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Sample records for alanine amino transferase

  1. 21 CFR 862.1030 - Alanine amino transferase (ALT/SGPT) test system.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Alanine amino transferase (ALT/SGPT) test system. 862.1030 Section 862.1030 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical...

  2. 21 CFR 862.1030 - Alanine amino transferase (ALT/SGPT) test system.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Alanine amino transferase (ALT/SGPT) test system. 862.1030 Section 862.1030 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical...

  3. 21 CFR 862.1030 - Alanine amino transferase (ALT/SGPT) test system.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Alanine amino transferase (ALT/SGPT) test system. 862.1030 Section 862.1030 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical...

  4. 21 CFR 862.1030 - Alanine amino transferase (ALT/SGPT) test system.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Alanine amino transferase (ALT/SGPT) test system. 862.1030 Section 862.1030 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical...

  5. 21 CFR 862.1030 - Alanine amino transferase (ALT/SGPT) test system.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Alanine amino transferase (ALT/SGPT) test system. 862.1030 Section 862.1030 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical...

  6. Effect of adrenergic agonists and antagonists on alanine amino transferase, fructose-1:6-bisphosphatase and glucose production in hepatocytes.

    PubMed

    Begum, N A; Datta, A G

    1992-08-18

    Using rat hepatocytes we confirmed our previous results that glucagon and beta-adrenergic agonists increased the enzyme activity of alanine aminotransferase (AAT) and propranolol abolished their effects. Only the enzyme activity was measured and other parameters like quantity of the enzyme or activation due to modification were not looked for. As in perfusion experiment phenylephrine and phenoxybenzamine (alpha-agonist and alpha-antagonist respectively) also alpha-antagonist respectively) also increased the AAT activity in isolated rat hepatocytes and propranolol reversed these effects. The additive effect of glucagon and phenoxybenzamine on AAT was also persistent in hepatocyte system. Fructose-1:6-bisphosphatase (Fru-P2-ase), another key enzyme in gluconeogenic pathway, was elevated by glucagon and other beta-adrenergic agonists both in liver perfusion and isolated hepatocyte experiments and was brought back to the normal level by propranolol. In this case also only the enzyme activity was measured and no other parameters were looked for. Unlike AAT this enzyme was not stimulated by phenylephrine or phenoxybenzamine. But AAT and Fru-P2-ase activities were increased significantly by adenylate cyclase activators like fluoride or forskolin. Thus, it appears that the regulation of fru-P2-ase by glucagon is purely a b-receptor mediated process whereas AAT activation shows a mixed type of regulation where some well known alpha-agonist and antagonists are behaving as beta-agonists. Results further indicate the presence of phosphodiesterase in hepatocyte membrane which was stimulated by glucagon and brought back to the normal level by propranolol. The different adrenergic compounds stated above, not only modified the activity of the above two enzymes but also stimulated glucose production by hepatocytes from alanine which was in turn abolished by propranolol as well as amino oxyacetate (AOA), a highly specified inhibitor of AAT. This confirm the participation of AAT in

  7. D-Amino acid dipeptide production utilizing D-alanine-D-alanine ligases with novel substrate specificity.

    PubMed

    Sato, Masaru; Kirimura, Kohtaro; Kino, Kuniki

    2005-06-01

    D-Alanine-D-alanine ligase (Ddl) is an important enzyme in the synthesis of bacterial peptidoglycan. The genes encoding Ddls from Escherichia coli K12 (EcDdlB), Oceanobacillus iheyensis JCM 11309 (OiDdl), Synechocystis sp. PCC 6803 (SsDdl) and Thermotoga maritima ATCC 43589 (TmDdl), the genomic DNA sequences of which have been determined, were cloned and the substrate specificities of these recombinant Ddls were investigated. Although OiDdl had a high substrate specificity for D-alanine; EcDdlB, SsDdl and TmDdl showed broad substrate specificities for D-serine, D-threonine, D-cysteine and glycine, in addition to D-alanine. Four D-amino acid dipeptides were produced using EcDdlB, and D-amino acid homo-dipeptides were successfully produced at high yields except for D-threonyl-D-threonine.

  8. A photoactivable amino acid based on a novel functional coumarin-6-yl-alanine.

    PubMed

    Fonseca, Andrea S C; Gonçalves, M Sameiro T; Costa, Susana P G

    2012-12-01

    A novel fluorescent amino acid, L-4-chloromethylcoumarin-6-yl-alanine, was obtained from tyrosine by a Pechmann reaction. The assembly of the heterocyclic ring at the tyrosine side chain could be achieved before or after incorporation of tyrosine into a dipeptide, and amino acid and dipeptide ester conjugates were obtained by coupling to a model N-protected alanine. The behaviour of one of the fluorescent conjugates towards irradiation was studied in a photochemical reactor at different wavelengths (254, 300, 350 and 419 nm). The photoreaction course in methanol/HEPES buffer solution (80:20) was followed by HPLC/UV monitoring. It was found that the novel unnatural amino acid could act as a fluorescent label, due to its fluorescence properties, and, more importantly, as a photoactivable unit, due to the short irradiation times necessary to cleave the ester bond between the model amino acid and the coumarin-6-yl-alanine.

  9. Equine endurance exercise alters serum branched-chain amino acid and alanine concentrations.

    PubMed

    Trottier, N L; Nielsen, B D; Lang, K J; Ku, P K; Schott, H C

    2002-09-01

    Six 2-year-old Arabian horses were used to determine whether 60 km prolonged endurance exercise (approximately 4 h) alters amino acid concentrations in serum and muscle, and the time required for serum amino acid concentrations to return to basal resting values. Blood and muscle samples were collected throughout exercise and during a 3 day recovery period. Isoleucine concentration in muscle tended to increase and leucine and valine did not change due to exercise. Serum alanine concentrations did not increase immediately after exercise, but increased at 24, 48 and 72 h postexercise. Serum isoleucine, leucine, and valine concentrations decreased after exercise and time required to reach pre-exercising concentrations was 48 h. In conclusion, endurance exercise in the horse decreases serum isoleucine, leucine, and valine concentrations, and increases serum alanine concentration. The decrease in serum branched-chain amino acid concentrations did not correspond to a measurable increase in total muscle branched-chain amino acid concentrations.

  10. Mapping of amino acid substitutions conferring herbicide resistance in wheat glutathione transferase.

    PubMed

    Govindarajan, Sridhar; Mannervik, Bengt; Silverman, Joshua A; Wright, Kathy; Regitsky, Drew; Hegazy, Usama; Purcell, Thomas J; Welch, Mark; Minshull, Jeremy; Gustafsson, Claes

    2015-03-20

    We have used design of experiments (DOE) and systematic variance to efficiently explore glutathione transferase substrate specificities caused by amino acid substitutions. Amino acid substitutions selected using phylogenetic analysis were synthetically combined using a DOE design to create an information-rich set of gene variants, termed infologs. We used machine learning to identify and quantify protein sequence-function relationships against 14 different substrates. The resulting models were quantitative and predictive, serving as a guide for engineering of glutathione transferase activity toward a diverse set of herbicides. Predictive quantitative models like those presented here have broad applicability for bioengineering.

  11. 21 CFR 862.1100 - Aspartate amino transferase (AST/SGOT) test system.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Aspartate amino transferase (AST/SGOT) test system. 862.1100 Section 862.1100 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical...

  12. 21 CFR 862.1100 - Aspartate amino transferase (AST/SGOT) test system.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Aspartate amino transferase (AST/SGOT) test system. 862.1100 Section 862.1100 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical...

  13. 21 CFR 862.1100 - Aspartate amino transferase (AST/SGOT) test system.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Aspartate amino transferase (AST/SGOT) test system. 862.1100 Section 862.1100 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical...

  14. 21 CFR 862.1100 - Aspartate amino transferase (AST/SGOT) test system.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Aspartate amino transferase (AST/SGOT) test system. 862.1100 Section 862.1100 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical...

  15. 21 CFR 862.1100 - Aspartate amino transferase (AST/SGOT) test system.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Aspartate amino transferase (AST/SGOT) test system. 862.1100 Section 862.1100 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical...

  16. Impact of charged amino acid substitution in the transmembrane domain of L-alanine exporter, AlaE, of Escherichia coli on the L-alanine export.

    PubMed

    Kim, Seryoung; Ihara, Kohei; Katsube, Satoshi; Ando, Tasuke; Isogai, Emiko; Yoneyama, Hiroshi

    2017-01-01

    The Escherichia coli alaE gene encodes the L-alanine exporter, AlaE, that catalyzes active export of L-alanine using proton electrochemical potential. The transporter comprises only 149 amino acid residues and four predicted transmembrane domains (TMs), which contain three charged amino acid residues. The AlaE-deficient L-alanine non-metabolizing cells (ΔalaE cells) appeared hypersusceptible to L-alanyl-L-alanine showing a minimum inhibitory concentration (MIC) of 2.5 µg/ml for the dipeptide due to a toxic accumulation of L-alanine. To elucidate the mechanism by which AlaE exports L-alanine, we replaced charged amino acid residues in the TMs, glutamic acid-30 (TM-I), arginine-45 (TM-II), and aspartic acid-84 (TM-III) with their respective charge-conserved amino acid or a net neutral cysteine. The ΔalaE cells producing R45K or R45C appeared hypersusceptible to the dipeptide, indicating that arginine-45 is essential for AlaE activity. MIC of the dipeptide in the ΔalaE cells expressing E30D and E30C was 156 µg/ml and >10,000 µg/ml, respectively, thereby suggesting that a negative charge at this position is not essential. The ΔalaE cells expressing D84E or D84C showed an MIC >10,000 and 78 µg/ml, respectively, implying that a negative charge is required at this position. These results were generally consistent with that of the L-alanine accumulation experiments in intact cells. We therefore concluded that charged amino acid residues (R45 and D84) in the AlaE transmembrane domain play a pivotal role in L-alanine export. Replacement of three cysteine residues at C22, C28 (both in TM-I), and C135 (C-terminal region) with alanine showed only a marginal effect on L-alanine export.

  17. Amino acid oxidation and alanine production in rat hemidiaphragm in vitro. Effects of dichloroacetate.

    PubMed Central

    Palmer, T N; Caldecourt, M A; Sugden, M C

    1984-01-01

    Dichloroacetate (an activator of pyruvate dehydrogenase) stimulates 14CO2 production from [U-14C]glucose, but not from [U-14C]glutamate, [U-14C]aspartate, [U-14C]- and [1-14C]-valine and [U-14C]- and [1-14C]-leucine. It is concluded (1) that pyruvate dehydrogenase is not rate-limiting in the oxidation to CO2 of amino acids that are metabolized to tricarboxylic acid-cycle intermediates, and (2) that carbohydrate (and not amino acids) is the main carbon precursor in alanine formation in muscle. PMID:6149743

  18. Probing the interaction of the amino acid alanine with the surface of ZnO(1010).

    PubMed

    Gao, Y K; Traeger, F; Shekhah, O; Idriss, H; Wöll, C

    2009-10-01

    The adsorption modes and stability of the amino acid alanine (NH(2)-CH(CH(3))-COOH) have been studied on the nonpolar single crystal surface of zinc oxide, ZnO(1010), experimentally by X-ray photoelectron spectroscopy (XPS) and computationally using density functional theory (DFT). Deposition at 200 K was found to lead to the formation of multilayers identified by an XPS N1s peak at 401.7 eV assigned to the NH(3)(+) group, a fingerprint of the zwitterionic structure of alanine in the solid state. Heating to 300 K resulted in the removal of most of the multilayers with the remaining surface coverage estimated to 0.4 with respect to Zn cations. At this temperature most of the alanine molecules are found to be deprotonated (dissociated), yielding a carboxylate species (NH(2)-CH(CH(3))-COO(-) (a) + OH (s); where O is surface oxygen, (a) for adsorbed and (s) for surface species). Further heating of the surface resulted in a gradual decrease of the surface coverage and by 500 K a large fraction of adsorbed alanine molecules have desorbed from the surface. Total energy DFT computations of different adsorbate species identified two stable dissociative adsorption modes: bidentate and monodentate. The bidentate species with adsorption energy of 1.75 eV was found to be more stable than the monodentate species by about 0.7 eV.

  19. Proteins with β-(thienopyrrolyl)alanines as alternative chromophores and pharmaceutically active amino acids

    PubMed Central

    Budisa, Nediljko; Alefelder, Stefan; Bae, Jae Hyun; Golbik, Ralph; Minks, Caroline; Huber, Robert; Moroder, Luis

    2001-01-01

    L-β-(Thieno[3,2-b]pyrrolyl)alanine and L-β-(thieno[2,3-b]pyrrolyl)alanine are mutually isosteric and pharmaceutically active amino acids that mimic tryptophan with the benzene ring in the indole moiety replaced by thiophene. Sulfur as a heteroatom causes physicochemical changes in these tryptophan surrogates that bring about completely new properties not found in the indole moiety. These synthetic amino acids were incorporated into recombinant proteins in response to the Trp UGG codons by fermentation in a Trp-auxotrophic Escherichia coli host strain using the selective pressure incorporation method. Related protein mutants expectedly retain the secondary structure of the native proteins but show significantly changed optical and thermodynamic properties. In this way, new spectral windows, fluorescence, polarity, thermodynamics, or pharmacological properties are inserted into proteins. Such an engineering approach by translational integration of synthetic amino acids with a priori defined properties, as shown in this study, proved to be a novel and useful tool for protein rational design. PMID:11420430

  20. A comparative study on the growth and characterization of nonlinear optical amino acid crystals: L-alanine (LA) and L-alanine alaninium nitrate (LAAN).

    PubMed

    Aravindan, A; Srinivasan, P; Vijayan, N; Gopalakrishnan, R; Ramasamy, P

    2008-11-15

    A comparative study on the properties of L-alanine and LAAN crystals has been made and discussed. It may be concluded that the protonation of the amino group in the L-alanine molecule is the key factor in increasing the relative SHG efficiency of LAAN. The protonation is justified by the crystal structure analysis, FTIR and photoluminescence studies. The factor group vibrations are compared and found that there is an increase in vibrational modes of LA when reacted with nitric acid forming LAAN.

  1. Gliotoxicity of the cyanotoxin, β-methyl-amino-L-alanine (BMAA).

    PubMed

    Chiu, Alexander S; Gehringer, Michelle M; Braidy, Nady; Guillemin, Gilles J; Welch, Jeffrey H; Neilan, Brett A

    2013-01-01

    The amino acid variant β-methyl-amino-L-alanine (BMAA) has long been associated with the increased incidence and progression of the amyotrophic lateral sclerosis/Parkinsonism dementia complex (ALS/PDC). Previous studies have indicated that BMAA damages neurons via excitotoxic mechanisms. We have challenged rat olfactory ensheathing cells (OECs) with exogenous BMAA and found it to be cytotoxic. BMAA also induces a significant increase in Ca2+ influx, enhanced production of reactive oxygen species (ROS), and disrupts mitochondrial activity in OECs. This is the first study investigating BMAA toxicity using pure glial cells. These findings align BMAA with the three proposed mechanisms of degeneration in ALS, those being non-cell autonomous death, excitotoxicity and mitochondrial dysfunction.

  2. The non-protein amino acid β-N-methylamino-L-alanine in Portuguese cyanobacterial isolates.

    PubMed

    Cervantes Cianca, Rosa C; Baptista, Mafalda S; Lopes, Viviana R; Vasconcelos, Vitor M

    2012-06-01

    The tailor made amino acid β-N-methyl-amino-L-alanine (BMAA) is a neurotoxin produced by cyanobacteria. It has been associated with certain forms of progressive neurodegenerative disease, including sporadic Amyotrophic Lateral Sclerosis and Alzheimer's disease. Some different reports of BMAA in cyanobacterial blooms from lakes, reservoirs, and other water resources have been made by different investigators. We here report the detection of BMAA of both free and protein-bound produced by cyanobacteria, belonging to the Chroococcales, Oscillatoriales and Nostocales ordered. We use a rapid and sensitive HPLC-FD method that utilizes methanol elution and the Waters AQC Tag chemistry. On other hand, we have used three different assay procedures for BMAA extraction from cyanobacteria: Trichloroacetic acid (TCA), Methanol/Acetone and hydrochloric acid (HCl). All assays let successfully detect BMAA in all cyanobacteria samples analyzed. Nevertheless, with TCA and HCl extraction procedures the highest BMAA values, for free as well as protein-bound BMAA were detected. BMAA content could not be related to the taxonomy of the isolates or to their geographical origin, and no correlation between free and protein-bound BMAA concentrations were observed within or between taxonomic groups. These data offer confirmation of the taxonomic and geographic ubiquity of BMAA from naturally occurring populations of cyanobacteria, for the first time reported for estuaries.

  3. Chiral selectivity of amino acid adsorption on chiral surfaces—The case of alanine on Pt

    SciTech Connect

    Franke, J.-H.; Kosov, D. S.

    2015-02-07

    We study the binding pattern of the amino acid alanine on the naturally chiral Pt surfaces Pt(531), Pt(321), and Pt(643). These surfaces are all vicinal to the (111) direction but have different local environments of their kink sites and are thus a model for realistic roughened Pt surfaces. Alanine has only a single methyl group attached to its chiral center, which makes the number of possible binding conformations computationally tractable. Additionally, only the amine and carboxyl group are expected to interact strongly with the Pt substrate. On Pt(531), we study the molecule in its pristine as well as its deprotonated form and find that the deprotonated one is more stable by 0.47 eV. Therefore, we study the molecule in its deprotonated form on Pt(321) and Pt(643). As expected, the oxygen and nitrogen atoms of the deprotonated molecule provide a local binding “tripod” and the most stable adsorption configurations optimize the interaction of this “tripod” with undercoordinated surface atoms. However, the interaction of the methyl group plays an important role: it induces significant chiral selectivity of about 60 meV on all surfaces. Hereby, the L-enantiomer adsorbs preferentially to the Pt(321){sup S} and Pt(643){sup S} surfaces, while the D-enantiomer is more stable on Pt(531){sup S}. The binding energies increase with increasing surface density of kink sites, i.e., they are largest for Pt(531){sup S} and smallest for Pt(643){sup S}.

  4. A versatile proline/alanine transporter in the unicellular pathogen Leishmania donovani regulates amino acid homoeostasis and osmotic stress responses.

    PubMed

    Inbar, Ehud; Schlisselberg, Doreen; Suter Grotemeyer, Marianne; Rentsch, Doris; Zilberstein, Dan

    2013-01-15

    Unlike all other organisms, parasitic protozoa of the family Trypanosomatidae maintain a large cellular pool of proline that, together with the alanine pool, serve as alternative carbon sources as well as reservoirs of organic osmolytes. These reflect adaptation to their insect vectors whose haemolymphs are exceptionally rich in the two amino acids. In the present study we identify and characterize a new neutral amino acid transporter, LdAAP24, that translocates proline and alanine across the Leishmania donovani plasma membrane. This transporter fulfils multiple functions: it is the sole supplier for the intracellular pool of proline and contributes to the alanine pool; it is essential for cell volume regulation after osmotic stress; and it regulates the transport and homoeostasis of glutamate and arginine, none of which are its substrates. Notably, we provide evidence that proline and alanine exhibit different roles in the parasitic response to hypotonic shock; alanine affects swelling, whereas proline influences the rate of volume recovery. On the basis of our data we suggest that LdAAP24 plays a key role in parasite adaptation to its varying environments in host and vector, a phenomenon essential for successful parasitism.

  5. Amino Acid Mixture Enriched With Arginine, Alanine, and Phenylalanine Stimulates Fat Metabolism During Exercise.

    PubMed

    Ueda, Keisuke; Nakamura, Yutaka; Yamaguchi, Makoto; Mori, Takeshi; Uchida, Masayuki; Fujita, Satoshi

    2016-02-01

    Although there have been many investigations of the beneficial effects of both exercise and amino acids (AAs), little is known about their combined effects on the single-dose ingestion of AAs for lipid metabolism during exercise. We hypothesize that taking a specific combination of AAs implicated in glucagon secretion during exercise may increase fat metabolism. We recently developed a new mixture, d-AA mixture (D-mix), that contains arginine, alanine, and phenylalanine to investigate fat oxidation. In a double-blind, placebo-controlled crossover study, 10 healthy male volunteers were randomized to ingest either D-mix (3 g/dose) or placebo. Subjects in each condition subsequently performed a physical task that included workload trials on a cycle ergometer at 50% of maximal oxygen consumption for 1 hr. After oral intake of D-mix, maximum serum concentrations of glycerol (9.32 ± 6.29 mg/L and 5.22 ± 2.22 mg/L, respectively; p = .028), free fatty acid level (0.77 ± 0.26 mEq/L and 0.63 ± 0.28 mEq/L, respectively; p = .022), and acetoacetic acid levels (37.9 ± 17.7 μmol/L and 30.3 ± 13.9 μmol/L, respectively; p = .040) were significantly higher than in the placebo groups. The area under the curve for glucagon during recovery was numerically higher than placebo (6.61 ± 1.33 μg/L · min and 6.06 ± 1.23 μg/L · min, respectively; p = .099). These results suggest that preexercise ingestion of D-mix may stimulate fat metabolism. Combined with exercise, the administration of AA mixtures could prove to be a useful nutritional strategy to maximize fat metabolism.

  6. Determination of the non protein amino acid β-N-methylamino-l-alanine in estuarine cyanobacteria by capillary electrophoresis.

    PubMed

    Baptista, Mafalda S; Cianca, Rosa C C; Lopes, Viviana R; Almeida, C Marisa R; Vasconcelos, Vitor M

    2011-10-01

    A capillary electrophoretic method for the determination of the amino acid β-N-methylamino-l-alanine (BMAA) was achieved using a fused-silica capillary column (50 cm × 75 μm I.D.) filled with 5 mM sodium tetraborate solution (pH 9), with an applied voltage of 25 kV, at 25 °C. The method was then applied in quantifying BMAA in eighteen strains of lyophilized estuarine cyanobacteria, following amino acid extraction using 0.1 M trichloroacetic acid and 6 M hydrochloric acid, sequentially.

  7. Theoretical and experimental study of valence photoelectron spectrum of D,L-alanine amino acid.

    PubMed

    Farrokhpour, H; Fathi, F; De Brito, A Naves

    2012-07-05

    In this work, the He-I (21.218 eV) photoelectron spectrum of D,L-alanine in the gas phase is revisited experimentally and theoretically. To support the experiment, the high level ab initio calculations were used to calculate and assign the photoelectron spectra of the four most stable conformers of gaseous alanine, carefully. The symmetry adapted cluster/configuration interaction (SAC-CI) method based on single and double excitation operators (SD-R) and its more accurate version, termed general-R, was used to separately calculate the energies and intensities of the ionization bands of the L- and D-alanine conformers. The intensities of ionization bands were calculated based on the monopole approximation. Also, natural bonding orbital (NBO) calculations were employed for better spectral band assignment. The relative electronic energy, Gibbs free energy, and Boltzmann population ratio of the conformers were calculated at the experimental temperature (403 K) using several theoretical methods. The theoretical photoelectron spectrum of alanine was calculated by summing over the spectra of individual D and L conformers weighted by different population ratios. Finally, the population ratio of the four most stable conformers of alanine was estimated from the experimental photoelectron spectrum using theoretical calculations for the first time.

