the results support the view that phenylalanine toxicity extends beyond early childhood and that some component of the toxicity is reversible, even in severely retarded patients with phenylketonuria.

  9. In vivo tyrosine hydroxylation in rat retina: effect of aspartame ingestion in rats pretreated with p-chlorophenylalanine.

    PubMed

    Fernstrom, J D; Fernstrom, M H; Massoudi, M S

    1991-04-01

    Rats were pretreated with p-chlorophenylalanine (PCPA) to inhibit hepatic phenylalanine hydroxylase. Two days later, oral aspartame (APM; aspartylphenylalanine methylester) administration substantially increased serum phenylalanine (Phe) concentrations and the ratio, in serum, of Phe to the sum of its competitors for transport into brain and retina (the other large neutral amino acids). Smaller changes occurred in serum tyrosine (Tyr) concentrations and in the ratio, in serum, of Tyr to the sum of its competitors. P-chlorophenylalanine-pretreated rats showed normal increases in retinal Tyr hydroxylation rate after Tyr injection, indicating that the enzyme was functionally normal. APM (0, 500, 1000, 1500 mg/kg body wt) intubation of PCPA-pretreated rats produced a dose-related increment in retinal Phe concentrations (up to six times normal values), no changes in retinal Tyr concentration, and no changes in retinal Tyr hydroxylation rate. The results thus indicate that very large increments in retinal Phe concentrations produced by enormous doses of APM do not modify Tyr hydroxylation in vivo.

  10. Tyrosine Phosphorylation of A17 during Vaccinia Virus Infection: Involvement of the H1 Phosphatase and the F10 Kinase

    PubMed Central

    Derrien, M.; Punjabi, A.; Khanna, M.; Grubisha, O.; Traktman, P.

    1999-01-01

    Vaccinia virus encodes two protein kinases (B1 and F10) and a dual-specificity phosphatase (VH1), suggesting that phosphorylation and dephosphorylation of substrates on serine/threonine and tyrosine residues are important in regulating diverse aspects of the viral life cycle. Using a recombinant in which expression of the H1 phosphatase can be regulated experimentally (vindH1), we have previously demonstrated that repression of H1 leads to the maturation of noninfectious virions that contain several hyperphosphorylated substrates (K. Liu et al., J. Virol. 69:7823–7834). In this report, we demonstrate that among these is a 25-kDa protein that is phosphorylated on tyrosine residues in H1-deficient virions and can be dephosphorylated by recombinant H1. We demonstrate that the 25-kDa phosphoprotein represents the product of the A17 gene and that A17 is phosphorylated on serine, threonine, and tyrosine residues during infection. Detection of phosphotyrosine within A17 is abrogated when Tyr203 (but not Tyr3, Tyr6, or Tyr7) is mutated to phenylalanine, suggesting strongly that this amino acid is the site of tyrosine phosphorylation. Phosphorylation of A17 fails to occur during nonpermissive infections performed with temperature-sensitive mutants defective in the F10 kinase. Our data suggest that this enzyme, which was initially characterized as a serine/threonine kinase, might in fact have dual specificity. This hypothesis is strengthened by the observation that Escherichia coli induced to express F10 contain multiple proteins which are recognized by antiphosphotyrosine antiserum. This study presents the first evidence for phosphotyrosine signaling during vaccinia virus infection and implicates the F10 kinase and the H1 phosphatase as the dual-specificity enzymes that direct this cycle of reversible phosphorylation. PMID:10438817

  11. Accurate determination of the amino acid content of selected feedstuffs.

    PubMed

    Rutherfurd, Shane M

    2009-01-01

    The accurate determination of the amino acid content is important. In the present study, a least-squares non-linear regression model of the amino acid content determined over multiple hydrolysis times was used to accurately determine the content of amino acids in five different feedstuffs. These values were compared with 24-h hydrolysis values determined for the same feedstuffs. Overall, approximately two-thirds of the amino acids determined in this study (aspartic acid, threonine, glutamic acid, proline, glycine, alanine, leucine, tyrosine, phenylalanine and arginine) using 24-h hydrolysis were in good agreement (<3% difference). When examined across feedstuffs, the concentration of serine was underestimated by the 24-h hydrolysis method by 4.8%, while the concentrations of histidine and lysine were overestimated by 3.9% and 3.1%, respectively.

  12. The transmembrane tyrosines Y56, Y91 and Y167 play important roles in determining the affinity and transport rate of the rabbit proton-coupled peptide transporter PepT1.

    PubMed

    Pieri, Myrtani; Gan, Christine; Bailey, Patrick; Meredith, David

    2009-11-01

    The mammalian proton-coupled peptide transporter PepT1 is widely accepted as the major route of uptake for dietary nitrogen, as well as being responsible for the oral absorption of a number of classes of drugs, including beta-lactam antibiotics and angiotensin-converting enzyme (ACE) inhibitors. Using site-directed mutagenesis and zero-trans transport assays, we investigated the role of conserved tyrosines in the transmembrane domains (TMDs) of rabbit PepT1 as predicted by hydropathy plots. All the individual TMD tyrosines were substituted with phenylalanine and shown to retain the ability to traffic to the plasma membrane of Xenopus laevis oocytes. These single substitutions of TMD tyrosines by phenylalanine residues did not affect the proton dependence of peptide uptake, with all retaining wild-type PepT1-like pH dependence. Individual mutations of four of the nine TMD residue tyrosines (Y64, Y287, Y345 and Y587) were without measurable effect on PepT1 function, whereas the other five (Y12, Y56, Y91, Y167 and Y345) were shown to result in altered transport function compared to the wild-type PepT1. Intriguingly, the affinity of Y56F-PepT1 was found to be dramatically increased (approximately 100-fold) in comparison to that of the wild-type rabbit PepT1. Y91 mutations also affected the substrate affinity of the transporter, which increased in line with the hydrophilicity of the substituted amino acid (F>Y>Q>R). Y167 was demonstrated to play a pivotal role in rabbit PepT1 function since Y167F, Y167R and Y167Q demonstrated very little transport function. These results are discussed with regard to a proposed mechanism for PepT1 substrate binding.

  13. Tyrosine monitoring in children with early and continuously treated phenylketonuria: results of an international practice survey.

    PubMed

    Sharman, Rachael; Sullivan, Karen A; Young, Ross McD; McGill, James J

    2010-12-01

    Investigations into the biochemical markers associated with executive function (EF) impairment in children with early and continuously treated phenylketonuria (ECT-PKU) remain largely phenylalanine-only focused, despite experimental data showing that a high phenylalanine:tyrosine (phe:tyr) ratio is more strongly associated with EF deficit than phe alone. A high phe:tyr ratio is hypothesized to lead to a reduction in dopamine synthesis within the brain, which in turn results in the development of EF impairment. This paper provides a snapshot of current practice in the monitoring and/or treatment of tyrosine levels in children with PKU, across 12 countries from Australasia, North America and Europe. Tyrosine monitoring in this population has increased over the last 5 years, with over 80% of clinics surveyed reporting routine monitoring of tyrosine levels in infancy alongside phe levels. Twenty-five percent of clinics surveyed reported actively treating/managing tyrosine levels (with supplemental tyrosine above that contained in PKU formulas) to ensure tyrosine levels remain within normal ranges. Anecdotally, supplemental tyrosine has been reported to ameliorate symptoms of both attention deficit hyperactivity disorder and depression in this population. EF assessment of children with ECT-PKU was likewise highly variable, with 50% of clinics surveyed reporting routine assessments of intellectual function. However when function was assessed, test instruments chosen tended towards global measures of IQ prior to school entry, rather than specific assessment of EF development. Further investigation of the role of tyrosine and its relationship with phe and EF development is needed to establish whether routine tyrosine monitoring and increased supplementation is recommended. PMID:20882350

  14. Diagnostic ions for the analysis of phenylalanine adducts of acrylamide and styrene by ESI-QTOF mass spectrometry.

    PubMed

    Chu, Fong Lam; Sleno, Lekha; Yaylayan, Varoujan Antranik

    2013-10-30

    To facilitate the detection of acrylamide or styrene adduct of amino acids by mass spectrometry based techniques, phenylalanine was used as a representative amino acid and pyrolysis was employed in conjunction with isotope labeling technique as a microscale sample preparation tool to generate the reaction products. The residues remaining after the pyrolysis of phenylalanine/styrene, phenylalanine/acrylamide, and phenylalanine/glucose mixtures at 250 °C were analyzed by electrospray quadrupole time-of-flight (ESI-QqTOF) mass spectrometry to identify the adducts. The phenylalanine/acrylamide adduct was independently synthesized for confirmation. Characteristic product ions in the tandem mass spectra were found at m/z 191 for the acrylamide adduct and at m/z 262 and 190 for its double-addition product. On the other hand, an ion at m/z 224 was shown to be diagnostic of the styrene adduct. The ability of the m/z 224 ion to predict the presence of styrene adduct in a heated phenylalanine/glucose model system was tested and verified. Detailed isotope labeling analysis of the phenylalanine/glucose model further indicated the formation of a novel adduct that was consistent with the reaction of the Amadori product with styrene. Such diagnostic ions that are needed to develop MS/MS-based screening methodologies may accelerate in the future the detection of Michael-type adducts in food.

  15. Age-Related Psychophysiological Vulnerability to Phenylalanine in Phenylketonuria

    PubMed Central

    Leuzzi, Vincenzo; Mannarelli, Daniela; Manti, Filippo; Pauletti, Caterina; Locuratolo, Nicoletta; Carducci, Carla; Carducci, Claudia; Vanacore, Nicola; Fattapposta, Francesco

    2014-01-01

    Background: Phenylketonuria (PKU) is caused by the inherited defect of the phenylalanine hydroxylase enzyme, which converts phenylalanine (Phe) into tyrosine (Tyr). Neonatal screening programs and early treatment have radically changed the natural history of PKU. Nevertheless, an increased risk of neurocognitive and psychiatric problems in adulthood remains a challenging aspect of the disease. In order to assess the vulnerability of complex skills to Phe, we explored: (a) the effect of a rapid increase in blood Phe levels on event-related potentials (ERP) in PKU subjects during their second decade of life; (b) the association (if existing) between psychophysiological and neurocognitive features. Methods: Seventeen early-treated PKU subjects, aged 10–20, underwent ERP [mismatch negativity, auditory P300, contingent negative variation (CNV), and Intensity Dependence of Auditory Evoked Potentials] recording before and 2 h after an oral loading of Phe. Neurocognitive functioning, historical and concurrent biochemical values of blood Phe, Tyr, and Phe/Tyr ratio, were all included in the statistical analysis. Results: Event-related potential components were normally detected in all the subjects. In subjects younger than 13 CNV amplitude, W2-CNV area, P3b latency, and reaction times in motor responses were negatively influenced by Phe-loading. Independently from the psychophysiological vulnerability, some neurocognitive skills were more impaired in younger patients. No correlation was found between biochemical alterations and neurocognitive and psychophysiological findings. Conclusion: The vulnerability of the emerging neurocognitive functions to Phe suggests a strict metabolic control in adolescents affected by PKU and a neurodevelopmental approach in the study of neurocognitive outcome in PKU. PMID:25003100

  16. Receptor Tyrosine Kinase and Tyrosine Kinase Inhibitors

    PubMed Central

    Mirshafiey, Abbas; Ghalamfarsa, Ghasem; Asghari, Babak

    2014-01-01

    Receptor tyrosine kinases (RTKs) are essential components of signal transduction pathways that mediate cell-to-cell communication and their function as relay points for signaling pathways. They have a key role in numerous processes that control cellular proliferation and differentiation, regulate cell growth and cellular metabolism, and promote cell survival and apoptosis. Recently, the role of RTKs including TCR, FLT-3, c-Kit, c-Fms, PDGFR, ephrin, neurotrophin receptor, and TAM receptor in autoimmune disorder, especially rheumatoid arthritis and multiple sclerosis has been suggested. In multiple sclerosis pathogenesis, RTKs and their tyrosine kinase enzymes are selective important targets for tyrosine kinase inhibitor (TKI) agents. TKIs, compete with the ATP binding site of the catalytic domain of several tyrosine kinases, and act as small molecules that have a favorable safety profile in disease treatment. Up to now, the efficacy of TKIs in numerous animal models of MS has been demonstrated, but application of these drugs in human diseases should be tested in future clinical trials. PMID:25337443

  17. The Conserved Phenylalanine in the Transmembrane Domain Enhances Heteromeric Interactions of Syndecans.

    PubMed

    Kwon, Mi-Jung; Park, Jisu; Jang, Sinae; Eom, Chi-Yong; Oh, Eok-Soo

    2016-01-01

    The transmembrane domain (TMD) of the syndecans, a family of transmembrane heparin sulfate proteoglycans, is involved in forming homo- and heterodimers and oligomers that transmit signaling events. Recently, we reported that the unique phenylalanine in TMD positively regulates intramolecular interactions of syndecan-2. Besides the unique phenylalanine, syndecan-2 contains a conserved phenylalanine (SDC2-Phe-169) that is present in all syndecan TMDs, but its function has not been determined. We therefore investigated the structural role of SDC2-Phe-169 in syndecan TMDs. Replacement of SDC2-Phe-169 by tyrosine (S2F169Y) did not affect SDS-resistant homodimer formation but significantly reduced SDS-resistant heterodimer formation between syndecan-2 and -4, suggesting that SDC2-Phe-169 is involved in the heterodimerization/oligomerization of syndecans. Similarly, in an in vitro binding assay, a syndecan-2 mutant (S2(F169Y)) showed a significantly reduced interaction with syndecan-4. FRET assays showed that heteromolecular interactions between syndecan-2 and -4 were reduced in HEK293T cells transfected with S2(F169Y) compared with syndecan-2. Moreover, S2(F169Y) reduced downstream reactions mediated by the heterodimerization of syndecan-2 and -4, including Rac activity, cell migration, membrane localization of PKCα, and focal adhesion formation. The conserved phenylalanine in syndecan-1 and -3 also showed heterodimeric interaction with syndecan-2 and -4. Taken together, these findings suggest that the conserved phenylalanine in the TMD of syndecans is crucial in regulating heteromeric interactions of syndecans.

  18. Identification of the Allosteric Site for Phenylalanine in Rat Phenylalanine Hydroxylase.

    PubMed

    Zhang, Shengnan; Fitzpatrick, Paul F

    2016-04-01

    Liver phenylalanine hydroxylase (PheH) is an allosteric enzyme that requires activation by phenylalanine for full activity. The location of the allosteric site for phenylalanine has not been established. NMR spectroscopy of the isolated regulatory domain (RDPheH(25-117) is the regulatory domain of PheH lacking residues 1-24) of the rat enzyme in the presence of phenylalanine is consistent with formation of a side-by-side ACT dimer. Six residues in RDPheH(25-117) were identified as being in the phenylalanine-binding site on the basis of intermolecular NOEs between unlabeled phenylalanine and isotopically labeled protein. The location of these residues is consistent with two allosteric sites per dimer, with each site containing residues from both monomers. Site-specific variants of five of the residues (E44Q, A47G, L48V, L62V, and H64N) decreased the affinity of RDPheH(25-117) for phenylalanine based on the ability to stabilize the dimer. Incorporation of the A47G, L48V, and H64N mutations into the intact protein increased the concentration of phenylalanine required for activation. The results identify the location of the allosteric site as the interface of the regulatory domain dimer formed in activated PheH.

  19. Free amino acid profiling in the giant puffball mushroom (Calvatia gigantea) using UPLC-MS/MS.

    PubMed

    Kıvrak, İbrahim; Kıvrak, Şeyda; Harmandar, Mansur

    2014-09-01

    Wild edible and medicinal mushroom, Calvatia gigantea, was quantitatively analyzed for the determination of its free amino acids using ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). The concentrations of total free amino acids, essential and non-essential amino acids were 199.65 mg/100 g, 113.69 mg/100 g, and 85.96 mg/100 g in C. gigantea, respectively. This study showed that C. gigantea, so called a giant puffball mushroom, has free amino acids content. The essential amino acids: tryptophan, isoleucine, valine, phenylalanine, leucine, threonine, lysine, histidine, methionine, and the non-essential amino acids: tyrosine, 4-hyrdroxy proline, arginine, proline, glycine, serine, alanine, glutamine, glutamic acid, aspargine, aspartic acid were detected.

  20. Plasma Amino Acid Concentrations Predict Mortality in Patients with End-Stage Liver Disease

    PubMed Central

    Kinny-Köster, Benedict; Bartels, Michael; Becker, Susen; Scholz, Markus; Thiery, Joachim

    2016-01-01

    Background The liver plays a key role in amino acid metabolism. In former studies, a ratio between branched-chain and aromatic amino acids (Fischer’s ratio) revealed associations with hepatic encephalopathy. Furthermore, low concentrations of branched-chain amino acids were linked to sarcopenia in literature. Encephalopathy and sarcopenia are known to dramatically worsen the prognosis. Aim of this study was to investigate a complex panel of plasma amino acids in the context of mortality in patients with end-stage liver disease. Methods 166 patients evaluated for orthotopic liver transplantation were included. 19 amino acids were measured from citrated plasma samples using mass spectrometry. We performed survival analysis for plasma amino acid constellations and examined the relationship to established mortality predictors. Results 33/166 (19.9%) patients died during follow-up. Lower values of valine (p<0.001), Fischer’s ratio (p<0.001) and valine to phenylalanine ratio (p<0.001) and higher values of phenylalanine (p<0.05) and tyrosine (p<0.05) were significantly associated with mortality. When divided in three groups, the tertiles discriminated cumulative survival for valine (p = 0.016), phenylalanine (p = 0.024) and in particular for valine to phenylalanine ratio (p = 0.003) and Fischer’s ratio (p = 0.005). Parameters were also significantly correlated with MELD and MELD-Na score. Conclusions Amino acids in plasma are valuable biomarkers to determine increased risk of mortality in patients with end-stage liver disease. In particular, valine concentrations and constellations composed of branched-chain and aromatic amino acids were strongly associated with prognosis. Due to their pathophysiological importance, the identified amino acids could be used to examine individual dietary recommendations to serve as potential therapeutic targets. PMID:27410482

  1. Ultrafast electron dynamics in phenylalanine initiated by attosecond pulses

    NASA Astrophysics Data System (ADS)

    Calegari, F.; Ayuso, D.; Trabattoni, A.; Belshaw, L.; De Camillis, S.; Anumula, S.; Frassetto, F.; Poletto, L.; Palacios, A.; Decleva, P.; Greenwood, J. B.; Martín, F.; Nisoli, M.

    2014-10-01

    In the past decade, attosecond technology has opened up the investigation of ultrafast electronic processes in atoms, simple molecules, and solids. Here, we report the application of isolated attosecond pulses to prompt ionization of the amino acid phenylalanine and the subsequent detection of ultrafast dynamics on a sub-4.5-femtosecond temporal scale, which is shorter than the vibrational response of the molecule. The ability to initiate and observe such electronic dynamics in polyatomic molecules represents a crucial step forward in attosecond science, which is progressively moving toward the investigation of more and more complex systems.

  2. First-principles study on the adsorption properties of phenylalanine on carbon graphitic structures

    NASA Astrophysics Data System (ADS)

    Kang, Seoung-Hun; Kwon, Dae-Gyeon; Park, Sora; Kwon, Young-Kyun

    2015-12-01

    Using ab-initio density functional theory, we investigate the binding properties of phenylalanine, an amino acid, on graphitic carbon structures, such as graphene, nanotubes, and their modified structures. We focus especially on the effect of the adsorbate on the geometrical and the electronic structures of the absorbents. The phenylalanine molecule is found to bind weakly on pristine graphitic structures with a binding energy of 40-70 meV and not to change the electronic configuration of the graphitic structures, implying that the phenylalanine molecule may not be detected on pristine graphitic structures. On the other hand, the phenylalanine molecule exhibits a substantial increase in its binding energy up to ~2.60 eV on the magnesium-decorated boron-doped graphitic structures. We discover that the Fermi level of the system, which was shifted below the Dirac point of the graphitic structures due to p-doping by boron substitution, can be completely restored to the Dirac point because of the amino acid adsorption. This behavior implies that such modified structures can be utilized to detect phenylalanine molecules.

  3. Ternary oxovanadium(IV) complexes with amino acid-Schiff base and polypyridyl derivatives: synthesis, characterization, and protein tyrosine phosphatase 1B inhibition.

    PubMed

    Lu, Liping; Yue, Jinjun; Yuan, Caixia; Zhu, Miaoli; Han, Hong; Liu, Zhiwei; Guo, Maolin

    2011-10-01

    To investigate the structure-activity relationship of vanadium complexes in inhibiting protein tyrosine phosphatase1B (PTP1B), eight mixed-ligand oxovanadium(IV) complexes, [V(IV)O(SalAla)(NN)] (H(2)SalAla for salicylidene alanine, NN for N,N'-donor heterocyclic base, namely, 2,2'-bipyridine (bpy, 1), 1,10-phenanthroline (phen, 2), dipyrido[3,2-d:2',3'-f]quinoxaline (dpq, 3), dipyrido[3,2-a:2',3'-c]phenazine (dppz, 4)), [V(IV)O(SalLys)(dpq)] (5), [V(IV)O(SalLys)(dppz)] (6), [V(IV)O(SalAsp)(dppz)], (7) and [V(IV)O(SalTrp)(dppz)] (8)), of which 3-8 are new, have been prepared and characterized by elemental analysis, infrared, UV-visible, electrospray ionization mass spectrometry and conductivity. The molar conductance data confirmed the non-electrolytic nature of the complexes in DMSO solution. The coordination in [V(IV)O (SalAla)(phen)] (2) was confirmed by X-ray crystal structure analysis. The oxidation state of V(IV) with d(1) configuration in 2 was confirmed by EPR. The speciation of VO-SalAla-phen in aqueous solution was investigated by potentiometric pH titrations. The results indicate that the main species are two ternary complexes at the pH range 7.0-7.4. Biochemical assays demonstrate that the mixed-ligand oxovanadium(IV) complexes are potent inhibitors of PTP1B with IC(50) values in the range of 62-597nM, approximately 3-10 fold weaker in potency than those of similar mixed-ligand oxovanadium(IV) complexes of salicylidene anthranilic acid (SAA) derivative with polypyridyl ligands, except complex 8, which exhibits comparable or better inhibition activity than those of the mixed-ligand oxovanadium(IV) complexes of SAA derivative with polypyridyl ligands. The results demonstrate that the structures of vanadium complexes influence the PTP1B inhibition activity. Kinetics assays reveal that complex 2 inhibits PTP1B in a competitive manner.

  4. Phenylalanine hydroxylase: function, structure, and regulation.

    PubMed

    Flydal, Marte I; Martinez, Aurora

    2013-04-01

    Mammalian phenylalanine hydroxylase (PAH) catalyzes the rate-limiting step in the phenylalanine catabolism, consuming about 75% of the phenylalanine input from the diet and protein catabolism under physiological conditions. In humans, mutations in the PAH gene lead to phenylketonuria (PKU), and most mutations are mainly associated with PAH misfolding and instability. The established treatment for PKU is a phenylalanine-restricted diet and, recently, supplementation with preparations of the natural tetrahydrobiopterin cofactor also shows effectiveness for some patients. Since 1997 there has been a significant increase in the understanding of the structure, catalytic mechanism, and regulation of PAH by its substrate and cofactor, in addition to improved correlations between genotype and phenotype in PKU. Importantly, there has also been an increased number of studies on the structure and function of PAH from bacteria and lower eukaryote organisms, revealing an additional anabolic role of the enzyme in the synthesis of melanin-like pigments. In this review, we discuss these recent studies, which contribute to define the evolutionary adaptation of the PAH structure and function leading to sophisticated regulation for effective catabolic processing of phenylalanine in mammalian organisms.

  5. Therapeutic implication of L-phenylalanine aggregation mechanism and its modulation by D-phenylalanine in phenylketonuria.

    PubMed

    Singh, Virender; Rai, Ratan Kumar; Arora, Ashish; Sinha, Neeraj; Thakur, Ashwani Kumar

    2014-01-27

    Self-assembly of phenylalanine is linked to amyloid formation toxicity in phenylketonuria disease. We are demonstrating that L-phenylalanine self-assembles to amyloid fibrils at varying experimental conditions and transforms to a gel state at saturated concentration. Biophysical methods including nuclear magnetic resonance, resistance by alpha-phenylglycine to fibril formation and preference of protected phenylalanine to self-assemble show that this behaviour of L-phenylalanine is governed mainly by hydrophobic interactions. Interestingly, D-phenylalanine arrests the fibre formation by L-phenylalanine and gives rise to flakes. These flakes do not propagate further and prevent fibre formation by L-phenylalanine. This suggests the use of D-phenylalanine as modulator of L-phenylalanine amyloid formation and may qualify as a therapeutic molecule in phenylketonuria.

  6. Amino acid profile during exercise and training in Standardbreds.

    PubMed

    Westermann, C M; Dorland, L; Wijnberg, I D; de Sain-van der Velden, M G M; van Breda, E; Barneveld, A; de Graaf-Roelfsema, E; Keizer, H A; van der Kolk, J H

    2011-08-01

    The objective of this study is to assess the influence of acute exercise, training and intensified training on the plasma amino acid profile. In a 32-week longitudinal study using 10 Standardbred horses, training was divided into four phases, including a phase of intensified training for five horses. At the end of each phase, a standardized exercise test, SET, was performed. Plasma amino acid concentrations before and after each SET were measured. Training significantly reduced mean plasma aspartic acid concentration, whereas exercise significantly increased the plasma concentrations of alanine, taurine, methionine, leucine, tyrosine and phenylalanine and reduced the plasma concentrations of glycine, ornithine, glutamine, citrulline and serine. Normally and intensified trained horses differed not significantly. It is concluded that amino acids should not be regarded as limiting training performance in Standardbreds except for aspartic acid which is the most likely candidate for supplementation. PMID:20863542

  7. Protein Quantification by Derivatization-Free High-Performance Liquid Chromatography of Aromatic Amino Acids

    PubMed Central

    Hesse, Almut

    2016-01-01

    Amino acid analysis is considered to be the gold standard for quantitative peptide and protein analysis. Here, we would like to propose a simple HPLC/UV method based on a reversed-phase separation of the aromatic amino acids tyrosine (Tyr), phenylalanine (Phe), and optionally tryptophan (Trp) without any derivatization. The hydrolysis of the proteins and peptides was performed by an accelerated microwave technique, which needs only 30 minutes. Two internal standard compounds, homotyrosine (HTyr) and 4-fluorophenylalanine (FPhe) were used for calibration. The limit of detection (LOD) was estimated to be 0.05 µM (~10 µg/L) for tyrosine and phenylalanine at 215 nm. The LOD for a protein determination was calculated to be below 16 mg/L (~300 ng BSA absolute). Aromatic amino acid analysis (AAAA) offers excellent accuracy and a precision of about 5% relative standard deviation, including the hydrolysis step. The method was validated with certified reference materials (CRM) of amino acids and of a pure protein (bovine serum albumin, BSA). AAAA can be used for the quantification of aromatic amino acids, isolated peptides or proteins, complex peptide or protein samples, such as serum or milk powder, and peptides or proteins immobilized on solid supports. PMID:27559481

  8. Protein Quantification by Derivatization-Free High-Performance Liquid Chromatography of Aromatic Amino Acids.

    PubMed

    Hesse, Almut; Weller, Michael G

    2016-01-01

    Amino acid analysis is considered to be the gold standard for quantitative peptide and protein analysis. Here, we would like to propose a simple HPLC/UV method based on a reversed-phase separation of the aromatic amino acids tyrosine (Tyr), phenylalanine (Phe), and optionally tryptophan (Trp) without any derivatization. The hydrolysis of the proteins and peptides was performed by an accelerated microwave technique, which needs only 30 minutes. Two internal standard compounds, homotyrosine (HTyr) and 4-fluorophenylalanine (FPhe) were used for calibration. The limit of detection (LOD) was estimated to be 0.05 µM (~10 µg/L) for tyrosine and phenylalanine at 215 nm. The LOD for a protein determination was calculated to be below 16 mg/L (~300 ng BSA absolute). Aromatic amino acid analysis (AAAA) offers excellent accuracy and a precision of about 5% relative standard deviation, including the hydrolysis step. The method was validated with certified reference materials (CRM) of amino acids and of a pure protein (bovine serum albumin, BSA). AAAA can be used for the quantification of aromatic amino acids, isolated peptides or proteins, complex peptide or protein samples, such as serum or milk powder, and peptides or proteins immobilized on solid supports. PMID:27559481

  9. Cloning and Expression Analysis of Phenylalanine Ammonia-Lyase Gene in the Mycelium and Fruit Body of the Edible Mushroom Flammulina velutipes.

    PubMed

    Yun, Yeo Hong; Koo, Ja Sun; Kim, Seong Hwan; Kong, Won Sik

    2015-09-01

    Phenylalanine ammonia-lyase (PAL) gene is known to be expressed in plants, and is involved in the differentiation, growth and synthesis of secondary metabolites. However, its expression in fungi remains to be explored. To understand its expression in mushroom fungi, the PAL gene of the edible mushroom Flammulina velutipes (Fvpal) was cloned and characterized. The cloned Fvpal consists of 2,175 bp, coding for a polypeptide containing 724 amino acids and having 11 introns. The translated amino acid sequence of Fvpal shares a high identity (66%) with that of ectomycorrhizal fungus Tricholoma matsutake. Distinctively, the Fvpal expression in the mycelium was higher in minimal medium supplemented with L-tyrosine than with other aromatic amino acids. During cultivation of the mushroom on sawdust medium, Fvpal expression in the fruit body correspondingly increased as the mushroom grew. In the fruiting body, Fvpal was expressed more in the stipe than in the pileus. These results suggest that F. velutipes PAL activity differs in the different organs of the mushroom. Overall, this is first report to show that the PAL gene expression is associated with mushroom growth in fungi. PMID:26539050

  10. Plasma phenylalanine levels in phenylketonuric heterozygous and normal adults administered aspartame at 34 mg/kg body weight.

    PubMed

    Stegink, L D; Koch, R; Blaskovics, M E; Filer, L J; Baker, G L; McDonnell, J E

    1981-01-01

    Following administration of aspartame (34 mg/kg body wt) in orange juice, plasma concentrations of free amino acids were measured in 12 female subjects known to be heterozygous for phenylketonuria and 22 normal subjects (12 male, 10 female). No change in fasting plasma aspartate concentrations were noted after aspartame loading in either group. In normal male subjects, the mean (+/-S.D.) plasma phenylalanine concentration increased from a fasting value of 5.86 +/- 1.25 mumol/dl. Plasma phenylalanine levels in normal female subjects increased from a mean fasting concentration of 4.83 +/- 0.84 mumol/dl to a men peak value of 8.95 +/- 1.49 mumol/dl suggesting a more rapid absorption, metabolism, and/or clearance of phenylalanine by females. In female heterozygous subjects, the mean peak plasma phenylalanine concentration was significantly higher than in normal females. Plasma phenylalanine values increased from a mean fasting value of 5.92 +/- 1.51 mumol/dl to a mean peak value of 15.1 +/- 4.76 mumol/dl. Similarly, the area under the plasma phenylalanine concentration-time curve was significantly greater in heterozygous female subjects (21.36 +/- 5.10 IU) than in normal female subjects (10.84 +/- 2.32 IU). However, peak plasma phenylalanine levels were well below those associated with toxic effects in all cases.

  11. Preparation of l-phenylalanine-imprinted solid-phase extraction sorbent by Pickering emulsion polymerization and the selective enrichment of l-phenylalanine from human urine.

    PubMed

    Li, Ji; Hu, Xiaoling; Guan, Ping; Zhang, Xiaoyan; Qian, Liwei; Zhang, Nan; Du, Chunbao; Song, Renyuan

    2016-05-01

    A novel l-phenylalanine molecularly imprinted solid-phase extraction sorbent was synthesized by the combination of Pickering emulsion polymerization and ion-pair dummy template imprinting. Compared to other polymerization methods, the molecularly imprinted polymers thus prepared exhibit a high specific surface, large pore diameter, and appropriate particle size. The key parameters for solid-phase extraction were optimized, and the result indicated that the molecularly imprinted polymer thus prepared exhibits a good recovery of 98.9% for l-phenylalanine. Under the optimized conditions of the procedure, an analytical method for l-phenylalanine was well established. By comparing the performance of the molecularly imprinted polymer and a commercial reverse-phase silica gel, the obtained molecularly imprinted polymer as an solid-phase extraction sorbent is more suitable, exhibiting high precision (relative standard deviation 3.2%, n = 4) and a low limit of detection (60.0 ± 1.9 nmol·L(-1) ) for the isolation of l-phenylalanine. Based on these results, the combination of the Pickering emulsion polymerization and ion-pair dummy template imprinting is effective for preparing selective solid-phase extraction sorbents for the separation of amino acids and organic acids from complex biological samples. PMID:26991761

  12. The rotational spectrum of tyrosine.

    PubMed

    Pérez, Cristóbal; Mata, Santiago; Cabezas, Carlos; López, Juan C; Alonso, José L

    2015-04-23

    In this work neutral tyrosine has been generated in the gas phase by laser ablation of solid samples, and its most abundant conformers characterized through their rotational spectra. Their identification has been made by comparison between the experimental and ab initio values of the rotational and quadrupole coupling constants. Both conformers are stabilized by an O-H•••N hydrogen bond established within the amino acid skeleton chain and an additional weak N-H•••π hydrogen bond. The observed conformers differ in the orientation of the phenolic -OH group.

  13. Construction and evaluation of a novel bifunctional phenylalanine-formate dehydrogenase fusion protein for bienzyme system with cofactor regeneration.

    PubMed

    Jiang, Wei; Fang, Bai-Shan

    2016-05-01

    Phenylalanine dehydrogenase (PheDH) plays an important role in enzymatic synthesis of L-phenylalanine for aspartame (sweetener) and detection of phenylketonuria (PKU), suggesting that it is important to obtain a PheDH with excellent characteristics. Gene fusion of PheDH and formate dehydrogenase (FDH) was constructed to form bifunctional multi-enzymes for bioconversion of L-phenylalanine coupled with coenzyme regeneration. Comparing with the PheDH monomer from Microbacterium sp., the bifunctional PheDH-FDH showed noteworthy stability under weakly acidic and alkaline conditions (pH 6.5-9.0). The bifunctional enzyme can produce 153.9 mM L-phenylalanine with remarkable performance of enantiomers choice by enzymatic conversion with high molecular conversion rate (99.87 %) in catalyzing phenylpyruvic acid to L-phenylalanine being 1.50-fold higher than that of the separate expression system. The results indicated the potential application of the PheDH and PheDH-FDH with coenzyme regeneration for phenylpyruvic acid analysis and L-phenylalanine biosynthesis in medical diagnosis and pharmaceutical field. PMID:26819086

  14. Construction and evaluation of a novel bifunctional phenylalanine-formate dehydrogenase fusion protein for bienzyme system with cofactor regeneration.

    PubMed

    Jiang, Wei; Fang, Bai-Shan

    2016-05-01

    Phenylalanine dehydrogenase (PheDH) plays an important role in enzymatic synthesis of L-phenylalanine for aspartame (sweetener) and detection of phenylketonuria (PKU), suggesting that it is important to obtain a PheDH with excellent characteristics. Gene fusion of PheDH and formate dehydrogenase (FDH) was constructed to form bifunctional multi-enzymes for bioconversion of L-phenylalanine coupled with coenzyme regeneration. Comparing with the PheDH monomer from Microbacterium sp., the bifunctional PheDH-FDH showed noteworthy stability under weakly acidic and alkaline conditions (pH 6.5-9.0). The bifunctional enzyme can produce 153.9 mM L-phenylalanine with remarkable performance of enantiomers choice by enzymatic conversion with high molecular conversion rate (99.87 %) in catalyzing phenylpyruvic acid to L-phenylalanine being 1.50-fold higher than that of the separate expression system. The results indicated the potential application of the PheDH and PheDH-FDH with coenzyme regeneration for phenylpyruvic acid analysis and L-phenylalanine biosynthesis in medical diagnosis and pharmaceutical field.

  15. Effect of L-aspartyl-L-phenylalanine methyl ester on leukotriene biosynthesis in macrophage cells.

    PubMed

    Hardcastle, J E; Bruch, R J

    1997-09-01

    Macrophage cells treated with L-aspartyl-L-phenylalanine methyl ester (aspartame) produced leukotrienes and other arachidonic acid metabolites. Leukotriene C4, leukotriene B4, and 15-hydroxyeicosatetraenoic acid were the major metabolites detected. The aspartame-treated macrophage cell cultures produced three times as much arachidonic acid metabolites as did the untreated control cell cultures. Leukotriene C4 was produced in the largest amount by the aspartame-treated cells.

  16. Biosynthesis of the Aromatic Amino Acids.

    PubMed

    Pittard, James; Yang, Ji

    2008-09-01

    This chapter describes in detail the genes and proteins of Escherichia coli involved in the biosynthesis and transport of the three aromatic amino acids tyrosine, phenylalanine, and tryptophan. It provides a historical perspective on the elaboration of the various reactions of the common pathway converting erythrose-4-phosphate and phosphoenolpyruvate to chorismate and those of the three terminal pathways converting chorismate to phenylalanine, tyrosine, and tryptophan. The regulation of key reactions by feedback inhibition, attenuation, repression, and activation are also discussed. Two regulatory proteins, TrpR (108 amino acids) and TyrR (513 amino acids), play a major role in transcriptional regulation. The TrpR protein functions only as a dimer which, in the presence of tryptophan, represses the expression of trp operon plus four other genes (the TrpR regulon). The TyrR protein, which can function both as a dimer and as a hexamer, regulates the expression of nine genes constituting the TyrR regulon. TyrR can bind each of the three aromatic amino acids and ATP and under their influence can act as a repressor or activator of gene expression. The various domains of this protein involved in binding the aromatic amino acids and ATP, recognizing DNA binding sites, interacting with the alpha subunit of RNA polymerase, and changing from a monomer to a dimer or a hexamer are all described. There is also an analysis of the various strategies which allow TyrR in conjunction with particular amino acids to differentially affect the expression of individual genes of the TyrR regulon. PMID:26443741

  17. A study of placental transfer mechanisms in nonhuman primates using (/sup 14/C)phenylalanine

    SciTech Connect

    Pueschel, S.M.; Boylan, J.M.; Jackson, B.T.; Piasecki, G.J.

    1982-02-01

    Placental transfer mechanisms were investigated in pregnant Macaca Fascicularis and Macaca mulatta during the gestational age of 120 to 130 days. These primates underwent an operative procedure that allowed continuous fetal blood sampling. The administration of (/sup 14/C)phenylalanine into the maternal circulation revealed a significant increase of radioactive material in the fetal circulation, indicating an active placental transport mechanism unidirectional to the fetus. When (/sup 14/C)phenylalanine was injected into the fetus, radioactive aromatic amino acids in the maternal circulation increased only slightly over time, resembling a simple diffusion process.

  18. Phenylalanine binding is linked to dimerization of the regulatory domain of phenylalanine hydroxylase.

    PubMed

    Zhang, Shengnan; Roberts, Kenneth M; Fitzpatrick, Paul F

    2014-10-28

    Analytical ultracentrifugation has been used to analyze the oligomeric structure of the isolated regulatory domain of phenylalanine hydroxylase. The protein exhibits a monomer-dimer equilibrium with a dissociation constant of ~46 μM; this value is unaffected by the removal of the 24 N-terminal residues or by phosphorylation of Ser16. In contrast, phenylalanine binding (Kd = 8 μM) stabilizes the dimer. These results suggest that dimerization of the regulatory domain of phenylalanine hydroxylase is linked to allosteric activation of the enzyme.

  19. Beta 2 integrin-dependent protein tyrosine phosphorylation and activation of the FGR protein tyrosine kinase in human neutrophils

    PubMed Central

    1994-01-01

    Stimulation of adherent human neutrophils (PMN) with tumor necrosis factor (TNF) triggers protein tyrosine phosphorylation (Fuortes, M., W. W. Jin, and C. Nathan. 1993. J. Cell Biol. 120:777-784). We investigated the dependence of this response on beta 2 integrins by using PMN isolated from a leukocyte adhesion deficiency (LAD) patient, which do not express beta 2 integrins, and by plating PMN on surface bound anti-beta 2 (CD18) antibodies. Protein tyrosine phosphorylation increased in PMN plated on fibrinogen and this phosphorylation was enhanced by TNF. Triggering of protein tyrosine phosphorylation did not occur in LAD PMN plated on fibrinogen either in the absence or the presence of TNF. Surface bound anti-CD18, but not isotype-matched anti- Class I major histocompatibility complex (MHC) antigens, antibodies triggered tyrosine phosphorylation in normal, but not in LAD PMN. As the major tyrosine phosphorylated proteins we found in our assay conditions migrated with an apparent molecular mass of 56-60 kD, we investigated whether beta 2 integrins are implicated in activation of members of the src family of intracellular protein-tyrosine kinases. We found that the fgr protein-tyrosine kinase (p58fgr) activity, and its extent of phosphorylation in tyrosine, in PMN adherent to fibrinogen, was enhanced by TNF. Activation of p58fgr in response to TNF was evident within 10 min of treatment and increased with times up to 30 min. Also other activators of beta 2 integrins such as phorbol-12- myristate 13-acetate (PMA), and formyl methionyl-leucyl-phenylalanine (FMLP), induced activation of p58fgr kinase activity. Activation of p58fgr kinase activity, and phosphorylation in tyrosine, did not occur in PMN of a LAD patient in response to TNF. Soluble anti-CD18, but not anti-Class I MHC antigens, antibodies inhibited activation of p58fgr kinase activity in PMN adherent to fibrinogen in response to TNF, PMA, and FMLP. These findings demonstrate that, in PMN, beta 2 integrins

  20. Aromatic amino acids as precursors of antimicrobial metabolites in Geotrichum candidum.

    PubMed

    Naz, Saima; Gueguen-Minerbe, Marielle; Cretenet, Marina; Vernoux, Jean-Paul

    2013-07-01

    Geotrichum candidum ATCC 204307 was previously found to generate phenyllactic acid (PLA) and indoleacetic acid (ILA) in complex culture media. In this study, a relationship between concentrations of PLA, ILA, and hydroxy PLA (OH-PLA) and initial concentrations of phenylalanine, tryptophan, and tyrosine, added respectively as unique sources of nitrogen in synthetic medium, was established. Phenylpyruvic acid (PPA), an intermediate compound of PLA metabolism, was able to induce not only PLA but also phenylethyl alcohol (PEA) production when used separately as initial substrate. Under pH, temperature, and salt concentrations used for cheese-making, phenylalanine was found to be the most efficient substrate for antimicrobial metabolite production. In excess of substrate, different yeast strains of Geotrichum candidum, Yarrowia lipolytica, Candida natalensis, and Candida catenulata were shown here to produce 1.6 ± 0.5-5.0 ± 0.2 mM of PLA from phenylalanine, 5.0 ± 0.1-10.9 ± 0.3 mM of ILA from tryptophan, and 1.3 ± 0.3-7.0 ± 0.02 of PLA and 0.1 ± 0.0-2.22 ± 0.09 mM of PEA from PPA. Geotrichum candidum ATCC 204307 was the highest producer. This is the first time these antimicrobial metabolites PLA, OH-PLA, ILA, and PEA are being reported as the reaction products of aromatic amino acids catabolism in G. candidum. PMID:23590565

  1. Phenylalanine and Phenylglycine Analogues as Arginine Mimetics in Dengue Protease Inhibitors.

    PubMed

    Weigel, Lena F; Nitsche, Christoph; Graf, Dominik; Bartenschlager, Ralf; Klein, Christian D

    2015-10-01

    Dengue virus is an increasingly global pathogen. One of the promising targets for antiviral drug discovery against dengue and related flaviviruses such as West Nile virus is the viral serine protease NS2B-NS3. We here report the synthesis and in vitro characterization of potent peptidic inhibitors of dengue virus protease that incorporate phenylalanine and phenylglycine derivatives as arginine-mimicking groups with modulated basicity. The most promising compounds were (4-amidino)-L-phenylalanine-containing inhibitors, which reached nanomolar affinities against dengue virus protease. The type and position of the substituents on the phenylglycine and phenylalanine side chains has a significant effect on the inhibitory activity against dengue virus protease and selectivity against other proteases. In addition, the non-natural, basic amino acids described here may have relevance for the development of other peptidic and peptidomimetic drugs such as inhibitors of the blood clotting cascade.

  2. Phenylalanine is required to promote specific developmental responses and prevents cellular damage in response to ultraviolet light in soybean (Glycine max) during the seed-to-seedling transition.

    PubMed

    Sullivan, Joe H; Muhammad, DurreShahwar; Warpeha, Katherine M

    2014-01-01

    UV-radiation elicits a suite of developmental (photomorphogenic) and protective responses in plants, but responses early post-germination have received little attention, particularly in intensively bred plants of economic importance. We examined germination, hypocotyl elongation, leaf pubescence and subcellular responses of germinating and/or etiolated soybean (Glycine max (L.) Merr.) seedlings in response to treatment with discrete wavelengths of UV-A or UV-B radiation. We demonstrate differential responses of germinating/young soybean seedlings to a range of UV wavelengths that indicate unique signal transduction mechanisms regulate UV-initiated responses. We have investigated how phenylalanine, a key substrate in the phenylpropanoid pathway, may be involved in these responses. Pubescence may be a key location for phenylalanine-derived protective compounds, as UV-B irradiation increased pubescence and accumulation of UV-absorbing compounds within primary leaf pubescence, visualized by microscopy and absorbance spectra. Mass spectrometry analysis of pubescence indicated that sinapic esters accumulate in the UV-irradiated hairs compared to unirradiated primary leaf tissue. Deleterious effects of some UV-B wavelengths on germination and seedling responses were reduced or entirely prevented by inclusion of phenylalanine in the growth media. Key effects of phenylalanine were not duplicated by tyrosine or tryptophan or sucrose, nor is the specificity of response due to the absorbance of phenylalanine itself. These results suggest that in the seed-to-seedling transition, phenylalanine may be a limiting factor in the development of initial mechanisms of UV protection in the developing leaf.

  3. Autistic children exhibit distinct plasma amino acid profile.

    PubMed

    Naushad, Shaik Mohammad; Jain, Jamal Md Nurul; Prasad, Chintakindi Krishna; Naik, Usha; Akella, Radha Rama Devi

    2013-10-01

    In order to ascertain whether autistic children display characteristic metabolic signatures that are of diagnostic value, plasma amino acid analyses were carried out on a cohort of 138 autistic children and 138 normal controls using reverse-phase HPLC. Pre-column derivatization of amino acids with phenyl isothiocyanate forms phenyl thio-carbamate derivates that have a lamba(max) of 254 nm, enabling their detection using photodiode array. Autistic children showed elevated levels of glutamic acid (120 +/- 89 vs. 83 +/- 35 micromol/L) and asparagine (85 +/- 37 vs. 47 +/- 19 micromol/L); lower levels of phenylalanine (45 +/- 20 vs. 59 +/- 18 micromol/L), tryptophan (24 +/- 11 vs. 41 +/- 16 micromol/L), methionine (22 +/- 9 vs. 28 +/- 9 micromol/L) and histidine (45 +/- 21 vs. 58 +/- 15 micromol/L). A low molar ratio of (tryptophan/large neutral amino acids) x 100 was observed in autism (5.4 vs 9.2), indicating lesser availability of tryptophan for neurotransmitter serotonin synthesis. To conclude, elevated levels of excitatory amino acids (glutamate and asparagine), decreased essential amino acids (phenylalanine, tryptophan and methionine) and decreased precursors of neurotransmitters (tyrosine and tryptophan) are the distinct characteristics of plasma amino acid profile of autistic children. Thus, such metabolic signatures might be useful tools for early diagnosis of autism.

  4. On the reported optical activity of amino acids in the Murchison meteorite

    NASA Technical Reports Server (NTRS)

    Bada, J. L.; Ho, M.-S.; Steinberg, S.; Cronin, J. R.; Kvenvolden, K. A.; Lawless, J. G.; Miller, S. L.; Oro, J.

    1983-01-01

    It is shown that the explanation of terrestrial contamination of the Murchison meteorite is consistent with the analysis of extracts from the meteorite reported by Engel and Nagy (EN) (1982) and is much more probable than their suggestion that the excess of L-enantiomers for several protein amino acids is due to asymmetric synthesis or decomposition. The low abundance of serine and threonine reported by EN may be due to their decomposition during the derivatization procedure, and the absence of methionine, tyrosine, and phenylalanine can be attributed to various causes. The amount of contamination in EN's extracts are estimated from a mass balance of the amino acid enantiomers, and it is found that the amino acids in the HCl could be due entirely to contamination while in the water extract the amount of contamination ranges from about 40 to 97 percent, depending on the amino acid. The argument that contaminants were preferentially extracted by EN's procedure cannot account for the failure to detect methionine, tyrosine, and phenylalanine.

  5. Standardization of formulations for the acute amino acid depletion and loading tests.

    PubMed

    Badawy, Abdulla A-B; Dougherty, Donald M

    2015-04-01

    The acute tryptophan depletion and loading and the acute tyrosine plus phenylalanine depletion tests are powerful tools for studying the roles of cerebral monoamines in behaviour and symptoms related to various disorders. The tests use either amino acid mixtures or proteins. Current amino acid mixtures lack specificity in humans, but not in rodents, because of the faster disposal of branched-chain amino acids (BCAAs) by the latter. The high content of BCAA (30-60%) is responsible for the poor specificity in humans and we recommend, in a 50g dose, a control formulation with a lowered BCAA content (18%) as a common control for the above tests. With protein-based formulations, α-lactalbumin is specific for acute tryptophan loading, whereas gelatine is only partially effective for acute tryptophan depletion. We recommend the use of the whey protein fraction glycomacropeptide as an alternative protein. Its BCAA content is ideal for specificity and the absence of tryptophan, tyrosine and phenylalanine render it suitable as a template for seven formulations (separate and combined depletion or loading and a truly balanced control). We invite the research community to participate in standardization of the depletion and loading methodologies by using our recommended amino acid formulation and developing those based on glycomacropeptide.

  6. Multiple tyrosine metabolites are GPR35 agonists

    PubMed Central

    Deng, Huayun; Hu, Haibei; Fang, Ye

    2012-01-01

    Both kynurenic acid and 2-acyl lysophosphatidic acid have been postulated to be the endogenous agonists of GPR35. However, controversy remains whether alternative endogenous agonists exist. The molecular targets accounted for many nongenomic actions of thyroid hormones are mostly unknown. Here we report the agonist activity of multiple tyrosine metabolites at the GPR35. Tyrosine metabolism intermediates that contain carboxylic acid and/or catechol functional groups were first selected. Whole cell dynamic mass redistribution (DMR) assays enabled by label-free optical biosensor were then used to characterize their agonist activity in native HT-29. Molecular assays including β-arrestin translocation, ERK phosphorylation and receptor internalization confirmed that GPR35 functions as a receptor for 5,6-dihydroxyindole-2-carboxylic acid, 3,3′,5′-triiodothyronine, 3,3′,5-triiodothyronine, gentisate, rosmarinate, and 3-nitrotyrosine. These results suggest that multiple tyrosine metabolites are alternative endogenous ligands of GPR35, and GPR35 may represent a druggable target for treating certain diseases associated with abnormality of tyrosine metabolism. PMID:22523636

  7. Reduction of L-phenylalanine in protein hydrolysates using L-phenylalanine ammonia-lyase from Rhodosporidium toruloides.

    PubMed

    Castañeda, María Teresita; Adachi, Osao; Hours, Roque Alberto

    2015-10-01

    L-Phenylalanine ammonia-lyase (PAL, EC 4.3.1.25) from Rhodosporidium toruloides was utilized to remove L-phenylalanine (L-Phe) from different commercial protein hydrolysates. A casein acid hydrolysate (CAH, L-Phe ~2.28 %) was employed as a model substrate. t-Cinnamic acid resulting from deamination of L-Phe was extracted, analyzed at λ = 290 nm, and used for PAL activity determination. Optimum reaction conditions, optimized using successive Doehlert design, were 35 mg mL(-1) of CAH and 800 mU mL(-1) of PAL, while temperature and pH were 42 °C and 8.7, respectively. Reaction kinetics of PAL with CAH was determined under optimized conditions. Then, removal of L-Phe from CAH was tested. Results showed that more than 92 % of initial L-Phe was eliminated. Similar results were obtained with other protein hydrolysates. These findings demonstrate that PAL is a useful biocatalyst for L-Phe removal from protein hydrolysates, which can be evaluated as potential ingredients in foodstuffs for PKU patients.

  8. Alterations in amino acid concentrations in the plasma and muscle in human subjects during 24 h of simulated adventure racing.