  8. Neurotoxic Non-proteinogenic Amino Acid β-N-Methylamino-L-alanine and Its Role in Biological Systems.

    PubMed

    Popova, A A; Koksharova, O A

    2016-08-01

    Secondary metabolites of photoautotrophic organisms have attracted considerable interest in recent years. In particular, molecules of non-proteinogenic amino acids participating in various physiological processes and capable of producing adverse ecological effects have been actively investigated. For example, the non-proteinogenic amino acid β-N-methylamino-L-alanine (BMAA) is neurotoxic to animals including humans. It is known that BMAA accumulation via the food chain can lead to development of neurodegenerative diseases in humans such as Alzheimer's and Parkinson's diseases as well as amyotrophic lateral sclerosis. Moreover, BMAA can be mistakenly incorporated into a protein molecule instead of serine. Natural sources of BMAA and methods for its detection are discussed in this review, as well as the role of BMAA in metabolism of its producers and possible mechanisms of toxicity of this amino acid in different living organisms.

  9. N- Trichloro- and dichloroacetyl amino acids and compounds of amino acids with halogeno acetic acids: 35Cl nuclear quadrupole resonance spectroscopy; crystal structure of N- trichloroacetyl- glycine, - DL-alanine, and - L-alanine

    NASA Astrophysics Data System (ADS)

    Dou, Shi-qi; Kehrer, Armin; Ofial, Armin R.; Weiss, Alarich

    1995-02-01

    The crystal structures of N- trichloroacetyl- glycine ( N- TCA- G), N-trichloroacetyl-dl-alanine ( N-TCA- dl-A ), and N-trichloroacetyl- l-alanine ( N-TCA- l-A ) were determined. In addition, the 35Cl NQR spectra of these N-trichloroacetyl amino acids, of N-trichloroacetyl- l-valine ( N-TCA- l-V ), and of N- dichloroacetyl- glycine and - L-alanine were measured, mostly as a function of temperature. Compounds of glycine and L-alanine with chlorodifluoroacetic acid, of glycine and L-leucine with monochloroacetic acid, of glycine and L-leucine with dichloroacetic acid, and of glycine and L-leucine with trichloroacetic acid were also studied using 35Cl NQR. The structures (in picometres and degrees) were found to be as follows. N- TCA- G: Pna2 1, Z = 8, a = 1641, b = 1002, c = 1018. N-TCA- dl-A : {C2}/{c}, Z = 8, a = 3280, b = 556, c = 1031, β = 96.68. N-TCA- l-A: P1 , Z = 2, a = 967, b = 949, c = 619, α = 74.97, β = 74.20, γ = 61.20. The 35Cl NQR frequencies (ν) were observed in the range 35-41 MHz, and decrease with increasing temperature. Some of the resonances bleach out at a temperature ( Tb) far below the melting temperature; this provides information about the crystal structures at 77 K. No phase transitions were observed by differential thermal analysis between 77 and 295 K. The crystal structures are discussed in connection with the NQR results, and conclusions are drawn about the structures of the compounds for which only 35Cl NQR data are available.

  10. The natural non-protein amino acid N-β-methylamino-L-alanine (BMAA) is incorporated into protein during synthesis.

    PubMed

    Glover, W Broc; Mash, Deborah C; Murch, Susan J

    2014-11-01

    N-β-methylamino-L-alanine (BMAA) is an amino acid produced by cyanobacteria and accumulated through trophic levels in the environment and natural food webs. Human exposure to BMAA has been linked to progressive neurodegenerative diseases, potentially due to incorporation of BMAA into protein. The insertion of BMAA and other non-protein amino acids into proteins may trigger protein misfunction, misfolding and/or aggregation. However, the specific mechanism by which BMAA is associated with proteins remained unidentified. Such studies are challenging because of the complexity of biological systems and samples. A cell-free in vitro protein synthesis system offers an excellent approach for investigation of changing amino acid composition in protein. In this study, we report that BMAA incorporates into protein as an error in synthesis when a template DNA sequence is used. Bicinchoninic acid assay of total protein synthesis determined that BMAA effectively substituted for alanine and serine in protein product. LC-MS/MS confirmed that BMAA was selectively inserted into proteins in place of other amino acids, but isomers N-(2-aminoethyl)glycine (AEG) and 2,4-diaminobutyric acid (DAB) did not share this characteristic. Incorporation of BMAA into proteins was significantly higher when genomic DNA from post-mortem brain was the template. About half of BMAA in the synthetic proteins was released with denaturation with sodium dodecylsulfonate and dithiothreitol, but the remaining BMAA could only be released by acid hydrolysis. Together these data demonstrate that BMAA is incorporated into the amino acid backbone of proteins during synthesis and also associated with proteins through non-covalent bonding.

  11. Intramolecular vibrations in low-frequency normal modes of amino acids: L-alanine in the neat solid state.

    PubMed

    Zhang, Feng; Wang, Houng-Wei; Tominaga, Keisuke; Hayashi, Michitoshi

    2015-03-26

    This paper presents a theoretical analysis of the low-frequency phonons of L-alanine by using the solid-state density functional theory at the Γ point. We are particularly interested in the intramolecular vibrations accessing low-frequency phonons via harmonic coupling with intermolecular vibrations. A new mode-analysis method is introduced to quantify the vibrational characteristics of such intramolecular vibrations. We find that the torsional motions of COO(-) are involved in low-frequency phonons, although COO(-) is conventionally assumed to undergo localized torsion. We also find the broad distributions of intramolecular vibrations relevant to important functional groups of amino acids, e.g., the COO(-) and NH3(+) torsions, in the low-frequency phonons. The latter finding is illustrated by the concept of frequency distribution of vibrations. These findings may lead to immediate implications in other amino acid systems.

  12. Efficient optical resolution of amino acid by alanine racemaze chiral analogue supported on mesoporous carbon

    NASA Astrophysics Data System (ADS)

    Jang, D.; Kim, K.; Park, D.; Kim, G.

    2012-09-01

    Optically pure D-amino acids are industrially important chiral building blocks for the synthesis of pharmaceuticals, food ingredients, and drug intermediates. Chemoenzymatic dynamic kinetic-resolution processes have recently been developed for deracemization of amino acids. S-ARCA would be a good candidate for the selective adsorption of D amino acid through the imine formation reaction. The organic phase containing S-ARCA adsorbent, TPPC or Ionic Liquid (as a phase transfer catalyst) in MC were coated on the surfaces of mesoporous carbon C-SBA-15(CMK). The aqueous solution of racemic D/L-amino acid and NaOH were added to the carbon support coated with ARCA. The D/L ratios on ARCA and in solution were determined with increasing reaction time. S-ARCA has a unique property for the selective adsorption of D- amino acid (up to 90% selcetivity) in the racemic mixture. The fixed bed reactor containing ARCA/carbon support was also adopted successfully for the selective separation of amino acid.

  13. First principles DFT study of weak C-H…O bonds in crystalline amino acids under pressure-alanine

    NASA Astrophysics Data System (ADS)

    Ramaniah, Lavanya M.; Kamal, C.; Sikka, S. K.

    2013-02-01

    Many crystalline solids containing C-H…O hydrogen bonds display blue shifting of the C-H stretching frequency under pressure. No agreed explanation is available for this. Here, we use first principles density functional theory, to determine the hydrogen atom positions to understand the cause of this blue shift. No neutron diffraction is feasible due to flux limitations for this purpose. As a first case, we have taken up the study of the amino acid, alanine. We find that the C_H_…O bond in it no longer remain isolated under compression as is case at ambient pressure. The hydrogen atom in the bond has now repulsive contacts with other atoms. This results in contraction of the C-H bond length and consequently to blue shifting as is found experimentally.

  14. Determination of β -Cyano-L-alanine, γ -Glutamyl- β -cyano-L-alanine, and Common Free Amino Acids in Vicia sativa (Fabaceae) Seeds by Reversed-Phase High-Performance Liquid Chromatography.

    PubMed

    Megías, Cristina; Cortés-Giraldo, Isabel; Girón-Calle, Julio; Vioque, Javier; Alaiz, Manuel

    2014-01-01

    A method for determination of β-cyano-L-alanine, γ-glutamyl-β-cyano-L-alanine and other free amino acids in Vicia sativa is presented. Seed extracts were derivatized by reaction with diethyl ethoxymethylenemalonate and analyzed by reverse-phase high-performance liquid chromatography. Calibration curves showed very good linearity of the response. The limit of detection and quantification was 0.15 and 0.50 μM, respectively. The method has high intra- (RSD = 0.28-0.31%) and interrepeatability (RSD = 2.76-3.08%) and remarkable accuracy with a 99% recovery in spiked samples. The method is very easy to carry out and allows for ready analysis of large number of samples using very basic HPLC equipment because the derivatized samples are very stable and have very good chromatographic properties. The method has been applied to the determination of γ-glutamyl-β-cyano-L-alanine, β-cyano-L-alanine, and common free amino acids in eight wild populations of V. sativa from southwestern Spain.

  15. Excitotoxic potential of the cyanotoxin β-methyl-amino-L-alanine (BMAA) in primary human neurons.

    PubMed

    Chiu, Alexander S; Gehringer, Michelle M; Braidy, Nady; Guillemin, Gilles J; Welch, Jeffrey H; Neilan, Brett A

    2012-11-01

    The toxicity of the cyanobacterial modified amino acid, BMAA, has been described in rat, mouse and leech neurons. Particular emphasis has been placed on the potential ability of BMAA to induce neuronal damage via excitotoxic mechanisms. Here we present data indicating that the effects observed on lower organisms are also evident in a human model. Our data indicates that BMAA induces increased intracellular Ca²⁺ influx, DNA damage, mitochondrial activity, lactate dehydrogenase (LDH) release and generation of reactive oxygen species (ROS). The amelioration of LDH release in the presence of the N-methyl-D-aspartate (NMDA) receptor antagonist MK801 indicates that the neurotoxic effects of BMAA are mediated via NMDA receptor activation. Additionally, we have shown that BMAA induces the expression of neuronal nitric oxide synthase (nNOS) and caspase-3 indicating that it can stimulate apoptosis in human neurons, presumably via activation of NMDA receptors.

  16. Quantitative proteomics analysis of zebrafish exposed to sub-lethal dosages of β-methyl-amino-L-alanine (BMAA)

    PubMed Central

    Frøyset, Ann Kristin; Khan, Essa Ahsan; Fladmark, Kari Espolin

    2016-01-01

    The non-protein amino acid β-methylamino-L-alanine (BMAA) is a neurotoxin present in microalgae and shown to accumulate in the food web. BMAA has been linked to the complex neurodegenerative disorder of Guam and to increased incidents sporadic ALS. Two main neurotoxic routes are suggested; an excitotoxic by acting as an agonist towards glutamate receptors and a metabolic by misincorporating into cellular proteins. We have used zebrafish, an increasingly used model for neurodegenerative diseases, to further identify signaling components involved in BMAA-induced toxicity. Zebrafish embryos were exposed to sub-lethal dosages of BMAA and a label-free proteomics analysis was conducted on larvae 4 days post fertilization. The exposed larvae showed no developmental abnormalities, but a reduced heart rate and increased expression of GSK3 isoforms. Search towards a reviewed database containing 2968 entries identified 480 proteins. Only 17 of these were regulated 2-fold or more in the exposed larvae. Seven of these proteins could be associated to glutamate receptor signaling and recycling. The remaining nine have all been linked to disturbance in protein homeostasis, reactive oxygen species (ROS) development or neuronal cell death. We also found that BMAA influenced the endocannabinoid system by up-regulation of fatty acid amide hydrolase (FAAH) and that FAAH inhibitor URB597 reduced the BMAA effect on heart rate and GSK3 expression. PMID:27404450

  17. Quantitative proteomics analysis of zebrafish exposed to sub-lethal dosages of β-methyl-amino-L-alanine (BMAA).

    PubMed

    Frøyset, Ann Kristin; Khan, Essa Ahsan; Fladmark, Kari Espolin

    2016-07-12

    The non-protein amino acid β-methylamino-L-alanine (BMAA) is a neurotoxin present in microalgae and shown to accumulate in the food web. BMAA has been linked to the complex neurodegenerative disorder of Guam and to increased incidents sporadic ALS. Two main neurotoxic routes are suggested; an excitotoxic by acting as an agonist towards glutamate receptors and a metabolic by misincorporating into cellular proteins. We have used zebrafish, an increasingly used model for neurodegenerative diseases, to further identify signaling components involved in BMAA-induced toxicity. Zebrafish embryos were exposed to sub-lethal dosages of BMAA and a label-free proteomics analysis was conducted on larvae 4 days post fertilization. The exposed larvae showed no developmental abnormalities, but a reduced heart rate and increased expression of GSK3 isoforms. Search towards a reviewed database containing 2968 entries identified 480 proteins. Only 17 of these were regulated 2-fold or more in the exposed larvae. Seven of these proteins could be associated to glutamate receptor signaling and recycling. The remaining nine have all been linked to disturbance in protein homeostasis, reactive oxygen species (ROS) development or neuronal cell death. We also found that BMAA influenced the endocannabinoid system by up-regulation of fatty acid amide hydrolase (FAAH) and that FAAH inhibitor URB597 reduced the BMAA effect on heart rate and GSK3 expression.

  18. Novel emissive bio-inspired non-proteinogenic coumarin-alanine amino acid: fluorescent probe for polyfunctional systems.

    PubMed

    Oliveira, Elisabete; Capelo, José Luis; Lima, João Carlos; Lodeiro, Carlos

    2012-10-01

    Two new bio-inspired non-proteinogenic compounds L1 and L2, containing coumarin and/or acridine chromophores and bearing as spacer an alanine amino acid were successfully synthesized and fully characterized by elemental analysis, (1)H and (13)C NMR, infrared spectroscopy (KBr discs), melting point, ESI-TOF (electrospray ionization-time of flight-mass), UV-vis absorption and emission spectroscopy, fluorescence quantum yields and lifetime measurements. A relative fluorescence quantum yield of 0.02 was determined for both compounds. In L2 the presence of an intramolecular energy transfer from the coumarin to the acridine unit was observed. L1 and L2 are quite sensitive to the basicity of the environment. At alkaline values both compounds show a strong quenching in the fluorescence emission, attributed to the photoinduced electron transfer (PET). However, both deprotonated forms recover the emission with the addition of Zn(2+), Cd(2+) and Al(3+) metal ions. As multifunctional emissive probes, the titration of L1 and L2 with lanthanides (III), Eu(3+) and Tb(3+) was also explored as new visible bio-probes in the absence and in the presence of liposomes. In a liposomal environment a lower energy transfer was observed.

  19. Oxidation of 3,4-dehydro-D-proline and other D-amino acid analogues by D-alanine dehydrogenase from Escherichia coli.

    PubMed

    Deutch, Charles E

    2004-09-15

    3,4-Dehydro-DL-proline is a toxic analogue of L-proline which has been useful in studying the uptake and metabolism of this key amino acid. When membrane fractions from Escherichia coli strain UMM5 (putA1::Tn5 proC24) lacking both L-proline dehydrogenase and L-Delta(1)-pyrroline-5-carboxylate reductase were incubated with 3,4-dehydro-DL-proline, pyrrole-2-carboxylate was formed. There was no enzyme activity with 3,4-dehydro-L-proline, but activity was restored after racemization of the substrate. Oxidation of 3,4-dehydro-DL-proline by membrane fractions from strain UMM5 was induced by growth in minimal medium containing D- or L-alanine, had a pH optimum of 9, and was competitively inhibited by D-alanine. An E. coli strain with no D-alanine dehydrogenase activity due to the dadA237 mutation was unable to oxidize either 3,4-dehydro-D-proline or D-alanine, as were spontaneous Dad(-) mutants of E. coli strain UMM5. Membrane fractions containing D-alanine dehydrogenase also catalyzed the oxidation of D-2-aminobutyrate, D-norvaline, D-norleucine, cis-4-hydroxy-D-proline, and DL-ethionine. These results indicate that d-alanine dehydrogenase is responsible for the residual 3,4-dehydro-DL-proline oxidation activity in putA proC mutants of E. coli and provide further evidence that this enzyme plays a general role in the metabolism of D-amino acids and their analogues.

  20. Enzymatic resolution for the preparation of enantiomerically enriched D-beta-heterocyclic alanine derivatives using Escherichia coli aromatic L-amino acid transaminase.

    PubMed

    Cho, Byung-Kwan; Park, Hyung-Yeon; Seo, Joo-Hyun; Kinnera, Koteshwar; Lee, Bon-Su; Kim, Byung-Gee

    2004-11-20

    An enzymatic resolution was carried out for the preparation of enriched beta-heterocyclic D-alanine derivatives using Escherichia coli aromatic L-amino acid transaminase. The excess of pyrazole, imidazole, or 1,2,4-triazole reacted with methyl-2-acetamidoacrylate in acetonitrile in the presence of potassium carbonate at 60 degrees C, directly leading to make the potassium salt of the corresponding N-acetyl-beta-heterocyclic alanine derivatives. After the acidic deprotection of the N-acetyl group, 10 mM of racemic pyrazolylalanine, triazolylalanine, and imidazolylalanine were resolved to D-pyrazolylalanine, D-triazolylalanine, and D-imidazolylalanine with 46% (85% ee), 42% (72% ee), and 48% (95% ee) conversion yield in 18 h, respectively, using E. coli aromatic L-amino acid transaminase (EC 2.6.1.5). Although the three beta-heterocyclic L-alanine derivatives have similar molecular structures, they showed different reaction rates and enantioselectivities. The relative reactivities of the transaminase toward the beta-heterocyclic L-alanine derivatives could be explained by the relationship between the substrate binding energy (E, kcal/mol) to the enzyme active site and the distance (delta, A) from the nitrogen of alpha-amino group of the substrates to the C4' carbon of PLP-Lys258 Schiff base. As the ratio of the substrate binding energy (E) to the distance (delta) becomes indicative value of k(cat)/K(M) of the enzyme to the substrate, the relative reactivities of the beta-heterocyclic L-alanine derivatives were successfully correlated with E/delta, and the relationship was confirmed by our experiments.

  1. Excess of L-alanine in amino acids synthesized in a plasma torch generated by a hypervelocity meteorite impact reproduced in the laboratory

    NASA Astrophysics Data System (ADS)

    Managadze, George G.; Engel, Michael H.; Getty, Stephanie; Wurz, Peter; Brinckerhoff, William B.; Shokolov, Anatoly G.; Sholin, Gennady V.; Terent'ev, Sergey A.; Chumikov, Alexander E.; Skalkin, Alexander S.; Blank, Vladimir D.; Prokhorov, Vyacheslav M.; Managadze, Nina G.; Luchnikov, Konstantin A.

    2016-10-01

    We present a laboratory reproduction of hypervelocity impacts of a carbon containing meteorite on a mineral substance representative of planetary surfaces. The physical conditions of the resulting impact plasma torch provide favorable conditions for abiogenic synthesis of protein amino acids: We identified glycine and alanine, and in smaller quantities serine, in the produced material. Moreover, we observe breaking of alanine mirror symmetry with L excess, which coincides with the bioorganic world. Therefore the selection of L-amino acids for the formation of proteins for living matter could have been the result from plasma processes occurring during the impact meteorites on the surface. This indicates that the plasma torch from meteorite impacts could play an important role in the formation of biomolecular homochirality. Thus, meteorite impacts possibly were the initial stage of this process and promoted conditions for the emergence of a living matter.

  2. Alanine water complexes.

    PubMed

    Vaquero, Vanesa; Sanz, M Eugenia; Peña, Isabel; Mata, Santiago; Cabezas, Carlos; López, Juan C; Alonso, José L

    2014-04-10

    Two complexes of alanine with water, alanine-(H2O)n (n = 1,2), have been generated by laser ablation of the amino acid in a supersonic jet containing water vapor and characterized using Fourier transform microwave spectroscopy. In the observed complexes, water molecules bind to the carboxylic group of alanine acting as both proton donors and acceptors. In alanine-H2O, the water molecule establishes two intermolecular hydrogen bonds forming a six-membered cycle, while in alanine-(H2O)2 the two water molecules establish three hydrogen bonds forming an eight-membered ring. In both complexes, the amino acid moiety is in its neutral form and shows the conformation observed to be the most stable for the bare molecule. The microsolvation study of alanine-(H2O)n (n = 1,2) can be taken as a first step toward understanding bulk properties at a microscopic level.

  3. The metabolism of the non-proteinogenic amino acid β-N-methylamino-L-alanine (BMAA) in the cyanobacterium Synechocystis PCC6803.

    PubMed

    Downing, Simoné; Downing, Timothy Grant

    2016-06-01

    The neurotoxic amino acid β-N-methylamino-L-alanine (BMAA) is produced by cyanobacteria under nitrogen starvation conditions and its metabolism is closely associated with cellular nitrogen control. Very little is known regarding the metabolism or biosynthesis of this amino acid in the producing organisms and current knowledge is limited to the spontaneous formation of carbamate adducts in the presence of aqueous carbon dioxide, the rapid removal of free cellular BMAA upon the addition of ammonia to nitrogen-starved cyanobacterial cultures, and the link between cellular nitrogen status and BMAA synthesis. Data presented here show that exogenous BMAA is readily metabolised by cyanobacteria during which, the primary amino group is rapidly transferred to other cellular amino acids. Furthermore, data suggest that BMAA is metabolised in cyanobacteria via a reversible transamination reaction. This study presents novel data on BMAA metabolism in cyanobacteria and provides the first proposed biosynthetic precursor to BMAA biosynthesis in cyanobacteria.

  4. Differential effect of beta-N-oxalylamino-L-alanine, the Lathyrus sativus neurotoxin, and (+/-)-alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate on the excitatory amino acid and taurine levels in the brain of freely moving rats.

    PubMed

    La Bella, V; Piccoli, F

    2000-05-01

    We studied the effect of beta-oxalylamino-L-alanine, a glutamate analog present in Lathyrus sativus seeds and implicated in the etiopathogenesis of neurolathyrism, and (+/-)-alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate on the extracellular levels of aspartate, glutamate and taurine in the primary motor cortex of freely moving rats. We found that while both neurotoxins increase the level of aspartate and glutamate, only (+/-)-alpha(-amino-3-hydroxy-5-methylisoxazole-4-propionate is able to modulate the level of taurine. GYKI-52466, a non-competitive non-NMDA antagonist, inhibited beta-oxalylamino-L-alanine-induced increase of aspartate, but not that of glutamate. Conversely, this antagonist proved to be very efficient in blocking the stimulating effect of (+/-)-alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate on all three amino acids. We suggest that beta-oxalylamino-L-alanine increases the level of glutamate in vivo by a mechanism not connected to its effect on the non-NMDA receptors, which might involve the inhibition of glutamate transport. This would allow the excitatory neurotransmitter to reach a concentration sufficient to stimulate the non-NMDA receptors, which in their turn mediate the specific release of aspartate. Although the role of aspartate as a neurotransmitter is still under discussion, it might indeed amplify the excitotoxic cascade through its action on NMDA receptors. We speculate that this sequence of events might represent an important step in the molecular cascade leading to the appearance of the selective motoneuron degeneration in neurolathyrism.

  5. Identification of the roles of individual amino acid residues of the helix E of the major antenna of photosystem II (LHCII) by alanine scanning mutagenesis.

    PubMed

    Liu, Cheng; Rao, Yan; Zhang, Lei; Yang, Chunhong

    2014-10-01

    The functions of the helix E (W97-F105), an amphiphilic lumenal 310 helix of the major antenna of photosystem II (LHCII), are still unidentified. To elucidate the roles of individual amino acid residue of the helix E, alanine scanning mutagenesis has been performed to mutate every residue of this domain to alanine. The influence of every alanine substitution on the structure and function of LHCII has been investigated biochemically and spectroscopically. The results show that all mutations have little impact on the pigment binding and configuration. However, many mutants presented decreased thermo- or photo-stability compared with the wild type, highlighting the significance of this helix to the stability of LHCII. The most critical residue for stability is W97. The mutant W97A yielded very fragile trimeric pigment protein complexes. The structural analysis revealed that the hydrogen bonding and aromatic interactions between W97, F195, F194 and a water molecule contributed greatly to the stability of LHCII. Moreover, Q103A and F105A have been identified to be able to reinforce the tendency of aggregation in vitro. The structural analysis suggested that the enhancement in aggregation formation for Q103A and F105A might be attributed to the changing hydrophobicity of the region.

  6. Amino acids (L-arginine and L-alanine) passivated CdS nanoparticles: Synthesis of spherical hierarchical structure and nonlinear optical properties

    NASA Astrophysics Data System (ADS)

    Talwatkar, S. S.; Tamgadge, Y. S.; Sunatkari, A. L.; Gambhire, A. B.; Muley, G. G.

    2014-12-01

    CdS nanoparticles (NPs) passivated with amino acids (L-alanine and L-arginine) having spherical hierarchical morphology were synthesized by room temperature wet chemical method. Synthesized NPs were characterized by ultraviolet-visible (UV-vis) spectroscopy to study the variation of band gaps with concentration of surface modifying agents. Increase in band gap has been observed with the increase in concentration of surface modifying agents and was found more prominent for CdS NPs passivated with L-alanine. Powder X-ray diffraction (XRD) and transmission electron microscopy (TEM) analysis were carried out for the study of crystal structure and morphology of CdS NPs. The average particle size of CdS NPs calculated from Debye-Scherer formula was found to less than 5 nm and agrees well with those determined from UV-vis spectra and TEM data. Fourier transform infrared (FT-IR) spectroscopy was performed to know the functional groups of the grown NPs. Peaks in FT-IR spectra indicate the formation of CdS NPs and capping with L-alanine and L-arginine. Photoluminescence spectra of these NPs were also studied. Finally, colloidal solution of CdS-PVAc was subjected to Z-scan experiment under low power cw laser illumination to characterize them for third order nonlinear optical properties. CdS-PVAc colloidal solution shows enhanced nonlinear absorption due to RSA and weak FCA on account of two photon absorption processes triggered by thermal effect.

  7. Amino acid residues in the GerAB protein important in the function and assembly of the alanine spore germination receptor of Bacillus subtilis 168.

    PubMed

    Cooper, Gareth R; Moir, Anne

    2011-05-01

    The paradigm gerA operon is required for endospore germination in response to c-alanine as the sole germinant, and the three protein products, GerAA, GerAB, and GerAC are predicted to form a receptor complex in the spore inner membrane. GerAB shows homology to the amino acid-polyamine-organocation (APC) family of single-component transporters and is predicted to be an integral membrane protein with 10 membrane-spanning helices. Site-directed mutations were introduced into the gerAB gene at its natural location on the chromosome. Alterations to some charged or potential helix-breaking residues within membrane spans affected receptor function dramatically. In some cases, this is likely to reflect the complete loss of the GerA receptor complex, as judged by the absence of the germinant receptor protein GerAC, which suggests that the altered GerAB protein itself may be unstable or that the altered structure destabilizes the complex. Mutants that have a null phenotype for Instituto de Biotecnología de León, INBIOTEC, Parque Científico de León, Av. Real, 1, 24006 León, Spain-alanine germination but retain GerAC protein at near-normal levels are more likely to define amino acid residues of functional, rather than structural, importance. Single-amino-acid substitutions in each of the GerAB and GerAA proteins can prevent incorporation of GerAC protein into the spore; this provides strong evidence that the proteins within a specific receptor interact and that these interactions are required for receptor assembly. The lipoprotein nature of the GerAC receptor subunit is also important; an amino acid change in the prelipoprotein signal sequence in the gerAC1 mutant results in the absence of GerAC protein from the spore.

  8. Limiting values of diffusion coefficients of glycine, alanine, [Formula: see text]-amino butyric acid, norvaline and norleucine in a relevant physiological aqueous medium.

    PubMed

    Rodriguez, Diana M; Verissimo, Luis M P; Barros, Marisa C F; Rodrigues, Daniela F S L; Rodrigo, Maria Melia; Esteso, Miguel A; Romero, Carmen M; Ribeiro, Ana C F

    2017-02-01

    The side chain effect on transport in ionic aqueous salt solutions was investigated for [Formula: see text]-amino acids glycine, alanine, [Formula: see text]-amino butyric acid, norvaline, and norleucine --that together define a chemical homologous series based on the length of the characteristic side chain which increases from zero to four carbons, respectively. Binary mutual diffusion coefficients at infinitesimal concentration in aqueous solutions of NaCl (0.15 mol kg (-1)) are measured by means of Taylor dispersion technique for this series and significant differences were found against previous published results for identical systems in pure water. In this way, NaCl effect on the transport of each amino acid is thus assessed and discussed in terms of salting-out effects. Also, solvated Stokes hydrodynamic radii were computed for the series showing comparable results in water and NaCl solution. The new information should prove useful in the design and characterization of transport-controlled systems in physiological and pharmacological studies.

  9. 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.

  10. Global cellular responses to β-methyl-amino-L-alanine (BMAA) by olfactory ensheathing glial cells (OEC).

    PubMed

    Chiu, Alexander S; Braidy, Nady; Marçal, Helder; Welch, Jeffrey H; Gehringer, Michelle M; Guillemin, Gilles J; Neilan, Brett A

    2015-06-01

    This study utilised a proteomics approach to identify any differential protein expression in a glial cell line, rat olfactory ensheathing cells (OECs), treated with the cyanotoxin β-methylamino-l-alanine (BMAA). Five proteins of interest were identified, namely Rho GDP-dissociation inhibitor 1 (RhoGDP1), Nck-associated protein 1 (NCKAP1), voltage-dependent anion-selective channel protein 1 (VDAC1), 3-hydroxyacyl-CoA dehydrogenase type-2 (3hCoAdh2), and ubiquilin-4 (UBQLN4). Four of these candidates, nuclear receptor subfamily 4 group A member 1 (Nur77), cyclophilin A (CyPA), RhoGDP1 and VDAC1, have been reported to be involved in cell growth. A microarray identified UBQLN4, palladin and CyPA, which have been implicated to have roles in excitotoxicity. Moreover, the NCKAP1, UBQLN4, CyPA and 3hCoAdh2 genes have been associated with abnormal protein aggregation. Differential expression of genes involved in mitochondrial activity, Nur77, 3hCoAdh2, VDAC1 and UBQLN4, were also identified. Confirmatory reverse transcription quantitative PCR (RT-qPCR) analysis of transcripts generated from the genes of interest corroborated the differential expression trends identified in the global protein analysis. BMAA induced cell cycle arrest in the G2/M phase of OEC and apoptosis after 48 h at concentrations of 250 μM and 500 μM. Collectively, this work advances our understanding of the mechanism of BMAA-mediated glial-toxicity in vitro.

  11. Protein association of the neurotoxin and non-protein amino acid BMAA (β-N-methylamino-L-alanine) in the liver and brain following neonatal administration in rats.

    PubMed

    Karlsson, Oskar; Jiang, Liying; Andersson, Marie; Ilag, Leopold L; Brittebo, Eva B

    2014-04-07

    The environmental neurotoxin β-N-methylamino-L-alanine (BMAA) is not an amino acid that is normally found in proteins. Our previous autoradiographic study of (3)H-labeled BMAA in adult mice unexpectedly revealed a tissue distribution similar to that of protein amino acids. The aim of this study was to characterize the distribution of free and protein-bound BMAA in neonatal rat tissues following a short exposure using autoradiographic imaging and ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). The autoradiographic imaging of (14)C-L-BMAA demonstrated a distinct uptake of radioactivity that was retained following acid extraction in tissues with a high rate of cell turnover and/or protein synthesis. The UHPLC-MS/MS analysis conclusively demonstrated a dose-dependent increase of protein-associated BMAA in neonatal rat tissues. The level of protein-associated BMAA in the liver was more than 10 times higher than that in brain regions not fully protected by the blood-brain barrier which may be due to the higher rate of protein synthesis in the liver. In conclusion, this study demonstrated that BMAA was associated with rat proteins suggesting that BMAA may be misincorporated into proteins. However, protein-associated BMAA seemed to be cleared over time, as none of the samples from adult rats had any detectable free or protein-associated BMAA.

  12. Potential transfer of neurotoxic amino acid β-N-methylamino-alanine (BMAA) from mother to infant during breast-feeding: Predictions from human cell lines.

    PubMed

    Andersson, Marie; Ersson, Lisa; Brandt, Ingvar; Bergström, Ulrika

    2017-04-01

    β-N-methylamino-alanine (BMAA) is a non-protein amino acid produced by cyanobacteria, diatoms and dinoflagellates. BMAA has potential to biomagnify in a terrestrial food chain, and to bioaccumulate in fish and shellfish. We have reported that administration of [(14)C]l-BMAA to lactating mice and rats results in a mother to off-spring transfer via the milk. A preferential enantiomer-specific uptake of [(14)C]l-BMAA has also been demonstrated in differentiated murine mammary epithelium HC11 cells. These findings, together with neurotoxic effects of BMAA demonstrated both in vitro and in vivo, highlight the need to determine whether such transfer could also occur in humans. Here, we used four cell lines of human origin to examine and compare the transport of the two BMAA enantiomers in vitro. The uptake patterns of [(14)C]l- and [(14)C]d-BMAA in the human mammary MCF7 cell line were in agreement with the results in murine HC11 cells, suggesting a potential secretion of BMAA into human breast milk. The permeability coefficients for both [(14)C]l- and [(14)C]d-BMAA over monolayers of human intestinal Caco2 cells supported an efficient absorption from the human intestine. As a final step, transport experiments confirmed that [(14)C]l-and [(14)C]d-BMAA can be taken up by human SHSY5Y neuroblastoma cells and even more efficiently by human U343 glioblastoma cells. In competition experiments with various amino acids, the ASCT2 specific inhibitor benzylserine was the most effective inhibitor of [(14)C]l-BMAA uptake tested here. Altogether, our results suggest that BMAA can be transferred from an exposed mother, via the milk, to the brain of the nursed infant.

  13. Three-dimensional structure of Schistosoma japonicum glutathione S-transferase fused with a six-amino acid conserved neutralizing epitope of gp41 from HIV

    NASA Technical Reports Server (NTRS)

    Lim, Kap; Ho, Joseph X.; Keeling, Kim; Gilliland, Gary L.; Ji, Xinhua; Rueker, Florian; Carter, Daniel C.

    1994-01-01

    The 3-dimensional crystal structure of glutathione S-transferase (GST) of Schistosoma japonicum (Sj) fused with a conserved neutralizing epitope on gp41 (glycoprotein, 41 kDa) of human immunodeficiency virus type 1 (HIV-1) was determined at 2.5 A resolution. The structure of the 3-3 isozyme rat GST of the mu gene class was used as a molecular replacement model. The structure consists of a 4-stranded beta-sheet and 3 alpha-helices in domain 1 and 5 alpha-helices in domain 2. The space group of the Sj GST crystal is P4(sub 3)2(sub 1)2 with unit cell dimensions of a = b = 94.7 A, and c = 58.1 A. The crystal has 1 GST monomer per asymmetric unit, and 2 monomers that form an active dimer are related by crystallographic 2-fold symmetry. In the binding site, the ordered structure of reduced glutathione is observed. The gp41 peptide (Glu-Leu-Asp-Lys-Trp-Ala) fused to the C-terminus of Sj GST forms a loop stabilized by symmetry-related GSTs. The Sj GST structure is compared with previously determined GST structures of mammalian gene classes mu, alpha, and pi. Conserved amino acid residues among the 4 GSTs that are important for hydrophobic and hydrophilic interactions for dimer association and glutathione binding are discussed.

  14. Chronic administration of a combination of six herbs inhibits the progression of hyperglycemia and decreases serum lipids and aspartate amino transferase activity in diabetic rats.

    PubMed

    Shafiee-Nick, Reza; Ghorbani, Ahmad; Vafaee Bagheri, Farzaneh; Rakhshandeh, Hassan

    2012-01-01

    The effects of a polyherbal compound, containing six plants (Allium sativum, Cinnamomum zeylanicum, Nigella sativa, Punica granatum, Salvia officinalis and Teucrium polium) were tested on biochemical parameters in streptozotocin-induced diabetic rats. Streptozotocin caused an approximately 3-fold increase in fasting blood sugar level after 2 days. The diabetic control rats showed further increase in blood glucose after 30 days (384 ± 25 mg/dl in day 30 versus 280 ± 12 mg/dl in day 2, P < 0.001). Administration of the compound blocked the increase of blood glucose (272 ± 7 and 269 ± 48 mg/dl at day 2 and day 30, respectively). Also, there was significant difference in the level of triglyceride (60 ± 9 versus 158 ± 37 mg/dl, P < 0.01), total cholesterol (55 ± 2 versus 97 ± 11 mg/dl, P < 0.01) and aspartate amino transferase activity (75 ± 12 versus 129 ± 18 U/L, P < 0.05) between treated rats and diabetic control group. In conclusion, the MSEC inhibited the progression of hyperglycemia and decreased serum lipids and hepatic enzyme activity in diabetic rats. Therefore, it has the potential to be used as a natural product for the management of diabetes.

  15. Chronic Administration of a Combination of Six Herbs Inhibits the Progression of Hyperglycemia and Decreases Serum Lipids and Aspartate Amino Transferase Activity in Diabetic Rats

    PubMed Central

    Shafiee-Nick, Reza; Vafaee Bagheri, Farzaneh; Rakhshandeh, Hassan

    2012-01-01

    The effects of a polyherbal compound, containing six plants (Allium sativum, Cinnamomum zeylanicum, Nigella sativa, Punica granatum, Salvia officinalis and Teucrium polium) were tested on biochemical parameters in streptozotocin-induced diabetic rats. Streptozotocin caused an approximately 3-fold increase in fasting blood sugar level after 2 days. The diabetic control rats showed further increase in blood glucose after 30 days (384 ± 25 mg/dl in day 30 versus 280 ± 12 mg/dl in day 2, P < 0.001). Administration of the compound blocked the increase of blood glucose (272 ± 7 and 269 ± 48 mg/dl at day 2 and day 30, respectively). Also, there was significant difference in the level of triglyceride (60 ± 9 versus 158 ± 37 mg/dl, P < 0.01), total cholesterol (55 ± 2 versus 97 ± 11 mg/dl, P < 0.01) and aspartate amino transferase activity (75 ± 12 versus 129 ± 18 U/L, P < 0.05) between treated rats and diabetic control group. In conclusion, the MSEC inhibited the progression of hyperglycemia and decreased serum lipids and hepatic enzyme activity in diabetic rats. Therefore, it has the potential to be used as a natural product for the management of diabetes. PMID:23304131

  16. Three-dimensional structure of Schistosoma japonicum glutathione S-transferase fused with a six-amino acid conserved neutralizing epitope of gp41 from HIV.

    PubMed Central

    Lim, K.; Ho, J. X.; Keeling, K.; Gilliland, G. L.; Ji, X.; Rüker, F.; Carter, D. C.

    1994-01-01

    The 3-dimensional crystal structure of glutathione S-transferase (GST) of Schistosoma japonicum (Sj) fused with a conserved neutralizing epitope on gp41 (glycoprotein, 41 kDa) of human immunodeficiency virus type 1 (HIV-1) (Muster T et al., 1993, J Virol 67:6642-6647) was determined at 2.5 A resolution. The structure of the 3-3 isozyme rat GST of the mu gene class (Ji X, Zhang P, Armstrong RN, Gilliland GL, 1992, Biochemistry 31:10169-10184) was used as a molecular replacement model. The structure consists of a 4-stranded beta-sheet and 3 alpha-helices in domain 1 and 5 alpha-helices in domain 2. The space group of the Sj GST crystal is P4(3)2(1)2, with unit cell dimensions of a = b = 94.7 A, and c = 58.1 A. The crystal has 1 GST monomer per asymmetric unit, and 2 monomers that form an active dimer are related by crystallographic 2-fold symmetry. In the binding site, the ordered structure of reduced glutathione is observed. The gp41 peptide (Glu-Leu-Asp-Lys-Trp-Ala) fused to the C-terminus of Sj GST forms a loop stabilized by symmetry-related GSTs. The Sj GST structure is compared with previously determined GST structures of mammalian gene classes mu, alpha, and pi. Conserved amino acid residues among the 4 GSTs that are important for hydrophobic and hydrophilic interactions for dimer association and glutathione binding are discussed. PMID:7538846

  17. Three-dimensional structure of Schistosoma japonicum glutathione S-transferase fused with a six-amino acid conserved neutralizing epitope of gp41 from HIV

    NASA Technical Reports Server (NTRS)

    Lim, K.; Ho, J. X.; Keeling, K.; Gilliland, G. L.; Ji, X.; Ruker, F.; Carter, D. C.

    1994-01-01

    The 3-dimensional crystal structure of glutathione S-transferase (GST) of Schistosoma japonicum (Sj) fused with a conserved neutralizing epitope on gp41 (glycoprotein, 41 kDa) of human immunodeficiency virus type 1 (HIV-1) (Muster T et al., 1993, J Virol 67:6642-6647) was determined at 2.5 A resolution. The structure of the 3-3 isozyme rat GST of the mu gene class (Ji X, Zhang P, Armstrong RN, Gilliland GL, 1992, Biochemistry 31:10169-10184) was used as a molecular replacement model. The structure consists of a 4-stranded beta-sheet and 3 alpha-helices in domain 1 and 5 alpha-helices in domain 2. The space group of the Sj GST crystal is P4(3)2(1)2, with unit cell dimensions of a = b = 94.7 A, and c = 58.1 A. The crystal has 1 GST monomer per asymmetric unit, and 2 monomers that form an active dimer are related by crystallographic 2-fold symmetry. In the binding site, the ordered structure of reduced glutathione is observed. The gp41 peptide (Glu-Leu-Asp-Lys-Trp-Ala) fused to the C-terminus of Sj GST forms a loop stabilized by symmetry-related GSTs. The Sj GST structure is compared with previously determined GST structures of mammalian gene classes mu, alpha, and pi. Conserved amino acid residues among the 4 GSTs that are important for hydrophobic and hydrophilic interactions for dimer association and glutathione binding are discussed.

  18. Catalytic Stereoinversion of L-Alanine to Deuterated D-Alanine.

    PubMed

    Moozeh, Kimia; So, Soon Mog; Chin, Jik

    2015-08-03

    A combination of an achiral pyridoxal analogue and a chiral base has been developed for catalytic deuteration of L-alanine with inversion of stereochemistry to give deuterated D-alanine under mild conditions (neutral pD and 25 °C) without the use of any protecting groups. This system can also be used for catalytic deuteration of D-alanine with retention of stereochemistry to give deuterated D-alanine. Thus a racemic mixture of alanine can be catalytically deuterated to give an enantiomeric excess of deuterated D-alanine. While catalytic deracemization of alanine is forbidden by the second law of thermodynamics, this system can be used for catalytic deracemization of alanine with deuteration. Such green and biomimetic approach to catalytic stereocontrol provides insights into efficient amino acid transformations.

  19. Genome-enabled determination of amino acid biosynthesis in Xanthomonas campestris pv. campestris and identification of biosynthetic pathways for alanine, glycine, and isoleucine by 13C-isotopologue profiling.

    PubMed

    Schatschneider, Sarah; Vorhölter, Frank-Jörg; Rückert, Christian; Becker, Anke; Eisenreich, Wolfgang; Pühler, Alfred; Niehaus, Karsten

    2011-10-01

    To elucidate the biosynthetic pathways for all proteinogenic amino acids in Xanthomonas campestris pv. campestris, this study combines results obtained by in silico genome analysis and by (13)C-NMR-based isotopologue profiling to provide a panoramic view on a substantial section of bacterial metabolism. Initially, biosynthesis pathways were reconstructed from an improved annotation of the complete genome of X. campestris pv. campestris B100. This metabolic reconstruction resulted in the unequivocal identification of biosynthesis routes for 17 amino acids in total: arginine, asparagine, aspartate, cysteine, glutamate, glutamine, histidine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine, and valine. Ambiguous pathways were reconstructed from the genome data for alanine, glycine, and isoleucine biosynthesis. (13)C-NMR analyses supported the identification of the metabolically active pathways. The biosynthetic routes for these amino acids were derived from the precursor molecules pyruvate, serine, and pyruvate, respectively. By combining genome analysis and isotopologue profiling, a comprehensive set of biosynthetic pathways covering all proteinogenic amino acids was unraveled for this plant pathogenic bacterium, which plays an important role in biotechnology as a producer of the exopolysaccharide xanthan. The data obtained lay ground for subsequent functional analyses in post-genomics and biotechnology, while the innovative combination of in silico and wet lab technology described here is promising as a general approach to elucidate metabolic pathways.