    PubMed

    Borgenvik, Marcus; Nordin, Marie; Mikael Mattsson, C; Enqvist, Jonas K; Blomstrand, Eva; Ekblom, Björn

    2012-10-01

    This investigation was designed to evaluate changes in plasma and muscle levels of free amino acids during an ultra-endurance exercise and following recovery. Nine male ultra-endurance trained athletes participated in a 24-h standardized endurance trial with controlled energy intake. The participants performed 12 sessions of running, kayaking and cycling (4 × each discipline). Blood samples were collected before, during and after exercise, as well as after 28 h of recovery. Muscle biopsies were taken before the test and after exercise, as well as after 28 h of recovery. During the 24-h exercise, plasma levels of branched-chain (BCAA), essential amino acids (EAA) and glutamine fell 13, 14 and 19% (P < 0.05), respectively, whereas their concentrations in muscle were unaltered. Simultaneously, tyrosine and phenylalanine levels rose 38 and 50% (P < 0.05) in the plasma and 66 and 46% (P < 0.05) in muscle, respectively. After the 24-h exercise, plasma levels of BCAA were positively correlated with muscle levels of glycogen (r (2) = 0.73, P < 0.05), as was the combined concentrations of muscle tyrosine and phenylalanine with plasma creatine kinase (R (2) = 0.55, P < 0.05). Following 28-h of recovery, plasma and muscle levels of amino acids had either returned to their initial levels or were elevated. In conclusion, ultra-endurance exercise caused significant changes elevations in plasma and muscle levels of tyrosine and phenylalanine, which suggest an increase in net muscle protein breakdown during exercise. There was a reduction in plasma concentrations of EAA and glutamine during exercise, whereas no changes were detected in their muscle concentration after exercise. PMID:22350359

  9. Influences of Various Amino Acids on Tryptophan-Mediated Control of the Tryptophan Biosynthetic Enzymes in Escherichia coli

    PubMed Central

    Stubbs, John D.; Stubbs, E. Ann

    1971-01-01

    Lysates of Escherichia coli Ymel obtained from cultures grown in the absence of tryptophan in minimal medium supplemented with 0.1% casein hydrolysate show an approximate fivefold increase in steady-state specific activity of both anthranilate synthetase and tryptophan synthetase A protein relative to cultures grown in nonsupplemented medium. In the presence of repressing levels of exogenous tryptophan, growth of cultures in casein hydrolysate-supplemented medium results in a noncoordinate enhancement of repression of 10-fold for anthranilate synthetase and twofold for tryptophan synthetase A protein. Similar, but less pronounced, effects are shown for strain W3110. Strains possessing tryptophan regulator gene mutations do not exhibit this first effect, but do yield an approximate twofold decrease in specific activity of both enzymes when grown in medium supplemented with tryptophan and casein hydrolysate. A stimulation of derepression of both enzymes in strain Ymel equivalent to that induced by casein hydrolysate can be reproduced by growth in minimal medium supplemented with threonine, phenylalanine, tyrosine, serine, glutamic acid, and glutamine. Doubling time in this medium is not significantly different from that in minimal medium. An enhancement of repression which partially mimics that observed on growth in medium supplemented with tryptophan plus casein hydrolysate is obtained when Ymel is grown on medium supplemented with tryptophan plus methionine. Threonine or phenylalanine plus tyrosine as separate medium supplements are independently capable of producing a 1.4-fold or 3.4-fold stimulation, respectively, but in combination only the phenylalanine plus tyrosine effect is manifested unless serine and glutamic acid or glutamine are included. Our data show that expression of the tryptophan biosynthetic enzymes can be significantly influenced in vivo as a result of growth in medium supplemented with a variety of amino acids. PMID:4945190

  10. Dietary amino acids and brain function.

    PubMed

    Fernstrom, J D

    1994-01-01

    Two groups of amino acids--the aromatic and the acidic amino acids--are reputed to influence brain function when their ingestion in food changes the levels of these amino acids in the brain. The aromatic amino acids (tryptophan, tyrosine, phenylalanine) are the biosynthetic precursors for the neurotransmitters serotonin, dopamine, and norepinephrine. Single meals, depending on their protein content, can rapidly influence uptake of aromatic amino acid into the brain and, as a result, directly modify their conversion to neurotransmitters. Such alterations in the production of transmitters can directly modify their release from neurons and, thus, influence brain function. The acidic amino acids glutamate and aspartate are themselves brain neurotransmitters. However, they do not have ready access to the brain from the circulation or the diet. As a result, the ingestion of proteins, which are naturally rich in aspartate and glutamate, has no effect on the level of acidic amino acid in the brain (or, thus, on brain function by this mechanism). Nevertheless, the food additives monosodium glutamate and aspartame (which contains aspartate) have been reputed to raise the level of acidic amino acid in the brain (when ingested in enormous amounts), to modify brain function, and even to cause neuronal damage. Despite such claims, a substantial body of published evidence clearly indicates that the brain is not affected by ingestion of aspartame and is affected by glutamate only when the amino acid is administered alone in extremely large doses. Therefore, when consumed in the diet neither compound presents a risk to normal brain function.

  11. Redox regulation of protein tyrosine phosphatase 1B (PTP1B): Importance of steric and electronic effects on the unusual cyclization of the sulfenic acid intermediate to a sulfenyl amide

    NASA Astrophysics Data System (ADS)

    Sarma, Bani Kanta

    2013-09-01

    The redox regulation of protein tyrosine phosphatase 1B (PTP1B) via the unusual transformation of its sulfenic acid (PTP1B-SOH) to a cyclic sulfenyl amide intermediate is studied by using small molecule chemical models. These studies suggest that the sulfenic acids derived from the H2O2-mediated reactions o-amido thiophenols do not efficiently cyclize to sulfenyl amides and the sulfenic acids produced in situ can be trapped by using methyl iodide. Theoretical calculations suggest that the most stable conformer of such sulfenic acids are stabilized by nO → σ*S-OH orbital interactions, which force the -OH group to adopt a position trans to the S⋯O interaction, leading to an almost linear arrangement of the O⋯S-O moiety and this may be the reason for the slow cyclization of such sulfenic acids to their corresponding sulfenyl amides. On the other hand, additional substituents at the 6-position of o-amido phenylsulfenic acids that can induce steric environment and alter the electronic properties around the sulfenic acid moiety by S⋯N or S⋯O nonbonded interactions destabilize the sulfenic acids by inducing strain in the molecule. This may lead to efficient the cyclization of such sulfenic acids. This model study suggests that the amino acid residues in the close proximity of the sulfenic acid moiety in PTP1B may play an important role in the cyclization of PTP1B-SOH to produce the corresponding sulfenyl amide.

  12. Density functional theory study of conformation-dependent properties of neutral and radical cationic L-tyrosine and L-tryptophan.

    PubMed

    Baek, K Y; Fujimura, Y; Hayashi, M; Lin, S H; Kim, S K

    2011-09-01

    Conformation-dependent properties of L-tyrosine and L-tryptophan in neutral and radical cations were studied by using the density functional theory (DFT) with a new density functional M05-2X. The results are compared with those obtained by using the conventional DFT (B3LYP). Results obtained by both types of DFT were in qualitative accord, including the existence of two conformational subgroups and their subgroup-dependent adiabatic ionization energy and hydrogen bonding. On the other hand, quantitative differences were found between the two DFT methods as well: the M05-2X method successfully reproduced experimental adiabatic ionization energy, whereas the B3LYP functional consistently yielded significantly lower values by 0.2-0.3 eV. More importantly, natural bond orbital (NBO) analysis for cationic conformers showed that all conformers of L-tyrosine and L-tryptophan undergo charge localization upon ionization regardless of the presence of intramolecular hydrogen bonding, unlike the case of L-phenylalanine that was treated earlier by other studies. Different degrees of charge localization among all three aromatic amino acids are explained by employing a simple model in which the aromatic amino acid is assumed to consist of two submoieties of distinct cationic core: the backbone and aromatic side chain. The difference in adiabatic ionization energy between these two submoieties is found to govern the degree of charge localization. PMID:21539381

  13. Euglena mitochondria and chloroplasts form tyrosine-O-sulfate

    SciTech Connect

    Saidha, T.; Hanfstingl, U.; Schiff, J.A. )

    1989-04-01

    Mitochondria from light-grown wild-type Euglena gracilis var. bacillaris Cori or dark-grown mutant W{sub 10}BSmL incubated with {sup 35}SO{sub 4}{sup 2{minus}} and ATP, or with {sup 14}C-tyrosine, non-radioactive sulfate and ATP accumulate a labeled compound in the medium. Since this compound shows exact coelectrophoresis with tyrosine-O-sulfate (TOS) at pH 2.0, 5.8 or 8.0., yields sulfate and tyrosine on acid hydrolysis, and treatment with aryl sulfatase from Aerobacter aerogenes yields sulfate and tyrosine but no tyrosine methyl ester, it is identified as TOS. No TOS is found outside purified developing chloroplasts incubated with {sup 35}SO{sub 4}{sup 2{minus}} and ATP, but both chloroplasts and mitochondria form to {sup 35}S externally when incubated with adenosine 3{prime} phosphate 5{prime}phospho({sup 35}S) sulfate (PAP{sup 35}S). Since no tyrosine need be added, tyrosine is provided from endogenous sources. Although TOS is found in the free pool of Euglena cells it cannot be detected in proteins of cells or mucus ruling our sulfation of tyrosine of protein or incorporation of TOS into proteins. The system forming TOS is membrane-bound and may be involved in tyrosine transport.

  14. The Effect of Copper And Zinc Nanoparticles on the Growth Parameters, Contents of Ascorbic Acid, and Qualitative Composition of Amino Acids and Acylcarnitines in Pistia stratiotes L. (Araceae)

    NASA Astrophysics Data System (ADS)

    Olkhovych, Olga; Volkogon, Mykola; Taran, Nataliya; Batsmanova, Lyudmyla; Kravchenko, Inna

    2016-04-01

    The paper covers the research of copper and zinc nanoparticle effect on the content of ascorbic acid, and quantitative and qualitative composition of amino acids and acylcarnitines in Pistia stratiotes L. plants. Plant exposition to copper nanoparticles led to the decrease in (1) the amount of ascorbic acid, (2) the total content of amino acids (by 25 %), and (3) the amount of all studied amino acids except for the glycine amino acid. At this, the amount of 5-oxoproline, arginine, leucine, ornithine, phenylalanine, proline, serine, and tyrosine was two times lower than in control plants. The reduction of the contents of 8 out of 12 investigated acylcarnitines (namely C0, C2, C3, C5, C6, C8, C16, C18:1) was observed in plants under the influence of copper nanoparticles. The result of plants incubation with zinc nanoparticles was the decrease in (1) the amount of ascorbic acid, (2) the total content of amino acids (by 15 %), (3) the content of leucine, methionine, phenylalanine, proline, and tyrosine (more than twice), and (4) the content of 10 acylcarnitines (C0, C2, C3, C4, C5, C10, C16, C18, C18:1, C18:2). The observed reduction in amino acid contents may negatively affect plants adaptive reactions associated with de novo synthesis of stress proteins. At the same time, the decrease in the content of acylcarnitines, responsible for fatty acid transportation, may lead to the changes in the activity and direction of lipid metabolism in plants and reduce plant's ability to use free fatty acids as the oxidation substrate for cell reparation.

  15. The Effect of Copper And Zinc Nanoparticles on the Growth Parameters, Contents of Ascorbic Acid, and Qualitative Composition of Amino Acids and Acylcarnitines in Pistia stratiotes L. (Araceae).

    PubMed

    Olkhovych, Olga; Volkogon, Mykola; Taran, Nataliya; Batsmanova, Lyudmyla; Kravchenko, Inna

    2016-12-01

    The paper covers the research of copper and zinc nanoparticle effect on the content of ascorbic acid, and quantitative and qualitative composition of amino acids and acylcarnitines in Pistia stratiotes L. plants. Plant exposition to copper nanoparticles led to the decrease in (1) the amount of ascorbic acid, (2) the total content of amino acids (by 25 %), and (3) the amount of all studied amino acids except for the glycine amino acid. At this, the amount of 5-oxoproline, arginine, leucine, ornithine, phenylalanine, proline, serine, and tyrosine was two times lower than in control plants. The reduction of the contents of 8 out of 12 investigated acylcarnitines (namely C0, C2, C3, C5, C6, C8, C16, C18:1) was observed in plants under the influence of copper nanoparticles. The result of plants incubation with zinc nanoparticles was the decrease in (1) the amount of ascorbic acid, (2) the total content of amino acids (by 15 %), (3) the content of leucine, methionine, phenylalanine, proline, and tyrosine (more than twice), and (4) the content of 10 acylcarnitines (C0, C2, C3, C4, C5, C10, C16, C18, C18:1, C18:2). The observed reduction in amino acid contents may negatively affect plants adaptive reactions associated with de novo synthesis of stress proteins. At the same time, the decrease in the content of acylcarnitines, responsible for fatty acid transportation, may lead to the changes in the activity and direction of lipid metabolism in plants and reduce plant's ability to use free fatty acids as the oxidation substrate for cell reparation. PMID:27107771

  16. The Effect of Copper And Zinc Nanoparticles on the Growth Parameters, Contents of Ascorbic Acid, and Qualitative Composition of Amino Acids and Acylcarnitines in Pistia stratiotes L. (Araceae).

    PubMed

    Olkhovych, Olga; Volkogon, Mykola; Taran, Nataliya; Batsmanova, Lyudmyla; Kravchenko, Inna

    2016-12-01

    The paper covers the research of copper and zinc nanoparticle effect on the content of ascorbic acid, and quantitative and qualitative composition of amino acids and acylcarnitines in Pistia stratiotes L. plants. Plant exposition to copper nanoparticles led to the decrease in (1) the amount of ascorbic acid, (2) the total content of amino acids (by 25 %), and (3) the amount of all studied amino acids except for the glycine amino acid. At this, the amount of 5-oxoproline, arginine, leucine, ornithine, phenylalanine, proline, serine, and tyrosine was two times lower than in control plants. The reduction of the contents of 8 out of 12 investigated acylcarnitines (namely C0, C2, C3, C5, C6, C8, C16, C18:1) was observed in plants under the influence of copper nanoparticles. The result of plants incubation with zinc nanoparticles was the decrease in (1) the amount of ascorbic acid, (2) the total content of amino acids (by 15 %), (3) the content of leucine, methionine, phenylalanine, proline, and tyrosine (more than twice), and (4) the content of 10 acylcarnitines (C0, C2, C3, C4, C5, C10, C16, C18, C18:1, C18:2). The observed reduction in amino acid contents may negatively affect plants adaptive reactions associated with de novo synthesis of stress proteins. At the same time, the decrease in the content of acylcarnitines, responsible for fatty acid transportation, may lead to the changes in the activity and direction of lipid metabolism in plants and reduce plant's ability to use free fatty acids as the oxidation substrate for cell reparation.

  17. Biosynthesis and Metabolic Fate of Phenylalanine in Conifers.

    PubMed

    Pascual, María B; El-Azaz, Jorge; de la Torre, Fernando N; Cañas, Rafael A; Avila, Concepción; Cánovas, Francisco M

    2016-01-01

    The amino acid phenylalanine (Phe) is a critical metabolic node that plays an essential role in the interconnection between primary and secondary metabolism in plants. Phe is used as a protein building block but it is also as a precursor for numerous plant compounds that are crucial for plant reproduction, growth, development, and defense against different types of stresses. The metabolism of Phe plays a central role in the channeling of carbon from photosynthesis to the biosynthesis of phenylpropanoids. The study of this metabolic pathway is particularly relevant in trees, which divert large amounts of carbon into the biosynthesis of Phe-derived compounds, particularly lignin, an important constituent of wood. The trunks of trees are metabolic sinks that consume a considerable percentage of carbon and energy from photosynthesis, and carbon is finally immobilized in wood. This paper reviews recent advances in the biosynthesis and metabolic utilization of Phe in conifer trees. Two alternative routes have been identified: the ancient phenylpyruvate pathway that is present in microorganisms, and the arogenate pathway that possibly evolved later during plant evolution. Additionally, an efficient nitrogen recycling mechanism is required to maintain sustained growth during xylem formation. The relevance of phenylalanine metabolic pathways in wood formation, the biotic interactions, and ultraviolet protection is discussed. The genetic manipulation and transcriptional regulation of the pathways are also outlined. PMID:27468292

  18. Characterization of mutations at the mouse phenylalanine hydroxylase locus

    SciTech Connect

    McDonald, J.D.; Charlton, C.K.

    1997-02-01

    Two genetic mouse models for human phenylketonuria have been characterized by DNA sequence analysis. For each, a distinct mutation was identified within the protein coding sequence of the phenylalanine hydroxylase gene. This establishes that the mutated locus is the same as that causing human phenylketonuria and allows a comparison between these mouse phenylketonuria models and the human disease. A genotype/phenotype relationship that is strikingly similar to the human disease emerges, underscoring the similarity of phenylketonuria in mouse and man. In PAH{sup ENU1}, the phenotype is mild. The Pah{sup enu1} mutation predicts a conservative valine to alanine amino acid substitution and is located in exon 3, a gene region where serious mutations are rare in humans. In PAH{sup ENU2} the phenotype is severe. The Pah{sup enu2} mutation predicts a radical phenylalanine to serine substitution and is located in exon 7, a gene region where serious mutations are common in humans. In PAH{sup ENU2}, the sequence information was used to devise a direct genotyping system based on the creation of a new Alw26I restriction endonuclease site. 26 refs., 2 figs., 1 tab.

  19. Biosynthesis and Metabolic Fate of Phenylalanine in Conifers

    PubMed Central

    Pascual, María B.; El-Azaz, Jorge; de la Torre, Fernando N.; Cañas, Rafael A.; Avila, Concepción; Cánovas, Francisco M.

    2016-01-01

    The amino acid phenylalanine (Phe) is a critical metabolic node that plays an essential role in the interconnection between primary and secondary metabolism in plants. Phe is used as a protein building block but it is also as a precursor for numerous plant compounds that are crucial for plant reproduction, growth, development, and defense against different types of stresses. The metabolism of Phe plays a central role in the channeling of carbon from photosynthesis to the biosynthesis of phenylpropanoids. The study of this metabolic pathway is particularly relevant in trees, which divert large amounts of carbon into the biosynthesis of Phe-derived compounds, particularly lignin, an important constituent of wood. The trunks of trees are metabolic sinks that consume a considerable percentage of carbon and energy from photosynthesis, and carbon is finally immobilized in wood. This paper reviews recent advances in the biosynthesis and metabolic utilization of Phe in conifer trees. Two alternative routes have been identified: the ancient phenylpyruvate pathway that is present in microorganisms, and the arogenate pathway that possibly evolved later during plant evolution. Additionally, an efficient nitrogen recycling mechanism is required to maintain sustained growth during xylem formation. The relevance of phenylalanine metabolic pathways in wood formation, the biotic interactions, and ultraviolet protection is discussed. The genetic manipulation and transcriptional regulation of the pathways are also outlined. PMID:27468292

  20. Genetic engineering activates biosynthesis of aromatic fumaric acid amides in the human pathogen Aspergillus fumigatus.

    PubMed

    Kalb, Daniel; Heinekamp, Thorsten; Lackner, Gerald; Scharf, Daniel H; Dahse, Hans-Martin; Brakhage, Axel A; Hoffmeister, Dirk

    2015-03-01

    The Aspergillus fumigatus nonribosomal peptide synthetase FtpA is among the few of this species whose natural product has remained unknown. Both FtpA adenylation domains were characterized in vitro. Fumaric acid was identified as preferred substrate of the first and both l-tyrosine and l-phenylalanine as preferred substrates of the second adenylation domain. Genetically engineered A. fumigatus strains expressed either ftpA or the regulator gene ftpR, encoded in the same cluster of genes, under the control of the doxycycline-inducible tetracycline-induced transcriptional activation (tet-on) cassette. These strains produced fumaryl-l-tyrosine and fumaryl-l-phenylalanine which were identified by liquid chromatography and high-resolution mass spectrometry. Modeling of the first adenylation domain in silico provided insight into the structural requirements to bind fumaric acid as peptide synthetase substrate. This work adds aromatic fumaric acid amides to the secondary metabolome of the important human pathogen A. fumigatus which was previously not known as a producer of these compounds.

  1. Genetic Engineering Activates Biosynthesis of Aromatic Fumaric Acid Amides in the Human Pathogen Aspergillus fumigatus

    PubMed Central

    Kalb, Daniel; Heinekamp, Thorsten; Lackner, Gerald; Scharf, Daniel H.; Dahse, Hans-Martin; Brakhage, Axel A.

    2014-01-01

    The Aspergillus fumigatus nonribosomal peptide synthetase FtpA is among the few of this species whose natural product has remained unknown. Both FtpA adenylation domains were characterized in vitro. Fumaric acid was identified as preferred substrate of the first and both l-tyrosine and l-phenylalanine as preferred substrates of the second adenylation domain. Genetically engineered A. fumigatus strains expressed either ftpA or the regulator gene ftpR, encoded in the same cluster of genes, under the control of the doxycycline-inducible tetracycline-induced transcriptional activation (tet-on) cassette. These strains produced fumaryl-l-tyrosine and fumaryl-l-phenylalanine which were identified by liquid chromatography and high-resolution mass spectrometry. Modeling of the first adenylation domain in silico provided insight into the structural requirements to bind fumaric acid as peptide synthetase substrate. This work adds aromatic fumaric acid amides to the secondary metabolome of the important human pathogen A. fumigatus which was previously not known as a producer of these compounds. PMID:25527545

  2. Oral aspartame and plasma phenylalanine: pharmacokinetic difference between rodents and man, and relevance to CNS effects of phenylalanine. Short note.

    PubMed

    Fernstrom, J D

    1989-01-01

    The ingestion of aspartame, a phenylalanine-containing dipeptide, raises plasma phenylalanine levels. These increments are much greater in humans than rats, because the rat hydroxylates phenylalanine five times faster than man. Accordingly, dose comparisons of aspartame (or phenylalanine) between humans and rats have usually been corrected by a factor of five. Recently, a correction factor of sixty has been proposed (Wurtman and Maher, 1987); the rationale is based on a novel calculation of competitive phenylalanine transport into brain. An analysis of the logic behind this postulation reveals there to be no basis for accepting the higher dose conversion of 60 between rat and man.

  3. Nuclear magnetic resonance studies of phenylalanine analog interactions with normal and sicklen hemoglobin

    SciTech Connect

    Lee, Y.H.

    1985-01-01

    Several phenylalanine derivatives have been found to inhibit the gelation of deoxygenated sickle hemoglobin (deoxy HbS). Proton and /sup 19/F-NMR techniques were used to monitor the interaction of selected phenylalanine derivatives with the Hb molecule by using fluorine containing phenylalanine derivatives, Hb labeled at the ..beta..93 position with N-(2,2,2-trifluoroethyl) iodoacetamide (IA-F/sub 3/), and by monitoring the relaxation rates of the C2 and C4 histidine protons. The results show that the /sup 19/F spin-spin relaxation times of L-phenylalanin-4-fluorobenzylamide (PheNBz1-F), which has a deoxy HbS antigelling activity comparable to that of the amino acid, tryptophan, are affected much more strongly by interaction with Hb than are those of glycin-4-fluorobenzylamide (GlyNBz1-F). In contrast, it is shown that N-(2,2,5,5-tetramethylpyrrolidin-1-oxy-3-carboxyl)-L-phenylalanine t-butyl ester (SL-Phe) exhibits specific binding to Hb, and an antigelling activity more than two orders of magnitude greater than that of phenylalanine. These results indicate that the fluorine nuclei strongly influenced by the presence of spin label nitroxide are located in a conformation within a few angstroms of the SL-Phe binding site. Proton NMR relaxation measurements of the C2 and C4 proton resonances from the ..beta..2, 4b143 and ..beta..146 histidine residues show significant and selective effects from the binding of SL-Phe to Hb, indicating that the SL-Phe binding site must be close to the side chains of these three residues. The strong antigelation activity of SL-Phe suggests that this binding site may be one of the intermolecular contact sites of importance to the deoxy HbS aggregation process.

  4. Beta-lactamase-catalyzed aminolysis of depsipeptides: Proof of the nonexistence of a specific D-phenylalanine/enzyme complex by double-label isotope trapping

    SciTech Connect

    Pazhanisamy, S.; Pratt, R.F. )

    1989-08-22

    The steady-state kinetics of the Enterobacter cloacae P99 beta-lactamase-catalyzed aminolysis of the depsipeptide m-(((phenylacetyl)glycyl)oxy)benzoic acid by D-phenylalanine were consistent with an ordered sequential mechanism with D-phenylalanine binding first. In terms of this mechanism, the kinetics data required that in 20 mM MOPS buffer, pH 7.5, the dissociation constant of the initially formed enzyme/D-phenylalanine complex be around 1.3 mM; at pH 9.0 in 0.1 M carbonate buffer, the complex should be somewhat more stable. Attempts to detect this complex in a binary mixture by spectroscopic methods (fluorescence, circular dichroic, and nuclear magnetic resonance spectra) failed. Kinetic methods were also unsuccessful--the presence of 20 mM D-phenylalanine did not appear to affect beta-lactamase activity nor inhibition of the enzyme by phenylmethanesulfonyl fluoride, phenylboronic acid, or (3-dansylamidophenyl)boronic acid. Equilibrium dialysis experiments appeared to indicate that the dissociation constant of any binary enzyme/D-phenylalanine complex must be somewhat higher than the kinetics allowed (greater than 2 mM). Since the kinetics also required that, at high depsipeptide concentrations, and again with the assumption of the ordered sequential mechanism, the reaction of the enzyme/D-phenylalanine complex to aminolysis products be faster than its reversion to enzyme and D-phenylalanine, a double-label isotope-trapping experiment was performed.

  5. Disruption of phenylalanine hydroxylase reduces adult lifespan and fecundity, and impairs embryonic development in parthenogenetic pea aphids

    PubMed Central

    Simonet, Pierre; Gaget, Karen; Parisot, Nicolas; Duport, Gabrielle; Rey, Marjolaine; Febvay, Gérard; Charles, Hubert; Callaerts, Patrick; Colella, Stefano; Calevro, Federica

    2016-01-01

    Phenylalanine hydroxylase (PAH) is a key tyrosine-biosynthetic enzyme involved in neurological and melanin-associated physiological processes. Despite extensive investigations in holometabolous insects, a PAH contribution to insect embryonic development has never been demonstrated. Here, we have characterized, for the first time, the PAH gene in a hemimetabolous insect, the aphid Acyrthosiphon pisum. Phylogenetic and sequence analyses confirmed that ApPAH is closely related to metazoan PAH, exhibiting the typical ACT regulatory and catalytic domains. Temporal expression patterns suggest that ApPAH has an important role in aphid developmental physiology, its mRNA levels peaking at the end of embryonic development. We used parental dsApPAH treatment to generate successful knockdown in aphid embryos and to study its developmental role. ApPAH inactivation shortens the adult aphid lifespan and considerably affects fecundity by diminishing the number of nymphs laid and impairing embryonic development, with newborn nymphs exhibiting severe morphological defects. Using single nymph HPLC analyses, we demonstrated a significant tyrosine deficiency and a consistent accumulation of the upstream tyrosine precursor, phenylalanine, in defective nymphs, thus confirming the RNAi-mediated disruption of PAH activity. This study provides first insights into the role of PAH in hemimetabolous insects and demonstrates that this metabolic gene is essential for insect embryonic development. PMID:27694983

  6. Selective separation of sodium ions from a mixture with phenylalanine by Donnan dialysis with a profiled sulfogroup cation exchange membrane

    NASA Astrophysics Data System (ADS)

    Vasil'eva, V. I.; Goleva, E. A.