  20. Inhibitors of alanine racemase enzyme: a review.

    PubMed

    Azam, Mohammed Afzal; Jayaram, Unni

    2016-08-01

    Alanine racemase is a fold type III PLP-dependent amino acid racemase enzyme catalysing the conversion of l-alanine to d-alanine utilised by bacterial cell wall for peptidoglycan synthesis. As there are no known homologs in humans, it is considered as an excellent antibacterial drug target. The standard inhibitors of this enzyme include O-carbamyl-d-serine, d-cycloserine, chlorovinyl glycine, alaphosphin, etc. d-Cycloserine is indicated for pulmonary and extra pulmonary tuberculosis but therapeutic use of drug is limited due to its severe toxic effects. Toxic effects due to off-target affinities of cycloserine and other substrate analogs have prompted new research efforts to identify alanine racemase inhibitors that are not substrate analogs. In this review, an updated status of known inhibitors of alanine racemase enzyme has been provided which will serve as a rich source of structural information and will be helpful in generating selective and potent inhibitor of alanine racemase.

  1. Glutathione Transferases

    PubMed Central

    Dixon, David P.; Edwards, Robert

    2010-01-01

    The 55 Arabidopsis glutathione transferases (GSTs) are, with one microsomal exception, a monophyletic group of soluble enzymes that can be divided into phi, tau, theta, zeta, lambda, dehydroascorbate reductase (DHAR) and TCHQD classes. The populous phi and tau classes are often highly stress inducible and regularly crop up in proteomic and transcriptomic studies. Despite much study on their xenobiotic-detoxifying activities their natural roles are unclear, although roles in defence-related secondary metabolism are likely. The smaller DHAR and lambda classes are likely glutathione-dependent reductases, the zeta class functions in tyrosine catabolism and the theta class has a putative role in detoxifying oxidised lipids. This review describes the evidence for the functional roles of GSTs and the potential for these enzymes to perform diverse functions that in many cases are not “glutathione transferase” activities. As well as biochemical data, expression data from proteomic and transcriptomic studies are included, along with subcellular localisation experiments and the results of functional genomic studies. PMID:22303257

  2. 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.

  3. Solved? The reductive radiation chemistry of alanine.

    PubMed

    Pauwels, Ewald; De Cooman, Hendrik; Waroquier, Michel; Hole, Eli O; Sagstuen, Einar

    2014-02-14

    The structural changes throughout the entire reductive radiation-induced pathway of l-α-alanine are solved on an atomistic level with the aid of periodic DFT and nudged elastic band (NEB) simulations. This yields unprecedented information on the conformational changes taking place, including the protonation state of the carboxyl group in the "unstable" and "stable" alanine radicals and the internal transformation converting these two radical variants at temperatures above 220 K. The structures of all stable radicals were verified by calculating EPR properties and comparing those with experimental data. The variation of the energy throughout the full radiochemical process provides crucial insight into the reason why these structural changes and rearrangements occur. Starting from electron capture, the excess electron quickly localizes on the carbon of a carboxyl group, which pyramidalizes and receives a proton from the amino group of a neighboring alanine molecule, forming a first stable radical species (up to 150 K). In the temperature interval 150-220 K, this radical deaminates and deprotonates at the carboxyl group, the detached amino group undergoes inversion and its methyl group sustains an internal rotation. This yields the so-called "unstable alanine radical". Above 220 K, triggered by the attachment of an additional proton on the detached amino group, the radical then undergoes an internal rotation in the reverse direction, giving rise to the "stable alanine radical", which is the final stage in the reductive radiation-induced decay of alanine.

  4. Production of Alanine by Fusarium moniliforme

    PubMed Central

    Carito, Sebastian L.; Pisano, Michael A.

    1966-01-01

    Fusarium moniliforme grown in a chemically defined medium in submerged culture accumulated amino acids extracellularly. Alanine and glutamic acid were present in greatest amounts, with traces of glycine, lysine, threonine, and valine detectable. Increasing the glucose and urea concentrations of the medium increased yields of alanine. Further increases in alanine production occurred with elevated levels of mineral salts in the medium, whereas the addition of a vitamin mixture proved to be inhibitory. Chemical changes resulting from the growth of F. moniliforme in the final fermentation medium disclosed maximal alanine production, mycelial weight, and glucose consumption after 72 hr of incubation at 28.5 C. Total soluble nitrogen, by contrast, was minimal at the same time period. The pH remained in the alkaline range throughout the fermentation. PMID:5914495

  5. Enzymatic Glycosylation by Transferases

    NASA Astrophysics Data System (ADS)

    Blixt, Ola; Razi, Nahid

    Glycosyltransferases are important biological catalysts in cellular systems generating complex cell surface glycans involved in adhesion and signaling processes. Recent advances in glycoscience have increased the demands to access significant amount of glycans representing the glycome. Glycosyltransferases are now playing a key role for in vitro synthesis of oligosaccharides and the bacterial genome are increasingly utilized for cloning and over expression of active transferases in glycosylation reactions. This chapter highlights the recent progress towards preparative synthesis of oligosaccharides representing terminal sequences of glycoproteins and glycolipids using recombinant transferases. Transferases are also being explored in the context of solid-phase synthesis, immobilized on resins and over expression in vivo by engineered bacteria.

  6. Production of D-Alanine by Corynebacterium fascians

    PubMed Central

    Yamada, Shigeki; Maeshima, Haruko; Wada, Mitsuru; Chibata, Ichiro

    1973-01-01

    A strain identified as Corynebacterium fascians was found to accumulate extracellular D-alanine from glycerol. Cultural conditions for the accumulation of D-alanine were investigated and, as a result, a yield of 7 g of D-alanine per liter was obtained after a 96-h incubation in a medium containing 5% glycerol, 4% (NH4)2HPO4, and 0.3% corn steep liquor. Optical purity of D-alanine was dependent upon the concentration of corn steep liquor. At the optimal condition, almost optically pure D-alanine was formed and readily isolated (5 g/liter) from the fermentation broth. The product was not contaminated with any detectable amount of other amino acids, except for glycine which was present at a concentration of less than 1 percent. PMID:4699220

  7. Atomic Layer Deposition of L-Alanine Polypeptide

    SciTech Connect

    Fu, Yaqin; Li, Binsong; Jiang, Ying-Bing; Dunphy, Darren R.; Tsai, Andy; Tam, Siu-Yue; Fan, Hongyou Y.; Zhang, Hongxia; Rogers, David; Rempe, Susan; Atanassov, Plamen; Cecchi, Joseph L.; Brinker, C. Jeffrey

    2014-10-30

    L-Alanine polypeptide thin films were synthesized via atomic layer deposition (ALD). Rather, instead of using an amino acid monomer as the precursor, an L-alanine amino acid derivatized with a protecting group was used to prevent self-polymerization, increase the vapor pressure, and allow linear cycle-by-cycle growth emblematic of ALD. Moreover, the successful deposition of a conformal polypeptide film has been confirmed by FTIR, TEM, and Mass Spectrometry, and the ALD process has been extended to polyvaline.

  8. Farnesyl transferase inhibitors as anticancer agents.

    PubMed

    Haluska, P; Dy, G K; Adjei, A A

    2002-09-01

    Protein farnesylation catalysed by the enzyme farnesyl protein transferase involves the addition of a 15-carbon farnesyl group to conserved amino acid residues at the carboxyl terminus of certain proteins. Protein substrates of farnesyl transferase include several G-proteins, which are critical intermediates of cell signalling and cytoskeletal organisation such as Ras, Rho, PxF and lamins A and B. Activated Ras proteins trigger a cascade of phosphorylation events through sequential activation of the PI3 kinase/AKT pathway, which is critical for cell survival, and the Raf/Mek/Erk kinase pathway that has been implicated in cell proliferation. Ras mutations which encode for constitutively activated proteins are found in 30% of human cancers. Because farnesylation of Ras is required for its transforming and proliferative activity, the farnesyl protein transferase inhibitors were designed as anticancer agents to abrogate Ras function. However, current evidence suggests that the anticancer activity of the farnesyl transferase inhibitors may not be simply due to Ras inhibition. This review will discuss available clinical data on three of these agents that are currently undergoing clinical trials.

  9. Synthesis and GGCT Inhibitory Activity of N-Glutaryl-L-alanine Analogues.

    PubMed

    Ii, Hiromi; Yoshiki, Tatsuhiro; Hoshiya, Naoyuki; Uenishi, Jun'ichi

    2016-01-01

    γ-Glutamylcyclotransferase (GGCT) is an important enzyme that cleaves γ-glutamyl-amino acid in the γ-glutamyl cycle to release 5-oxoproline and amino acid. Eighteen N-acyl-L-alanine analogues including eleven new compounds have been synthesized and examined for their inhibitory activity against recombinant human GGCT protein. Simple N-glutaryl-L-alanine was found to be the most potent inhibitor for GGCT. Other N-glutaryl-L-alanine analogues having methyl and dimethyl substituents at the 2-position were moderately effective, while N-(3R-aminoglutary)-L-alanine, the substrate having an (R)-amino group at the 3-position or N-(N-methyl-3-azaglutaryl)-L-alanine, the substrate having an N-methyl substituent on the 3-azaglutaryl carbon, in constract, exhibited excellent inhibition properties.

  10. Mitochondrial carnitine palmitoyl transferase-II inactivity aggravates lipid accumulation in rat hepatocarcinogenesis

    PubMed Central

    Gu, Juan-Juan; Yao, Min; Yang, Jie; Cai, Yin; Zheng, Wen-Jie; Wang, Li; Yao, Deng-Bing; Yao, Deng-Fu

    2017-01-01

    AIM To investigate the dynamic alteration of mitochondrial carnitine palmitoyl transferase II (CPT-II) expression during malignant transformation of rat hepatocytes. METHODS Sprague-Dawley male rats were fed with normal, high fat (HF), and HF containing 2-fluorenylacetamide (2-FAA) diet, respectively. According to the Hematoxylin and Eosin staining of livers, rats were divided into control, fatty liver, degeneration, precancerous, and cancerous groups. Liver lipids were dyed with Oil Red O, CPT-II alterations were analyzed by immunohistochemistry, and compared with CPT-II specific concentration (μg/mg protein). Levels of total cholesterol (Tch), triglyceride (TG), and amino-transferases [alanine aminotransferase (ALT), aspartate aminotransferase (AST)] were determined by the routine methods. RESULTS After intake of HF and/or HF+2-FAA diets, the rat livers showed mass lipid accumulation. The lipid level in the control group was significantly lower than that in other groups. The changes of serum TG and Tch levels were abnormally increasing, 2-3 times more than those in the controls (P < 0.05). During the rat liver morphological changes from normal to cancer development process with hepatocyte injury, serum AST and ALT levels were significantly higher (4-8 times, P < 0.05) than those in the control group. The specific concentration of CPT-II in liver tissues progressively decreased during hepatocyte malignant transformation, with the lowest CPT-II levels in the cancer group than in any of the other groups (P < 0.05). CONCLUSION Low CPT-II expression might lead to abnormal hepatic lipid accumulation, which should promote the malignant transformation of hepatocytes. PMID:28127199

  11. Optical and Spectral Studies on β Alanine Metal Halide Hybrid Crystals

    NASA Astrophysics Data System (ADS)

    Sweetlin, M. Daniel; Selvarajan, P.; Perumal, S.; Ramalingom, S.

    2011-10-01

    We have synthesized and grown β alanine metal halide hybrid crystals viz. β alanine cadmium chloride (BACC), an amino acid transition metal halide complex crystal and β alanine potassium chloride (BAPC), an amino acid alkali metal halide complex crystal by slow evaporation method. The grown crystals were found to be transparent and have well defined morphology. The optical characteristics of the grown crystals were carried out with the help of UV-Vis Spectroscopy. The optical transmittances of the spectrums show that BAPC is more transparent than BACC. The Photoluminescence of the materials were determined by the Photoluminescent Spectroscopy

  12. [Effects of ß-alanine supplementation on athletic performance].

    PubMed

    Domínguez, Raúl; Hernández Lougedo, Juan; Maté-Muñoz, José Luis; Garnacho-Castaño, Manuel Vicente

    2014-10-06

    Carnosine, dipeptide formed by amino acids ß-alanine and L-histidine, has important physiological functions among which its antioxidant and related memory and learning. However, in connection with the exercise, the most important functions would be associated with muscle contractility, improving calcium sensitivity in muscle fibers, and the regulatory function of pH. Thus, it is proposed that carnosine is the major intracellular buffer, but could contribute to 7-10% in buffer or buffer capacity. Since carnosine synthesis seems to be limited by the availability of ß-alanine supplementation with this compound has been gaining increasing popularity among the athlete population. Therefore, the objective of this study literature review was to examine all those research works have shown the effect of ß-alanine supplementation on athletic performance. Moreover, it also has attempted to establish a specific dosage that maximizing the potential benefits, minimize paresthesia, the main side effect presented in response to supplementation.

  13. Alanine aminotransferase controls seed dormancy in barley

    PubMed Central

    Sato, Kazuhiro; Yamane, Miki; Yamaji, Nami; Kanamori, Hiroyuki; Tagiri, Akemi; Schwerdt, Julian G.; Fincher, Geoffrey B.; Matsumoto, Takashi; Takeda, Kazuyoshi; Komatsuda, Takao

    2016-01-01

    Dormancy allows wild barley grains to survive dry summers in the Near East. After domestication, barley was selected for shorter dormancy periods. Here we isolate the major seed dormancy gene qsd1 from wild barley, which encodes an alanine aminotransferase (AlaAT). The seed dormancy gene is expressed specifically in the embryo. The AlaAT isoenzymes encoded by the long and short dormancy alleles differ in a single amino acid residue. The reduced dormancy allele Qsd1 evolved from barleys that were first domesticated in the southern Levant and had the long dormancy qsd1 allele that can be traced back to wild barleys. The reduced dormancy mutation likely contributed to the enhanced performance of barley in industrial applications such as beer and whisky production, which involve controlled germination. In contrast, the long dormancy allele might be used to control pre-harvest sprouting in higher rainfall areas to enhance global adaptation of barley. PMID:27188711

  14. Plant glutathione transferases

    PubMed Central

    Dixon, David P; Lapthorn, Adrian; Edwards, Robert

    2002-01-01

    The soluble glutathione transferases (GSTs, EC 2.5.1.18) are encoded by a large and diverse gene family in plants, which can be divided on the basis of sequence identity into the phi, tau, theta, zeta and lambda classes. The theta and zeta GSTs have counterparts in animals but the other classes are plant-specific and form the focus of this article. The genome of Arabidopsis thaliana contains 48 GST genes, with the tau and phi classes being the most numerous. The GST proteins have evolved by gene duplication to perform a range of functional roles using the tripeptide glutathione (GSH) as a cosubstrate or coenzyme. GSTs are predominantly expressed in the cytosol, where their GSH-dependent catalytic functions include the conjugation and resulting detoxification of herbicides, the reduction of organic hydroperoxides formed during oxidative stress and the isomerization of maleylacetoacetate to fumarylacetoacetate, a key step in the catabolism of tyrosine. GSTs also have non-catalytic roles, binding flavonoid natural products in the cytosol prior to their deposition in the vacuole. Recent studies have also implicated GSTs as components of ultraviolet-inducible cell signaling pathways and as potential regulators of apoptosis. Although sequence diversification has produced GSTs with multiple functions, the structure of these proteins has been highly conserved. The GSTs thus represent an excellent example of how protein families can diversify to fulfill multiple functions while conserving form and structure. PMID:11897031

  15. Determination of D- and L-alanine concentrations using a pyruvic acid sensor.

    PubMed

    Inaba, Yohei; Hamada-Sato, Naoko; Kobayashi, Takeshi; Imada, Chiaki; Watanabe, Etsuo

    2003-08-01

    The concentrations of D- and L-alanine in bivalves are useful as indicators of environmental pollution. Amino acid oxidase with a low substrate specificity catalyzes the oxidation of various amino acids. Among the various amino acids, pyruvic acid can be generated from alanine only by the catalytic oxidative reaction of this oxidase. Therefore, in this study, the concentrations of D- and L-alanine were determined from the concentration of pyruvic acid, which was determined from the consumption of oxygen based on the oxidative reaction of pyruvate oxidase. From this point of view, there is a very strong possibility that biosensors utilizing enzymes with a low substrate specificity can be developed. The results obtained were as follows. (1) The optimum conditions for the use of pyruvic acid sensor were as follows: temperature of 25 degrees C, pH of 6.8, flow rate of 0.1 ml/min, thiamin diphosphate concentration of 1.5 mM, and injection volume of 50 microl. (2) D-Alanine and L-alanine optimally reacted with D- and L-amino acid oxidase at 30 degrees C, pH 8.2, for 30 min and at 37 degrees C, pH 7.8, for 90 min, respectively. (3) The linear relationships between the concentrations of D- and L-alanine and the output of the sensor were obtained at 3.56-106.8 microg of D-alanine and 5.34-71.3 microg of L-alanine. (4) The concentrations of D- and L-alanine in Meretrix iusoria, Patinopecten yessonsi, and Corbicula leana obtained by the proposed assay were in good agreement with those determined by a conventional method.

  16. Molecular Cloning of Adenosinediphosphoribosyl Transferase.

    DTIC Science & Technology

    1987-09-08

    ACCESSION NO.D,. 03261102F 2312 A~5 11. TITLE (include Securqt Classification) 0 Molecular Cloning of Adenosinediphosphoribosyl Transferase 12. PERSONAL...I’:- AFOSR.Tlt. 8 7 - 0 9 8,2 0IL * pi AFOSR- 85 -0377 PROGRESS REPORT Molecular Cloning of Adenosinediphosphoribosyl Transferase 5." Period of...Pharmacology and the Cardiovascular Research Institute September 8, 1987 .’, 5.’- "’S ". -f, AFOSR - 85 -0377 PROGRESS REPORT Molecular Cloning of

  17. Blood Test: Comprehensive Metabolic Panel (CMP)

    MedlinePlus

    ... a decrease in kidney function. Alkaline phosphatase (ALP) , alanine amino transferase (ALT) , aspartate amino transferase (AST) , and ... Test: Glucose Kidneys and Urinary Tract Blood Test: Alanine Aminotransferase (ALT, or SGPT) Blood Test: Basic Metabolic ...

  18. How similar is the electronic structures of β-lactam and alanine?

    NASA Astrophysics Data System (ADS)

    Chatterjee, Subhojyoti; Ahmed, Marawan; Wang, Feng

    2016-02-01

    The C1s spectra of β-lactam i.e. 2-azetidinone (C3H5NO), a drug and L-alanine (C3H7NO2), an amino acid, exhibit striking similarities, which may be responsible for the competition between 2-azetidinone and the alanyl-alanine moiety in biochemistry. The present study is to reveal the degree of similarities and differences between their electronic structures of the two model molecular pairs. It is found that the similarities in C1s and inner valence binding energy spectra are due to their bonding connections but other properties such as ring structure (in 2-azetidinone) and chiral carbon (alanine) can be very different. Further, the inner valence region of ionization potential greater than 18 eV for 2-azetidinone and alanine is also significantly similar. Finally the strained lactam ring exhibits more chemical reactivity measured at all non-hydrogen atoms by Fukui functions with respect to alanine.

  19. Physiological hypercortisolemia increases proteolysis, glutamine, and alanine production

    SciTech Connect

    Darmaun, D.; Matthews, D.E.; Bier, D.M. Cornell Univ. Medical College, New York, NY )

    1988-09-01

    Physiological elevations of plasma cortisol levels, as are encountered in stress and severe trauma, were produced in six normal subjects by infusing them with hydrocortisone for 64 h. Amino acid kinetics were measured in the postabsorptive state using three 4-h infusions of L-(1-{sup 13}C)leucine, L-phenyl({sup 2}H{sub 5})phenylalanine, L-(2-{sup 15}N)glutamine, and L-(1-{sup 13}C)alanine tracers (1) before, (2) at 12 h, and (3) at 60 h of cortisol infusion. Before and throughout the study, the subjects ate a normal diet of adequate protein and energy intake. The cortisol infusion raised plasma cortisol levels significantly from 10 {plus minus} 1 to 32 {plus minus} 4 {mu}g/dl, leucine flux from 83 {plus minus} 3 to 97 {plus minus} 3 {mu}mol{center dot}kg{sup {minus}1}{center dot}h{sup {minus}1}, and phenylalanine flux from 34 {plus minus} 1 to 39 {plus minus} 1 (SE) {mu}mol{center dot}kg{sup {minus}1}{center dot}h{sup {minus}1} after 12 h of cortisol infusion. These increases were maintained until the cortisol infusion was terminated. These nearly identical 15% increases in two different essential amino acid appearance rates are reflective of increased whole body protein breakdown. Glutamine flux rose by 12 h of cortisol infusion and remained elevated at the same level at 64 h. The increase in flux was primarily due to a 55% increase in glutamine de novo synthesis. Alanine flux increased with acute hypercortisolemia and increased further at 60 h of cortisol infusion, a result primarily of increased alanine de novo synthesis. Insulin, alanine, and lactate plasma levels responded similarly with significant rises between the acute and chronic periods of cortisol infusion. Thus hypercortisolemia increases both protein breakdown and the turnover of important nonessential amino acids for periods of up to 64 h.

  20. Structures of yeast glutathione-S-transferase Gtt2 reveal a new catalytic type of GST family

    PubMed Central

    Ma, Xiao-Xiao; Jiang, Yong-Liang; He, Yong-Xing; Bao, Rui; Chen, Yuxing; Zhou, Cong-Zhao

    2009-01-01

    Glutathione-S-transferases (GSTs) are ubiquitous detoxification enzymes that catalyse the conjugation of electrophilic substrates to glutathione. Here, we present the crystal structures of Gtt2, a GST of Saccharomyces cerevisiae, in apo and two ligand-bound forms, at 2.23 Å, 2.20 Å and 2.10 Å, respectively. Although Gtt2 has the overall structure of a GST, the absence of the classic catalytic essential residues—tyrosine, serine and cysteine—distinguishes it from all other cytosolic GSTs of known structure. Site-directed mutagenesis in combination with activity assays showed that instead of the classic catalytic residues, a water molecule stabilized by Ser129 and His123 acts as the deprotonator of the glutathione sulphur atom. Furthermore, only glycine and alanine are allowed at the amino-terminus of helix-α1 because of stereo-hindrance. Taken together, these results show that yeast Gtt2 is a novel atypical type of cytosolic GST. PMID:19851333

  1. Formation of simple biomolecules from alanine in ocean by impacts

    NASA Astrophysics Data System (ADS)

    Umeda, Y.; Sekine, T.; Furukawa, Y.; Kakegawa, T.; Kobayashi, T.