    2013-11-01

    The possibility of separating ions of metal from a mixture with ampholyte (an amino acid) by Donnan dialysis with an MK-40 sulfogroup cation exchange membrane is demonstrated. Conditions ensuring the selectivity and intensity of the mass transfer of sodium ions from a mixture with bipolar phenylalanine ions into a diffusate containing hydrochloric acid through a cation exchange membrane are found.

  7. Mutagenic effect by phenylalanine during gamma-irradiation of plasmid DNA in aqueous solution under oxic conditions.

    PubMed

    Reitsma-Wijker, C A; Slotman, B J; Lafleur, M V

    2000-11-01

    Irradiation of DNA in aqueous solution or in cells with gamma-rays results in different mutational spectra, indicating that in both situations different patterns of DNA damages are induced. One of the causes for these different types of damages might be the formation of secondary, organic radicals, if cells are irradiated. Some organic compounds, including the amino acid phenylalanine, are well known to produce radicals during irradiation. Under oxic conditions these secondary radicals react with oxygen, thus forming peroxyl radicals which can be very harmful to DNA, and which may, therefore, induce DNA damage leading to mutations. This study examines the influence of the presence of phenylalanine during gamma-irradiation of DNA in aqueous solution under oxic conditions. The results indicate that the formation of phenylalanine radicals influences the types of induced mutations in the gamma-radiation-induced mutation spectrum. The most prominent difference is the increase in G:C to T:A transversions and the decrease in G:C to A:T transitions in the presence of phenylalanine. Further, it appears that the gamma-radiation-induced mutation spectrum after irradiation of DNA in aqueous solution is more comparable to the intracellular gamma-radiation-induced mutation spectrum in E. coli cells, if phenylalanine is present during irradiation. Therefore, these results suggest that the presence of phenylalanine during irradiation of DNA in aqueous solution gives a better impression of gamma-radiation-induced mutations in bacterial systems than water only. PMID:11035161

  8. Improving the Production of L-Phenylalanine by Identifying Key Enzymes Through Multi-Enzyme Reaction System in Vitro.

    PubMed

    Ding, Dongqin; Liu, Yongfei; Xu, Yiran; Zheng, Ping; Li, Haixing; Zhang, Dawei; Sun, Jibin

    2016-08-25

    L-Phenylalanine (L-Phe) is an important amino acid used in both food and medicinal applications. We developed an in vitro system that allowed a direct, quantitative investigation of phenylalanine biosynthesis in E. coli. Here, the absolute concentrations of six enzymes (AroK, AroL, AroA, AroC, PheA and TyrB) involved in the shikimate (SHIK) pathway were determined by a quantitative proteomics approach and in vitro enzyme titration experiments. The reconstitution of an in vitro reaction system for these six enzymes was established and their effects on the phenylalanine production were tested. The results showed that the yield of phenylalanine increased 3.0 and 2.1 times when the concentrations of shikimate kinase (AroL) and 5-enolpyruvoyl shikimate 3-phosphate (EPSP) synthase (AroA) were increased 2.5 times. Consistent results were obtained from in vivo via the overexpression of AroA in a phenylalanine-producing strain, and the titer of phenylalanine reached 62.47 g/l after 48 h cultivation in a 5-liter jar fermentor. Our quantitative findings provide a practical method to detect the potential bottleneck in a specific metabolic pathway to determine which gene products should be targeted to improve the yield of the desired product.

  9. Improving the Production of L-Phenylalanine by Identifying Key Enzymes Through Multi-Enzyme Reaction System in Vitro.

    PubMed

    Ding, Dongqin; Liu, Yongfei; Xu, Yiran; Zheng, Ping; Li, Haixing; Zhang, Dawei; Sun, Jibin

    2016-01-01

    L-Phenylalanine (L-Phe) is an important amino acid used in both food and medicinal applications. We developed an in vitro system that allowed a direct, quantitative investigation of phenylalanine biosynthesis in E. coli. Here, the absolute concentrations of six enzymes (AroK, AroL, AroA, AroC, PheA and TyrB) involved in the shikimate (SHIK) pathway were determined by a quantitative proteomics approach and in vitro enzyme titration experiments. The reconstitution of an in vitro reaction system for these six enzymes was established and their effects on the phenylalanine production were tested. The results showed that the yield of phenylalanine increased 3.0 and 2.1 times when the concentrations of shikimate kinase (AroL) and 5-enolpyruvoyl shikimate 3-phosphate (EPSP) synthase (AroA) were increased 2.5 times. Consistent results were obtained from in vivo via the overexpression of AroA in a phenylalanine-producing strain, and the titer of phenylalanine reached 62.47 g/l after 48 h cultivation in a 5-liter jar fermentor. Our quantitative findings provide a practical method to detect the potential bottleneck in a specific metabolic pathway to determine which gene products should be targeted to improve the yield of the desired product. PMID:27558633

  10. Improving the Production of L-Phenylalanine by Identifying Key Enzymes Through Multi-Enzyme Reaction System in Vitro

    PubMed Central

    Ding, Dongqin; Liu, Yongfei; Xu, Yiran; Zheng, Ping; Li, Haixing; Zhang, Dawei; Sun, Jibin

    2016-01-01

    L-Phenylalanine (L-Phe) is an important amino acid used in both food and medicinal applications. We developed an in vitro system that allowed a direct, quantitative investigation of phenylalanine biosynthesis in E. coli. Here, the absolute concentrations of six enzymes (AroK, AroL, AroA, AroC, PheA and TyrB) involved in the shikimate (SHIK) pathway were determined by a quantitative proteomics approach and in vitro enzyme titration experiments. The reconstitution of an in vitro reaction system for these six enzymes was established and their effects on the phenylalanine production were tested. The results showed that the yield of phenylalanine increased 3.0 and 2.1 times when the concentrations of shikimate kinase (AroL) and 5-enolpyruvoyl shikimate 3-phosphate (EPSP) synthase (AroA) were increased 2.5 times. Consistent results were obtained from in vivo via the overexpression of AroA in a phenylalanine-producing strain, and the titer of phenylalanine reached 62.47 g/l after 48 h cultivation in a 5-liter jar fermentor. Our quantitative findings provide a practical method to detect the potential bottleneck in a specific metabolic pathway to determine which gene products should be targeted to improve the yield of the desired product. PMID:27558633

  11. The M1 family of vertebrate aminopeptidases: role of evolutionarily conserved tyrosines in the enzymatic mechanism of aminopeptidase B.

    PubMed

    Cadel, Sandrine; Darmon, Cécile; Pernier, Julien; Hervé, Guy; Foulon, Thierry

    2015-02-01

    Aminopeptidase B (Ap-B), a member of the M1 family of Zn(2+)-aminopeptidases, removes basic residues at the NH2-terminus of peptides and is involved in the in vivo proteolytic processing of miniglucagon and cholecystokinin-8. M1 enzymes hydrolyze numerous different peptides and are implicated in many physiological functions. As these enzymes have similar catalytic mechanisms, their respective substrate specificity and/or catalytic efficiency must be based on subtle structural differences at or near the catalytic site. This leads to the hypothesis that each primary structure contains a consensus structural template, strictly necessary for aminopeptidase activity, and a specific amino acid environment localized in or outside the catalytic pocket that finely tunes the substrate specificity and catalytic efficiency of each enzyme. A multiple sequence alignment of M1 peptidases from vertebrates allowed to identify conserved tyrosine amino acids, which are members of this catalytic backbone. In the present work, site-directed mutagenesis and 3D molecular modeling of Ap-B were used to specify the role of four fully (Y281, Y229, Y414, and Y441) and one partially (Y409) conserved residues. Tyrosine to phenylalanine mutations allowed confirming the influence of the hydroxyl groups on the enzyme activity. These groups are implicated in the reaction mechanism (Y414), in substrate specificity and/or catalytic efficiency (Y409), in stabilization of essential amino acids of the active site (Y229, Y409) and potentially in the maintenance of its structural integrity (Y281, Y441). The importance of hydrogen bonds is verified by the Y229H substitution, which preserves the enzyme activity. These data provide new insights into the catalytic mechanism of Ap-B in the M1 family of aminopeptidases.

  12. The extraction of aromatic amino acids with binary and ternary mixtures of hydrophilic solvents

    NASA Astrophysics Data System (ADS)

    Mokshina, N. Ya.; Pakhomova, O. A.; Niftaliev, S. I.

    2007-11-01

    The extraction of tyrosine and phenylalanine with binary and ternary mixtures of hydrophilic solvents from aqueous salt solutions was studied, and several tendencies were observed. Simplex-lattice planning of experiment was used for the optimization of the composition of solvent mixtures. It was shown that the extraction systems developed could be employed for the almost complete extraction of tyrosine and phenylalanine from aqueous solutions.

  13. Biochemical properties and crystal structure of a β-phenylalanine aminotransferase from Variovorax paradoxus.

    PubMed

    Crismaru, Ciprian G; Wybenga, Gjalt G; Szymanski, Wiktor; Wijma, Hein J; Wu, Bian; Bartsch, Sebastian; de Wildeman, Stefaan; Poelarends, Gerrit J; Feringa, Ben L; Dijkstra, Bauke W; Janssen, Dick B

    2013-01-01

    By selective enrichment, we isolated a bacterium that can use β-phenylalanine as a sole nitrogen source. It was identified by 16S rRNA gene sequencing as a strain of Variovorax paradoxus. Enzyme assays revealed an aminotransferase activity. Partial genome sequencing and screening of a cosmid DNA library resulted in the identification of a 1,302-bp aminotransferase gene, which encodes a 46,416-Da protein. The gene was cloned and overexpressed in Escherichia coli. The recombinant enzyme was purified and showed a specific activity of 17.5 U mg(-1) for (S)-β-phenylalanine at 30°C and 33 U mg(-1) at the optimum temperature of 55°C. The β-specific aminotransferase exhibits a broad substrate range, accepting ortho-, meta-, and para-substituted β-phenylalanine derivatives as amino donors and 2-oxoglutarate and pyruvate as amino acceptors. The enzyme is highly enantioselective toward (S)-β-phenylalanine (enantioselectivity [E], >100) and derivatives thereof with different substituents on the phenyl ring, allowing the kinetic resolution of various racemic β-amino acids to yield (R)-β-amino acids with >95% enantiomeric excess (ee). The crystal structures of the holoenzyme and of the enzyme in complex with the inhibitor 2-aminooxyacetate revealed structural similarity to the β-phenylalanine aminotransferase from Mesorhizobium sp. strain LUK. The crystal structure was used to rationalize the stereo- and regioselectivity of V. paradoxus aminotransferase and to define a sequence motif with which new aromatic β-amino acid-converting aminotransferases may be identified.

  14. Adsorption dynamics of L-glutamic acid copolymers at a heptane/water interface.

    PubMed

    Beverung, C J; Radke, C J; Blanch, H W

    1998-02-16

    Random copolymers of glutamic acid (glu-ala, glu-leu, glu-phe, glu-tyr) were employed to investigate the relationship between side chain structure and peptide charge on adsorption behavior at an oil/water boundary. Adsorption of a series of glutamate copolymers at a heptane/water interface was examined by the dynamic pendant-drop method to determine interfacial tension. Incorporation of leucine or phenylalanine into a glutamate copolymer results in greater tension reduction than incorporation of alanine or tyrosine. These effects are amplified at pH values near the isoelectric point of glutamate, where macroscopic adsorbed films of glu-leu and glu-phe exhibit gel-like properties in response to interfacial area compression. Differences in interfacial tension behavior of glu-tyr and glu-phe indicate the importance of the tyrosine p-hydroxyl group on adsorption and aggregation at the oil/water interface. PMID:9540205

  15. Novel post-translational incorporation of tyrosine in PMA-activated polymorphonuclear leukocytes (PMN)

    SciTech Connect

    Nath, J.; Oliver, C.; Ohno, Y.; Gallin, J.I.

    1986-03-05

    During studies undertaken to determine whether stimulation of tubulin tyrosinolation occurs in PMA-activated PMN, a distinctly different and novel post-translational incorporation of tyrosine into multiple PMN proteins was observed. The reaction also occurred in organelle-depleted neutrophil cytoplasts and was highly exaggerated in organelle-enriched karyogranuloplasts. The incorporation was specific for tyrosine, did not require extracellular Ca/sup 2 +/ and was inhibited in the presence of a variety of reducing agents, intracellular scavengers of oxygen radicals and inhibitors of peroxidase-mediated reactions. The PMA-induced incorporation of tyrosine was completely absent in PMN from patients with chronic granulomatous disease, but occurred normally in PMN of a patient with myeloperoxidase deficiency. Moreover, the incorporation of tyrosine was blocked by N-acetyl-L-tyrosine but not by phenylalanine suggesting a requirement for the phenolic group. A two-fold increase in stable protein carbonyl derivatives was demonstrated suggesting an increased oxidative modification of the proteins. SDS urea PAGE and reversed phase HPLC did not reveal any detectable changes in the extent of protein cross-linking. The PMN tyrosine pool was approximately 900 ..mu..M and yet only 1 ..mu..M tyrosine was added in these experiments. The functional significance of this reaction is not yet clear.

  16. Electronic structure of aromatic amino acids studied by soft x-ray spectroscopy

    NASA Astrophysics Data System (ADS)

    Zhang, Wenhua; Carravetta, Vincenzo; Plekan, Oksana; Feyer, Vitaliy; Richter, Robert; Coreno, Marcello; Prince, Kevin C.

    2009-07-01

    The electronic structure of phenylalanine, tyrosine, tryptophan, and 3-methylindole in the gas phase was investigated by x-ray photoemission spectroscopy (XPS) and near edge x-ray absorption fine structure (NEXAFS) spectroscopy at the C, N, and O K-edges. The XPS spectra have been calculated for the four principal conformers of each amino acid, and the spectra weighted by the Boltzmann population ratios calculated from published free energies. Instead of the single peaks expected from the stoichiometry of the compounds, the N 1s core level spectra of phenylalanine and tryptophan show features indicating that more than one conformer is present. The calculations reproduce the experimental features. The C and O 1s spectra do not show evident effects due to conformational isomerism. The calculations predict that such effects are small for carbon, and for oxygen it appears that only broadening occurs. The carbon K-edge NEXAFS spectra of these aromatic amino acids are similar to the published data of the corresponding molecules in the solid state, but show more structure due to the higher resolution in the present study. The N K-edge spectra of tryptophan and 3-methylindole differ from phenylalanine and tyrosine, as the first two both contain a nitrogen atom located in a pyrrole ring. The nitrogen K-edge NEXAFS spectra of aromatic amino acids do not show any measurable effects due to conformational isomerism, in contrast to the photoemission results. Calculations support this result and show that variations of the vertical excitation energies of different conformers are small, and cannot be resolved in the present experiment. The O NEXAFS spectra of these three aromatic compounds are very similar to other, simpler amino acids, which have been studied previously.

  17. Tyrosine promotes oxidative stress in cerebral cortex of young rats.

    PubMed

    Sgaravatti, Angela M; Vargas, Bethânia A; Zandoná, Bernardo R; Deckmann, Kátia B; Rockenbach, Francieli J; Moraes, Tarsila B; Monserrat, José M; Sgarbi, Mirian B; Pederzolli, Carolina D; Wyse, Angela T S; Wannmacher, Clóvis M D; Wajner, Moacir; Dutra-Filho, Carlos Severo

    2008-10-01

    Tyrosine accumulates in inborn errors of tyrosine catabolism, especially in tyrosinemia type II, where tyrosine levels are highly elevated in tissues and physiological fluids of affected patients. In tyrosinemia type II, high levels of tyrosine are correlated with eyes, skin and central nervous system disturbances. Considering that the mechanisms of brain damage in these disorders are poorly known, in the present study, we investigated whether oxidative stress is elicited by l-tyrosine in cerebral cortex homogenates of 14-day-old Wistar rats. The in vitro effect of 0.1-4.0mM l-tyrosine was studied on the following oxidative stress parameters: total radical-trapping antioxidant potential (TRAP), total antioxidant reactivity (TAR), ascorbic acid content, reduced glutathione (GSH) content, spontaneous chemiluminescence, thiobarbituric acid-reactive substances (TBA-RS), thiol-disulfide redox state (SH/SS ratio), protein carbonyl content, formation of DNA-protein cross-links, and the activities of the enzymes superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), and glucose-6-phosphate dehydrogenase (G6PDH). TRAP, TAR, ascorbic acid content, SH/SS ratio and CAT activity were significantly diminished, while formation of DNA-protein cross-link was significantly enhanced by l-tyrosine in vitro. In contrast, l-tyrosine did not affect the other parameters of oxidative stress evaluated. These results indicate that l-tyrosine decreases enzymatic and non-enzymatic antioxidant defenses, changes the redox state and stimulates DNA damage in cerebral cortex of young rats in vitro. This suggests that oxidative stress may represent a pathophysiological mechanism in tyrosinemic patients, in which this amino acid accumulates.

  18. Charge Migration in Phenylalanine Initiated by Attosecond Pulses

    NASA Astrophysics Data System (ADS)

    Greenwood, Jason; Trabattoni, Andrea; Ayuso, David; Belshaw, Louise; de Camillis, Simone; Anumula, Sunil; Frassetto, Fabio; Poletto, Luca; Palacios, Alicia; Declava, Piero; Martin, Fernando; Calegari, Francesca; Nisoli, Mauro

    2015-05-01

    In the past few years attosecond techniques have been implemented for the investigation of ultrafast dynamics in molecules. The generation of isolated attosecond pulses characterized by a relatively high photon flux has opened up new possibilities in the study of molecular dynamics. We report on experimental and theoretical results of ultrafast charge dynamics in a biochemically relevant molecule, namely, the amino acid phenylalanine. The data represent the first experimental demonstration of the generation and observation of a charge migration process in a complex molecule, where electron dynamics precede nuclear motion. The application of attosecond technology to the investigation of electron dynamics in biologically relevant molecules represents a multidisciplinary work, which can open new research frontiers: those in which few femtosecond and even sub-femtosecond electron processes determine the fate of biomolecules.

  19. Phenylalanine-based polyarylacetylenes as enantiomer-differentiating alignment media.

    PubMed

    Krupp, Alexis; Reggelin, Michael

    2012-12-01

    Lyotropic liquid crystalline phases of a phenylalanine-based polyacetylene are introduced as new enantiodifferentiating alignment media. Based on the unusual temperature dependence of the quadrupolar splitting of the (2)H-signal of the solvent (CDCl(3)), three distinct states of the phase with different orientational properties can be identified. This offers the opportunity to measure multiple alignment data sets without changing the sample. Unexpectedly, the largest difference in the orientation of the enantiomers of isopinocampheol was found in the high temperature domain of the phase. This is even more astonishing because the helical structure of the polymer backbone seems to break down at temperatures above approximately 10 °C, leaving the stereogenic centers of the amino acid moieties in the repeating units as the only reason for the enantiodifferentiation.

  20. 3,4-Dihydroxy-l-phenylalanine as a biomarker of oxidative damage in proteins: improved detection using cloud-point extraction and HPLC.

    PubMed

    McPherson, Peter A C; Türemen, Bryn T

    2014-09-26

    Oxidized protein adducts are formed under conditions of oxidative stress and may represent a valuable biomarker for a variety of diseases which share this common aetiology. A suitable candidate biomarker for oxidized proteins is protein-bound 3,4-dihydroxyl-l-phenylalanine (l-DOPA), which is formed on 3'-hydroxylation of tyrosine residues by hydroxyl radicals. Existing methodologies to measure protein-bound l-DOPA employ lengthy acid hydrolysis steps (ca. 16h) which may cause artifactual protein oxidation, followed by HPLC with detection based on the intrinsic fluorescence of l-DOPA. We report a novel method for the measurement of protein-bound l-DOPA which involves rapid hydrolysis followed by pre-column concentration of 6-aminoquinolyl-derivatives using cloud-point extraction. The derivatized material is resolved by reversed-phase HPLC in less than 30min and has derivatization chemistry compatible with both UV and fluorescent detection, providing detection down to the femtomole level. The method provides identical results to those found with highly specific ELISA-based techniques and requires only basic instrumentation. The stability of the 6-aminoquinolyl-derivatives together with the fast and sensitive nature of the assay will be appealing to those who require large sample throughput.

  1. A new approach for quantitative analysis of L-phenylalanine using a novel semi-sandwich immunometric assay.

    PubMed

    Kubota, Kazuyuki; Mizukoshi, Toshimi; Miyano, Hiroshi

    2013-10-01

    Here, we describe a novel method for L-phenylalanine analysis using a sandwich-type immunometric assay approach for use as a new method for amino acid analysis. To overcome difficulties of the preparation of high-affinity and selectivity monoclonal antibodies against L-phenylalanine and the inability to use sandwich-type immunometric assays due to their small molecular weight, three procedures were examined. First, amino groups of L-phenylalanine were modified by "N-Fmoc-L-cysteine" (FC) residues and the derivative (FC-Phe) was used as a hapten. Immunization of mice with bovine serum albumin/FC-Phe conjugate successfully yielded specific monoclonal anti-FC-Phe antibodies. Second, a new derivatization reagent, "biotin linker conjugate of FC-Phe N-succinimidyl ester" (FC(Biotin)-NHS), was synthesized to convert L-phenylalanine to FC-(Biotin)-Phe as a hapten structure. The biotin moiety linked to the thiol group of cysteine formed a second binding site for streptavidin/horseradish peroxidase (HRP) conjugates for optical detection. Third, a new semi-sandwich-type immunometric assay was established using pre-derivatized L-phenylalanine, the monoclonal anti-FC-Phe antibody, and streptavidin/HRP conjugate (without second antibody). Using the new "semi-sandwich" immunometric assay system, a detection limit of 35 nM (60 amol per analysis) and a detection range of 0.1-20 μM were attained using a standard L-phenylalanine solution. Rat plasma samples were analyzed to test reliability. Intra-day assay precision was within 6% of the coefficient of variation; inter-day variation was 0.1%. The recovery rates were from 92.4 to 123.7%. This is the first report of the quantitative determination of L-phenylalanine using a reliable semi-sandwich immunometric assay approach and will be applicable to the quantitative determination of other amino acids.

  2. 3, 4-dihydroxy-L-phenylalanine-derived melanin from Yarrowia lipolytica mediates the synthesis of silver and gold nanostructures

    PubMed Central

    2013-01-01

    Background Nanobiotechnology applies the capabilities of biological systems in generating a variety of nano-sized structures. Plants, algae, fungi and bacteria are some systems mediating such reactions. In fungi, the synthesis of melanin is an important strategy for cell-survival under metal-stressed conditions. Yarrowia lipolytica, the biotechnologically significant yeast also produces melanin that sequesters heavy metal ions. The content of this cell-associated melanin is often low and precursors such as L-tyrosine or 3, 4-dihydroxy-L-phenylalanine (L-DOPA) can enhance its production. The induced melanin has not been exploited for the synthesis of nanostructures. In this investigation, we have employed L-DOPA-melanin for the facile synthesis of silver and gold nanostructures. The former have been used for the development of anti-fungal paints. Methods Yarrowia lipolytica NCIM 3590 cells were incubated with L-DOPA for 18 h and the resultant dark pigment was subjected to physical and chemical analysis. This biopolymer was used as a reducing and stabilizing agent for the synthesis of silver and gold nanostructures. These nanoparticles were characterized by UV-Visible spectra, X-ray diffraction (XRD) studies, and electron microscopy. Silver nanoparticles were evaluated for anti-fungal activity. Results The pigment isolated from Y. lipolytica was identified as melanin. The induced pigment reduced silver nitrate and chloroauric acid to silver and gold nanostructures, respectively. The silver nanoparticles were smaller in size (7 nm) and displayed excellent anti-fungal properties towards an Aspergillus sp. isolated from a wall surface. An application of these nanoparticles as effective paint-additives has been demonstrated. Conclusion The yeast mediated enhanced production of the metal-ion-reducing pigment, melanin. A simple and rapid method for the extracellular synthesis of nanoparticles with paint-additive-application was developed. PMID:23363424

  3. Poly-dopamine-beta-cyclodextrin: A novel nanobiopolymer towards sensing of some amino acids at physiological pH.

    PubMed

    Hasanzadeh, Mohammad; Sadeghi, Sattar; Bageri, Leyla; Mokhtarzadeh, Ahad; Karimzadeh, Ayub; Shadjou, Nasrin; Mahboob, Soltanali

    2016-12-01

    A novel nanobiopolymer film was electrodeposited on the surface of glassy carbon through cyclic voltammetry from dopamine, β-cyclodextrin, and phosphate buffer solution in physiological pH (7.40). The electrochemical behavior of polydopamine-Beta-cyclodextrin modified glassy carbon electrode was investigated for electro-oxidation and determination of some amino acids (l-Cysteine, l-Tyrosine, l-Glycine, and l-Phenylalanine). The modified electrode was applied for selected amino acid detection at physiological pH using cyclic voltammetry, differential pulse voltammetry and chronoamperometry, chronocoulometery. The linear concentration range of the proposed sensor for the l-Glycine, l-Cysteine, l-Tyrosine, and l-Phenylalanine were 0.2-70, 0.06-0.2, 0.01-0.1, and 0.2-10μM, while low limit of quantifications were 0.2, 0.06, 0.01, and 0.2μM, respectively. The modified electrode shows many advantages as an amino acid sensor such as simple preparation method without using any specific electron transfer mediator or specific reagent, good sensitivity, short response time, and long term stability. PMID:27612722

  4. Poly-dopamine-beta-cyclodextrin: A novel nanobiopolymer towards sensing of some amino acids at physiological pH.

    PubMed

    Hasanzadeh, Mohammad; Sadeghi, Sattar; Bageri, Leyla; Mokhtarzadeh, Ahad; Karimzadeh, Ayub; Shadjou, Nasrin; Mahboob, Soltanali

    2016-12-01

    A novel nanobiopolymer film was electrodeposited on the surface of glassy carbon through cyclic voltammetry from dopamine, β-cyclodextrin, and phosphate buffer solution in physiological pH (7.40). The electrochemical behavior of polydopamine-Beta-cyclodextrin modified glassy carbon electrode was investigated for electro-oxidation and determination of some amino acids (l-Cysteine, l-Tyrosine, l-Glycine, and l-Phenylalanine). The modified electrode was applied for selected amino acid detection at physiological pH using cyclic voltammetry, differential pulse voltammetry and chronoamperometry, chronocoulometery. The linear concentration range of the proposed sensor for the l-Glycine, l-Cysteine, l-Tyrosine, and l-Phenylalanine were 0.2-70, 0.06-0.2, 0.01-0.1, and 0.2-10μM, while low limit of quantifications were 0.2, 0.06, 0.01, and 0.2μM, respectively. The modified electrode shows many advantages as an amino acid sensor such as simple preparation method without using any specific electron transfer mediator or specific reagent, good sensitivity, short response time, and long term stability.

  5. Metabolism of aspartame and its L-phenylalanine methyl ester decomposition product by the porcine gut.

    PubMed

    Burgert, S L; Andersen, D W; Stegink, L D; Takeuchi, H; Schedl, H P

    1991-06-01

    The intestinal metabolism of aspartame (N-L-alpha-aspartyl-L-phenylalanine methyl ester; APM) and its L-phenylalanine methyl ester (PME) decomposition product was evaluated in six young pigs. Equimolar doses (2.5 mmol/kg body weight) of APM, PME, and L-phenylalanine (PHE) administered to the proximal jejunum produced similar increases in portal blood PHE concentrations. Methanol, nondetectable in portal blood after PHE ingestion, increased markedly after administration of either APM or PME. Portal blood aspartate concentrations were unchanged after PME and PHE administration, but increased significantly after APM administration. Increases in portal blood PHE concentrations were significantly greater than were increases in aspartate concentrations following APM administration. Neither APM, PME, nor aspartyl-phenylalanine (AspPhe) were detected in portal or vena caval blood following administration of any test compound. Steady-state perfusion of the small intestine with APM showed a net intraluminal appearance rate of AspPhe at 36% of the disappearance rate of APM. During steady-state PME perfusion, PHE had a significantly greater net appearance rate than during APM perfusion. Methanol appearance rates were slightly, but not significantly, higher during PME than during APM perfusions. The data suggest that (1) APM is hydrolyzed to AspPhe and both APM and PME are hydrolyzed to their constituent amino acids and and methanol prior to entering the portal circulation; (2) AspPhe is an important intraluminal intermediate in aspartame metabolism; and (3) aspartate is rapidly metabolized by the enterocyte.

  6. Virulent Hessian fly larvae manipulate the free amino acid content of host wheat plants.

    PubMed

    Saltzmann, Kurt D; Giovanini, Marcelo P; Zheng, Cheng; Williams, Christie E

    2008-11-01

    Gall-forming insects induce host plants to form specialized structures (galls) that provide immature life stages of the insect access to host plant nutrients and protection from natural enemies. Feeding by larvae of the Hessian fly (Mayetiola destructor Say) causes susceptible host wheat plants to produce a gall-like nutritive tissue that supports larval growth and development. To determine if changes in host plant free amino acid levels are associated with virulent Biotype L Hessian fly larval feeding, we quantified free amino acid levels in crown tissues of susceptible Newton wheat plants 1, 4, and 7 days after Hessian fly egg hatch. Hessian fly-infested susceptible plants were more responsive than resistant plants or uninfested controls, showing higher concentrations of alanine, glutamic acid, glycine, phenylalanine, proline, and serine 4 days after egg hatch. This 4-day post-hatch time point corresponds to the maturation of nutritive tissue cells in susceptible plants and the onset of rapid larval growth. By 7 days after egg hatch, when virulent second instars are actively feeding on the contents of nutritive tissue cells, the aromatic amino acids phenylalanine and tyrosine were more abundant compared to uninfested controls, but the levels of other free amino acids were no longer elevated. Changes in free amino acid abundance described in this report were associated with increased levels of mRNA encoded by wheat genes involved in amino acid synthesis and transport.

  7. Effect of thenardite on the direct detection of aromatic amino acids: implications for the search for life in the solar system

    NASA Astrophysics Data System (ADS)

    Doc Richardson, C.; Hinman, Nancy W.; Scott, Jill R.

    2009-10-01

    With the discovery of Na-sulphate minerals on Mars and Europa, recent studies using these minerals have focused on their ability to assist in the detection of bio/organic signatures. This study further investigates the ability of thenardite (Na2SO4) to effectively facilitate the ionization and identification of aromatic amino acids (phenylalanine, tyrosine and tryptophan) using a technique called geomatrix-assisted laser desorption/ionization in conjunction with a Fourier transform ion cyclotron resonance mass spectrometry. This technique is based on the ability of a mineral host to facilitate desorption and ionization of bio/organic molecules for detection. Spectra obtained from each aromatic amino acid alone and in combination with thenardite show differences in ionization mechanism and fragmentation patterns. These differences are due to chemical and structural differences between the aromatic side chains of their respective amino acid. Tyrosine and tryptophan when combined with thenardite were observed to undergo cation-attachment ([M+Na]+), due to the high alkali ion affinity of their aromatic side chains. In addition, substitution of the carboxyl group hydrogen by sodium led to formation of [M-H+Na]Na+ peaks. In contrast, phenylalanine mixed with thenardite showed no evidence of Na+ attachment. Understanding how co-deposition of amino acids with thenardite can affect the observed mass spectra is important for future exploration missions that are likely to use laser desorption mass spectrometry to search for bio/organic compounds in extraterrestrial environments.

  8. Effect of Thenardite on the Direct Detection of Aromatic Amino Acids: Implications for the Search for Life in the Solar System

    SciTech Connect

    C. Doc Richardson; Nancy W. Hinman; Jill R. Scott

    2009-10-01

    With the discovery of Na-sulfate minerals on Mars and Europa, recent studies using these minerals have focused on their ability to assist in the detection of bio/organic signatures. This study further investigates the ability of thenardite (Na2SO4) to effectively facilitate the ionization and identification of aromatic amino acids (phenylalanine, tyrosine, and tryptophan) using a technique called geomatrix-assisted laser desorption/ionization (GALDI) in conjunction with a Fourier transform mass spectrometry (FTICR-MS). This technique is based on the ability of a mineral host to facilitate the ionization and detection of bio/organic molecules. Spectra obtained from each aromatic amino acid alone and in combination with thenardite show differences in ionization mechanism and fragmentation patterns. These differences are due to chemical and structural differences between the aromatic side chains of their respective amino acid. Tyrosine and tryptophan when combined with thenardite were observed to undergo cation-attachment ([M+Na]+), due to the high alkali affinity of their aromatic side chains. Subsequent cation substitution of the carboxyl group led to formation double cation-attached peaks ([M-H+Na]Na+). In contrast, phenylalanine mixed with thenardite showed no evidence of Na+ interaction. Understanding how codeposition of amino acids with thenardite can affect the observed mass spectra is important for future exploration missions that are likely to use laser desorption mass spectrometry to search for bio/organic compounds in extraterrestrial environments.

  9. [Investigation of fluorescent components of drug "heparin" and its complexing with phenylalanine, tyrosine and tryptophan].

    PubMed

    Astakhov, S S; Iunusov, V M; Sultanbaev, M V; Akhmadeeva, G Kh; Iunusov, M S

    2013-01-01

    Using spectral-luminescent and spectrophotometric methods a composition of aminoacidic (AA) and protein components of commercial drug of heparin (Hep) as well as its interaction with Trp, Tyr and Phe was studied. The impurities of aromatic aminoacids Phe, Tyr and elastin protein was revealed in drug of heparin. The quenching of fluorescence (FL) of Trp, Tyr and Phe with heparin was studied and the Stern-Volmer Constants (K) showing the stability of its complexes with aminoacids were determined. The stability of complexes AA...Hep increases in the series K (Trp...Hep) = 19 +/- 2 M(-1) < K (Tyr...Hep) = 39 +/- 3 M(-1) < K (Phe...Hep) = 710 +/- 70 M(-1), that, probably, determines dominating contribution of Phe impurity in heparin and the absence of that of tryptophan. It was assumed, that animal elastin, which is different from human one can provoke allergic reactions up to anaphylactic shock.

  10. Optimal serum phenylalanine for adult patients with phenylketonuria.

    PubMed

    Okano, Yoshiyuki; Nagasaka, Hironori

    2013-12-01

    High serum phenylalanine in adult patients with phenylketonuria (PKU) causes neuropsychological and psychosocial problems that can be resolved by phenylalanine-restricted diet. Therefore, PKU patients must continue to adhere to phenylalanine-restricted diet for life, although the optimal serum phenylalanine level in later life has yet to be established. The purpose of this review was to establish the optimal serum phenylalanine level in later life of PKU patients. We evaluated oxidative stress status, nitric oxide metabolism, cholesterol-derived oxysterols, vitamin D and bone status, and magnetic resonance imaging (MRI) in adult PKU patients according to serum phenylalanine level. Oxidative stress increased markedly at serum phenylalanine of 700-800 μmol/L. Serum phenylalanine higher than 700-850 μmol/L correlated with the disturbance of nitric oxide regulatory system. Adult PKU patients had poor vitamin D status and exhibited predominance of bone resorption over bone formation. In the brain, the levels of 24S-hydroxycholesterol, a marker of brain cholesterol elimination, were low at serum phenylalanine levels exceeding 650 μmol/L. MRI studies showed high signal intensity in deep white matter on T2-weighted and FLAIR images of PKU patients with serum phenylalanine greater than 500 μmol/L, with decreased apparent diffusion coefficients. Changes in most parameters covering the entire body organs in adult PKU were almost acceptable below 700-800 μmol/L of phenylalanine level. However, the optimal serum phenylalanine level should be 500 μmol/L or less in later life for the brain to be safe.

  11. Reaction of alpha-tubulin with iodotyrosines catalyzed by tubulin:tyrosine ligase: carboxy-terminal labeling of tubulin with ( sup 125 I)monoiodotyrosine

    SciTech Connect

    Joniau, M.; Coudijzer, K.; De Cuyper, M. )

    1990-02-01

    We have studied the capacity of different iodinated derivatives of phenylalanine and tyrosine to inhibit the incorporation of (3H)tyrosine into tubulin catalyzed by tubulin:tyrosine ligase. In contrast to thyronine and its iodinated derivatives, iodotyrosines were efficient inhibitors. That they also functioned as substrates of the enzyme was shown by the effective incorporation of (125I)mono- and diiodotyrosine into tubulin. The label was shown to be located at the carboxy terminus. Labeling by this method conserves the polymerization capacity of tubulin in contrast with classical radioiodination methods involving oxidation.

  12. A new precursor for the preparation of 6-[18F]-fluoro-L-m-tyrosine (FMT): Efficient synthesis and comparison of radiolabeling

    SciTech Connect

    VanBrocklin, Henry F.; Blagoev, Milan; Hoepping, Alexander; O'Neil, James P.; Klose, Manuela; Schubiger, Pius A.; Ametamey, Simon

    2004-01-09

    For the electrophilic preparation of 6-[18F]-Fluoro-L-m-tyrosine (FMT), a PET tracer for measuring changes in dopaminergic function in movement disorders, a novel precursor, N-(tert-butoxycarbonyl)-3-(tert-butoxycarbonyloxy)-6-trimethylstannnyl-L-phenylalanine ethyl ester, was synthesized in four steps and 26 percent yield starting from L-m-tyrosine. FMT produced by two methods at two institutions was comparable in decay corrected yield, 25-26 percent, and quality (chemical, enantiomeric, and radiochemical purity and specific activity) as that obtained with the original N-trifluoroacetyl-3-acetyl-6-trimethylstannyl-L-m-tyrosine ethyl ester FMT precursor.

  13. Dopamine release in rat striatum - Physiological coupling to tyrosine supply

    NASA Technical Reports Server (NTRS)

    During, Matthew J.; Acworth, Ian N.; Wurtman, Richard J.

    1989-01-01

    Intracerebral microdialysis was used to monitor dopamine release in rat striatal extracellular fluid following the intraperitoneal administration of dopamine's precursor amino acid, L-tyrosine. Dopamine concentrations in dialysates increased transiently after tyrosine (50-100 mg/kg) administration. Pretreatment with haloperidol or the partial lesioning of nigrostriatal neurons enhanced the effect of tyrosine on dopamine release, and haloperidol also prolonged this effect. These data suggest that nigrostriatal dopaminergic neurons are responsive to changes in precursor availability under basal conditions, but that receptor-mediated feedback mechanisms limit the magnitude and duration of this effect.

  14. Light-activated amino acid transport in Halobacterium halobium envelope vesicles

    NASA Technical Reports Server (NTRS)

    Macdonald, R. E.; Lanyi, J. K.

    1977-01-01

    Vesicles prepared from Halobacterium halobium cell envelopes accumulate amino acids in response to light-induced electrical and chemical gradients. Nineteen of 20 commonly occurring amino acids have been shown to be actively accumulated by these vesicles in response to illumination or in response to an artificially created Na+ gradient. On the basis of shared common carriers the transport systems can be divided into eight classes, each responsible for the transport of one or several amino acids: arginine, lysine, histidine; asparagine, glutamine; alanine, glycine, threonine, serine; leucine, valine, isoleucine, methionine; phenylalanine, tyrosine, tryptophan; aspartate; glutamate; proline. Available evidence suggests that these carriers are symmetrical in that amino acids can be transported equally well in both directions across the vesicle membranes. A tentative working model to account for these observations is presented.

  15. Chemical modification of amino acids by atmospheric-pressure cold plasma in aqueous solution

    NASA Astrophysics Data System (ADS)

    Takai, Eisuke; Kitamura, Tsuyoshi; Kuwabara, Junpei; Ikawa, Satoshi; Yoshizawa, Shunsuke; Shiraki, Kentaro; Kawasaki, Hideya; Arakawa, Ryuichi; Kitano, Katsuhisa

    2014-07-01

    Plasma medicine is an attractive new research area, but the principles of plasma modification of biomolecules in aqueous solution remain elusive. In this study, we investigated the chemical effects of atmospheric-pressure cold plasma on 20 naturally occurring amino acids in aqueous solution. High-resolution mass spectrometry revealed that chemical modifications of 14 amino acids were observed after plasma treatment: (i) hydroxylation and nitration of aromatic rings in tyrosine, phenylalanine and tryptophan; (ii) sulfonation and disulfide linkage formation of thiol groups in cysteine; (iii) sulfoxidation of methionine and (iv) amidation and ring-opening of five-membered rings in histidine and proline. A competitive reaction experiment using 20 amino acids demonstrated that sulfur-containing and aromatic amino acids were preferentially decreased by the plasma treatment. These data provide fundamental information for elucidating the mechanism of protein inactivation for biomedical plasma applications.

  16. Tetrazine-Containing Amino Acid for Peptide Modification and Live Cell Labeling.

    PubMed

    Ni, Zhongqiu; Zhou, Lanxia; Li, Xu; Zhang, Jing; Dong, Shouliang

    2015-01-01

    A novel amino acid derivative 3-(4-(1, 2, 4, 5-tetrazine-3-yl) phenyl)-2-aminopropanoic acid was synthesized in this study. The compound possessed better water-solubility and was synthesized more easily compared with the well-known and commercially available 3-(p-benzylamino)-1, 2, 4, 5-tetrazine. Tetrazine-containing amino acid showed excellent stability in biological media and might be used for cancer cell labeling. Moreover, the compound remained relatively stable in 50% TFA/DCM with little decomposition after prolonged exposure at room temperature. The compound could be utilized as phenylalanine or tyrosine analogue in peptide modification, and the tetrazine-containing peptide demonstrated more significant biological activity than that of the parent peptide. The combination of tetrazine group and amino acid offered broad development prospects of the bioorthogonal labeling and peptide synthesis.

  17. Tetrazine-Containing Amino Acid for Peptide Modification and Live Cell Labeling

    PubMed Central

    Ni, Zhongqiu; Zhou, Lanxia; Li, Xu; Zhang, Jing; Dong, Shouliang

    2015-01-01

    A novel amino acid derivative 3-(4-(1, 2, 4, 5-tetrazine-3-yl) phenyl)-2-aminopropanoic acid was synthesized in this study. The compound possessed better water-solubility and was synthesized more easily compared with the well-known and commercially available 3-(p-benzylamino)-1, 2, 4, 5-tetrazine. Tetrazine-containing amino acid showed excellent stability in biological media and might be used for cancer cell labeling. Moreover, the compound remained relatively stable in 50% TFA/DCM with little decomposition after prolonged exposure at room temperature. The compound could be utilized as phenylalanine or tyrosine analogue in peptide modification, and the tetrazine-containing peptide demonstrated more significant biological activity than that of the parent peptide. The combination of tetrazine group and amino acid offered broad development prospects of the bioorthogonal labeling and peptide synthesis. PMID:26536589

  18. Effects of hemorrhagic hypotension on tyrosine concentrations in rat spinal cord and plasma

    NASA Technical Reports Server (NTRS)

    Conlay, L. A.; Maher, T. J.; Roberts, C. H.; Wurtman, R. J.

    1988-01-01

    Tyrosine is the precursor for catecholamine neurotransmitters. When catecholamine-containing neurons are physiologically active (as sympathoadrenal cells are in hypotension), tyrosine administration increases catecholamine synthesis and release. Since hypotension can alter plasma amino acid composition, the effects of an acute hypotensive insult on tyrosine concentrations in plasma and spinal cord were examined. Rats were cannulated and bled until the systolic blood pressure was 50 mmHg, or were kept normotensive for 1 h. Tyrosine and other large neutral amino acids (LNAA) known to compete with tyrosine for brain uptake were assayed in plasma and spinal cord. The rate at which intra-arterial (H-3)tyrosine disappeared from the plasma was also estimated in hemorrhaged and control rats. In plasma of hemorrhaged animals, both the tyrosine concentration and the tyrosine/LNAA ratio was elevated; moreover, the disappearance of (H-3)tyrosine was slowed. Tyrosine concentrations also increased in spinal cords of hemorrhaged-hypotensive rats when compared to normotensive controls. Changes in plasma amino acid patterns may thus influence spinal cord concentrations of amino acid precursors for neurotransmitters during the stress of hemorrhagic shock.

  19. Interaction of tryptophan and phenylalanine with metal ferrocyanides and its relevance in chemical evolution.

    PubMed

    Ali, Shah Raj; Alam, Tanveer; Kamaluddin

    2004-01-01

    The interaction of two naturally occurring aromatic alpha-amino acids, namely, tryptophan and phenylalanine, with zinc, nickel, cobalt, and copper ferrocyanides has been studied. Both amino acids showed a high adsorption affinity toward metal ferrocyanides at neutral pH (7.0). Adsorption trends followed the Langmuir adsorption isotherm. Values of the Langmuir constants K(L) and X(m) suggest tryptophan is a better adsorbate than phenylalanine. Zinc ferrocyanide showed the highest adsorption, while the minimum adsorption was found in the case of copper ferrocyanide. Infrared spectral studies of adsorbate, adsorbent, and adsorption adducts indicate that adsorption occurs because of the interaction of adsorbate molecules with outer divalent metal ions present in the lattice of metal ferrocyanides. The present investigation supports the hypothesis that metal ferrocyanides might have concentrated the biomonomers on their surface in primeval seas during the course of chemical evolution.

  20. Experiments on the origins of optical activity. [in amino acids

    NASA Technical Reports Server (NTRS)

    Bonner, W. A.; Flores, J. J.

    1975-01-01

    An investigation was conducted concerning the asymmetric adsorption of phenylalanine enantiomers by kaolin. No preferential adsorption of either phenylalanine enantiomer could be detected and there was no resolution of the racemic phenylalanine by kaolin. The attempted asymmetric polymerization of aspartic acid by kaolin is also discussed along with a strontium-90 bremsstrahlung radiolysis of leucine.

  1. Protein nutrition of Tenebrio molitor L. XX. Growth response of larvae to graded levels of amino acids.

    PubMed

    John, A M; Davis, G R; Sosulski, F W

    1979-12-01

    Larvae of Tenebrio molitor L;, Gembloux strain, race F, having an average initial weight of 10 mg were reared for 4 weeks at 27 +/- 0.25 degrees C and 65 +/- 5% relative humidity on diets containing an amino-acid mixture resembling the composition of larval tissues; Each of 14 amino acids was tested individually at levels of 0, 10, 20, 50, 100 and 200% of the amount found in larval tissues, while the concentration of other amino acids remained constant. Diets were maintained isonitrogenous at 1.6% nitrogen with supplementary glutamic acid, glycine, serine and tyrosine; Maximum fresh weight gains and dry weights were achieved by larvae fed reference levels of all essential amino acids except phenylalanine, threonine and tryptophan. Maximum growth was achieved by larvae fed 50% of the phenylalanine level found in larval tissues, Threonine and tryptophan were the limiting amino acids in this study and are probably required in the diet in excess of twice the concentration occuring in larval tissues, Probable ranges for quantitative amino-acid requirements of T. molitor were determined and suggestions were made for improving the nutritional adequacy of the amino-acid mixture.

  2. The carboxyl terminal tyrosine 417 residue of NOK has an autoinhibitory effect on NOK-mediated signaling transductions

    SciTech Connect

    Li Yinghua; Zhong Shan; Rong Zhili; Ren Yongming; Li Zhiyong; Zhang Shuping; Chang Zhijie; Liu Li . E-mail: Liu_Li@mail.tsinghua.edu.cn

    2007-05-04

    Receptor protein tyrosine kinases (RPTKs) are essential mediators of cell growth, differentiation, migration, and metabolism. Recently, a novel RPTK named NOK has been cloned and characterized. In current study, we investigated the role of the carboxyl terminal tyrosine 417 residue of NOK in the activations of different signaling pathways. A single tyrosine to phenylalanine point mutation at Y417 site (Y417 F) not only dramatically enhanced the NOK-induced activation of extracellular signal-regulated kinase (ERK), but also markedly promoted the NOK-mediated activation of both signal transducer and activator of transcription 1 and 3 (STAT1 and 3). Moreover, the proliferation potential of NIH3T3-NOK (Y417F) stable cells were significantly elevated as compared with that of NIH3T3-NOK. Overall, our results demonstrate that the tyrosine Y417 residue at the carboxyl tail of NOK exhibits an autoinhibitory role in NOK-mediated signaling transductions.

  3. Identification of nitrated tyrosine residues of protein kinase G-Iα by mass spectrometry.

    PubMed

    Lu, Jingshan; Yao, Ikuko; Shimojo, Masahito; Katano, Tayo; Uchida, Hitoshi; Setou, Mitsutoshi; Ito, Seiji

    2014-02-01

    The nitration of tyrosine to 3-nitrotyrosine is an oxidative modification of tyrosine by nitric oxide and is associated with many diseases, and targeting of protein kinase G (PKG)-I represents a potential therapeutic strategy for pulmonary hypertension and chronic pain. The direct assignment of tyrosine residues of PKG-I has remained to be made due to the low sensitivity of the current proteomic approach. In order to assign modified tyrosine residues of PKG-I, we nitrated purified PKG-Iα expressed in insect Sf9 cells by use of peroxynitrite in vitro and analyzed the trypsin-digested fragments by matrix-assisted laser desorption/ionization-time of flight mass spectrometry and liquid chromatography-tandem mass spectrometry. Among the 21 tyrosine residues of PKG-Iα, 16 tyrosine residues were assigned in 13 fragments; and six tyrosine residues were nitrated, those at Y71, Y141, Y212, Y336, Y345, and Y567, in the peroxynitrite-treated sample. Single mutation of tyrosine residues at Y71, Y212, and Y336 to phenylalanine significantly reduced the nitration of PKG-Iα; and four mutations at Y71, Y141, Y212, and Y336 (Y4F mutant) reduced it additively. PKG-Iα activity was inhibited by peroxynitrite in a concentration-dependent manner from 30 μM to 1 mM, and this inhibition was attenuated in the Y4F mutant. These results demonstrated that PKG-Iα was nitrated at multiple tyrosine residues and that its activity was reduced by nitration of these residues.

  4. Enhanced rosmarinic acid production in cultured plants of two species of Mentha.

    PubMed

    Roy, Debleena; Mukhopadhyay, Sandip

    2012-11-01

    In the present investigation an attempt has been made to enhance rosmarinic acid level in plants, grown in vitro, of 2 species of Mentha in presence of 2 precursors in the nutrient media during culture. For in vitro culture establishment and shoot bud multiplication, MS basal media were used supplemented with different concentrations and combinations of different growth regulator like NAA (alpha-napthaleneacetic acid), BAP (6-benzylaminopurine). The medium containing NAA (0.25 mg/L) and BAP (2.5 mg/L) gave the highest potentiality of shoot formation (average 58.0 numbers of shoots) per explant for Mentha piperita L. and the medium containing BAP (2.0 mg/L) gave the highest potentiality of shoot (average 19.2 numbers of shoots) formation per explant for Mentha arvensis L. The complete plants were regenerated in above mentioned media after 8 weeks of subculture. For in vitro enhancement of rosmarinic acid production, the 2 precursors tyrosine (Tyr) and phenylalanine (Phe) were added in the nutrient media at different levels (0.5 mg/L to 15.0 mg/L). Tyrosine was found to be very effective for augmenting rosmarinic acid content in Mentha piperita L. It nearly increased the production up to 1.77 times. In case of Mentha arvensis L., phenylalanine significantly affected the production of rosmarinic acid and the production was nearly 2.03 times more than the control. No significant increase in biomass was observed after addition of these precursors indicating that the added amino acids acting as precursors for rosmarinic acid synthesis were readily utilized in producing rosmarinic acid without promoting growth. Total protein profile also revealed the presence of a specific band in polyacrylamide gel electrophoresis.

  5. Kinetic Characterization of O-Phospho-L-Tyrosine Phosphohydrolase Activity of Two Fungal Phytases.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Fungal phytases belonging to 'Histidine Acid Phosphatase' or HAP class of phosphomonoesterase that catalyzes the hydrolysis of phytic acid could also hydrolyze O-phospho-tyrosine. Two phytases from Aspergillus niger and Aspergillus awamori with pH optima 2.5 were tested for phospho-tyrosine hydrola...

  6. Strecker type degradation of phenylalanine by 4-hydroxy-2-nonenal in model systems.

    PubMed

    Hidalgo, Francisco J; Gallardo, Emerenciana; Zamora, Rosario

    2005-12-28

    The reaction of 4-hydroxy-2-nonenal, an oxidative stress product, with phenylalanine in acetonitrile-water (2:1, 1:1, and 1:2) at 37, 60, and 80 degrees C was investigated to determine whether 4-hydroxy-2-alkenals degrade amino acids, analogously to 4,5-epoxy-2-alkenals, and to compare the reactivities of both hydroxyalkenals and epoxyalkenals for production of Strecker aldehydes. In addition to the formation of N-substituted 2-pentylpyrrole and 2-pentylfuran, the studied hydroxyalkenal also degraded phenylalanine to phenylacetaldehyde with a reaction yield of 17%. The reaction mechanism is suggested to be produced through the corresponding imine, which is then decarboxylated and hydrolyzed. This reaction also produced a conjugated amine, which both may be one of the origins of the produced 2-pentyl-1H-pyrrole and may contribute to the development of browning in these reactions. 4-Hydroxy-2-nonenal and 4,5-epoxy-2-decenal degraded phenylalanine in an analogous extent, which is likely a consequence of the similarity of the degradation mechanisms involved. These results suggest that different lipid oxidation products are able to degrade amino acids; therefore, the Strecker type degradation of amino acids produced by oxidized lipids may be quantitatively significant in foods. PMID:16366724

  7. Structural basis for ligand-dependent dimerization of phenylalanine hydroxylase regulatory domain

    PubMed Central

    Patel, Dipali; Kopec, Jolanta; Fitzpatrick, Fiona; McCorvie, Thomas J.; Yue, Wyatt W.