    2013-12-01

    The biomolecules on the Earth are thought either to have originated from the extraterrestrial parts carried with flying meteorites or to have been formed from the inorganic materials on the Earth through given energy. From the standpoint to address the importance of impact energy, it is required to simulate experimentally the chemical reactions during impacts, because violent impacts may have occurred 3.8-4.0 Gyr ago to create biomolecules initially. It has been demonstrated that shock reactions among ocean (H2O), atmospheric nitrogen, and meteoritic constitution (Fe) can induce locally reduction environment to form simple bioorganic molecules such as ammonia and amino acid (Nakazawa et al., 2005; Furukawa et al., 2009). We need to know possible processes for alanine how chemical reactions proceed during repeated impacts and how complicated biomolecules are formed. Alanine can be formed from glycine (Umeda et al., in preparation). In this study, we carried out shock recovery experiments at pressures of 4.4-5.7 GPa to investigate the chemical reactions of alanine. Experiments were carried out with a propellant gun. Stainless steel containers (30 mm in diameter, 30 mm long) with 13C-labeled alanine aqueous solution immersed in olivine or hematite powders were used as targets. Air gap was present in the sample room (18 mm in diameter, 2 mm thick) behind the sample. The powder, solution, and air represent meteorite, ocean, and atmosphere on early Earth, respectively. Two powders of olivine and hematite help to keep the oxygen fugacity low and high during experiments, respectively in order to investigate the effect of oxygen fugacity on chemical processes of alanine. The recovered containers, after cleaned completely, were immersed into liquid nitrogen to freeze sample solution and then we drilled on the impact surface to extract water-soluble run products using pure water. Thus obtained products were analyzed by LC/MS for four amino acids (glycine, alanine, valine, and

  2. Glutathione transferases and neurodegenerative diseases.

    PubMed

    Mazzetti, Anna Paola; Fiorile, Maria Carmela; Primavera, Alessandra; Lo Bello, Mario

    2015-03-01

    There is substantial agreement that the unbalance between oxidant and antioxidant species may affect the onset and/or the course of a number of common diseases including Parkinson's and Alzheimer's diseases. Many studies suggest a crucial role for oxidative stress in the first phase of aging, or in the pathogenesis of various diseases including neurological ones. Particularly, the role exerted by glutathione and glutathione-related enzymes (Glutathione Transferases) in the nervous system appears more relevant, this latter tissue being much more vulnerable to toxins and oxidative stress than other tissues such as liver, kidney or muscle. The present review addresses the question by focusing on the results obtained by specimens from patients or by in vitro studies using cells or animal models related to Parkinson's and Alzheimer's diseases. In general, there is an association between glutathione depletion and Parkinson's or Alzheimer's disease. In addition, a significant decrease of glutathione transferase activity in selected areas of brain and in ventricular cerebrospinal fluid was found. For some glutathione transferase genes there is also a correlation between polymorphisms and onset/outcome of neurodegenerative diseases. Thus, there is a general agreement about the protective effect exerted by glutathione and glutathione transferases but no clear answer about the mechanisms underlying this crucial role in the insurgence of neurodegenerative diseases.

  3. Degradation of glycine and alanine on irradiated quartz.

    PubMed

    Pawlikowski, Maciej; Benko, Aleksandra; Wróbel, Tomasz P

    2013-04-01

    Recent researches suggest participation of minerals in the formation of life under primordial conditions. Among all of the minerals, quartz seems to be one of the most probable to take part in such processes. However, an external source of energy is needed, e.g. electric discharge. A device simulating the proposed conditions was designed and was used to simulate prebiotic conditions. Investigation of processes occurring during the stimulation of quartz with electric discharge was studied by means of Ultraviolet-visible (UV-VIS) spectroscopy, in order to monitor the generation kinetics of free radicals. Additionally, infrared spectroscopy was applied to identify chemical reaction products created in a solution of alanine or glycine, in the presence of quartz treated with electric discharge. Formation of increased amounts of free radicals, compared to experiments performed without quartz and/or amino acid, is reported, along with identification of possible degradation products of alanine. No synthetic reactions were observed.

  4. Crystal Structures of Aedes Aegypt Alanine Glyoxylate Aminotransferase

    SciTech Connect

    Han,Q.; Robinson, H.; Gao, Y.; Vogelaar, N.; Wilson, S.; Rizzi, M.; Li, J.

    2006-01-01

    Mosquitoes are unique in having evolved two alanine glyoxylate aminotransferases (AGTs). One is 3-hydroxykynurenine transaminase (HKT), which is primarily responsible for catalyzing the transamination of 3-hydroxykynurenine (3-HK) to xanthurenic acid (XA). Interestingly, XA is used by malaria parasites as a chemical trigger for their development within the mosquito. This 3-HK to XA conversion is considered the major mechanism mosquitoes use to detoxify the chemically reactive and potentially toxic 3-HK. The other AGT is a typical dipteran insect AGT and is specific for converting glyoxylic acid to glycine. Here we report the 1.75{angstrom} high-resolution three-dimensional crystal structure of AGT from the mosquito Aedes aegypti (AeAGT) and structures of its complexes with reactants glyoxylic acid and alanine at 1.75 and 2.1{angstrom} resolution, respectively. This is the first time that the three-dimensional crystal structures of an AGT with its amino acceptor, glyoxylic acid, and amino donor, alanine, have been determined. The protein is dimeric and adopts the type I-fold of pyridoxal 5-phosphate (PLP)-dependent aminotransferases. The PLP co-factor is covalently bound to the active site in the crystal structure, and its binding site is similar to those of other AGTs. The comparison of the AeAGT-glyoxylic acid structure with other AGT structures revealed that these glyoxylic acid binding residues are conserved in most AGTs. Comparison of the AeAGT-alanine structure with that of the Anopheles HKT-inhibitor complex suggests that a Ser-Asn-Phe motif in the latter may be responsible for the substrate specificity of HKT enzymes for 3-HK.

  5. Crystal structures of Aedes aegypti alanine glyoxylate aminotransferase.

    PubMed

    Han, Qian; Robinson, Howard; Gao, Yi Gui; Vogelaar, Nancy; Wilson, Scott R; Rizzi, Menico; Li, Jianyong

    2006-12-01

    Mosquitoes are unique in having evolved two alanine glyoxylate aminotransferases (AGTs). One is 3-hydroxykynurenine transaminase (HKT), which is primarily responsible for catalyzing the transamination of 3-hydroxykynurenine (3-HK) to xanthurenic acid (XA). Interestingly, XA is used by malaria parasites as a chemical trigger for their development within the mosquito. This 3-HK to XA conversion is considered the major mechanism mosquitoes use to detoxify the chemically reactive and potentially toxic 3-HK. The other AGT is a typical dipteran insect AGT and is specific for converting glyoxylic acid to glycine. Here we report the 1.75A high-resolution three-dimensional crystal structure of AGT from the mosquito Aedes aegypti (AeAGT) and structures of its complexes with reactants glyoxylic acid and alanine at 1.75 and 2.1A resolution, respectively. This is the first time that the three-dimensional crystal structures of an AGT with its amino acceptor, glyoxylic acid, and amino donor, alanine, have been determined. The protein is dimeric and adopts the type I-fold of pyridoxal 5-phosphate (PLP)-dependent aminotransferases. The PLP co-factor is covalently bound to the active site in the crystal structure, and its binding site is similar to those of other AGTs. The comparison of the AeAGT-glyoxylic acid structure with other AGT structures revealed that these glyoxylic acid binding residues are conserved in most AGTs. Comparison of the AeAGT-alanine structure with that of the Anopheles HKT-inhibitor complex suggests that a Ser-Asn-Phe motif in the latter may be responsible for the substrate specificity of HKT enzymes for 3-HK.

  6. Effect of abomasal glucose infusion on alanine metabolism and urea production in sheep.

    PubMed

    Obitsu, T; Bremner, D; Milne, E; Lobley, G E

    2000-08-01

    The effect of abomasal infusion of glucose (120 kJ/d per kg body weight (BW)0.75, 758 mmol/d) on urea production, plasma alanine-N flux rate and the conversion of alanine-N to urea was studied in sheep offered a low-N diet at limited energy intake (500 kJ/d per kg BW0.75), based on hay and grass pellets. Glucose provision reduced urinary N (P = 0.040) and urea (P = 0.009) elimination but this was offset by poorer N digestibility. Urea-N production was significantly reduced (822 v. 619 mmol/d, P = 0.024) by glucose while plasma alanine-N flux rate was elevated (295 v. 342 mmol/d, P = 0.011). The quantity of urea-N derived from alanine tended to be decreased by glucose (127 v. 95 mmol/d) but the fraction of urea production from alanine was unaltered (15%). Plasma urea and alanine concentrations (plus those of the branched chain amino acids) decreased in response to exogenous glucose, an effect probably related to enhanced anabolic usage of amino acids and lowered urea production.

  7. [Raman scattering study of DL-alanine].

    PubMed

    Gong, Yan; Wang, Wen-qing

    2006-01-01

    Studies of Raman vibration spectra are useful to obtaining information on biomolecular crystals. The cell dimensions of the L- and DL-alanine crystals are nearly identical, and both structures belong to the orthorhombic system, but the space group is P2(1) 2(1) 2(1) for the L-isomer, and Pna2(1) for the racemate crystal. The Raman spectrum of L-alanine has been measured by many authors. The present work is focusing on the Raman scattering study of DL-alanine powder. Based on the analysis of the differences between DL-alanine and L-alanine Raman spectra, the authors obtained indispensable information on hydrogen bond and the motion of the molecular conformation in alanine crystals.

  8. Alanine racemase mutants of Mycobacterium tuberculosis require D-alanine for growth and are defective for survival in macrophages and mice.

    PubMed

    Awasthy, Disha; Bharath, Sowmya; Subbulakshmi, Venkita; Sharma, Umender

    2012-02-01

    Alanine racemase (Alr) is an essential enzyme in most bacteria; however, some species (e.g. Listeria monocytogenes) can utilize d-amino acid transaminase (Dat) to generate d-alanine, which renders Alr non-essential. In addition to the conflicting reports on gene knockout of alr in Mycobacterium smegmatis, a recent study concluded that depletion of Alr does not affect the growth of M. smegmatis. In order to get an unambiguous answer on the essentiality of Alr in Mycobacterium tuberculosis and validate it as a drug target in vitro and in vivo, we have inactivated the alr gene of M. tuberculosis and found that it was not possible to generate an alr knockout in the absence of a complementing gene copy or d-alanine in the growth medium. The growth kinetics of the alr mutant revealed that M. tuberculosis requires very low amounts of d-alanine (5-10 µg ml(-1)) for optimum growth. Survival kinetics of the mutant in the absence of d-alanine indicated that depletion of this amino acid results in rapid loss of viability. The alr mutant was found to be defective for growth in macrophages. Analysis of phenotype in mice suggested that non-availability of d-alanine in mice leads to clearance of bacteria followed by stabilization of bacterial number in lungs and spleen. Additionally, reversal of d-cycloserine inhibition in the presence of d-alanine in M. tuberculosis suggested that Alr is the primary target of d-cycloserine. Thus, Alr of M. tuberculosis is a valid drug target and inhibition of Alr alone should result in loss of viability in vitro and in vivo.

  9. Vibrational dynamics of crystalline L-alanine

    SciTech Connect

    Bordallo, H.N.; Eckert, J.; Barthes, M.

    1997-11-01

    The authors report a new, complete vibrational analysis of L-alanine and L-alanine-d{sub 4} which utilizes IINS intensities in addition to frequency information. The use of both isotopomers resulted in a self-consistent force field for and assignment of the molecular vibrations in L-alanine. Some details of the calculation as well as a comparison of calculated and observed IINS spectra are presented. The study clarifies a number of important issues on the vibrational dynamics of this molecule and presents a self-consistent force field for the molecular vibrations in crystalline L-alanine.

  10. Interactions of L-alanine with alumina as studied by vibrational spectroscopy.

    PubMed

    Garcia, Ana R; de Barros, Ricardo Brito; Fidalgo, Alexandra; Ilharco, Laura M

    2007-09-25

    The interactions of L-alanine with gamma- and alpha-alumina have been investigated by diffuse reflectance infrared Fourier transform spectroscopy (DRIFTS). L-alanine/alumina samples were dried from aqueous suspensions, at 36.5 degrees C, with two amino acid concentrations (0.4 and 0.8 mmol g-1) and at different pH values (1, 6, and 13). The vibrational spectra proved that the nature of L-alanine interactions with both aluminas is the same (hydrogen bonding), although the groups involved depend on the L-alanine form and on alumina surface groups, both controlled by the pH. For samples prepared at pH 1, cationic L-alanine [CH3CH(NH3+)COOH] displaces physisorbed water from alumina, and strong hydrogen bonds are established between the carbonyl groups of alanine, as electron donors, and the surface Al-OH2+ groups of alumina. This occurs at the expense of alanine dimer dissociation and breaking of intramolecular bonds. When samples are prepared at pH 6, the interacting groups are Al-OH2+ and the carboxylate groups of zwitterionic L-alanine [CH3CH(NH3+)COO-]. The affinity of L-alanine toward alumina decreases, as the strong NH3+...-OOC intermolecular hydrogen bonds prevail over the interactions with alumina. Thus, for a load of 0.8 mmol g-1, phase segregation is observed. On alpha-alumina, crystal deposition is even observed for a load of 0.4 mmol g-1. At pH 13, the carboxylate groups of anionic L-alanine [CH3CH(NH2)COO-] are not affected by alumina. Instead, hydrogen bond interactions occur between NH2 and the Al-OH surface groups of the substrate. Complementary N2 adsorption-desorption isotherms showed that adsorption of L-alanine occurs onto the alumina pore network for samples prepared at pH 1 and 13, whereas at pH 6 the amino acid/alumina interactions are not strong enough to promote adsorption. The mesoporous structure and the high specific surface area of gamma-alumina make it a more efficient substrate for adsorption of L-alanine. For each alumina, however, it is

  11. Microbial production of amino acids in Japan.

    PubMed

    Kumagai, H

    2000-01-01

    The microbial biotechnology of amino acids production which was developed and industrialized in Japan have been summarized. The amino acids include L-glutamic acid, L-lysine, L-threonine, L-aspartic acid, L-alanine, L-cysteine, L-dihydroxyphenylalanine, D-p-hydroxyphenyl-glycine, and hydroxy-L-proline.

  12. Protein-based peptide-bond formation by aminoacyl-tRNA protein transferase.

    PubMed

    Watanabe, Kazunori; Toh, Yukimatsu; Suto, Kyoko; Shimizu, Yoshihiro; Oka, Natsuhisa; Wada, Takeshi; Tomita, Kozo

    2007-10-18

    Eubacterial leucyl/phenylalanyl-tRNA protein transferase (LF-transferase) catalyses peptide-bond formation by using Leu-tRNA(Leu) (or Phe-tRNA(Phe)) and an amino-terminal Arg (or Lys) of a protein, as donor and acceptor substrates, respectively. However, the catalytic mechanism of peptide-bond formation by LF-transferase remained obscure. Here we determine the structures of complexes of LF-transferase and phenylalanyl adenosine, with and without a short peptide bearing an N-terminal Arg. Combining the two separate structures into one structure as well as mutation studies reveal the mechanism for peptide-bond formation by LF-transferase. The electron relay from Asp 186 to Gln 188 helps Gln 188 to attract a proton from the alpha-amino group of the N-terminal Arg of the acceptor peptide. This generates the attacking nucleophile for the carbonyl carbon of the aminoacyl bond of the aminoacyl-tRNA, thus facilitating peptide-bond formation. The protein-based mechanism for peptide-bond formation by LF-transferase is similar to the reverse reaction of the acylation step observed in the peptide hydrolysis reaction by serine proteases.

  13. Glucose and Alanine Metabolism in Children with Maple Syrup Urine Disease

    PubMed Central

    Haymond, Morey W.; Ben-Galim, Ehud; Strobel, Karen E.

    1978-01-01

    In vitro studies have suggested that catabolism of branched chain amino acids is linked with alanine and glutamine formed in, and released from, muscle. To explore this possibility in vivo, static and kinetic studies were performed in three patients with classical, and one patient with partial, branched chain α-ketoacid decarboxylase deficiency (maple syrup urine disease, MSUD) and compared to similar studies in eight age-matched controls. The subjects underwent a 24-30-h fast, and a glucose-alanine flux study using stable isotopes. Basal plasma leucine concentrations were elevated (P <0.001) in patients with MSUD (1,140±125 μM vs. 155±18 μM in controls); and in contrast to the controls, branched chain amino acid concentrations in plasma increased during the fast in the MSUD patients. Basal plasma alanine concentrations were lower (P <0.01) in patients with classical MSUD (153±8 μM vs. 495±27 μM in controls). This discrepancy was maintained throughout the fast despite a decrease in alanine concentrations in both groups. Plasma alanine and leucine concentrations in the patient with partial MSUD were intermediate between those of the controls and the subjects with the classical form of the disease. Circulating ketone bodies and glucoregulatory hormones concentrations were similar in the MSUD and normal subjects during the fast. Alanine flux rates in two patients with classical MSUD (3.76 and 4.00 μmol/Kg per min) and the patient with partial MSUD (5.76 μmol/Kg per min) were clearly lower than those of the controls (11.72±2.53 [SD] μmol/Kg per min). After short-term starvation, glucose flux and fasting concentrations were similar in the MSUD patients and normal subjects. These data indicate that branched chain amino acid catabolism is an important rate limiting event for alanine production in vivo. PMID:670400

  14. Mutation of glycine receptor subunit creates beta-alanine receptor responsive to GABA.

    PubMed

    Schmieden, V; Kuhse, J; Betz, H

    1993-10-08

    The amino acid at position 160 of the ligand-binding subunit, alpha 1, is an important determinant of agonist and antagonist binding to the glycine receptor. Exchange of the neighboring residues, phenylalanine at position 159 and tyrosine at position 161, increased the efficacy of amino acid agonists. Whereas wild-type alpha 1 channels expressed in Xenopus oocytes required 0.7 millimolar beta-alanine for a half-maximal response, the doubly mutated (F159Y,Y161F) alpha 1 subunit had an affinity for beta-alanine (which was more potent than glycine) that was 110-fold that of the wild type. Also, gamma-aminobutyric acid and D-serine, amino acids that do not activate wild-type alpha 1 receptors, efficiently gated the mutant channel. Thus, aromatic hydroxyl groups are crucial for ligand discrimination at inhibitory amino acid receptors.

  15. Pancreatic stellate cells support tumour metabolism through autophagic alanine secretion.

    PubMed

    Sousa, Cristovão M; Biancur, Douglas E; Wang, Xiaoxu; Halbrook, Christopher J; Sherman, Mara H; Zhang, Li; Kremer, Daniel; Hwang, Rosa F; Witkiewicz, Agnes K; Ying, Haoqiang; Asara, John M; Evans, Ronald M; Cantley, Lewis C; Lyssiotis, Costas A; Kimmelman, Alec C

    2016-08-25

    Pancreatic ductal adenocarcinoma (PDAC) is an aggressive disease characterized by an intense fibrotic stromal response and deregulated metabolism. The role of the stroma in PDAC biology is complex and it has been shown to play critical roles that differ depending on the biological context. The stromal reaction also impairs the vasculature, leading to a highly hypoxic, nutrient-poor environment. As such, these tumours must alter how they capture and use nutrients to support their metabolic needs. Here we show that stroma-associated pancreatic stellate cells (PSCs) are critical for PDAC metabolism through the secretion of non-essential amino acids (NEAA). Specifically, we uncover a previously undescribed role for alanine, which outcompetes glucose and glutamine-derived carbon in PDAC to fuel the tricarboxylic acid (TCA) cycle, and thus NEAA and lipid biosynthesis. This shift in fuel source decreases the tumour’s dependence on glucose and serum-derived nutrients, which are limited in the pancreatic tumour microenvironment. Moreover, we demonstrate that alanine secretion by PSCs is dependent on PSC autophagy, a process that is stimulated by cancer cells. Thus, our results demonstrate a novel metabolic interaction between PSCs and cancer cells, in which PSC-derived alanine acts as an alternative carbon source. This finding highlights a previously unappreciated metabolic network within pancreatic tumours in which diverse fuel sources are used to promote growth in an austere tumour microenvironment.

  16. Characterization of psychrophilic alanine racemase from Bacillus psychrosaccharolyticus.

    PubMed

    Okubo, Y; Yokoigawa, K; Esaki, N; Soda, K; Kawai, H

    1999-03-16

    A psychrophilic alanine racemase gene from Bacillus psychrosaccharolyticus was cloned and expressed in Escherichia coli SOLR with a plasmid pYOK3. The gene starting with the unusual initiation codon GTG showed higher preference for codons ending in A or T. The enzyme purified to homogeneity showed the high catalytic activity even at 0 degrees C and was extremely labile over 35 degrees C. The enzyme was found to have a markedly large Km value (5.0 microM) for the pyridoxal 5'-phosphate (PLP) cofactor in comparison with other reported alanine racemases, and was stabilized up to 50 degrees C in the presence of excess amounts of PLP. The low affinity of the enzyme for PLP may be related to the thermolability, and may be related to the high catalytic activity, initiated by the transaldimination reaction, at low temperature. The enzyme has a distinguishing hydrophilic region around the residue no. 150 in the deduced amino acid sequence (383 residues), whereas the corresponding regions of other Bacillus alanine racemases are hydrophobic. The position of the region in the three dimensional structure of C atoms of the enzyme was predicted to be in a surface loop surrounding the active site. The region may interact with solvent and reduce the compactness of the active site.

  17. Importance of intrahepatic mechanisms to gluconeogenesis from alanine during exercise and recovery

    SciTech Connect

    Wasserman, D.H.; Williams, P.E.; Lacy, D.B.; Green, D.R.; Cherrington, A.D.

    1988-04-01

    These studies were performed to assess the importance of intrahepatic mechanisms to gluconeogenesis in the dog during 150 min of treadmill exercise and 90 min of recovery. Sampling catheters were implanted in an artery and portal and hepatic veins 16 days before experimentation. Infusions of (U-/sup 14/C)alanine, (3-/sup 3/H)glucose, and indocyanine green were used to assess gluconeogenesis. During exercise, a decline in arterial and portal vein plasma alanine and in hepatic blood flow led to a decrease in hepatic alanine delivery. During recovery, hepatic blood flow was restored to basal, causing an increase in hepatic alanine delivery beyond exercise rates but still below resting rates. Hepatic fractional alanine extraction increased from 0.26 +/- 0.02 at rest to 0.64 +/- 0.03 during exercise and remained elevated during recovery. Net hepatic alanine uptake was 2.5 +/- 0.2 mumol.kg-1.min-1 at rest and remained unchanged during exercise but was increased during recovery. The conversion rate of (/sup 14/C)alanine to glucose had increased by 248 +/- 38% by 150 min of exercise and had increased further during recovery. The efficiency with which alanine was channeled into glucose in the liver was accelerated to a rate of 338 +/- 55% above basal by 150 min of exercise but declined slightly during recovery. In conclusion, 1) gluconeogenesis from alanine is accelerated during exercise, due to an increase in the hepatic fractional extraction of the amino acid and through intrahepatic mechanisms that more efficiently channel it into glucose.

  18. Substrate Specificity of the Aspartate:Alanine Antiporter (AspT) of Tetragenococcus halophilus in Reconstituted Liposomes*

    PubMed Central

    Sasahara, Ayako; Nanatani, Kei; Enomoto, Masaru; Kuwahara, Shigefumi; Abe, Keietsu

    2011-01-01

    The aspartate:alanine antiporter (AspT) of the lactic acid bacterium Tetragenococcus halophilus is a member of the aspartate:alanine exchanger (AAEx) transporter family. T. halophilus AspT catalyzes the electrogenic exchange of l-aspartate1− with l-alanine0. Although physiological functions of AspT were well studied, l-aspartate1−:l-alanine0 antiport mechanisms are still unsolved. Here we report that the binding sites of l-aspartate and l-alanine are independently present in AspT by means of the kinetic studies. We purified His6-tagged T. halophilus AspT and characterized its kinetic properties when reconstituted in liposomes (Km = 0.35 ± 0.03 mm for l-aspartate, Km = 0.098 ± 0 mm for d-aspartate, Km = 26 ± 2 mm for l-alanine, Km = 3.3 ± 0.2 mm for d-alanine). Competitive inhibition by various amino acids of l-aspartate or l-alanine in self-exchange reactions revealed that l-cysteine selectively inhibited l-aspartate self-exchange but only weakly inhibited l-alanine self-exchange. Additionally, l-serine selectively inhibited l-alanine self-exchange but barely inhibited l-aspartate self-exchange. The aspartate analogs l-cysteine sulfinic acid, l-cysteic acid, and d-cysteic acid competitively and strongly inhibited l-aspartate self-exchange compared with l-alanine self-exchange. Taken together, these kinetic data suggest that the putative binding sites of l-aspartate and l-alanine are independently located in the substrate translocation pathway of AspT. PMID:21719707

  19. Substrate specificity of the aspartate:alanine antiporter (AspT) of Tetragenococcus halophilus in reconstituted liposomes.

    PubMed

    Sasahara, Ayako; Nanatani, Kei; Enomoto, Masaru; Kuwahara, Shigefumi; Abe, Keietsu

    2011-08-19

    The aspartate:alanine antiporter (AspT) of the lactic acid bacterium Tetragenococcus halophilus is a member of the aspartate:alanine exchanger (AAEx) transporter family. T. halophilus AspT catalyzes the electrogenic exchange of L-aspartate(1-) with L-alanine(0). Although physiological functions of AspT were well studied, L-aspartate(1-):L-alanine(0) antiport mechanisms are still unsolved. Here we report that the binding sites of L-aspartate and L-alanine are independently present in AspT by means of the kinetic studies. We purified His(6)-tagged T. halophilus AspT and characterized its kinetic properties when reconstituted in liposomes (K(m) = 0.35 ± 0.03 mm for L-aspartate, K(m) = 0.098 ± 0 mm for D-aspartate, K(m) = 26 ± 2 mm for L-alanine, K(m) = 3.3 ± 0.2 mm for D-alanine). Competitive inhibition by various amino acids of L-aspartate or L-alanine in self-exchange reactions revealed that L-cysteine selectively inhibited L-aspartate self-exchange but only weakly inhibited L-alanine self-exchange. Additionally, L-serine selectively inhibited L-alanine self-exchange but barely inhibited L-aspartate self-exchange. The aspartate analogs L-cysteine sulfinic acid, L-cysteic acid, and D-cysteic acid competitively and strongly inhibited L-aspartate self-exchange compared with L-alanine self-exchange. Taken together, these kinetic data suggest that the putative binding sites of L-aspartate and L-alanine are independently located in the substrate translocation pathway of AspT.

  20. Radiolysis of alanine adsorbed in a clay mineral

    SciTech Connect

    Aguilar-Ovando, Ellen Y.; Negron-Mendoza, Alicia

    2013-07-03

    Optical activity in molecules is a chemical characteristic of living beings. In this work, we examine the hypothesis of the influence of different mineral surfaces on the development of a specific chirality in organic molecules when subjected to conditions simulating the primitive Earth during the period of chemical evolution. By using X-ray diffraction techniques and HPLC/ELSD to analyze aqueous suspensions of amino acids adsorbed on minerals irradiated in different doses with a cobalt-60 gamma source, the experiments attempt to prove the hypothesis that some solid surfaces (like clays and meteorite rocks) may have a concentration capacity and protective role against external sources of ionizing radiation (specifically {gamma}-ray) for some organic compounds (like some amino acids) adsorbed on them. Preliminary results show a slight difference in the adsorption and radiolysis of the D-and L-alanine.

  1. Hibiscus cannabinus feruloyl-coa:monolignol transferase

    SciTech Connect

    Wilkerson, Curtis; Ralph, John; Withers, Saunia; Mansfield, Shawn D.

    2016-11-15

    The invention relates to isolated nucleic acids encoding a feruloyl-CoA:monolignol transferase and feruloyl-CoA:monolignol transferase enzymes. The isolated nucleic acids and/or the enzymes enable incorporation of monolignol ferulates into the lignin of plants, where such monolignol ferulates include, for example, p-coumaryl ferulate, coniferyl ferulate, and/or sinapyl ferulate. The invention also includes methods and plants that include nucleic acids encoding a feruloyl-CoA:monolignol transferase enzyme and/or feruloyl-CoA:monolignol transferase enzymes.

  2. Effects of high-salinity seawater acclimation on the levels of D-alanine in the muscle and hepatopancreas of kuruma prawn, Marsupenaeus japonicus.

    PubMed

    Yoshikawa, Naoko; Yokoyama, Masahumi

    2015-12-10

    Changes in D- and L-alanine contents were determined in the muscle and hepatopancreas of kuruma prawn Marsupenaeus japonicus, during acclimation from seawater containing 100% salinity to artificial seawater containing 150% salinity. In the hepatopancreas, contents of both amino acids increased by approximately threefold. The activity of alanine racemase, which catalyzes the interconversion of D- and L-alanine, also increased in the high-salinity seawater. In addition, the expression of the gene encoding alanine racemase increased in the hepatopancreas with an increase in the alanine racemase activity. These data indicate that the biosynthesis of D- and L-alanine is controlled by the gene expression level of alanine racemase, and D-alanine in the hepatopancreas functions as a major osmolyte for isosmotic regulation. In contrast, the content of D-alanine and alanine racemase activity did not change in the muscle during hyper-osmotic acclimation. Therefore, we suggest that D-alanine, which exists in the several tissues of M. japonicus, is considered to be utilized in some different physiological phenomena in different tissues.

  3. Nonprotein Amino Acids from Spark Discharges and Their Comparison with the Murchison Meteorite Amino Acids

    PubMed Central

    Wolman, Yecheskel; Haverland, William J.; Miller, Stanley L.

    1972-01-01

    All the nonprotein amino acids found in the Murchison meteorite are products of the action of electric discharge on a mixture of methane, nitrogen, and water with traces of ammonia. These amino acids include α-amino-n-butyric acid, α-aminoisobutyric acid, norvaline, isovaline, pipecolic acid, β-alanine, β-amino-n-butyric acid, β-aminoisobutyric acid, γ-aminobutyric acid, sarcosine, N-ethylglycine, and N-methylalanine. In addition, norleucine, alloisoleucine, N-propylglycine, N-isopropylglycine, N-methyl-β-alanine, N-ethyl-β-alanine α,β-diaminopropionic acid, isoserine, α,γ-diaminobutyric acid, and α-hydroxy-γ-aminobutyric acid are produced by the electric discharge, but have not been found in the meteorite. PMID:16591973

  4. Alanine increases blood pressure during hypotension

    NASA Technical Reports Server (NTRS)

    Conlay, L. A.; Maher, T. J.; Wurtman, R. J.

    1990-01-01

    The effect of L-alanine administration on blood pressure (BP) during haemorrhagic shock was investigated using anesthetized rats whose left carotid arteries were cannulated for BP measurement, blood removal, and drug administration. It was found that L-alanine, in doses of 10, 25, 50, 100, and 200 mg/kg, increased the systolic BP of hypotensive rats by 38 to 80 percent (while 100 mg/kg pyruvate increased BP by only 9.4 mmhg, not significantly different from saline). The results suggest that L-alanine might influence cardiovascular function.

  5. Characteristics of the transport of alanine, serine and glutamine across the plasma membrane of isolated rat liver cells.

    PubMed Central

    Joseph, S K; Bradford, N M; McGivan, J D

    1978-01-01

    1. Alanine, glutamine and serine were actively accumulated in liver cells isolated from starved rats. 2. This accumulation was inhibited when either Na+ or HCO3- ions were omitted from the incubation medium. In general the degree of dependence on Na+ was quantitatively similar to that on HCO3-. 3. The apparent Km values for the transport of all three amino acids were in the range 3--5mM with Vmax. values in the range 15--25nmol/min per mg of cell protein at 37 degrees C. 4. Alanine and serine transport were mutually competitive; glutamine inhibited the transport of alanine and serine non-competitively. 5. The initial rate of transport of these amino acids was inhibited when the intracellular content of ATP was decreased. 6. Ouabain inhibited the rate of alanine transport without inhibiting the rate of alanine metabolism. 7. It is concluded that a minimum of three transport systems must be postulated to exist in the liver cell plasma membrane to account for the transport of alanine, serine and glutamine. The rate of transport of these amino acids in isolated hepatocytes is unlikely to limit the rate at which they are metabolized. PMID:747655

  6. Ultraviolet radiation induces stress in etiolated Landoltia punctata, as evidenced by the presence of alanine, a universal stress signal: a ¹⁵N NMR study.

    PubMed

    Monselise, E B-I; Levkovitz, A; Kost, D

    2015-01-01

    Analysis with (15) N NMR revealed that alanine, a universal cellular stress signal, accumulates in etiolated duckweed plants exposed to 15-min pulsed UV light, but not in the absence of UV irradiation. The addition of 10 mm vitamin C, a radical scavenger, reduced alanine levels to zero, indicating the involvement of free radicals. Free D-alanine was detected in (15) N NMR analysis of the chiral amino acid content, using D-tartaric acid as solvent. The accumulation of D-alanine under stress conditions presents a new perspective on the biochemical processes taking place in prokaryote and eukaryote cells.

  7. Nonprotein Amino Acids in the Murchison Meteorite

    PubMed Central

    Kvenvolden, Keith A.; Lawless, James G.; Ponnamperuma, Cyril

    1971-01-01

    Twelve nonprotein amino acids appear to be present in the Murchison meteorite. The identity of eight of them has been conclusively established as N-methylglycine, β-alanine, 2-methylalanine, α-amino-n-butyric acid, β-amino-n-butyric acid, γ-amino-n-butyric acid, isovaline, and pipecolic acid. Tentative evidence is presented for the presence of N-methylalanine, N-ethylglycine, β-aminoisobutyric acid, and norvaline. These amino acids appear to be extraterrestrial in origin and may provide new evidence for the hypothesis of chemical evolution. PMID:16591908

  8. Survivability and reactivity of glycine and alanine in early oceans: effects of meteorite impacts.

    PubMed

    Umeda, Yuhei; Fukunaga, Nao; Sekine, Toshimori; Furukawa, Yoshihiro; Kakegawa, Takeshi; Kobayashi, Takamichi; Nakazawa, Hiromoto

    2016-01-01

    Prebiotic oceans might have contained abundant amino acids, and were subjected to meteorite impacts, especially during the late heavy bombardment. It is so far unknown how meteorite impacts affected amino acids in the early oceans. Impact experiments were performed under the conditions where glycine was synthesized from carbon, ammonia, and water, using aqueous solutions containing (13)C-labeled glycine and alanine. Selected amino acids and amines in samples were analyzed with liquid chromatography-mass spectrometry (LC/MS). In particular, the (13)C-labeled reaction products were analyzed to distinguish between run products and contaminants. The results revealed that both amino acids survived partially in the early ocean through meteorite impacts, that part of glycine changed into alanine, and that large amounts of methylamine and ethylamine were formed. Fast decarboxylation was confirmed to occur during such impact processes. Furthermore, the formation of n-butylamine, detected only in the samples recovered from the solutions with additional nitrogen and carbon sources of ammonia and benzene, suggests that chemical reactions to form new biomolecules can proceed through marine impacts. Methylamine and ethylamine from glycine and alanine increased considerably in the presence of hematite rather than olivine under similar impact conditions. These results also suggest that amino acids present in early oceans can contribute further to impact-induced reactions, implying that impact energy plays a potential role in the prebiotic formation of various biomolecules, although the reactions are complicated and depend upon the chemical environments as well.

  9. Repeated Supramaximal Exercise-Induced Oxidative Stress: Effect of β-Alanine Plus Creatine Supplementation

    PubMed Central

    Belviranli, Muaz; Okudan, Nilsel; Revan, Serkan; Balci, Serdar; Gokbel, Hakki

    2016-01-01

    Background: Carnosine is a dipeptide formed from the β-alanine and histidine amino acids and found in mainly in the brain and muscle, especially fast twitch muscle. Carnosine and creatine has an antioxidant effect and carnosine accounts for about 10% of the muscle's ability to buffer the H+ ions produced by exercise. Objectives: The aim of the study was to investigate the effects of beta alanine and/or creatine supplementation on oxidant and antioxidant status during repeated Wingate tests (WTs). Patients and Methods: Forty four sedentary males participated in the study. Participants performed three 30s WTs with 2 minutes rest between exercise bouts. After the first exercise session, the subjects were assigned to one of four groups: Placebo, Creatine, Beta-alanine and Beta-alanine plus creatine. Participants ingested twice per day for 22 consecutive days, then four times per day for the following 6 days. After the supplementation period the second exercise session was applied. Blood samples were taken before and immediately after the each exercise session for the analysis of oxidative stress and antioxidant markers. Results: Malondialdehyde levels and superoxide dismutase activities were affected by neither supplementation nor exercise. During the pre-supplementation session, protein carbonyl reduced and oxidized glutathione (GSH and GSSG) levels increased immediately after the exercise. However, during the post-supplementation session GSH and GSSG levels increased in beta-alanine and beta-alanine plus creatine groups immediately after the exercise compared to pre-exercise. In addition, during the post-supplementation session total antioxidant capacity increased in beta-alanine group immediately after the exercise. Conclusions: Beta-alanine supplementation has limited antioxidant effect during the repeated WTs. PMID:27217925

  10. Effects of endogenous D-alanine synthesis and autoinhibition of Bacillus anthracis germination on in vitro and in vivo infections.

    PubMed

    McKevitt, Matthew T; Bryant, Katie M; Shakir, Salika M; Larabee, Jason L; Blanke, Steven R; Lovchik, Julie; Lyons, C Rick; Ballard, Jimmy D

    2007-12-01

    Bacillus anthracis transitions from a dormant spore to a vegetative bacillus through a series of structural and biochemical changes collectively referred to as germination. The timing of germination is important during early steps in infection and may determine if B. anthracis survives or succumbs to responsive macrophages. In the current study experiments determined the contribution of endogenous D-alanine production to the efficiency and timing of B. anthracis spore germination under in vitro and in vivo conditions. Racemase-mediated production of endogenous D-alanine by B. anthracis altered the kinetics for initiation of germination over a range of spore densities and exhibited a threshold effect wherein small changes in spore number resulted in major changes in germination efficiency. This threshold effect correlated with D-alanine production, was prevented by an alanine racemase inhibitor, and required L-alanine. Interestingly, endogenous production of inhibitory levels of D-alanine was detected under experimental conditions that did not support germination and in a germination-deficient mutant of B. anthracis. Racemase-dependent production of D-alanine enhanced survival of B. anthracis during interaction with murine macrophages, suggesting a role for inhibition of germination during interaction with these cells. Finally, in vivo experiments revealed an approximately twofold decrease in the 50% lethal dose of B. anthracis spores administered in the presence of D-alanine, indicating that rates of germination may be directly influenced by the levels of this amino acid during early stages of disease.

  11. Fragmentation of α- and β-alanine molecules by ions at Bragg-peak energies

    NASA Astrophysics Data System (ADS)

    Bari, S.; Sobocinski, P.; Postma, J.; Alvarado, F.; Hoekstra, R.; Bernigaud, V.; Manil, B.; Rangama, J.; Huber, B.; Schlathölter, T.

    2008-02-01

    The interaction of keV He+, He2+, and O5+ ions with isolated α and β isomers of the amino acid alanine was studied by means of high resolution coincidence time-of-flight mass spectrometry. We observed a strong isomer dependence of characteristic fragmentation channels which manifests in strongly altered branching ratios. Despite the ultrashort initial perturbation by the incoming ion, evidence for molecular rearrangement leading to the formation of H3+ was found. The measured kinetic energies of ionic alanine fragments can be sufficient to induce secondary damage to DNA in a biological environment.

  12. On the fragmentation of biomolecules: Fragmentation of alanine dipeptide along the polypeptide chain

    SciTech Connect

    Solov'yov, I. A. Yakubovich, A. V.; Solov'yov, A. V.; Greiner, W.

    2006-09-15

    The interaction potential between amino acids in alanine dipeptide has been studied for the first time taking into account exact molecular geometry. Ab initio calculation has been performed in the framework of density functional theory taking into account all electrons in the system. The fragmentation of dipeptide along the polypeptide chain, as well as the interaction between alanines, has been considered. The energy of the system has been analyzed as a function of the distance between fragments for all possible dipeptide fragmentation channels. Analysis of the energy barriers makes it possible to estimate the characteristic fragmentation times and to determine the degree of applicability of classical electrodynamics for describing the system energy.

  13. The determination of tRNALeu recognition nucleotides for Escherichia coli L/F transferase

    PubMed Central

    Fung, Angela Wai Shan; Leung, Charles Chung Yun; Fahlman, Richard Peter

    2014-01-01

    Escherichia coli leucyl/phenylalanyl-tRNA protein transferase catalyzes the tRNA-dependent post-translational addition of amino acids onto the N-terminus of a protein polypeptide substrate. Based on biochemical and structural studies, the current tRNA recognition model by L/F transferase involves the identity of the 3′ aminoacyl adenosine and the sequence-independent docking of the D-stem of an aminoacyl-tRNA to the positively charged cluster on L/F transferase. However, this model does not explain the isoacceptor preference observed 40 yr ago. Using in vitro-transcribed tRNA and quantitative MALDI-ToF MS enzyme activity assays, we have confirmed that, indeed, there is a strong preference for the most abundant leucyl-tRNA, tRNALeu (anticodon 5′-CAG-3′) isoacceptor for L/F transferase activity. We further investigate the molecular mechanism for this preference using hybrid tRNA constructs. We identified two independent sequence elements in the acceptor stem of tRNALeu (CAG)—a G3:C70 base pair and a set of 4 nt (C72, A4:U69, C68)—that are important for the optimal binding and catalysis by L/F transferase. This maps a more specific, sequence-dependent tRNA recognition model of L/F transferase than previously proposed. PMID:24935875

  14. Effects of changes in cell volume on the rates of glutamine and alanine release from rat skeletal muscle in vitro.

    PubMed

    Parry-Billings, M; Bevan, S J; Opara, E; Newsholme, E A

    1991-06-01

    The effect of changes in cell volume on the rates of release of glutamine and alanine from muscle and on the concentrations of these amino acids in muscle were investigated by using an isolated preparation of rat skeletal muscle incubated in the presence of hypo- and hyper-osmotic media. Changes in cell volume were associated with changes in the rates of release of glutamine and alanine from muscle: incubation in hypo-osmotic medium decreased the rates of release of glutamine and alanine, and incubation in hyperosmotic medium increased these rates. These changes were rapidly reversed by a change in osmoticity of the medium. Despite marked changes in cell volume, the concentrations of these amino acids in muscle were maintained. It is suggested that cell volume may play a role in the regulation of amino acid metabolism in skeletal muscle.

  15. Feruloyl-CoA:monolignol transferase

    DOEpatents

    Wilkerson, Curtis; Ralph, John; Withers, Saunia; Mansfield, Shawn D.

    2016-09-13

    The invention relates to nucleic acids encoding a feruloyl-CoA:monolignol transferase and the feruloyl-CoA:monolignol transferase enzyme that enables incorporation of monolignol ferulates, for example, including p-coumaryl ferulate, coniferyl ferulate, and sinapyl ferulate, into the lignin of plants.

  16. Feruloyl-CoA:monolignol transferase

    DOEpatents

    Wilkerson, Curtis; Ralph, John; Withers, Saunia; Mansfield, Shawn D.

    2016-11-08

    The invention relates to nucleic acids encoding a feruloyl-CoA:monolignol transferase and the feruloyl-CoA:monolignol transferase enzyme that enables incorporation of monolignol ferulates, for example, including p-coumaryl ferulate, coniferyl ferulate, and sinapyl ferulate, into the lignin of plants.

  17. Isotope labeling studies on the formation of multiple addition products of alanine in the pyrolysis residue of glucose/alanine mixtures by high-resolution ESI-TOF-MS.

    PubMed

    Chu, Fong Lam; Sleno, Lekha; Yaylayan, Varoujan A

    2011-11-09

    Pyrolysis was used as a microscale sample preparation tool to generate glucose/alanine reaction products to minimize the use of expensive labeled precursors in isotope labeling studies. The residue remaining after the pyrolysis at 250 °C was analyzed by electrospray time-of-flight mass spectrometry (ESI-TOF-MS). It was observed that a peak at m/z 199.1445 in the ESI-TOF-MS spectrum appeared only when the model system contained at least 2-fold excess alanine. The accurate mass determination indeed indicated the presence of two nitrogen atoms in the molecular formula (C(10)H(18)N(2)O(2)). To verify the origin of the carbon atoms in this unknown compound, model studies with [(13)U(6)]glucose, [(13)C-1]alanine, [(13)C-2]alanine, [(13)C-3]alanine, and [(15)N]alanine were also performed. Glucose furnished six carbon atoms, and alanine provides four carbon (2 × C-2 and 2 × C-3) and two nitrogen atoms. When commercially available fructosylalanine (N-attached to C-1) was reacted with only 1 mol of alanine, a peak at m/z 199.1445 was once again observed. In addition, when 3-deoxyglucosone (3-DG) was reacted with a 2-fold excess of alanine, a peak at m/z 199.1433 was also generated, confirming the points of attachment of the two amino acids at C-1 and C-2 atoms of 3-DG. These studies have indicated that amino acids can undergo multiple addition reactions with 1,2-dicarbonyl compounds such as 3-deoxyglucosone and eventually form a tetrahydropyrazine moiety.