    2016-01-01

    The multi-domain enzyme phenylalanine hydroxylase (PAH) catalyzes the hydroxylation of dietary I-phenylalanine (Phe) to I-tyrosine. Inherited mutations that result in PAH enzyme deficiency are the genetic cause of the autosomal recessive disorder phenylketonuria. Phe is the substrate for the PAH active site, but also an allosteric ligand that increases enzyme activity. Phe has been proposed to bind, in addition to the catalytic domain, a site at the PAH N-terminal regulatory domain (PAH-RD), to activate the enzyme via an unclear mechanism. Here we report the crystal structure of human PAH-RD bound with Phe at 1.8 Å resolution, revealing a homodimer of ACT folds with Phe bound at the dimer interface. This work delivers the structural evidence to support previous solution studies that a binding site exists in the RD for Phe, and that Phe binding results in dimerization of PAH-RD. Consistent with our structural observation, a disease-associated PAH mutant impaired in Phe binding disrupts the monomer:dimer equilibrium of PAH-RD. Our data therefore support an emerging model of PAH allosteric regulation, whereby Phe binds to PAH-RD and mediates the dimerization of regulatory modules that would bring about conformational changes to activate the enzyme. PMID:27049649

  8. Thermodynamic description of the ion exchange and superequivalent sorption of phenylalanine by KU-2-8 cationite

    NASA Astrophysics Data System (ADS)

    Khokhlova, O. N.; Khokhlov, V. Yu.; Trunaeva, E. S.

    2015-02-01

    The coefficients and thermodynamic equilibrium constants, activity coefficients of the sorbent phase, and integral and differential Gibbs energies of the ion-exchange and superequivalent sorption of phenylalanine by strongly acidic KU-2-8 cationites in the H form are calculated using a new thermodynamic approach that allows us to describe the simultaneous exchange and superequivalent sorption of substances by ionites.

  9. Thermodynamics of the complex formation of copper(II) with L-phenylalanine in aqueous ethanol solutions

    NASA Astrophysics Data System (ADS)

    Burov, D. M.; Ledenkov, S. F.; Vandyshev, V. N.

    2013-05-01

    Constants of the acid dissociation and complexation of L-phenylalanine (HPhe) with copper(II) ions are determined by potentiometry in aqueous ethanol solutions containing 0 to 0.7 molar fraction of alcohol. Changes in the Gibbs energy for the transfer from water to a binary solvent of L-phenylalanine, Phe- anion, and [CuPhe]+ complex are calculated. It is found that the weakening of solvation of the ligand donor groups in solvents with high ethanol contents is accompanied by an increase in the stability of [CuPhe]+ complex.

  10. Effect of different postharvest temperatures on the accumulation of sugars, organic acids, and amino acids in the juice sacs of Satsuma mandarin (Citrus unshiu Marc.) fruit.

    PubMed

    Matsumoto, Hikaru; Ikoma, Yoshinori

    2012-10-01

    To elucidate the effect of different postharvest temperatures on the accumulation of sugars, organic acids, and amino acids and to determine the best temperature to minimize their postharvest change, their content after harvest was investigated at 5, 10, 20, and 30 °C for 14 days in the juice sacs of Satsuma mandarin (Citrus unshiu Marc. cv. Aoshima-unshiu) fruit. In all sugars, the changes were negligible at all temperatures. Organic acids decreased slightly at all temperatures, with the exception of malic acid at 30 °C, which increased slightly. Two amino acids, ornithine and glutamine, increased at 5 °C, but they did not increase at other temperatures. In 11 amino acids (phenylalanine, tryptophan, tyrosine, isoleucine, leucine, valine, threonine, lysine, methionine, histidine, and γ-amino butyric acid), the content was higher at 20 and 30 °C than at other temperatures. Thus, the content of amino acids was more variable than that of sugars and organic acids in response to temperatures. Moreover, amino acids responded to temperature differently: two amino acids were cold responsive, and 11 were heat-responsive. The best temperature to minimize the postharvest changes in amino acid profiles in the juice sacs of Aoshima-unshiu was 10 °C. The responsiveness to temperatures in two cold-responsive (ornithine and glutamine) and five heat-responsive (phenylalanine, tryptophan, valine, lysine, and histidine) amino acids was conserved among three different Satsuma mandarin cultivars, Aoshima-unshiu (late-maturing cultivar), Silverhill (midmaturing cultivar), and Miyagawa-wase (early-maturing cultivar). The metabolic responsiveness to temperature stress was discussed on the basis of the changes in the amino acid profile.

  11. Isolation and some effects of functional, low-phenylalanine kappa-casein expressed in the milk of transgenic rabbits.

    PubMed

    Baranyi, Mária; Hiripi, László; Szabó, László; Catunda, Ana Paula; Harsányi, Ibolya; Komáromy, Péter; Bosze, Zsuzsanna

    2007-02-01

    Patients suffering certain metabolic diseases (e.g. phenylketonuria) need a low-phenylalanine diet throughout their lives. Transgenic rabbits were created to express low-phenylalanine kappa-casein in their milk. The aim was to demonstrate for the first time the feasibility of producing a modified milk protein in addition to normal milk proteins. A gene construct containing the coding region of the rabbit kappa-casein gene was modified by site-specific oligonucleotide directed mutagenesis. Four of the five phenylalanine amino acids present in the mature protein were mutated and the gene construct was used to create two transgenic rabbit lines. The transgenic rabbits produced the recombinant kappa-casein at a high level in their milk causing a reduction in the average size of the casein micelles. The low-phenylalanine kappa-casein was digestible with chymosin and it was separated from its native counterpart and from the other milk proteins by a one-step HPLC method on a reversed-phase column. In the future, low-phenylalanine casein produced in transgenic animals could be used as dietary replacements to meet the special requirements of certain consumer groups.

  12. A resolution approach of racemic phenylalanine with aqueous two-phase systems of chiral tropine ionic liquids.

    PubMed

    Wu, Haoran; Yao, Shun; Qian, Guofei; Yao, Tian; Song, Hang

    2015-10-30

    Aqueous two-phase systems (ATPS) based on tropine type chiral ionic liquids and inorganic salt solution were designed and prepared for the enantiomeric separation of racemic phenylalanine. The phase behavior of IL-based ATPS was comprehensive investigated, and phase equilibrium data were correlated by Merchuk equation. Various factors were also systematically investigated for their influence on separation efficiency. Under the appropriate conditions (0.13g/g [C8Tropine]pro, 35mg/g Cu(Ac)2, 20mg/g d,l-phenylalanine, 0.51g/g H2O and 0.30g/g K2HPO4), the enantiomeric excess value of phenylalanine in solid phase (mainly containing l-enantiomer) was 65%. Finally, the interaction mechanism was studied via 1D and 2D NMR. The results indicate that d-enantiomer of phenylalanine interacts more strongly with chiral ILs and Cu(2+) based on the chiral ion-pairs space coordination mechanism, which makes it tend to remain in the top IL-rich phase. By contrast, l-enantiomer is transferred into the solid phase. Above chiral ionic liquids aqueous two-phase systems have demonstrated obvious resolution to racemic phenylalanine and could be promising alterative resolution approach for racemic amino acids in aqueous circumstance.

  13. Transcriptome sequencing revealed the transcriptional organization at ribosome-mediated attenuation sites in Corynebacterium glutamicum and identified a novel attenuator involved in aromatic amino acid biosynthesis.

    PubMed

    Neshat, Armin; Mentz, Almut; Rückert, Christian; Kalinowski, Jörn

    2014-11-20

    The Gram-positive bacterium Corynebacterium glutamicum belongs to the order Corynebacteriales and is used as a producer of amino acids at industrial scales. Due to its economic importance, gene expression and particularly the regulation of amino acid biosynthesis has been investigated extensively. Applying the high-resolution technique of transcriptome sequencing (RNA-seq), recently a vast amount of data has been generated that was used to comprehensively analyze the C. glutamicum transcriptome. By analyzing RNA-seq data from a small RNA cDNA library of C. glutamicum, short transcripts in the known transcriptional attenuators sites of the trp operon, the ilvBNC operon and the leuA gene were verified. Furthermore, whole transcriptome RNA-seq data were used to elucidate the transcriptional organization of these three amino acid biosynthesis operons. In addition, we discovered and analyzed the novel attenuator aroR, located upstream of the aroF gene (cg1129). The DAHP synthase encoded by aroF catalyzes the first step in aromatic amino acid synthesis. The AroR leader peptide contains the amino acid sequence motif F-Y-F, indicating a regulatory effect by phenylalanine and tyrosine. Analysis by real-time RT-PCR suggests that the attenuator regulates the transcription of aroF in dependence of the cellular amount of tRNA loaded with phenylalanine when comparing a phenylalanine-auxotrophic C. glutamicum mutant fed with limiting and excess amounts of a phenylalanine-containing dipeptide. Additionally, the very interesting finding was made that all analyzed attenuators are leaderless transcripts. PMID:24910972

  14. Derepression of certain aromatic amino acid biosynthetic enzymes of Escherichia coli K-12 by growth in Fe3+-deficient medium.

    PubMed Central

    McCray, J W; Herrmann, K M

    1976-01-01

    3-Deoxy-arabino-heptulosonic acid 7-phosphate synthase, prephenate dehydratase, tryptophan synthase, and 2,3-dihydroxybenzoylserine synthase enzyme activities are derepressed in wild-type Escherichia coli K-12 cells grown on Fe3+-deficient medium. This derepression is reversed when FeSO4 is added to the growth medium. Addition of shikimic acid to the Fe3+-deficient growth medium caused repression of the first three enzyme activities but not of 2,3-dihydroxybenzoylserine synthase activity. Addition of 2,3-dihydroxybenzoic acid to the Fe3+-deficient growth medium has no effect on any of the above-mentioned enzyme activities. The Fe3+ deficiency-mediated derepression of 3-deoxyarabino-heptulosonic acid 7-phosphate synthase activity is due to an elevation of the tyrosine-sensitive isoenzyme; the phenylalanine-sensitive isoenzyme is not derepressed under these conditions. PMID:1383

  15. The development and amino acid binding ability of nano-materials based on azo derivatives: theory and experiment.

    PubMed

    Shang, Xuefang; Du, Jinge; Yang, Wancai; Liu, Yun; Fu, Zhiyuan; Wei, Xiaofang; Yan, Ruifang; Yao, Ningcong; Guo, Yaping; Zhang, Jinlian; Xu, Xiufang

    2014-05-01

    Two nano-material-containing azo groups have been designed and developed, and the binding ability of nano-materials with various amino acids has been characterized by UV-vis and fluorescence titrations. Results indicated that two nano-materials showed the strongest binding ability for homocysteine among twenty normal kinds of amino acids (alanine, valine, leucine, isoleucine, methionine, aspartic acid, glutamic acid, arginine, glycine, serine, threonine, asparagine, phenylalanine, histidine, tryptophan, proline, lysine, glutamine, tyrosine and homocysteine). The reason for the high sensitivity for homocysteine was that two nano-materials containing an aldehyde group reacted with SH in homocysteine and afforded very stable thiazolidine derivatives. Theoretical investigation further illustrated the possible binding mode in host-guest interaction and the roles of molecular frontier orbitals in molecular interplay. Thus, the two nano-materials can be used as optical sensors for the detection of homocysteine.

  16. Quantification of Five Clinically Important Amino Acids by HPLC-Triple TOF™ 5600 Based on Pre-column Double Derivatization Method.

    PubMed

    Deng, Shuang; Scott, David; Garg, Uttam

    2016-01-01

    Phenylalanine, tyrosine, glycine, cystine, and phosphoethanolamine are commonly measured amino acids in various physiological fluids to diagnose or follow-up various inborn errors of metabolism. The gold standard method for the amino acids quantitation has been ion exchange chromatography with ninhydrin post-column derivatization. However, this method is very laborious and time consuming. In recent years, liquid-chromatography mass spectrometry is being increasingly used for the assay of amino acids. Pre-column butyl derivatization with reverse phase chromatography has been widely used for mass spectrometry analysis of amino acids. Phosphoethanolamine is not butylated and cannot be measured by this method. Nevertheless, phosphoethanolamine can be dansyl-derivatized using dansyl chloride. We developed a double derivatization method by using butanol and dansyl chloride to derivatize carboxylic and amino groups separately, and then combining the derivatives to simultaneously measure these five amino acids using TOF-MS scan. Stable isotope-labeled internal standards were used. PMID:26602116

  17. Quantification of Five Clinically Important Amino Acids by HPLC-Triple TOF™ 5600 Based on Pre-column Double Derivatization Method.

    PubMed

    Deng, Shuang; Scott, David; Garg, Uttam

    2016-01-01

    Phenylalanine, tyrosine, glycine, cystine, and phosphoethanolamine are commonly measured amino acids in various physiological fluids to diagnose or follow-up various inborn errors of metabolism. The gold standard method for the amino acids quantitation has been ion exchange chromatography with ninhydrin post-column derivatization. However, this method is very laborious and time consuming. In recent years, liquid-chromatography mass spectrometry is being increasingly used for the assay of amino acids. Pre-column butyl derivatization with reverse phase chromatography has been widely used for mass spectrometry analysis of amino acids. Phosphoethanolamine is not butylated and cannot be measured by this method. Nevertheless, phosphoethanolamine can be dansyl-derivatized using dansyl chloride. We developed a double derivatization method by using butanol and dansyl chloride to derivatize carboxylic and amino groups separately, and then combining the derivatives to simultaneously measure these five amino acids using TOF-MS scan. Stable isotope-labeled internal standards were used.

  18. Amino acid contents along the visual and equatorial axes of a pig lens by Raman spectroscopy

    NASA Astrophysics Data System (ADS)

    Medina-Gutiérrez, C.; Frausto-Reyes, C.; Quintanar-Stephano, J. L.; Sato-Berrú, R.

    2004-08-01

    Using near infrared Raman microspectroscopy with laser light of 830 nm, the distribution of amino acids along the visual and equatorial axes of a normal pig lens was studied. The classification of pig lens Raman spectra in these axes was performed using principal component analysis and linear discriminant analysis. The analysis of the scattered light selectively collected from point to point, along the visual axis, indicated that the tyrosine and tryptophan increases and then, at ˜4 mm position, decreases. Moreover, in the equatorial plane, the nuclear part has the highest concentration of these amino acids. However, the phenylalanine content increases from anterior to posterior cortex of the lens as long as in the equatorial axis it slightly increases and then at ˜2-2.3 mm position, decreases. The changes in amino acid conformation along the visual axis, similarly to the changes in protein conformation, may explain the refractive gradient of the lens.

  19. Adsorption and properties of aromatic amino acids on single-walled carbon nanotubes

    NASA Astrophysics Data System (ADS)

    Wang, Cuihong; Li, Shuang; Zhang, Ruiqin; Lin, Zijing

    2012-02-01

    We investigated the adsorption of three aromatic amino acids--phenylalanine, tyrosine, and tryptophan--on the sidewalls of a number of representative single-walled carbon nanotubes (SWNTs) using density-functional tight-binding calculations, complemented by an empirical dispersion correction. The armchair (n, n) SWNTs (n = 3-12) and zigzag (n, 0) SWNTs (n = 4-12) were thoroughly examined. We found that the most stable amino acid/SWNT complexes for different SWNTs have similar local structures, and that the distance between the amino acid and SWNT is about 3 Å. Owing to the π-π and H-π stacking interactions, the benzene and indole rings are not exactly parallel to the SWNTs but instead lie at a small angle. We also investigated the diameter and chirality dependences of binding energies and found that SWNT (5, 0) has an especially large binding energy that can be used for SWNT identification or selection.

  20. Amino acid contents along the visual and equatorial axes of a pig lens by Raman spectroscopy.

    PubMed

    Medina-Gutiérrez, C; Frausto-Reyes, C; Quintanar-Stephano, J L; Sato-Berrú, R

    2004-08-01

    Using near infrared Raman microspectroscopy with laser light of 830 nm, the distribution of amino acids along the visual and equatorial axes of a normal pig lens was studied. The classification of pig lens Raman spectra in these axes was performed using principal component analysis and linear discriminant analysis. The analysis of the scattered light selectively collected from point to point, along the visual axis, indicated that the tyrosine and tryptophan increases and then, at approximately 4 mm position, decreases. Moreover, in the equatorial plane, the nuclear part has the highest concentration of these amino acids. However, the phenylalanine content increases from anterior to posterior cortex of the lens as long as in the equatorial axis it slightly increases and then at approximately 2-2.3 mm position, decreases. The changes in amino acid conformation along the visual axis, similarly to the changes in protein conformation, may explain the refractive gradient of the lens.

  1. The Value of 5-Aminolevulinic Acid in Low-grade Gliomas and High-grade Gliomas Lacking Glioblastoma Imaging Features: An Analysis Based on Fluorescence, Magnetic Resonance Imaging, 18F-Fluoroethyl Tyrosine Positron Emission Tomography, and Tumor Molecular Factors

    PubMed Central

    Jaber, Mohammed; Wölfer, Johannes; Ewelt, Christian; Holling, Markus; Hasselblatt, Martin; Niederstadt, Thomas; Zoubi, Tarek; Weckesser, Matthias

    2015-01-01

    BACKGROUND: Approximately 20% of grade II and most grade III gliomas fluoresce after 5-aminolevulinic acid (5-ALA) application. Conversely, approximately 30% of nonenhancing gliomas are actually high grade. OBJECTIVE: The aim of this study was to identify preoperative factors (ie, age, enhancement, 18F-fluoroethyl tyrosine positron emission tomography [18F-FET PET] uptake ratios) for predicting fluorescence in gliomas without typical glioblastomas imaging features and to determine whether fluorescence will allow prediction of tumor grade or molecular characteristics. METHODS: Patients harboring gliomas without typical glioblastoma imaging features were given 5-ALA. Fluorescence was recorded intraoperatively, and biopsy specimens collected from fluorescing tissue. World Health Organization (WHO) grade, Ki-67/MIB-1 index, IDH1 (R132H) mutation status, O6-methylguanine DNA methyltransferase (MGMT) promoter methylation status, and 1p/19q co-deletion status were assessed. Predictive factors for fluorescence were derived from preoperative magnetic resonance imaging and 18F-FET PET. Classification and regression tree analysis and receiver-operating-characteristic curves were generated for defining predictors. RESULTS: Of 166 tumors, 82 were diagnosed as WHO grade II, 76 as grade III, and 8 as glioblastomas grade IV. Contrast enhancement, tumor volume, and 18F-FET PET uptake ratio >1.85 predicted fluorescence. Fluorescence correlated with WHO grade (P < .001) and Ki-67/MIB-1 index (P < .001), but not with MGMT promoter methylation status, IDH1 mutation status, or 1p19q co-deletion status. The Ki-67/MIB-1 index in fluorescing grade III gliomas was higher than in nonfluorescing tumors, whereas in fluorescing and nonfluorescing grade II tumors, no differences were noted. CONCLUSION: Age, tumor volume, and 18F-FET PET uptake are factors predicting 5-ALA-induced fluorescence in gliomas without typical glioblastoma imaging features. Fluorescence was associated with an increased

  2. Valine and phenylalanine as precursors in the biosynthesis of alkamides in Acmella radicans.

    PubMed

    Cortez-Espinosa, Nancy; Aviña-Verduzco, Judit A; Ramírez-Chávez, Enrique; Molina-Torres, Jorge; Ríos-Chávez, Patricia

    2011-06-01

    Acmella radicans (Asteraceae) produces at least seven alkamides, most with either an isobutyl- or phenylethyl group as the amine moiety. These moieties suggest that the amino acids valine and phenylalanine are the biosynthetic precursors of these alkamides. On the basis of labeled feeding experiments using either L-[2H8]valine or L-[2H8]phenylalanine we present evidence for the involvement of these two amino acids in the biosynthesis of (2E,6Z,8E)-N-isobutyl-2,6,8-decatrienamide (affinin) (1), (2Z,4E)-N-(2-phenylethyl)-2,4-octadienamide (2), (2E)-N-(2-phenylethyl)-nona-2-en-6,8-diynamide (3), and 3-phenyl-N-(2-phenylethyl)-2-propenamide (4). Alkamides were isolated from young A. radicans plants and analyzed by gas chromatography-mass spectrometry (GC-MS). Additionally, in cell free in vitro experiments based on isobutyl and phenylethylamide biosynthesis, using a colorimetric assay and GC-MS, valine and phenylalanine decarboxylase activities were assayed in the soluble extract of A. radicans leaves.

  3. Quasiracemate Crystal Structures of Magainin 2 Derivatives Support the Functional Significance of the Phenylalanine Zipper Motif

    PubMed Central

    Hayouka, Zvi; Thomas, Nicole C.; Mortenson, David E.; Satyshur, Kenneth A.; Weisblum, Bernard; Forest, Katrina T.; Gellman, Samuel H.

    2016-01-01

    Quasiracemic crystallography has been used to explore the significance of homochiral and heterochiral associations in a set of host-defense peptide derivatives. The previously reported racemic crystal structure of a magainin 2 derivative displayed a homochiral dimer association featuring a “phenylalanine zipper” notable for the dual roles of phenylalanines in mediating dimerization and formation of an exposed hydrophobic swath. This motif is seen as well in two new quasiracemate crystals that contain the D form of the magainin 2 derivative along with an L-peptide in which one Ala has been replaced by a β-amino acid residue. This structural trend supports the hypothesis that the Phe zipper motif has functional significance. PMID:26369301

  4. N-(L-2-aminopentanoyl)-L-phenylalanine dihydrate, a hydrophobic dipeptide with a nonproteinogenic residue.

    PubMed

    Görbitz, Carl Henrik; Yadav, Vitthal N

    2013-09-01

    The title dipeptide, better known as L-norvalyl-L-phenylalanine {systematic name: (S)-2-[(S)-2-aminopentanamido]-3-phenylpropanoic acid dihydrate}, C14H20N2O3·2H2O, has a nonproteinogenic N-terminal residue. In the solid state, it takes on a molecular conformation typical for one of the three classes of nanoporous dipeptides, but like two related compounds with a hydrophobic N-terminal residue and a C-terminal L-phenylalanine, it fails to form channels or pores. Instead, the crystal structure is divided into distinct hydrophobic and hydrophilic layers, the latter encompassing cocrystallized water molecules connecting the charged N- and C-terminal groups.

  5. Reanalysis of the effects of phenylalanine, alanine, and aspartame on food intake in human subjects.

    PubMed

    Rogers, P J; Blundell, J E

    1994-08-01

    In 1987 Ryan-Harshman et al. reported finding no effects on food intake after administering high doses (up to 10.08 g) of phenylalanine and aspartame in capsules to human volunteers. However, this is contrary to the results of other studies, and trends in their tabulated data suggest that certain effects may have been overlooked. This is confirmed by a reanalysis of the raw data (available from a Ph.D. thesis: Ryan-Harshman, 1987) that can be interpreted as showing a dose-related suppression of food intake by phenylalanine. Furthermore, the data are consistent with an anorexic action of aspartame and perhaps also of alanine (which was designated as the placebo treatment by Ryan-Harshman et al.). These, together with other findings, suggest that the appetite effects of amino acids and small peptides should be investigated further. In addition to its theoretical importance, such work may have potential for therapeutic applications.

  6. Role of the distal phenylalanine 54 on the structure, stability, and ligand binding of Coprinus cinereus peroxidase.

    PubMed

    Neri, F; Indiani, C; Baldi, B; Vind, J; Welinder, K G; Smulevich, G

    1999-06-15

    Resonance Raman and electronic absorption spectra obtained at various pH values for the Fe3+ form of distal F54 mutants of Coprinus cinereus peroxidase are reported, together with the Fe2+ form and fluoride and imidazole adducts at pH 6.0, 5.0, and 10.5, respectively. The distal phenylalanine residue has been replaced by the small aliphatic residues glycine and valine and the hydrogen-bonding aromatic residues tyrosine and tryptophan (F54G, -V, -Y, and -W, respectively). These mutations resulted in transitions between ferric high-spin five-coordinate and six-coordinate forms, and caused a decrease of the pKa of the alkaline transition together with a higher tendency for unfolding. The mutations also alter the ability of the proteins to bind fluoride in such a way that those that are six-coordinate at pH 5.0 bind more strongly than both wild-type CIP and F54Y which are five-coordinate at this pH value. The data provide evidence that the architecture of the distal pocket of CIP is altered by the mutations. Direct evidence is provided that the distal phenylalanine plays an important role in controlling the conjugation between the vinyl double bonds and the porphyrin macrocycle, as indicated by the reorientation of the vinyl groups upon mutation of phenylalanine with the small aliphatic side chains of glycine and valine residues. Furthermore, it appears that the presence of the hydrogen-bonding tyrosine or tryptophan in the cavity increases the pKa of the distal histidine for protonation compared with that of wild-type CIP.

  7. Behavioral and cognitive effects of tyrosine intake in healthy human adults.

    PubMed

    Hase, Adrian; Jung, Sophie E; aan het Rot, Marije

    2015-06-01

    The amino acid tyrosine is the precursor to the catecholamine neurotransmitters dopamine and norepinephrine. Increasing tyrosine uptake may positively influence catecholamine-related psychological functioning. We conducted a systematic review to examine the effects of tyrosine on behavior and cognition. Fifteen studies were reviewed. All studies except one involved tyrosine loading during a single test session. In most behavioral studies, there were no significant effects of tyrosine on exercise performance. In contrast, cognitive studies employing neuropsychological measures found that tyrosine loading acutely counteracts decrements in working memory and information processing that are induced by demanding situational conditions such as extreme weather or cognitive load. The buffering effects of tyrosine on cognition may be explained by tyrosine's ability to neutralize depleted brain catecholamine levels. There is evidence that tyrosine may benefit healthy individuals exposed to demanding situational conditions. For future research we recommend moving from studying the acute effects of a single tyrosine load in small samples to studying the behavioral and cognitive effects of tyrosine in larger groups over multiple weeks.

  8. Bulkiness or aromatic nature of tyrosine-143 of actin is important for the weak binding between F-actin and myosin-ADP-phosphate

    SciTech Connect

    Gomibuchi, Yuki; Uyeda, Taro Q.P.; Wakabayashi, Takeyuki

    2013-11-29

    Highlights: •The effect of mutation of Tyr143 that becomes more exposed on assembly was examined. •Mutation of tyrosine-143 of Dictyostelium actin changed actin polymerizability. •The bulkiness or aromatic nature of Tyr143 is important for the weak binding. •The weak interaction between myosin and actin strengthened by Tyr143Trp mutation. -- Abstract: Actin filaments (F-actin) interact with myosin and activate its ATPase to support force generation. By comparing crystal structures of G-actin and the quasi-atomic model of F-actin based on high-resolution cryo-electron microscopy, the tyrosine-143 was found to be exposed more than 60 Å{sup 2} to the solvent in F-actin. Because tyrosine-143 flanks the hydrophobic cleft near the hydrophobic helix that binds to myosin, the mutant actins, of which the tyrosine-143 was replaced with tryptophan, phenylalanine, or isoleucine, were generated using the Dictyostelium expression system. It polymerized significantly poorly when induced by NaCl, but almost normally by KCl. In the presence of phalloidin and KCl, the extents of the polymerization of all the mutant actins were comparable to that of the wild-type actin so that the actin-activated myosin ATPase activity could be reliably compared. The affinity of skeletal heavy meromyosin to F-actin and the maximum ATPase activity (V{sub max}) were estimated by a double reciprocal plot. The Tyr143Trp-actin showed the higher affinity (smaller K{sub app}) than that of the wild-type actin, with the V{sub max} being almost unchanged. The K{sub app} and V{sub max} of the Tyr143Phe-actin were similar to those of the wild-type actin. However, the activation by Tyr143Ile-actin was much smaller than the wild-type actin and the accurate determination of K{sub app} was difficult. Comparison of the myosin ATPase activated by the various mutant actins at the same concentration of F-actin showed that the extent of activation correlates well with the solvent-accessible surface areas (ASA

  9. Phenylalanine Is Required to Promote Specific Developmental Responses and Prevents Cellular Damage in Response to Ultraviolet Light in Soybean (Glycine max) during the Seed-to-Seedling Transition

    PubMed Central

    Sullivan, Joe H.; Muhammad, DurreShahwar; Warpeha, Katherine M.