  18. Prebiotic Amino Acid Thioester Synthesis: Thiol-Dependent Amino Acid Synthesis from Formose substrates (Formaldehyde and Glycolaldehyde) and Ammonia

    NASA Technical Reports Server (NTRS)

    Weber, Arthur L.

    1998-01-01

    Formaldehyde and glycolaldehyde (substrates of the formose autocatalytic cycle) were shown to react with ammonia yielding alanine and homoserine under mild aqueous conditions in the presence of thiol catalysts. Since similar reactions carried out without ammonia yielded alpha-hydroxy acid thioesters, the thiol-dependent synthesis of alanine and homoserine is presumed to occur via amino acid thioesters-intermediates capable of forming peptides. A pH 5.2 solution of 20 mM formaldehyde, 20 mM glycolaldehyde, 20 mM ammonium chloride, 23 mM 3-mercaptopropionic acid, and 23 mM acetic acid that reacted for 35 days at 40 C yielded (based on initial formaldehyde) 1.8% alanine and 0.08% homoserine. In the absence of thiol catalyst, the synthesis of alanine and homoserine was negligible. Alanine synthesis required both formaldehyde and glycolaldehyde, but homoserine synthesis required only glycolaldehyde. At 25 days the efficiency of alanine synthesis calculated from the ratio of alanine synthesized to formaldehyde reacted was 2.1%, and the yield (based on initial formaldehyde) of triose and tetrose intermediates involved in alanine and homoserine synthesis was 0.3 and 2.1%, respectively. Alanine synthesis was also seen in similar reactions containing only 10 mM each of aldehyde substrates, ammonia, and thiol. The prebiotic significance of these reactions that use the formose reaction to generate sugar intermediates that are converted to reactive amino acid thioesters is discussed.

  19. 21 CFR 862.1535 - Ornithine carbamyl transferase test system.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... carbamyl transferase (OCT) in serum. Ornithine carbamyl transferase measurements are used in the diagnosis and treatment of liver diseases, such as infectious hepatitis, acute cholecystitis (inflammation...

  20. Gamma-glutamyl transferase and cardiovascular disease

    PubMed Central

    Kastrati, Adnan

    2016-01-01

    Gamma-glutamyl transferase (GGT) is an enzyme located on the external surface of cellular membranes. GGT contributes in maintaining the physiological concentrations of cytoplasmic glutathione and cellular defense against oxidative stress via cleavage of extracellular glutathione and increased availability of amino acids for its intracellular synthesis. Increased GGT activity is a marker of antioxidant inadequacy and increased oxidative stress. Ample evidence suggests that elevated GGT activity is associated with increased risk of cardiovascular disease (CVD) such as coronary heart disease (CHD), stroke, arterial hypertension, heart failure, cardiac arrhythmias and all-cause and CVD-related mortality. The evidence is weaker for an association between elevated GGT activity and acute ischemic events and myocardial infarction. The risk for CVD or CVD-related mortality mediated by GGT may be explained by the close correlation of GGT with conventional CVD risk factors and various comorbidities, particularly non-alcoholic fatty liver disease, alcohol consumption, oxidative stress, metabolic syndrome, insulin resistance and systemic inflammation. The finding of GGT activity in atherosclerotic plaques and correlation of intra-plaque GGT activity with histological indexes of plaque instability may suggest a participation of GGT in the pathophysiology of CVD, particularly atherosclerosis. However, whether GGT has a direct role in the pathophysiology of CVD or it is an epiphenomenon of coexisting CVD risk factors or comorbidities remains unknown and Hill’s criteria of causality relationship between GGT and CVD are not fulfilled. The exploration whether GGT provides prognostic information on top of the information provided by known cardiovascular risk factors regarding the CVD or CVD-related outcome and exploration of molecular mechanisms of GGT involvement in the pathophysiology of CVD and eventual use of interventions to reduce circulating GGT activity remain a duty of

  1. Probing the Catalytic Charge-Relay System in Alanine Racemase with Genetically Encoded Histidine Mimetics.

    PubMed

    Sharma, Vangmayee; Wang, Yane-Shih; Liu, Wenshe R

    2016-12-16

    Histidine is a unique amino acid with an imidazole side chain in which both of the nitrogen atoms are capable of serving as a proton donor and proton acceptor in hydrogen bonding interactions. In order to probe the functional role of histidine involved in hydrogen bonding networks, fine-tuning the hydrogen bonding potential of the imidazole side chain is required but not feasible through traditional mutagenesis methods. Here, we show that two close mimetics of histidine, 3-methyl-histidine and thiazole alanine, can be genetically encoded using engineered pyrrolysine incorporation machinery. Replacement of the three histidine residues predicted to be involved in an extended charge-relay system in alanine racemase with 3-methyl-histidine or thiazole alanine shows a dramatic loss in the enzyme's catalytic efficiency, implying the role of this extended charge-relay system in activating the active site residue Y265, a general acid/base catalyst in the enzyme.

  2. Paradox of mistranslation of serine for alanine caused by AlaRS recognition dilemma.

    PubMed

    Guo, Min; Chong, Yeeting E; Shapiro, Ryan; Beebe, Kirk; Yang, Xiang-Lei; Schimmel, Paul

    2009-12-10

    Mistranslation arising from confusion of serine for alanine by alanyl-tRNA synthetases (AlaRSs) has profound functional consequences. Throughout evolution, two editing checkpoints prevent disease-causing mistranslation from confusing glycine or serine for alanine at the active site of AlaRS. In both bacteria and mice, Ser poses a bigger challenge than Gly. One checkpoint is the AlaRS editing centre, and the other is from widely distributed AlaXps-free-standing, genome-encoded editing proteins that clear Ser-tRNA(Ala). The paradox of misincorporating both a smaller (glycine) and a larger (serine) amino acid suggests a deep conflict for nature-designed AlaRS. Here we show the chemical basis for this conflict. Nine crystal structures, together with kinetic and mutational analysis, provided snapshots of adenylate formation for each amino acid. An inherent dilemma is posed by constraints of a structural design that pins down the alpha-amino group of the bound amino acid by using an acidic residue. This design, dating back more than 3 billion years, creates a serendipitous interaction with the serine OH that is difficult to avoid. Apparently because no better architecture for the recognition of alanine could be found, the serine misactivation problem was solved through free-standing AlaXps, which appeared contemporaneously with early AlaRSs. The results reveal unconventional problems and solutions arising from the historical design of the protein synthesis machinery.

  3. The effect of immunonutrition (glutamine, alanine) on fracture healing

    PubMed Central

    Küçükalp, Abdullah; Durak, Kemal; Bayyurt, Sarp; Sönmez, Gürsel; Bilgen, Muhammed S.

    2014-01-01

    Background There have been various studies related to fracture healing. Glutamine is an amino acid with an important role in many cell and organ functions. This study aimed to make a clinical, radiological, and histopathological evaluation of the effects of glutamine on fracture healing. Methods Twenty rabbits were randomly allocated into two groups of control and immunonutrition. A fracture of the fibula was made to the right hind leg. All rabbits received standard food and water. From post-operative first day for 30 days, the study group received an additional 2 ml/kg/day 20% L-alanine L-glutamine solution via a gastric catheter, and the control group received 2 ml/kg/day isotonic via gastric catheter. At the end of 30 days, the rabbits were sacrificed and the fractures were examined clinically, radiologically, and histopathologically in respect to the degree of union. Results Radiological evaluation of the control group determined a mean score of 2.5 according to the orthopaedists and 2.65 according to the radiologists. In the clinical evaluation, the mean score was 1.875 for the control group and 2.0 for the study group. Histopathological evaluation determined a mean score of 8.5 for the control group and 9.0 for the study group. Conclusion One month after orally administered glutamine–alanine, positive effects were observed on fracture healing radiologically, clinically, and histopathologically, although no statistically significant difference was determined.

  4. Extraterrestrial Amino Acids in Orgueil and Ivuna: Tracing the Parent Body of CI Type Carbonaceous Chondrites

    NASA Technical Reports Server (NTRS)

    Meyer, Michael (Technical Monitor); Ehrenfreund, Pascale; Glavin, Daniel P.; Bota, Oliver; Cooper, George; Bada, Jeffrey

    2001-01-01

    Amino acid analyses using HPLC of pristine interior pieces of the CI carbonaceous chondrites Orgueil and Ivuna have found that beta-alanine, glycine, and gamma-amino-n-butyric acid (ABA) are the most abundant amino acids in these two meteorites, with concentrations ranging from approx. 600 to 2,000 parts per billion (ppb). Other alpha-amino acids such as alanine, alpha-ABA, alpha-aminoisobutyric acid (AIB), and isovaline are present only in trace amounts (less than 200 ppb). Carbon isotopic measurements of beta-alanine and glycine and the presence of racemic (D/L 1) alanine and beta-ABA in Orgueil suggest that these amino acids are extraterrestrial in origin. In comparison to the CM carbonaceous chondrites Murchison and Murray, the amino acid composition of the CIs is strikingly distinct, suggesting that these meteorites came from a different type of parent body, possibly an extinct comet, than did the CM carbonaceous chondrites.

  5. Extraterrestrial amino acids in Orgueil and Ivuna: Tracing the parent body of CI type carbonaceous chondrites

    PubMed Central

    Ehrenfreund, Pascale; Glavin, Daniel P.; Botta, Oliver; Cooper, George; Bada, Jeffrey L.

    2001-01-01

    Amino acid analyses using HPLC of pristine interior pieces of the CI carbonaceous chondrites Orgueil and Ivuna have found that β-alanine, glycine, and γ-amino-n-butyric acid (ABA) are the most abundant amino acids in these two meteorites, with concentrations ranging from ≈600 to 2,000 parts per billion (ppb). Other α-amino acids such as alanine, α-ABA, α-aminoisobutyric acid (AIB), and isovaline are present only in trace amounts (<200 ppb). Carbon isotopic measurements of β-alanine and glycine and the presence of racemic (D/L ≈ 1) alanine and β-ABA in Orgueil suggest that these amino acids are extraterrestrial in origin. In comparison to the CM carbonaceous chondrites Murchison and Murray, the amino acid composition of the CIs is strikingly distinct, suggesting that these meteorites came from a different type of parent body, possibly an extinct comet, than did the CM carbonaceous chondrites. PMID:11226205

  6. Enantiomeric Excesses of Acid Labile Amino Acid Precursors of the Murchison Meteorite

    NASA Technical Reports Server (NTRS)

    Pizzarello, Sandra

    1998-01-01

    Amino acids present in carbonaceous chondrite are extracted in water in part as free compounds and in approximately equal part as acid labile precursors. On the assumption that they would be free of contamination, the precursors of two Murchison amino acids that have terrestrial occurrence, alanine and glutamic acid, have been targeted for analysis of their enantiomeric ratios. Pyroglutamic acid, the precursor of glutamic acid, was found with an L-enantiomeric excess comparable to that of the free acid, while alanine's precursor, N-acetyl alanine, appears approximately racemic. Also alpha-imino propioacetic acid, a proposed end product of alanine synthesis in the meteorite, was analyzed and found racemic.

  7. First-principles study of fluorination of L-Alanine

    NASA Astrophysics Data System (ADS)

    Sreepad, H. R.; Ravi, H. R.; Ahmed, Khaleel; Dayananda, H. M.; Umakanth, K.; Manohara, B. M.

    2013-02-01

    First-principles calculations based on Density Functional Theory have been done on effect of fluorination of an important amino acid - L-Alanine. Its structure has been simulated. The unit cell is orthorhombic with lattice parameters a=5.90Å, b=13.85Å and c=5.75Å with volume 470 (Å)3. Bond lengths and bond angles have been estimated. Electronic Density of States calculations show that the material has a band gap of 4.47eV. Electronic band structure indicates that the material can be effectively used for NLO applications. The electronic contribution to the dielectric constant has been calculated and its average value comes out to be 2.165.

  8. Effects of glycine, beta-alanine and diazepam upon morphine-tolerant-dependent mice.

    PubMed

    Contreras, E; Tamayo, L

    1980-05-01

    The effects in mice of glycine, beta-alanine and diazepam on the analgesic response to morphine, on the intensity of tolerance and on the physical dependence on the analgesic have been examined. The two amino acids increased the analgesic response to morphine in a dose-related manner. However, both compounds were ineffective in the analgesic test (hot plate) when administered without morphine. Diazepam was ineffective in the analgesic test and it did not alter morphine analgesia, except when administered in a high dose which decreased and analgesic response. Glycine, either in single or repeated doses, did not modify tolerance to morphine, whereas beta-alanine induced a dose-related partial antagonism, which promptly reached a plateau. Diazepam induced a small decrease in the intensity of tolerance to the analgesic. The abstinence syndrome to morphine, induced by naloxone administration to primed mice, was reduced by single doses of glycine or beta-alanine. Diazepam behaved as a weak inhibitor of the abstinence syndrome when administered at a high dose. The potentiation of morphine analgesia and the antagonism of the abstinence syndrome induced by the amino acids may be related to their hyperpolarizing action in the c.n. system. The effects of beta-alanine on morphine tolerance cannot be explained by the same mechanism.

  9. Stereochemistry of amino acids in surface samples of a marine sediment

    USGS Publications Warehouse

    Pollock, G.E.; Kvenvolden, K.A.

    1978-01-01

    In two surface samples of marine sediment, the percentages of d-alanine and d-aspartic acid are significantly higher than the other d-amino acids and are similar to the range found in soils. The percentage of d-glutamic acid is also higher than the other amino acids but less than d-alanine and d-aspartic acid. These d-amino acids may come mainly from bacteria. ?? 1978.

  10. Stereochemistry of amino acids in surface samples of a marine sediment

    NASA Technical Reports Server (NTRS)

    Pollock, G. E.; Kvenvolden, K. A.

    1978-01-01

    In two surface samples of marine sediment, the percentages of D-alanine and D-aspartic acid are significantly higher than the other D-amino acids and are similar to the range found in soils. The percentage of D-glutamic acid is also higher than the other amino acids but less than D-alanine and D-aspartic acid. These D-amino acids may come mainly from bacteria.

  11. Solid phase synthesis and binding affinity of peptidyl transferase transition state mimics containing 2′-OH at P-site position A76

    PubMed Central

    Weinger, Joshua S.; Kitchen, David; Scaringe, Stephen A.; Strobel, Scott A.; Muth, Gregory W.

    2004-01-01

    All living cells are dependent on ribosomes to catalyze the peptidyl transfer reaction, by which amino acids are assembled into proteins. The previously studied peptidyl transferase transition state analog CC-dA-phosphate-puromycin (CCdApPmn) has important differences from the transition state, yet current models of the ribosomal active site have been heavily influenced by the properties of this molecule. One significant difference is the substitution of deoxyadenosine for riboadenosine at A76, which mimics the 3′ end of a P-site tRNA. We have developed a solid phase synthetic approach to produce inhibitors that more closely match the transition state, including the critical P-site 2′-OH. Inclusion of the 2′-OH or an even bulkier OCH3 group causes significant changes in binding affinity. We also investigated the effects of changing the A-site amino acid side chain from phenylalanine to alanine. These results indicate that the absence of the 2′-OH is likely to play a significant role in the binding and conformation of CCdApPmn in the ribosomal active site by eliminating steric clash between the 2′-OH and the tetrahedral phosphate oxygen. The conformation of the actual transition state must allow for the presence of the 2′-OH, and transition state mimics that include this critical hydroxyl group must bind in a different conformation from that seen in prior analog structures. These new inhibitors will provide valuable insights into the geometry and mechanism of the ribosomal active site. PMID:14999092

  12. Exogenous amino acids as fuel in shock.

    PubMed

    Daniel, A M; Kapadia, B; MacLean, L D

    1982-01-01

    It has been suggested that in shock branched-chain amino acids are preferentially oxidized resulting in continued proteolysis and stimulated gluconeogenesis. To determine if exogenous amino acids could be used as fuel in shock, dogs rendered hypotensive by controlled cardiac tamponade and normotensive controls were infused with amino acid mixtures and individual amino acids. When Nephramine, a mixture rich in branched-chain amino acids, was infused, plasma alpha-amino nitrogen levels rose but urea output did not increase in either the control state or in shock, suggesting that these amino acids were not rapidly deaminated to serve as fuels. Travasol, which in addition contained large amounts of alanine and glycine, tripled urea output in the controls and doubled it in shock. The limit of urea production was reached in both groups at 35 mumoles urea/minute/kg. In the Travasol-infused animals plasma alpha-amino nitrogen levels were maintained in normotension but rose sharply in shock. When glycine alone was infused into five dogs in shock urea production rate was 30.6 + 2.1 mumoles/minute/kg; with alanine the same value was 22.5 + 2.2 mumoles/minute/kg. In both cases plasma alpha-amino nitrogen levels were high, suggesting that transport of these amino acids into the cell was slow in shock. In four dogs in shock glycine-14C was added to the glycine infusate as a tracer. At radioactive equilibrium 28% of the label infused appeared in CO2; another 22% appeared in glucose. It is concluded that of all the amino acids tested only glycine and alanine are deaminated rapidly enough to serve as exogenous fuels in shock.

  13. A general and practical palladium-catalyzed monoarylation of β-methyl C(sp³)-H of alanine.

    PubMed

    Chen, Kai; Zhang, Shuo-Qing; Xu, Jing-Wen; Hu, Fang; Shi, Bing-Feng

    2014-11-21

    A palladium-catalyzed monoarylation of β-methyl C(sp(3))-H of an alanine derivative with aryl iodides using an 8-aminoquinoline auxiliary is described. The reaction is highly efficient, scalable and compatible with a variety of functional groups with complete retention of chirality, providing a general and practical access to various β-aryl-α-amino acids. The synthetic potential of this protocol is further demonstrated in the sequential synthesis of diverse β-branched α-amino acids.

  14. 21 CFR 582.5118 - Alanine.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 6 2012-04-01 2012-04-01 false Alanine. 582.5118 Section 582.5118 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS SUBSTANCES GENERALLY RECOGNIZED AS SAFE Nutrients and/or Dietary Supplements...

  15. 21 CFR 582.5118 - Alanine.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 6 2014-04-01 2014-04-01 false Alanine. 582.5118 Section 582.5118 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS SUBSTANCES GENERALLY RECOGNIZED AS SAFE Nutrients and/or Dietary Supplements...

  16. 21 CFR 582.5118 - Alanine.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 6 2011-04-01 2011-04-01 false Alanine. 582.5118 Section 582.5118 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS SUBSTANCES GENERALLY RECOGNIZED AS SAFE Nutrients and/or Dietary Supplements...

  17. 21 CFR 582.5118 - Alanine.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 6 2013-04-01 2013-04-01 false Alanine. 582.5118 Section 582.5118 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS SUBSTANCES GENERALLY RECOGNIZED AS SAFE Nutrients and/or Dietary Supplements...

  18. 21 CFR 582.5118 - Alanine.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Alanine. 582.5118 Section 582.5118 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS SUBSTANCES GENERALLY RECOGNIZED AS SAFE Nutrients and/or Dietary Supplements...

  19. Indigenous amino acids in primitive CR meteorites

    NASA Astrophysics Data System (ADS)

    Martins, Z.; Alexander, C. M. O. D.; Orzechowska, G. E.; Fogel, M. L.; Ehrenfreund, P.

    CR chondrites are among the most primitive meteorites. In this paper, we report the first measurements of amino acids in Antarctic CR meteorites. Three CRs, Elephant Moraine (EET) 92042, Graves Nunataks (GRA) 95229, and Grosvenor Mountains (GRO) 95577, were analyzed for their amino acid content using high-performance liquid chromatography with UV fluorescence detection (HPLC-FD) and gas chromatography-mass spectrometry (GC-MS). Our data show that EET 92042 and GRA 95229 are the most amino acid-rich chondrites ever analyzed, with total amino acid concentrations ranging from 180 ppm to 249 ppm. The most abundant amino acids present in the EET 92042 and GRA 95229 meteorites are the α-amino acids glycine, isovaline, α-aminoisobutyric acid (α-AIB), and alanine, with δ13C values ranging from +31.6‰ to +50.5‰. The carbon isotope results together with racemic enantiomeric ratios determined for most amino acids strongly indicate an extraterrestrial origin for these compounds. Compared to Elephant Moraine (EET) 92042 and GRA 95229, the more aqueously altered GRO 95577 is depleted in amino acids. In both CRs and CMs, the absolute amino acid abundances appear to be related to the degree of aqueous alteration in their parent bodies. In addition, the relative abundances of α-AIB and β-alanine in the Antarctic CRs also appear to depend on the degree of aqueous alteration.

  20. Effect of glucose, independent of changes in insulin and glucagon secretion, on alanine metabolism in the conscious dog.

    PubMed Central

    Shulman, G I; Lacy, W W; Liljenquist, J E; Keller, U; Williams, P E; Cherrington, A D

    1980-01-01

    To study the effects of hyperglycemia on the metabolism of alanine and lactate independent of changes in plasma insulin and glucagon, glucose was infused into five 36-h-fasted dogs along with somatostatin and constant replacement amounts of both insulin and glucagon. Hepatic uptakes of alanine and lactate were calculated using the arteriovenous difference technique. [14C]Alanine was infused to measure the conversion of alanine and lactate into glucose. Hyperglycemia (delta 115 mg/dl) of 2 h duration caused the plasma alanine level to increase by over 50%. This change was caused by an increase in the inflow of alanine into plasma since the net hepatic uptake of the amino acid did not change. Taken together, the above findings indicate that glucose per se can significantly impair the fractional extraction of alanine by the liver. Hepatic extraction of lactate was also affected by hyperglycemia and had fallen to zero within 90 min of starting the glucose infusion. This fall was associated with a doubling of arterial lactate level. Conversion of [14C]-alanine and [14C]lactate into [14C]glucose was suppressed by 60 +/- 11% after 2 h of hyperglycemia, and because this fall could not be entirely accounted for by decreased lactate extraction an inhibitory effect of glucose on gluconeogenesis within the liver is suggested. These studies indicate that the plasma glucose level per se can be an important determinant of the level of alanine and lactate in plasma as well as the rate at which they are converted to glucose. PMID:7356691

  1. Distinguishing the cyanobacterial neurotoxin β-N-methylamino-L-alanine (BMAA) from other diamino acids.

    PubMed

    Banack, S A; Metcalf, J S; Spáčil, Z; Downing, T G; Downing, S; Long, A; Nunn, P B; Cox, P A

    2011-04-01

    β-N-methylamino-L-alanine (BMAA) is produced by diverse taxa of cyanobacteria, and has been detected by many investigators who have searched for it in cyanobacterial blooms, cultures and collections. Although BMAA is distinguishable from proteinogenic amino acids and its isomer 2,4-DAB using standard chromatographic and mass spectroscopy techniques routinely used for the analysis of amino acids, we studied whether BMAA could be reliably distinguished from other diamino acids, particularly 2,6-diaminopimelic acid which has been isolated from the cell walls of many bacterial species. We used HPLC-FD, UHPLC-UV, UHPLC-MS, and triple quadrupole tandem mass spectrometry (UHPLC-MS/MS) to differentiate BMAA from the diamino acids 2,6-diaminopimelic acid, N-2(amino)ethylglycine, lysine, ornithine, 2,4-diaminosuccinic acid, homocystine, cystine, tryptophan, as well as other amino acids including asparagine, glutamine, and methionine methylsulfonium.