    2014-01-01

    UV-radiation elicits a suite of developmental (photomorphogenic) and protective responses in plants, but responses early post-germination have received little attention, particularly in intensively bred plants of economic importance. We examined germination, hypocotyl elongation, leaf pubescence and subcellular responses of germinating and/or etiolated soybean (Glycine max (L.) Merr.) seedlings in response to treatment with discrete wavelengths of UV-A or UV-B radiation. We demonstrate differential responses of germinating/young soybean seedlings to a range of UV wavelengths that indicate unique signal transduction mechanisms regulate UV-initiated responses. We have investigated how phenylalanine, a key substrate in the phenylpropanoid pathway, may be involved in these responses. Pubescence may be a key location for phenylalanine-derived protective compounds, as UV-B irradiation increased pubescence and accumulation of UV-absorbing compounds within primary leaf pubescence, visualized by microscopy and absorbance spectra. Mass spectrometry analysis of pubescence indicated that sinapic esters accumulate in the UV-irradiated hairs compared to unirradiated primary leaf tissue. Deleterious effects of some UV-B wavelengths on germination and seedling responses were reduced or entirely prevented by inclusion of phenylalanine in the growth media. Key effects of phenylalanine were not duplicated by tyrosine or tryptophan or sucrose, nor is the specificity of response due to the absorbance of phenylalanine itself. These results suggest that in the seed-to-seedling transition, phenylalanine may be a limiting factor in the development of initial mechanisms of UV protection in the developing leaf. PMID:25549094

  10. Phenylalanine is required to promote specific developmental responses and prevents cellular damage in response to ultraviolet light in soybean (Glycine max) during the seed-to-seedling transition.

    PubMed

    Sullivan, Joe H; Muhammad, DurreShahwar; Warpeha, Katherine M

    2014-01-01

    UV-radiation elicits a suite of developmental (photomorphogenic) and protective responses in plants, but responses early post-germination have received little attention, particularly in intensively bred plants of economic importance. We examined germination, hypocotyl elongation, leaf pubescence and subcellular responses of germinating and/or etiolated soybean (Glycine max (L.) Merr.) seedlings in response to treatment with discrete wavelengths of UV-A or UV-B radiation. We demonstrate differential responses of germinating/young soybean seedlings to a range of UV wavelengths that indicate unique signal transduction mechanisms regulate UV-initiated responses. We have investigated how phenylalanine, a key substrate in the phenylpropanoid pathway, may be involved in these responses. Pubescence may be a key location for phenylalanine-derived protective compounds, as UV-B irradiation increased pubescence and accumulation of UV-absorbing compounds within primary leaf pubescence, visualized by microscopy and absorbance spectra. Mass spectrometry analysis of pubescence indicated that sinapic esters accumulate in the UV-irradiated hairs compared to unirradiated primary leaf tissue. Deleterious effects of some UV-B wavelengths on germination and seedling responses were reduced or entirely prevented by inclusion of phenylalanine in the growth media. Key effects of phenylalanine were not duplicated by tyrosine or tryptophan or sucrose, nor is the specificity of response due to the absorbance of phenylalanine itself. These results suggest that in the seed-to-seedling transition, phenylalanine may be a limiting factor in the development of initial mechanisms of UV protection in the developing leaf. PMID:25549094

  11. Genomic Characterization of Phenylalanine Ammonia Lyase Gene in Buckwheat.

    PubMed

    Thiyagarajan, Karthikeyan; Vitali, Fabio; Tolaini, Valentina; Galeffi, Patrizia; Cantale, Cristina; Vikram, Prashant; Singh, Sukhwinder; De Rossi, Patrizia; Nobili, Chiara; Procacci, Silvia; Del Fiore, Antonella; Antonini, Alessandro; Presenti, Ombretta; Brunori, Andrea

    2016-01-01

    Phenylalanine Ammonia Lyase (PAL) gene which plays a key role in bio-synthesis of medicinally important compounds, Rutin/quercetin was sequence characterized for its efficient genomics application. These compounds possessing anti-diabetic and anti-cancer properties and are predominantly produced by Fagopyrum spp. In the present study, PAL gene was sequenced from three Fagopyrum spp. (F. tataricum, F. esculentum and F. dibotrys) and showed the presence of three SNPs and four insertion/deletions at intra and inter specific level. Among them, the potential SNP (position 949th bp G>C) with Parsimony Informative Site was selected and successfully utilised to individuate the zygosity/allelic variation of 16 F. tataricum varieties. Insertion mutations were identified in coding region, which resulted the change of a stretch of 39 amino acids on the putative protein. Our Study revealed that autogamous species (F. tataricum) has lower frequency of observed SNPs as compared to allogamous species (F. dibotrys and F. esculentum). The identified SNPs in F. tataricum didn't result to amino acid change, while in other two species it caused both conservative and non-conservative variations. Consistent pattern of SNPs across the species revealed their phylogenetic importance. We found two groups of F. tataricum and one of them was closely related with F. dibotrys. Sequence characterization information of PAL gene reported in present investigation can be utilized in genetic improvement of buckwheat in reference to its medicinal value. PMID:26990297

  12. Genomic Characterization of Phenylalanine Ammonia Lyase Gene in Buckwheat

    PubMed Central

    Thiyagarajan, Karthikeyan; Vitali, Fabio; Tolaini, Valentina; Galeffi, Patrizia; Cantale, Cristina; Vikram, Prashant; Singh, Sukhwinder; De Rossi, Patrizia; Nobili, Chiara; Procacci, Silvia; Del Fiore, Antonella; Antonini, Alessandro; Presenti, Ombretta; Brunori, Andrea

    2016-01-01

    Phenylalanine Ammonia Lyase (PAL) gene which plays a key role in bio-synthesis of medicinally important compounds, Rutin/quercetin was sequence characterized for its efficient genomics application. These compounds possessing anti-diabetic and anti-cancer properties and are predominantly produced by Fagopyrum spp. In the present study, PAL gene was sequenced from three Fagopyrum spp. (F. tataricum, F. esculentum and F. dibotrys) and showed the presence of three SNPs and four insertion/deletions at intra and inter specific level. Among them, the potential SNP (position 949th bp G>C) with Parsimony Informative Site was selected and successfully utilised to individuate the zygosity/allelic variation of 16 F. tataricum varieties. Insertion mutations were identified in coding region, which resulted the change of a stretch of 39 amino acids on the putative protein. Our Study revealed that autogamous species (F. tataricum) has lower frequency of observed SNPs as compared to allogamous species (F. dibotrys and F. esculentum). The identified SNPs in F. tataricum didn’t result to amino acid change, while in other two species it caused both conservative and non-conservative variations. Consistent pattern of SNPs across the species revealed their phylogenetic importance. We found two groups of F. tataricum and one of them was closely related with F. dibotrys. Sequence characterization information of PAL gene reported in present investigation can be utilized in genetic improvement of buckwheat in reference to its medicinal value. PMID:26990297

  13. Targeting of AIDS related encephalopathy using phenylalanine anchored lipidic nanocarrier.

    PubMed

    Vyas, Anil; Jain, Ankit; Hurkat, Pooja; Jain, Ashish; Jain, Sanjay K

    2015-07-01

    Transport of the anti-HIV agents across the blood-brain barrier (BBB) is a prerequisite to treat acquired immunodeficiency syndrome (AIDS) related encephalopathy. In the present study, we explored facilitated transport of efavirenz (EFV, a non-nucleoside reverse transcriptase inhibitor) across BBB using phenylalanine anchored solid lipid nanoparticles (PA-SLN). PA (amino acid micro-nutrient) was used as a ligand which facilitated carrier mediated transport (CMT) via l-amino acid transporter i.e. LAT1 to traverse BBB. PA was coupled to SLN via amide linkage using carbodiimide chemistry and coupling was confirmed by comparative infrared spectroscopic analysis. SLNs (SLN and PA-SLN) were nanometric in size (around 150nm) and possessed good entrapment efficiency (around 70%). In vitro drug release revealed controlled release pattern for more than 24h. In vivo studies showed 2-3-folds and 7-8-folds accumulation of PA-SLN in brain as compared to SLN and EFV, respectively. Further, transcytosis studies confirmed capability of PA-SLN to cross BBB i.e. 10-fold higher transcytosis potential as compared to EFV. Fluorescence microscopic imaging reassured enhanced brain localization of PA-SLN. Thus, PA-SLN improved the EFV bioavailability and maintained therapeutic levels in the brain for an extended period of time that can result in significant eradication of the viral load therein. Such nutrient mediated drug targeting could bring forth advances in biocompatible and biodegradable drug delivery systems. PMID:25988279

  14. Genomic Characterization of Phenylalanine Ammonia Lyase Gene in Buckwheat.

    PubMed

    Thiyagarajan, Karthikeyan; Vitali, Fabio; Tolaini, Valentina; Galeffi, Patrizia; Cantale, Cristina; Vikram, Prashant; Singh, Sukhwinder; De Rossi, Patrizia; Nobili, Chiara; Procacci, Silvia; Del Fiore, Antonella; Antonini, Alessandro; Presenti, Ombretta; Brunori, Andrea

    2016-01-01

    Phenylalanine Ammonia Lyase (PAL) gene which plays a key role in bio-synthesis of medicinally important compounds, Rutin/quercetin was sequence characterized for its efficient genomics application. These compounds possessing anti-diabetic and anti-cancer properties and are predominantly produced by Fagopyrum spp. In the present study, PAL gene was sequenced from three Fagopyrum spp. (F. tataricum, F. esculentum and F. dibotrys) and showed the presence of three SNPs and four insertion/deletions at intra and inter specific level. Among them, the potential SNP (position 949th bp G>C) with Parsimony Informative Site was selected and successfully utilised to individuate the zygosity/allelic variation of 16 F. tataricum varieties. Insertion mutations were identified in coding region, which resulted the change of a stretch of 39 amino acids on the putative protein. Our Study revealed that autogamous species (F. tataricum) has lower frequency of observed SNPs as compared to allogamous species (F. dibotrys and F. esculentum). The identified SNPs in F. tataricum didn't result to amino acid change, while in other two species it caused both conservative and non-conservative variations. Consistent pattern of SNPs across the species revealed their phylogenetic importance. We found two groups of F. tataricum and one of them was closely related with F. dibotrys. Sequence characterization information of PAL gene reported in present investigation can be utilized in genetic improvement of buckwheat in reference to its medicinal value.

  15. AMINO ACIDS AND HEMOGLOBIN PRODUCTION IN ANEMIA

    PubMed Central

    Whipple, G. H.; Robscheit-Robbins, F. S.

    1940-01-01

    Certain individual amino acids when given to standard anemic dogs cause an increase in new hemoglobin production. Occasional negative experiments are recorded. Glycine, glutamic acid, aspartic acid, cystine, histidine, phenylalanine, and proline when given in 1 gm. doses daily for 2 weeks, increase hemoglobin output on the average 23 to 25 gm. above the control level. This reaction amounts to 25 to 30 per cent of the new hemoglobin produced by the feeding of 300 gm. liver daily for 2 weeks—a standard liver test. Alanine, valine, isoleucine, and arginine in the same dosage increase the hemoglobin output on the average 13 to 17 gm. per 2 weeks over the control level. Leucine, methionine, lysine, tryptophane, and tyrosine fall in a middle group with hemoglobin output of about 20 gm. Isovaleric acid, β-hydroxybutyric acid, glutaric acid, and asparagine have shown positive effects and the butyrate is unusually potent for hemoglobin production (Table 2). The isomeric and dl-synthetic forms of the amino acids are as effectively utilized in this reaction as are the natural forms. PMID:19870982

  16. Conservative Tryptophan Mutants of the Protein Tyrosine Phosphatase YopH Exhibit Impaired WPD-Loop Function and Crystallize with Divanadate Esters in Their Active Sites

    PubMed Central

    Moise, Gwendolyn; Gallup, Nathan M.; Alexandrova, Anastassia N.; Hengge, Alvan C.; Johnson, Sean J.

    2016-01-01

    Catalysis in protein tyrosine phosphatases (PTPs) involves movement of a protein loop called the WPD loop that brings a conserved aspartic acid into the active site to function as a general acid. Mutation of the tryptophan in the WPD loop of the PTP YopH to any other residue with a planar, aromatic side chain (phenylalanine, tyrosine, or histidine) disables general acid catalysis. Crystal structures reveal these conservative mutations leave this critical loop in a catalytically unproductive, quasi-open position. Although the loop positions in crystal structures are similar for all three conservative mutants, the reasons inhibiting normal loop closure differ for each mutant. In the W354F and W354Y mutants, steric clashes result from six-membered rings occupying the position of the five-membered ring of the native indole side chain. The histidine mutant dysfunction results from new hydrogen bonds stabilizing the unproductive position. The results demonstrate how even modest modifications can disrupt catalytically important protein dynamics. Crystallization of all the catalytically compromised mutants in the presence of vanadate gave rise to vanadate dimers at the active site. In W354Y and W354H, a divanadate ester with glycerol is observed. Such species have precedence in solution and are known from the small molecule crystal database. Such species have not been observed in the active site of a phosphatase, as a functional phosphatase would rapidly catalyze their decomposition. The compromised functionality of the mutants allows the trapping of species that undoubtedly form in solution and are capable of binding at the active sites of PTPs, and, presumably, other phosphatases. In addition to monomeric vanadate, such higher-order vanadium-based molecules are likely involved in the interaction of vanadate with PTPs in solution. PMID:26445170

  17. Conservative tryptophan mutants of the protein tyrosine phosphatase YopH exhibit impaired WPD-loop function and crystallize with divanadate esters in their active sites.

    PubMed

    Moise, Gwendolyn; Gallup, Nathan M; Alexandrova, Anastassia N; Hengge, Alvan C; Johnson, Sean J

    2015-10-27

    Catalysis in protein tyrosine phosphatases (PTPs) involves movement of a protein loop called the WPD loop that brings a conserved aspartic acid into the active site to function as a general acid. Mutation of the tryptophan in the WPD loop of the PTP YopH to any other residue with a planar, aromatic side chain (phenylalanine, tyrosine, or histidine) disables general acid catalysis. Crystal structures reveal these conservative mutations leave this critical loop in a catalytically unproductive, quasi-open position. Although the loop positions in crystal structures are similar for all three conservative mutants, the reasons inhibiting normal loop closure differ for each mutant. In the W354F and W354Y mutants, steric clashes result from six-membered rings occupying the position of the five-membered ring of the native indole side chain. The histidine mutant dysfunction results from new hydrogen bonds stabilizing the unproductive position. The results demonstrate how even modest modifications can disrupt catalytically important protein dynamics. Crystallization of all the catalytically compromised mutants in the presence of vanadate gave rise to vanadate dimers at the active site. In W354Y and W354H, a divanadate ester with glycerol is observed. Such species have precedence in solution and are known from the small molecule crystal database. Such species have not been observed in the active site of a phosphatase, as a functional phosphatase would rapidly catalyze their decomposition. The compromised functionality of the mutants allows the trapping of species that undoubtedly form in solution and are capable of binding at the active sites of PTPs, and, presumably, other phosphatases. In addition to monomeric vanadate, such higher-order vanadium-based molecules are likely involved in the interaction of vanadate with PTPs in solution. PMID:26445170

  18. Arabidopsis thaliana NIP7;1: An Anther-Specific Boric Acid Transporter of the Aquaporin Superfamily Regulated by an Unusual Tyrosine in Helix 2 of the Transport Pore

    SciTech Connect

    Li, Tian; Choi, Won-Gyu; Baudry, Jerome Y; Roberts, Daniel M

    2011-01-01

    Plant nodulin-26 intrinsic proteins (NIPs) are members of the aquaporin superfamily that serve as multifunctional transporters of uncharged metabolites. In Arabidopsis thaliana, a specific NIP pore subclass, known as the NIP II proteins, is represented by AtNIP5;1 and AtNIP6;1, which encode channel proteins expressed in roots and leaf nodes, respectively, that participate in the transport of the critical cell wall nutrient boric acid. Modeling of the protein encoded by the AtNIP7;1 gene shows that it is a third member of the NIP II pore subclass in Arabidopsis. However, unlike AtNIP5;1 and AtNIP6;1 proteins, which form constitutive boric acid channels, AtNIP7;1 forms a channel with an extremely low intrinsic boric acid transport activity. Molecular modeling and molecular dynamics simulations of AtNIP7;1 suggest that a conserved tyrosine residue (Tyr81) located in transmembrane helix 2 adjacent to the aromatic arginine (ar/R) pore selectivity region stabilizes a closed pore conformation through interaction with the canonical Arg220 in ar/R region. Substitution of Tyr81 with a Cys residue, characteristic of established NIP boric acid channels, results in opening of the AtNIP7;1 pore that acquires a robust, transport activity for boric acid as well as other NIP II test solutes (glycerol and urea). Substitution of a Phe for Tyr81 also opens the channel, supporting the prediction from MD simulations that hydrogen bond interaction between the Tyr81 phenol group and the ar/R Arg may contribute to the stabilization of a closed pore state. Expression analyses show that AtNIP7;1 is selectively expressed in developing anther tissues of young floral buds of A. thaliana, principally in developing pollen grains of stage 9 11 anthers. Because boric acid is both an essential nutrient as well as a toxic compound at high concentrations, it is proposed that Tyr81 modulates transport and may provide an additional level of regulation for this transporter in male gametophyte development.

  19. Arabidopsis thaliana NIP7;1: an anther-specific boric acid transporter of the aquaporin superfamily regulated by an unusual tyrosine in helix 2 of the transport pore.

    PubMed

    Li, Tian; Choi, Won-Gyu; Wallace, Ian S; Baudry, Jerome; Roberts, Daniel M

    2011-08-01

    Plant nodulin-26 intrinsic proteins (NIPs) are members of the aquaporin superfamily that serve as multifunctional transporters of uncharged metabolites. In Arabidopsis thaliana, a specific NIP pore subclass, known as the NIP II proteins, is represented by AtNIP5;1 and AtNIP6;1, which encode channel proteins expressed in roots and leaf nodes, respectively, that participate in the transport of the critical cell wall nutrient boric acid. Modeling of the protein encoded by the AtNIP7;1 gene shows that it is a third member of the NIP II pore subclass in Arabidopsis. However, unlike AtNIP5;1 and AtNIP6;1 proteins, which form constitutive boric acid channels, AtNIP7;1 forms a channel with an extremely low intrinsic boric acid transport activity. Molecular modeling and molecular dynamics simulations of AtNIP7;1 suggest that a conserved tyrosine residue (Tyr81) located in transmembrane helix 2 adjacent to the aromatic arginine (ar/R) pore selectivity region stabilizes a closed pore conformation through interaction with the canonical Arg220 in ar/R region. Substitution of Tyr81 with a Cys residue, characteristic of established NIP boric acid channels, results in opening of the AtNIP7;1 pore that acquires a robust, transport activity for boric acid as well as other NIP II test solutes (glycerol and urea). Substitution of a Phe for Tyr81 also opens the channel, supporting the prediction from MD simulations that hydrogen bond interaction between the Tyr81 phenol group and the ar/R Arg may contribute to the stabilization of a closed pore state. Expression analyses show that AtNIP7;1 is selectively expressed in developing anther tissues of young floral buds of A. thaliana, principally in developing pollen grains of stage 9-11 anthers. Because boric acid is both an essential nutrient as well as a toxic compound at high concentrations, it is proposed that Tyr81 modulates transport and may provide an additional level of regulation for this transporter in male gametophyte development.

  20. Fasting and postprandial phenylalanine and leucine kinetics in liver cirrhosis.

    PubMed

    Tessari, P; Inchiostro, S; Barazzoni, R; Zanetti, M; Orlando, R; Biolo, G; Sergi, G; Pino, A; Tiengo, A

    1994-07-01

    To investigate body protein turnover and the pathogenesis of increased concentration of plasma phenylalanine in liver cirrhosis, we have studied phenylalanine and leucine kinetics in cirrhotic (diabetic and nondiabetic) patients, and in normal subjects, both in the postabsorptive state and during a mixed meal, using combined intravenous and oral isotope infusions. Postabsorptive phenylalanine concentration and whole body rate of appearance (Ra) were approximately 40% greater (P < 0.05) in patients than in controls. Leucine concentrations were comparable, but intracellular leucine Ra was also increased (P < 0.05), suggesting increased whole body protein breakdown. Postprandial phenylalanine Ra was also greater (P < 0.05) in the patients. This difference was due to a diminished fractional splanchnic uptake of the dietary phenylalanine (approximately 40% lower in the cirrhotics vs. controls, P < or = 0.05). Postprandial leucine Ra was also increased in the patients, but splanchnic uptake of dietary leucine was normal. Thus both increased body protein breakdown and decreased splanchnic extraction of dietary phenylalanine can account for the increased phenylalanine concentrations in liver cirrhosis.

  1. Effects of phenylalanine on reproductive performance and teratogenesis in mice.

    PubMed

    Oliveira, R J; Pesarini, J R; Mauro, M O; Fronza, L S; Victorelli, S G; Cantero, W B; Sena, M C; Antoniolli, A C M B

    2014-07-25

    We evaluated the effects of phenylalanine on reproductive performance and teratogenesis in mice, as well as we assessed its protective effect in mice treated with an acute dose of cyclophosphamide. Animals were divided into 6 experimental groups (females N = 15/group, males N = 5/group): G1, the negative control group, phosphate-buffered saline; G2, the positive control group, 35 mg cyclophosphamide/kg body weight (b.w.); G3 and G4 received phenylalanine at doses of 150 and 300 mg/ kg b.w., respectively; G5 and G6 received phenylalanine at doses of 150 and 300 mg/kg b.w. co-administered with cyclophosphamide at a dose of 35 mg/kg b.w., respectively. Pregnant mice received phenylalanine from 8-12 days of pregnancy and cyclophosphamide on the 10th day of treatment or the respective vehicles. In animals treated with cyclophosphamide, offspring fetal weight significantly decreased. The G5 and G6 groups, which received cyclophosphamide co-administered with phenylalanine, showed a smaller reduction in weight. Based on this analysis, the offspring from groups G2, G5, and G6 showed low weight due to pregnancy age. Moreover, at the doses used, phenylalanine did not interfere with embryo-fetal development. However, further studies are necessary to increase the understanding of the effects of phenylalanine on mouse reproductive performance and teratogenesis.

  2. Characterization of the Tyrosine Kinase-Regulated Proteome in Breast Cancer by Combined use of RNA interference (RNAi) and Stable Isotope Labeling with Amino Acids in Cell Culture (SILAC) Quantitative Proteomics*

    PubMed Central

    Stebbing, Justin; Zhang, Hua; Xu, Yichen; Grothey, Arnhild; Ajuh, Paul; Angelopoulos, Nicos; Giamas, Georgios

    2015-01-01

    Tyrosine kinases (TKs) are central regulators in cellular activities and perturbations of TK signaling contribute to oncogenesis. However, less than half of the TKs have been thoroughly studied and a global functional analysis of their proteomic portrait is lacking. Here we conducted a combined approach of RNA interference (RNAi) and stable isotope labeling with amino acids in cell culture (SILAC)-based quantitative proteomics to decode the TK-regulated proteome and associated signaling dynamics. As a result, a broad proteomic repertoire modulated by TKs was revealed, upon silencing of the 65 TKs expressed in MCF7 breast cancer cells. This yielded 10 new distinctive TK clusters according to similarity in TK-regulated proteome, each characterized by a unique signaling signature in contrast to previous classifications. We provide functional analyses and identify critical pathways for each cluster based on their common downstream targets. Analysis of different breast cancer subtypes showed distinct correlations of each cluster with clinical outcome. From the significantly up- and down-regulated proteins, we identified a number of markers of drug sensitivity and resistance. These data supports the role of TKs in regulating major aspects of cellular activity, but also reveals redundancy in signaling, explaining why kinase inhibitors alone often fail to achieve their clinical aims. The TK-SILACepedia provides a comprehensive resource for studying the global function of TKs in cancer. PMID:26089344

  3. Characterization of the Tyrosine Kinase-Regulated Proteome in Breast Cancer by Combined use of RNA interference (RNAi) and Stable Isotope Labeling with Amino Acids in Cell Culture (SILAC) Quantitative Proteomics.

    PubMed

    Stebbing, Justin; Zhang, Hua; Xu, Yichen; Grothey, Arnhild; Ajuh, Paul; Angelopoulos, Nicos; Giamas, Georgios

    2015-09-01

    Tyrosine kinases (TKs) are central regulators in cellular activities and perturbations of TK signaling contribute to oncogenesis. However, less than half of the TKs have been thoroughly studied and a global functional analysis of their proteomic portrait is lacking. Here we conducted a combined approach of RNA interference (RNAi) and stable isotope labeling with amino acids in cell culture (SILAC)-based quantitative proteomics to decode the TK-regulated proteome and associated signaling dynamics. As a result, a broad proteomic repertoire modulated by TKs was revealed, upon silencing of the 65 TKs expressed in MCF7 breast cancer cells. This yielded 10 new distinctive TK clusters according to similarity in TK-regulated proteome, each characterized by a unique signaling signature in contrast to previous classifications. We provide functional analyses and identify critical pathways for each cluster based on their common downstream targets. Analysis of different breast cancer subtypes showed distinct correlations of each cluster with clinical outcome. From the significantly up- and down-regulated proteins, we identified a number of markers of drug sensitivity and resistance. These data supports the role of TKs in regulating major aspects of cellular activity, but also reveals redundancy in signaling, explaining why kinase inhibitors alone often fail to achieve their clinical aims. The TK-SILACepedia provides a comprehensive resource for studying the global function of TKs in cancer. PMID:26089344

  4. Simultaneous determination of cysteine, uric acid and tyrosine using Au-nanoparticles/poly(E)-4-(p-tolyldiazenyl)benzene-1,2,3-triol film modified glassy carbon electrode.

    PubMed

    Taei, M; Hasanpour, F; Salavati, H; Banitaba, S H; Kazemi, F

    2016-02-01

    A novel Au nanoparticles/poly(E)-4-(p-tolyldiazenyl)benzene-1,2,3-triol (AuNPs/PTAT) film modified glassy carbon electrode (AuNPs/PTAT/GCE) was fabricated for the simultaneous determination of three antioxidants named, cysteine (Cys), uric acid (UA) and tyrosine (Tyr). The bare glassy carbon electrode (GCE) fails to separate the oxidation peak potentials of these molecules, while PTAT film modified electrode can resolve them. Electrochemical impedance spectroscopy (EIS) study indicates that the charge transfer resistance of bare electrode increased as (E)-4-(p-tolyldiazenyl)benzene-1,2,3-triol was electropolymerized at the bare electrode. Furthermore, EIS exhibits enhancement of electron transfer kinetics between analytes and electrode after electrodeposition of Au nanoparticles. Differential pulse voltammetry results show that the electrocatalytic current increases linearly in the ranges of 2-540μmolL(-1) for Cys, 5-820μmolL(-1) for UA and 10-560μmolL(-1) for Tyr with detection limits (S/N=3) of 0.04μmolL(-1), 0.1μmolL(-1) and 2μmolL(-1) for Cys, UA and Tyr, respectively. The proposed method was successfully applied for simultaneous determination of Cys, UA and Tyr in human urine samples.

  5. Biodistribution of phenylboric acid derivative entrapped lipiodol and 4-borono-2-18F-fluoro-L-phenylalanine-fructose in GP7TB liver tumor bearing rats for BNCT.

    PubMed

    Liao, A H; Chou, F I; Kuo, Y C; Chen, H W; Kai, J J; Chang, C W; Chen, F D; Hwang, J J

    2010-03-01

    A new phenylboric acid derivative entrapped lipiodol (PBAD-lipiodol) was developed as a boron carrier for the boron neutron capture therapy (BNCT) of hepatoma in Taiwan. The biodistribution of both PBAD-lipiodol and BPA-fructose was assayed in GP7TB hepatoma-bearing rat model. The highest uptake of PBAD-lipiodol was found at 2h post injection. The application of BNCT for the hepatoma treatment in tumor-bearing rats is suggested to be 2-4h post PBAD-lipiodol injection.

  6. Tyrosine phosphorylation and bacterial virulence

    PubMed Central

    Whitmore, Sarah E; Lamont, Richard J

    2012-01-01

    Protein phosphorylation on tyrosine has emerged as a key device in the control of numerous cellular functions in bacteria. In this article, we review the structure and function of bacterial tyrosine kinases and phosphatases. Phosphorylation is catalyzed by autophosphorylating adenosine triphosphate-dependent enzymes (bacterial tyrosine (BY) kinases) that are characterized by the presence of Walker motifs. The reverse reaction is catalyzed by three classes of enzymes: the eukaryotic-like phosphatases (PTPs) and dual-specific phosphatases; the low molecular weight protein-tyrosine phosphatases (LMW-PTPs); and the polymerase–histidinol phosphatases (PHP). Many BY kinases and tyrosine phosphatases can utilize host cell proteins as substrates, thereby contributing to bacterial pathogenicity. Bacterial tyrosine phosphorylation/dephosphorylation is also involved in biofilm formation and community development. The Porphyromonas gingivalis tyrosine phosphatase Ltp1 is involved in a restraint pathway that regulates heterotypic community development with Streptococcus gordonii. Ltp1 is upregulated by contact with S. gordonii and Ltp1 activity controls adhesin expression and levels of the interspecies signal AI-2. PMID:22388693

  7. Changes in free amino acids and polyamine levels in Satsuma leaves in response to Asian citrus psyllid infestation and water stress.

    PubMed

    Malik, Nasir S A; Perez, Jose L; Kunta, Madhurababu; Patt, Joseph M; Mangan, Robert L

    2014-12-01

    The effects of biotic and abiotic stresses on changes in amino acids and polyamine levels in Satsuma orange (Citrus unshiu; cultivar Owari) leaves were investigated. Asian citrus psyllids Diaphorina citri (Kuwayama) (ACP) infestation was used to induce biotic stress while a water deficit was imposed to induce abiotic stress. Potted trees were infested by placing 50 psyllids on 3 citrus leaves enclosed in nylon mesh bags for 5 d. A parallel set of plants were kept water stressed by maintaining the soil at 20% water holding capacity for 5 d. Levels of total free amino acids were higher in water stressed and ACP infested leaves. Polyamine putrescine increased in infested leaves but not in water stressed leaves. Proline was the most abundant amino acid and its levels significantly increased by both biotic and abiotic stresses. Proline levels in infested leaves were significantly higher than the water stressed leaves. Histidine, methionine, asparagine, arginine, serine, and leucine levels also increased significantly in infested leaves, but in water stressed leaves only leucine, methionine, and threonine increased. Levels of amino acids, such as tyrosine, isoleucine, phenylalanine, glutamic acid, and alanine, declined in infested leaves. Under water stress asparagine, phenylalanine, serine, and histidine also declined compared to controls. This indicates that while proteolysis occurred under both stresses, metabolic conversion of amino acids was different under the two stresses. In ACP infested leaves some amino acids may be used as feeding material and/or converted into secondary metabolites for defense. PMID:24178691

  8. Changes in free amino acids and polyamine levels in Satsuma leaves in response to Asian citrus psyllid infestation and water stress.

    PubMed

    Malik, Nasir S A; Perez, Jose L; Kunta, Madhurababu; Patt, Joseph M; Mangan, Robert L

    2014-12-01

    The effects of biotic and abiotic stresses on changes in amino acids and polyamine levels in Satsuma orange (Citrus unshiu; cultivar Owari) leaves were investigated. Asian citrus psyllids Diaphorina citri (Kuwayama) (ACP) infestation was used to induce biotic stress while a water deficit was imposed to induce abiotic stress. Potted trees were infested by placing 50 psyllids on 3 citrus leaves enclosed in nylon mesh bags for 5 d. A parallel set of plants were kept water stressed by maintaining the soil at 20% water holding capacity for 5 d. Levels of total free amino acids were higher in water stressed and ACP infested leaves. Polyamine putrescine increased in infested leaves but not in water stressed leaves. Proline was the most abundant amino acid and its levels significantly increased by both biotic and abiotic stresses. Proline levels in infested leaves were significantly higher than the water stressed leaves. Histidine, methionine, asparagine, arginine, serine, and leucine levels also increased significantly in infested leaves, but in water stressed leaves only leucine, methionine, and threonine increased. Levels of amino acids, such as tyrosine, isoleucine, phenylalanine, glutamic acid, and alanine, declined in infested leaves. Under water stress asparagine, phenylalanine, serine, and histidine also declined compared to controls. This indicates that while proteolysis occurred under both stresses, metabolic conversion of amino acids was different under the two stresses. In ACP infested leaves some amino acids may be used as feeding material and/or converted into secondary metabolites for defense.

  9. Phenylalanine ab initio models for the simulation of skin natural moisturizing factor.

    PubMed

    Carvalho, B G; Raniero, L J; Martin, A A; Favero, P P

    2013-04-01

    In this study, we evaluated models that can be used to simulate amino acids in biological environments via density functional theory (DFT). The goal was to obtain realistic representations that combine computational economy and result quality when compared to experimental data. We increased the complexity of the models by using a model of an amino acid in a vacuum, followed by a water-solvated amino acid model. To consider pH variation, we simulated zwitterionic and nonionic amino acid configurations. The amino acid chosen for testing was phenylalanine, an aromatic amino acid present in high concentrations in the natural moisturizing factor of skin that plays a fundamental role in ultraviolet protection and vitiligo disease. To validate the models, vibrational modes and electronic properties were calculated and compared to experimental results.

  10. Phenylalanine ab initio models for the simulation of skin natural moisturizing factor

    NASA Astrophysics Data System (ADS)

    Carvalho, B. G.; Raniero, L. J.; Martin, A. A.; Favero, P. P.

    2013-04-01

    In this study, we evaluated models that can be used to simulate amino acids in biological environments via density functional theory (DFT). The goal was to obtain realistic representations that combine computational economy and result quality when compared to experimental data. We increased the complexity of the models by using a model of an amino acid in a vacuum, followed by a water-solvated amino acid model. To consider pH variation, we simulated zwitterionic and nonionic amino acid configurations. The amino acid chosen for testing was phenylalanine, an aromatic amino acid present in high concentrations in the natural moisturizing factor of skin that plays a fundamental role in ultraviolet protection and vitiligo disease. To validate the models, vibrational modes and electronic properties were calculated and compared to experimental results.

  11. Effective atomic numbers and electron densities of bacteriorhodopsin and its comprising amino acids in the energy range 1 keV-100 GeV

    NASA Astrophysics Data System (ADS)

    Ahmadi, Morteza; Lunscher, Nolan; Yeow, John T. W.

    2013-04-01

    Recently, there has been an interest in fabrication of X-ray sensors based on bacteriorhodopsin, a proton pump protein in cell membrane of Halobacterium salinarium. Therefore, a better understanding of interaction of X-ray photons with bacteriorhodopsin is required. We use WinXCom program to calculate the mass attenuation coefficient of bacteriorhodopsin and its comprising amino acids for photon energies from 1 keV to 100 GeV. These amino acids include alanine, arginine, asparagine, aspartic acid, glutamine, glutamic acid, glycine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine, valine, Asx1, Asx2, Glx1 and Glx2. We then use that data to calculate effective atomic number and electron densities for the same range of energy. We also emphasize on two ranges of energies (10-200 keV and 1-20 MeV) in which X-ray imaging and radiotherapy machines work.

  12. The combination of 4-anilinoquinazoline and cinnamic acid: a novel mode of binding to the epidermal growth factor receptor tyrosine kinase.

    PubMed

    Li, Dong-Dong; Lv, Peng-Cheng; Zhang, Hui; Zhang, Hong-Jia; Hou, Ya-Ping; Liu, Kai; Ye, Yong-Hao; Zhu, Hai-Liang

    2011-08-15

    A novel type of cinnamic acid quinazoline amide derivatives (20-42), which designed the combination between quinazoline as the backbone and various substituted cinnamic acid as the side chain, have been synthesized and their biological activities were evaluated within cytotoxicity assay firstly and then potent EGFR inhibitory activity. Compound 42 demonstrated the most potent inhibitory activity (IC(50)=0.94 μM for EGFR), which could be optimized as a potential EGFR inhibitor in the further study. Docking simulation was performed to position compound 42 into the EGFR active site to determine the probable binding model. Analysis of the binding conformation of 42 in active site displayed compound 42 was stabilized by hydrogen bonding interactions with Lys822, which was different from other derivatives. In the further study, Compounds 43 and 44 had been synthesized and their biological activities were also evaluated, which were the same as that we expected. Compound 43 has demonstrated significant EGFR (IC(50)=0.12 μM) and tumor growth inhibitory activity as a potential anticancer agent.

  13. Growth response of Avena sativa in amino-acids-rich soils converted from phenol-contaminated soils by Corynebacterium glutamicum.

    PubMed

    Lee, Soo Youn; Kim, Bit-Na; Choi, Yong Woo; Yoo, Kye Sang; Kim, Yang-Hoon; Min, Jiho

    2012-04-01

    The biodegradation of phenol in laboratory-contaminated soil was investigated using the Gram-positive soil bacterium Corynebacterium glutamicum. This study showed that the phenol degradation caused by C. glutamicum was greatly enhanced by the addition of 1% yeast extract. From the toxicity test using Daphnia magna, the soil did not exhibit any hazardous effects after the phenol was removed using C. glutamicum. Additionally, the treatment of the phenolcontaminated soils with C. glutamicum increased various soil amino acid compositions, such as glycine, threonine, isoleucine, alanine, valine, leucine, tyrosine, and phenylalanine. This phenomenon induced an increase in the seed germination rate and the root elongation of Avena sativa (oat). This probably reflects that increased soil amino acid composition due to C. glutamicum treatment strengthens the plant roots. Therefore, the phenol-contaminated soil was effectively converted through increased soil amino acid composition, and additionally, the phenol in the soil environment was biodegraded by C. glutamicum.

  14. Expression and Properties of the Highly Alkalophilic Phenylalanine Ammonia-Lyase of Thermophilic Rubrobacter xylanophilus

    PubMed Central

    Kovács, Klaudia; Bánóczi, Gergely; Varga, Andrea; Szabó, Izabella; Holczinger, András; Hornyánszky, Gábor; Zagyva, Imre

    2014-01-01

    The sequence of a phenylalanine ammonia-lyase (PAL; EC: 4.3.1.24) of the thermophilic and radiotolerant bacterium Rubrobacter xylanophilus (RxPAL) was identified by screening the genomes of bacteria for members of the phenylalanine ammonia-lyase family. A synthetic gene encoding the RxPAL protein was cloned and overexpressed in Escherichia coli TOP 10 in a soluble form with an N-terminal His6-tag and the recombinant RxPAL protein was purified by Ni-NTA affinity chromatography. The activity assay of RxPAL with l-phenylalanine at various pH values exhibited a local maximum at pH 8.5 and a global maximum at pH 11.5. Circular dichroism (CD) studies showed that RxPAL is associated with an extensive α-helical character (far UV CD) and two distinctive near-UV CD peaks. These structural characteristics were well preserved up to pH 11.0. The extremely high pH optimum of RxPAL can be rationalized by a three-dimensional homology model indicating possible disulfide bridges, extensive salt-bridge formation and an excess of negative electrostatic potential on the surface. Due to these properties, RxPAL may be a candidate as biocatalyst in synthetic biotransformations leading to unnatural l- or d-amino acids or as therapeutic enzyme in treatment of phenylketonuria or leukemia. PMID:24475062

  15. Changes on grape phenolic composition induced by grapevine foliar applications of phenylalanine and urea.

    PubMed

    Portu, J; López-Alfaro, I; Gómez-Alonso, S; López, R; Garde-Cerdán, T

    2015-08-01

    Grapevines may require the input of nitrogen to grow and to guarantee an appropriate grape composition. Recently there has been a growing interest in foliar fertilization, which entails a fast and efficient assimilation of the products. The aim of this work was to study the influence of foliar applications of phenylalanine and urea, at two different doses, on grape anthocyanins, flavonols, flavan-3-ols, phenolic acids, and stilbenes. All treatments were applied at veraison and one week later at doses of 0.9 and 1.5 kg N/ha. The results showed that the synthesis of phenolic compounds was favoured by foliar applications of phenylalanine and urea. The application of the lowest dose of urea was the most effective treatment, increasing the content of several anthocyanins and flavonols. Moreover, none of the foliar treatments worsened the grape phenolic composition. In conclusion, foliar application of phenylalanine and especially urea, could be an interesting management tool for improving grape quality and their health-promoting properties.

  16. Design and Characterization of Auxotrophy-Based Amino Acid Biosensors

    PubMed Central

    Bertels, Felix; Merker, Holger; Kost, Christian

    2012-01-01

    Efficient and inexpensive methods are required for the high-throughput quantification of amino acids in physiological fluids or microbial cell cultures. Here we develop an array of Escherichia coli biosensors to sensitively quantify eleven different amino acids. By using online databases, genes involved in amino acid biosynthesis were identified that – upon deletion – should render the corresponding mutant auxotrophic for one particular amino acid. This rational design strategy suggested genes involved in the biosynthesis of arginine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, threonine, tryptophan, and tyrosine as potential genetic targets. A detailed phenotypic characterization of the corresponding single-gene deletion mutants indeed confirmed that these strains could neither grow on a minimal medium lacking amino acids nor transform any other proteinogenic amino acid into the focal one. Site-specific integration of the egfp gene into the chromosome of each biosensor decreased the detection limit of the GFP-labeled cells by 30% relative to turbidometric measurements. Finally, using the biosensors to determine the amino acid concentration in the supernatants of two amino acid overproducing E. coli strains (i.e. ΔhisL and ΔtdcC) both turbidometrically and via GFP fluorescence emission and comparing the results to conventional HPLC measurements confirmed the utility of the developed biosensor system. Taken together, our study provides not only a genotypically and phenotypically well-characterized set of publicly available amino acid biosensors, but also demonstrates the feasibility of the rational design strategy used. PMID:22829942

  17. Phenylalanine prevents acute poisoning by ochratoxina in mice.

    PubMed

    Creppy, E E; Schlegel, M; Röschenthaler, R; Dirheimer, G

    1980-07-01

    Ochratoxin-A (OT-A) a mycotoxin isolated from Aspergillus ochraceus is toxic for animals and man causing a fatal chronic kidney disease and liver damages. OT-A is a competitive inhibitor of phenylalanine in the phenylalanyl-tRNA synthetase-catalyzed reaction thus inhibiting protein synthesis. This inhibition can be reversed by phenylalanine. When injected intraperitoneally (i.p.) to mice a dose of 0.8 mg of OT-A is lethal within 24h. However, when this lethal dose is injected together with 1 mg of phenylalanine 100% of the animals survive. When phenylalanine was injected 30 min after OT-A the doses required for the survival of 92% of the animals had to be about 25 times higher. PMID:7414623

  18. Effects of phenylalanine and threonine oligopeptides on milk protein synthesis in cultured bovine mammary epithelial cells.

    PubMed

    Zhou, M M; Wu, Y M; Liu, H Y; Liu, J X

    2015-04-01

    This study was conducted to investigate the effects of phenylalanine (Phe) and threonine (Thr) oligopeptides on αs1 casein gene expression and milk protein synthesis in bovine mammary epithelial cells. Primary mammary epithelial cells were obtained from Holstein dairy cows and incubated in Dulbecco's modified Eagle's medium-F12 medium (DMEM/F12) containing lactogenic hormones (prolactin and glucocorticoids). Free Phe (117 μg/ml) was substituted partly with peptide-bound Phe (phenylalanylphenylalanine, phenylalanyl threonine, threonyl-phenylalanyl-phenylalanine) in the experimental media. After incubation with experimental medium, cells were collected for gene expression analysis and medium was collected for milk protein or amino acid determination. The results showed that peptide-bound Phe at 10% (11.7 μg/ml) significantly enhanced αs1 casein gene expression and milk protein synthesis as compared with equivalent amount of free Phe. When 10% Phe was replaced by phenylalanylphenylalanine, the disappearance of most essential amino acids increased significantly, and gene expression of peptide transporter 2 and some amino acid transporters was significantly enhanced. These results indicate that the Phe and Thr oligopeptides are important for milk protein synthesis, and peptide-bound amino acids could be utilised more efficiently in milk protein synthesis than the equivalent amount of free amino acids.

  19. Phenylalanine functionalized zwitterionic monolith for hydrophobic interaction electrochromatography.

    PubMed

    Wang, Jiabin; Jia, Wenchao; Lin, Xucong; Wu, Xiaoping; Xie, Zenghong

    2013-12-01

    A novel phenylalanine (Phe) functionalized zwitterionic monolith for hydrophobic electrochromatography was prepared by a two-step procedure involving the synthesis of glycidyl methacrylate based polymer monolith and subsequent on-column chemical modification with Phe via ring-opening reaction of epoxides. Benefitting from the hydrophobicity of both methacrylate-based matrix and aromatic group of Phe, this monolith could exhibit good hydrophobic interaction for the separation. Typical RP chromatographic behavior was observed toward various solutes. The well-controlled cathodic or anodic EOF of the prepared column could be facilely switched by altering the pH values of running buffers. The separation mechanism of this Phe functionalized zwitterionic monolith is discussed in detail. Two mixed-mode mechanisms of RP/cation exchange and RP/anion exchange could be further realized on the same monolith in different pH condition of the mobile phase. Versatile separation capabilities of neutral, basic, and acidic analytes have been successfully achieved in this zwitterionic monolith by CEC method.

  20. Tyrosine aminotransferase activity in the benzene intoxicated rat

    SciTech Connect

    Bong, M.; Michalska, A.; Laskowska-Klita, T.; Szymczyk, T.

    1985-01-01

    The toxic effect of hydrocarbon solvents on hepatic metabolism manifests itself by changes in the enzymatic pattern of blood serum. Changes in the activity of phosphatases as well as leucine aminopeptidase, glutamine aminotransferase, sorbitol dehydrogenase and ..gamma..-glutamyltransferase were observed in rats intoxicated with different fractions of benzene. Therefore it seemed reasonable to investigate the effect of benzene fraction of petroleum on cellular metabolism. The results of the present work concern the activity of tyrosine aminotransferase, the enzyme involved in catabolism of aromatic amino acid which is known to be under both hormonal and stress dependent control. Changes in tyrosine aminotransferase activity effect the level of tyrosine oxidation as well as the metabolic conversion of this amino acid into tyramine, tyroxin, adrenaline and noradrenaline.

  1. Tyrosine motifs are required for prestin basolateral membrane targeting

    PubMed Central

    Zhang, Yifan; Moeini-Naghani, Iman; Bai, JunPing; Santos-Sacchi, Joseph; Navaratnam, Dhasakumar S.

    2015-01-01

    ABSTRACT Prestin is targeted to the lateral wall of outer hair cells (OHCs) where its electromotility is critical for cochlear amplification. Using MDCK cells as a model system for polarized epithelial sorting, we demonstrate that prestin uses tyrosine residues, in a YXXΦ motif, to target the basolateral surface. Both Y520 and Y667 are important for basolateral targeting of prestin. Mutation of these residues to glutamine or alanine resulted in retention within the Golgi and delayed egress from the Golgi in Y667Q. Basolateral targeting is restored upon mutation to phenylalanine suggesting the importance of a phenol ring in the tyrosine side chain. We also demonstrate that prestin targeting to the basolateral surface is dependent on AP1B (μ1B), and that prestin uses transferrin containing early endosomes in its passage from the Golgi to the basolateral plasma membrane. The presence of AP1B (μ1B) in OHCs, and parallels between prestin targeting to the basolateral surface of OHCs and polarized epithelial cells suggest that outer hair cells resemble polarized epithelia rather than neurons in this important phenotypic measure. PMID:25596279

  2. Novel Anthraquinone-based Derivatives as Potent Inhibitors for Receptor Tyrosine Kinases

    PubMed Central

    Stasevych, M.; Zvarych, V.; Lunin, V.; Halenova, T.; Savchuk, O.; Dudchak, O.; Vovk, M.; Novikov, V.

    2015-01-01

    The influence of new derivatives of 9,10-anthraquinone with benzoylthiourea, thiazole, triazole and amino acid fragments on the activity of membrane-associated tyrosine kinases was investigated. Inhibitors of protein tyrosine kinase activity of the membrane fraction, as promising agents to search for new potential anticancer agents among the studied compounds, were discovered. PMID:26798182

  3. On the asymmetric adsorption of phenylalanine enantiomers by kaolin.

    NASA Technical Reports Server (NTRS)

    Bonner, W. A.; Flores, J.

    1973-01-01

    The attempt is described to verify a recent report that kaolin adsorbs D- and L-phenylalanine enantiomers to different extents from aqueous solutions at both pH 5.8 and pH 2. No evidence whatsoever could be found for the differential adsorption of D- versus L-phenylalanine by kaolin from either pH 6 or pH 2 solutions.

  4. Neutron inelastic scattering by amino acids

    SciTech Connect

    Thaper, C.L.; Sinha, S.K.; Dasannacharya, B.A.

    1982-01-01

    Inelastic neutron scattering experiments on normal, N-deuterated glycine, normal and N-deuterated alanine, L-valine, L-tyrosine and, L-phenylalanine at 100 K, are reported. Coupling of the external modes to different hydrogens is discussed.

  5. Tyrosine phosphorylation of WW proteins

    PubMed Central

    Reuven, Nina; Shanzer, Matan

    2015-01-01

    A number of key regulatory proteins contain one or two copies of the WW domain known to mediate protein–protein interaction via proline-rich motifs, such as PPxY. The Hippo pathway components take advantage of this module to transduce tumor suppressor signaling. It is becoming evident that tyrosine phosphorylation is a critical regulator of the WW proteins. Here, we review the current knowledge on the involved tyrosine kinases and their roles in regulating the WW proteins. PMID:25627656

  6. Further analysis of the short-term inhibition of food intake in humans by the dipeptide L-aspartyl-L-phenylalanine methyl ester (aspartame).

    PubMed

    Rogers, P J; Keedwell, P; Blundell, J E

    1991-04-01

    It was reported previously that the dipeptide sweetener aspartame suppresses food intake in humans by a postingestive action. The present study examined the hypothesis that this is due to an effect of phenylalanine, one of the primary breakdown products of aspartame (phenylalanine is a potent releaser of the so-called satiety hormone cholecystokinin, CCK). Capsulated aspartame (400 mg) administered to human volunteers reduced food intake by 15% (253 kcal) in a lunchtime test meal begun 1 hour later. However, neither phenylalanine (200 mg) nor the other constituent amino acid of aspartame, aspartic acid (200 mg), altered intake compared with placebo. Despite the large effect on food intake there were no treatment differences in pre- or postmeal ratings of motivation to eat. This suggests that aspartame may act to intensify the satiating effects of ingested food. Although high doses of phenylalanine reduce food intake, an individual action of phenylalanine cannot account for the potent anorexic effect of aspartame. In discussing alternative mechanisms it is noted that the amino acid sequence of aspartame (Asp-Phe) is the same as the C-terminal dipeptide of CCK. A direct action of aspartame at CCK receptors appears to be unlikely; however, aspartame might act as CCK releaser. Further studies are required to elucidate the mechanism of aspartame's anorexic action and perhaps to evaluate its therapeutic potential as an antiobesity agent.

  7. 21 CFR 172.320 - Amino acids.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ...-Leucine L-Lysine DL-Methionine (not for infant foods) L-Methionine L-Phenylalanine L-Proline L-Serine L... Aminoacetic acid (glycine) L-Leucine DL-Methionine L-Methionine L-Tryptophan L-Phenylalanine L-Proline L... DL-Methionine 3.1 L-Phenylalanine 5.8 L-Proline 4.2 L-Serine 8.4 L-Threonine 5.0 L-Tryptophan 1.6...

  8. 21 CFR 172.320 - Amino acids.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ...-Leucine L-Lysine DL-Methionine (not for infant foods) L-Methionine L-Phenylalanine L-Proline L-Serine L... Aminoacetic acid (glycine) L-Leucine DL-Methionine L-Methionine L-Tryptophan L-Phenylalanine L-Proline L... DL-Methionine 3.1 L-Phenylalanine 5.8 L-Proline 4.2 L-Serine 8.4 L-Threonine 5.0 L-Tryptophan 1.6...

  9. 21 CFR 172.320 - Amino acids.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ...-Leucine L-Lysine DL-Methionine (not for infant foods) L-Methionine L-Phenylalanine L-Proline L-Serine L... Aminoacetic acid (glycine) L-Leucine DL-Methionine L-Methionine L-Tryptophan L-Phenylalanine L-Proline L... DL-Methionine 3.1 L-Phenylalanine 5.8 L-Proline 4.2 L-Serine 8.4 L-Threonine 5.0 L-Tryptophan 1.6...

  10. Glucose-mediated tyrosine nitration in adipocytes: Target