  2. Synthesis and evaluation of a novel series of farnesyl protein transferase inhibitors as non-peptidic CAAX tetrapeptide analogues.

    PubMed

    Perez, Michel; Maraval, Catherine; Dumond, Stephan; Lamothe, Marie; Schambel, Philippe; Etiévant, Chantal; Hill, Bridget

    2003-04-17

    A novel series of compounds, derived from 4-amino-phenyl piperazine, has been designed to selectively inhibit farnesyl protein transferase (FPTase) as CAAX tetrapeptide analogues. Certain of these compounds were shown to possess low nanomolar inhibitory activity both against the isolated enzyme and in cultured cells.

  3. Amino acids

    MedlinePlus

    ... page: //medlineplus.gov/ency/article/002222.htm Amino acids To use the sharing features on this page, please enable JavaScript. Amino acids are organic compounds that combine to form proteins . ...

  4. BarR, an Lrp-type transcription factor in Sulfolobus acidocaldarius, regulates an aminotransferase gene in a β-alanine responsive manner.

    PubMed

    Liu, Han; Orell, Alvaro; Maes, Dominique; van Wolferen, Marleen; Lindås, Ann-Christin; Bernander, Rolf; Albers, Sonja-Verena; Charlier, Daniel; Peeters, Eveline

    2014-05-01

    In archaea, nothing is known about the β-alanine degradation pathway or its regulation. In this work, we identify and characterize BarR, a novel Lrp-like transcription factor and the first one that has a non-proteinogenic amino acid ligand. BarR is conserved in Sulfolobus acidocaldarius and Sulfolobus tokodaii and is located in a divergent operon with a gene predicted to encode β-alanine aminotransferase. Deletion of barR resulted in a reduced exponential growth rate in the presence of β-alanine. Furthermore, qRT-PCR and promoter activity assays demonstrated that BarR activates the expression of the adjacent aminotransferase gene, but only upon β-alanine supplementation. In contrast, auto-activation proved to be β-alanine independent. Heterologously produced BarR is an octamer in solution and forms a single complex by interacting with multiple sites in the 170 bp long intergenic region separating the divergently transcribed genes. In vitro, DNA binding is specifically responsive to β-alanine and site-mutant analyses indicated that β-alanine directly interacts with the ligand-binding pocket. Altogether, this work contributes to the growing body of evidence that in archaea, Lrp-like transcription factors have physiological roles that go beyond the regulation of α-amino acid metabolism.

  5. 21 CFR 172.540 - DL-Alanine.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 3 2010-04-01 2009-04-01 true DL-Alanine. 172.540 Section 172.540 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD FOR HUMAN... Agents and Related Substances § 172.540 DL-Alanine. DL-Alanine (a racemic mixture of D- and...

  6. On the existence of ‘L-alanine cadmium bromide'

    NASA Astrophysics Data System (ADS)

    Srinivasan, Bikshandarkoil R.

    2013-12-01

    It is argued that the recently reported nonlinear optical crystal L-alanine cadmium bromide, grown by slow solvent evaporation method at room temperature [P. Ilayabarathi, J. Chandrasekaran, Spectrochim. Acta 96A (2012) 684-689] is the well-known L-alanine crystal. The isolation of L-alanine crystal is explained due to fractional crystallization.

  7. On the existence of 'L-alanine cadmium bromide'.

    PubMed

    Srinivasan, Bikshandarkoil R

    2013-12-01

    It is argued that the recently reported nonlinear optical crystal L-alanine cadmium bromide, grown by slow solvent evaporation method at room temperature [P. Ilayabarathi, J. Chandrasekaran, Spectrochim. Acta 96A (2012) 684-689] is the well-known L-alanine crystal. The isolation of L-alanine crystal is explained due to fractional crystallization.

  8. Alanine Aminotransferase Variants Conferring Diverse NUE Phenotypes in Arabidopsis thaliana

    PubMed Central

    McAllister, Chandra H.; Good, Allen G.

    2015-01-01

    Alanine aminotransferase (AlaAT, E.C. 2.6.1.2), is a pyridoxal-5’-phosphate-dependent (PLP) enzyme that catalyzes the reversible transfer of an amino group from alanine to 2-oxoglutarate to produce glutamate and pyruvate, or vice versa. It has been well documented in both greenhouse and field studies that tissue-specific over-expression of AlaAT from barley (Hordeum vulgare, HvAlaAT) results in a significant increase in plant NUE in both canola and rice. While the physical phenotypes associated with over-expression of HvAlaAT have been well characterized, the role this enzyme plays in vivo to create a more N efficient plant remains unknown. Furthermore, the importance of HvAlaAT, in contrast to other AlaAT enzyme homologues in creating this phenotype has not yet been explored. To address the role of AlaAT in NUE, AlaAT variants from diverse sources and different subcellular locations, were expressed in the wild-type Arabidopsis thaliana Col-0 background and alaat1;2 (alaat1-1;alaat2-1) knockout background in various N environments. The analysis and comparison of both the physical and physiological properties of AlaAT over-expressing transgenic plants demonstrated significant differences between plants expressing the different AlaAT enzymes under different external conditions. This analysis indicates that the over-expression of AlaAT variants other than HvAlaAT in crop plants could further increase the NUE phenotype(s) previously observed. PMID:25830496

  9. Alanine aminotransferase variants conferring diverse NUE phenotypes in Arabidopsis thaliana.

    PubMed

    McAllister, Chandra H; Good, Allen G

    2015-01-01

    Alanine aminotransferase (AlaAT, E.C. 2.6.1.2), is a pyridoxal-5'-phosphate-dependent (PLP) enzyme that catalyzes the reversible transfer of an amino group from alanine to 2-oxoglutarate to produce glutamate and pyruvate, or vice versa. It has been well documented in both greenhouse and field studies that tissue-specific over-expression of AlaAT from barley (Hordeum vulgare, HvAlaAT) results in a significant increase in plant NUE in both canola and rice. While the physical phenotypes associated with over-expression of HvAlaAT have been well characterized, the role this enzyme plays in vivo to create a more N efficient plant remains unknown. Furthermore, the importance of HvAlaAT, in contrast to other AlaAT enzyme homologues in creating this phenotype has not yet been explored. To address the role of AlaAT in NUE, AlaAT variants from diverse sources and different subcellular locations, were expressed in the wild-type Arabidopsis thaliana Col-0 background and alaat1;2 (alaat1-1;alaat2-1) knockout background in various N environments. The analysis and comparison of both the physical and physiological properties of AlaAT over-expressing transgenic plants demonstrated significant differences between plants expressing the different AlaAT enzymes under different external conditions. This analysis indicates that the over-expression of AlaAT variants other than HvAlaAT in crop plants could further increase the NUE phenotype(s) previously observed.

  10. Roles for glutathione transferases in antioxidant recycling

    PubMed Central

    Dixon, David P; Steel, Patrick G

    2011-01-01

    Uniquely among the plant glutathione transferases, two classes possess a catalytic cysteine capable of performing glutathione-dependent reductions. These are the dehydroascorbate reductases (DHARs) and the lambda-class glutathione transferases (GSTLs). Using immobilized GSTLs probed with crude plant extracts we have identified flavonols as high affinity ligands and subsequently demonstrated a novel glutathione-dependent role for these enzymes in recycling oxidized quercetin. By comparing the activities of DHARs and GSTLs we now propose a unified catalytic mechanism that suggests oxidized anthocyanidins and tocopherols may be alternative polyphenolic substrates of GSTLs. PMID:21778824

  11. ANTIGENICITY OF POLYPEPTIDES (POLY ALPHA AMINO ACIDS)

    PubMed Central

    Pinchuck, Paul; Maurer, Paul H.

    1965-01-01

    The response of mice to synthetic linear polypeptides of known composition but random sequence has been studied. Neither Swiss mice nor a number of inbred strains could respond to copolymers of only 2 amino acids (G60L40, G60A40, G90T10). Upon introduction of as little as 4 mole per cent of a third amino acid, good immune responses were obtained, regardless of the nature of the third amino acid. The level of the immune response to a series of glu-lys-ala polymers increased with increasing alanine content of the polymer. PMID:5849232

  12. Amino acids in the Yamato carbonaceous chrondrite from Antarctica

    NASA Technical Reports Server (NTRS)

    Shimoyama, A.; Ponnamperuma, C.; Yanai, K.

    1979-01-01

    Evidence for the presence of amino acids of extraterrestrial origin in the Antarctic Yamato carbonaceous chrondrite is presented. Hydrolyzed and nonhydrolyzed water-extracted amino acid samples from exterior, middle and interior portions of the meteorite were analyzed by an amino acid analyzer and by gas chromatography of N-TFA-isopropyl amino acid derivatives. Nine protein and six nonprotein amino acids were detected in the meteorite at abundances between 34 and less than one nmole/g, with equal amounts in interior and exterior portions. Nearly equal abundances of the D and L enantiomers of alanine, aspartic acid and glutamic acid were found, indicating the abiotic, therefore extraterrestrial, origin of the amino acids. The Antarctic environment and the uniformity of protein amino acid abundances are discussed as evidence against the racemization of terrestrially acquired amino acids, and similarities between Yamato amino acid compositions and the amino acid compositions of the Murchison and Murray type II carbonaceous chrondrites are indicated.

  13. Protein Homeostasis Defects of Alanine-Glyoxylate Aminotransferase: New Therapeutic Strategies in Primary Hyperoxaluria Type I

    PubMed Central

    Pey, Angel L.; Albert, Armando; Salido, Eduardo

    2013-01-01

    Alanine-glyoxylate aminotransferase catalyzes the transamination between L-alanine and glyoxylate to produce pyruvate and glycine using pyridoxal 5′-phosphate (PLP) as cofactor. Human alanine-glyoxylate aminotransferase is a peroxisomal enzyme expressed in the hepatocytes, the main site of glyoxylate detoxification. Its deficit causes primary hyperoxaluria type I, a rare but severe inborn error of metabolism. Single amino acid changes are the main type of mutation causing this disease, and considerable effort has been dedicated to the understanding of the molecular consequences of such missense mutations. In this review, we summarize the role of protein homeostasis in the basic mechanisms of primary hyperoxaluria. Intrinsic physicochemical properties of polypeptide chains such as thermodynamic stability, folding, unfolding, and misfolding rates as well as the interaction of different folding states with protein homeostasis networks are essential to understand this disease. The view presented has important implications for the development of new therapeutic strategies based on targeting specific elements of alanine-glyoxylate aminotransferase homeostasis. PMID:23956997

  14. Positron and electron scattering by glycine and alanine: Shape resonances and methylation effect

    NASA Astrophysics Data System (ADS)

    Nunes, Fernanda B.; Bettega, Márcio H. F.; Sanchez, Sergio d'Almeida

    2016-12-01

    We report integral cross sections (ICSs) for both positron and electron scattering by glycine and alanine amino acids. These molecules differ only by a methyl group. We computed the scattering cross sections using the Schwinger multichannel method for both glycine and alanine in different levels of approximation for both projectiles. The alanine ICSs are greater in magnitude than the glycine ICSs for both positron and electron scattering, probably due to the larger size of the molecule. In electron scattering calculations, we found two resonances for each molecule. Glycine presents one at 1.8 eV, and another centered at around 8.5 eV, in the static-exchange plus polarization (SEP) approximation. The ICS for alanine shows one resonance at 2.5 eV and another at around 9.5 eV, also in SEP approximation. The results are in good agreement with most of the data present in the literature. The comparison of the electron scattering ICSs for both molecules indicates that the methylation of glycine destabilizes the resonances, shifting them to higher energies.

  15. Response of amino acids in hindlimb muscles to recovery from hypogravity and unloading by tail-cast suspension

    NASA Technical Reports Server (NTRS)

    Tischler, M. E.; Henriksen, E. J.; Jacob, S.; Cook, P. H.

    1985-01-01

    Concentrations of glutamine, glutamate, aspartate (+ asparagine) and alanine were compared in hindlimb muscles of SL-3 and ground control rats. Alanine was lower in the soleus of flown rats but not of suspended animals, with no response in other muscles except a slight increase in the unloaded plantaris. With recovery, alanine in the soleus was elevated. Since no differences in alanine metabolism were found by isolated muscle, changes in muscle alanine are probably due to altered body use of this amino acid leading to varied plasma levels.

  16. Barrier-Free Intermolecular Proton Transfer Induced by Excess Electron Attachment to the Complex of Alanine with Uracil

    SciTech Connect

    Dabkowska, Iwona; Rak, Janusz; Gutowski, Maciej S.; Nilles, J.M.; Stokes, Sarah; Bowen, Kit H.

    2004-04-01

    The photoelectron spectrum of the uracil-alanine anionic complex (UA)- has been recorded with 2.540 eV photons. This spectrum reveals a broad feature with a maximum between 1.6-2.1 eV. The vertical electron detachment energy is too large to be attributed to an (UA)- anionic complex in which an intact uracil anion is solvated by alanine, or vice versa. The neutral and anionic complexes of uracil and alanine were studied at the B3LYP and second order Moeller-Plesset level of theory with 6-31++G** basis sets. The neutral complexes form cyclic hydrogen bonds and the three most stable neutral complexes are bound by 0.72, 0.61 and 0.57 eV. The electron hole in complexes of uracil with alaninie is localized on uracil, but the formation of a complex with alanine strongly modulates the vertical ionization energy of uracil. The theoretical results indicate that the excess electron in (UA)- occupies a p* orbital localized on uracil. The excess electron attachment to the complex can induce a barrier-free proton transfer (BFPT) from the carboxylic group of alanine to the O8 atom of uracil. As a result, the four most stable structures of the uracil-alanine anionic complex can be characterized as the neutral radical of hydrogenated uracil solvated by the anion of deprotonated alanine. Our current results for the anionic complex of uracil with alanine are similar to our previous results for the anion of uracil with glycine [Eur. Phys. J. D 20, 431 (2002)], and together they indicate that the BFPT process is not very sensitive to the nature of the amino acid's hydrophobic residual group. The BFPT to the O8 atom of uracil may be relevant to the damage suffered by nucleic acid bases due to exposure to low energy electrons.

  17. Amino acids in the Martian meteorite Nakhla.

    PubMed

    Glavin, D P; Bada, J L; Brinton, K L; McDonald, G D

    1999-08-03

    A suite of protein and nonprotein amino acids were detected with high-performance liquid chromatography in the water- and acid-soluble components of an interior fragment of the Martian meteorite Nakhla, which fell in Egypt in 1911. Aspartic and glutamic acids, glycine, alanine, beta-alanine, and gamma-amino-n-butyric acid (gamma-ABA) were the most abundant amino acids detected and were found primarily in the 6 M HCl-hydrolyzed, hot water extract. The concentrations ranged from 20 to 330 parts per billion of bulk meteorite. The amino acid distribution in Nakhla, including the D/L ratios (values range from <0.1 to 0.5), is similar to what is found in bacterially degraded organic matter. The amino acids in Nakhla appear to be derived from terrestrial organic matter that infiltrated the meteorite soon after its fall to Earth, although it is possible that some of the amino acids are endogenous to the meteorite. The rapid amino acid contamination of Martian meteorites after direct exposure to the terrestrial environment has important implications for Mars sample-return missions and the curation of the samples from the time of their delivery to Earth.

  18. Amino acids in the Martian meteorite Nakhla

    NASA Technical Reports Server (NTRS)

    Glavin, D. P.; Bada, J. L.; Brinton, K. L.; McDonald, G. D.

    1999-01-01

    A suite of protein and nonprotein amino acids were detected with high-performance liquid chromatography in the water- and acid-soluble components of an interior fragment of the Martian meteorite Nakhla, which fell in Egypt in 1911. Aspartic and glutamic acids, glycine, alanine, beta-alanine, and gamma-amino-n-butyric acid (gamma-ABA) were the most abundant amino acids detected and were found primarily in the 6 M HCl-hydrolyzed, hot water extract. The concentrations ranged from 20 to 330 parts per billion of bulk meteorite. The amino acid distribution in Nakhla, including the D/L ratios (values range from <0.1 to 0.5), is similar to what is found in bacterially degraded organic matter. The amino acids in Nakhla appear to be derived from terrestrial organic matter that infiltrated the meteorite soon after its fall to Earth, although it is possible that some of the amino acids are endogenous to the meteorite. The rapid amino acid contamination of Martian meteorites after direct exposure to the terrestrial environment has important implications for Mars sample-return missions and the curation of the samples from the time of their delivery to Earth.

  19. Amino Acids in the Martian Meteorite Nakhla

    NASA Astrophysics Data System (ADS)

    Glavin, Daniel P.; Bada, Jeffrey L.; Brinton, Karen L. F.; McDonald, Gene D.

    1999-08-01

    A suite of protein and nonprotein amino acids were detected with high-performance liquid chromatography in the water- and acid-soluble components of an interior fragment of the Martian meteorite Nakhla, which fell in Egypt in 1911. Aspartic and glutamic acids, glycine, alanine, β -alanine, and γ -amino-n-butyric acid (γ -ABA) were the most abundant amino acids detected and were found primarily in the 6 M HCl-hydrolyzed, hot water extract. The concentrations ranged from 20 to 330 parts per billion of bulk meteorite. The amino acid distribution in Nakhla, including the D/L ratios (values range from <0.1 to 0.5), is similar to what is found in bacterially degraded organic matter. The amino acids in Nakhla appear to be derived from terrestrial organic matter that infiltrated the meteorite soon after its fall to Earth, although it is possible that some of the amino acids are endogenous to the meteorite. The rapid amino acid contamination of Martian meteorites after direct exposure to the terrestrial environment has important implications for Mars sample-return missions and the curation of the samples from the time of their delivery to Earth.

  20. 3-Hydroxykynurenine transaminase identity with alanine glyoxylate transaminase. A probable detoxification protein in Aedes aegypti.

    PubMed

    Han, Qian; Fang, Jianmin; Li, Jianyong

    2002-05-03

    This study describes the functional characterization of a specific mosquito transaminase responsible for catalyzing the transamination of 3-hydroxykynurenine (3-HK) to xanthurenic acid (XA). The enzyme was purified from Aedes aegypti larvae by ammonium sulfate fractionation, heat treatment, and various chromatographic techniques, plus non-denaturing electrophoresis. The purified transaminase has a relative molecular mass of 42,500 by SDS-PAGE. N-terminal and internal sequencing of the purified protein and its tryptic fragments resolved a partial N-terminal sequence of 19 amino acid residues and 3 partial internal peptide sequences with 7, 10, and 7 amino acid residues. Using degenerate primers based on the partial internal sequences for PCR amplification and cDNA library screening, a full-length cDNA clone with a 1,167-bp open reading frame was isolated. Its deduced amino acid sequence consists of 389 amino acid residues with a predicted molecular mass of 43,239 and shares 45-46% sequence identity with mammalian alanine glyoxylate transaminases. Northern analysis shows the active transcription of the enzyme in larvae and developing eggs. Substrate specificity analysis of this mosquito transaminase demonstrates that the enzyme is active with 3-HK, kynurenine, or alanine substrates. The enzyme has greater affinity and catalytic efficiency for 3-HK than for kynurenine and alanine. The biochemical characteristics of the enzyme in conjunction with the profiles of 3-HK transaminase activity and XA accumulation during mosquito development clearly point out its physiological function in the 3-HK to XA pathway. Our data suggest that the mosquito transaminase was evolved in a manner precisely reflecting the physiological requirement of detoxifying 3-HK produced in the tryptophan oxidation pathway in the mosquito.

  1. Unusual hydroxyl migration in the fragmentation of β-alanine dication in the gas phase.

    PubMed

    Piekarski, Dariusz Grzegorz; Delaunay, Rudy; Maclot, Sylvain; Adoui, Lamri; Martín, Fernando; Alcamí, Manuel; Huber, Bernd A; Rousseau, Patrick; Domaracka, Alicja; Díaz-Tendero, Sergio

    2015-07-14

    We present a combined experimental and theoretical study of the fragmentation of doubly positively charged β-alanine molecules in the gas phase. The dissociation of the produced dicationic molecules, induced by low-energy ion collisions, is analysed by coincidence mass spectrometric techniques; the coupling with ab initio molecular dynamics simulations allows rationalisation of the experimental observations. The present strategy gives deeper insights into the chemical mechanisms of multiply charged amino acids in the gas phase. In the case of the β-alanine dication, in addition to the expected Coulomb explosion and hydrogen migration processes, we have found evidence of hydroxyl-group migration, which leads to unusual fragmentation products, such as hydroxymethyl cation, and is necessary to explain some of the observed dominant channels.

  2. Chiral effects on helicity studied via the energy landscape of short (D, L)-alanine peptides.

    PubMed

    Neelamraju, Sridhar; Oakley, Mark T; Johnston, Roy L

    2015-10-28

    The homochirality of natural amino acids facilitates the formation of regular secondary structures such as α-helices and β-sheets. Here, we study the relationship between chirality and backbone structure for the example of hexa-alanine. The most stable stereoisomers are identified through global optimisation. Further, the energy landscape, a database of connected low-energy local minima and transition points, is constructed for various neutral and zwitterionic stereoisomers of hexa-alanine. Three order parameters for partial helicity are applied and metric disconnectivity graphs are presented with partial helicity as a metric. We also apply the Zimm-Bragg model to derive average partial helicities for Ace-(L-Ala)6-NHMe, Ace-(D-Ala-L-Ala)3-NHMe, and Ace-(L-Ala)3-(D-Ala)3-NHMe from the database of local minima and compare with previous studies.

  3. Membrane topology of the electrogenic aspartate-alanine antiporter AspT of Tetragenococcus halophilus.

    PubMed

    Nanatani, Kei; Ohonishi, Fumito; Yoneyama, Hiroshi; Nakajima, Tasuku; Abe, Keietsu

    2005-03-04

    AspT is an electrogenic aspartate:alanine exchange protein that represents the vectorial component of a proton-motive metabolic cycle found in some strains of Tetragenococcus halophilus. AspT is the sole member of a new family, the Aspartate: Alanine Exchanger (AAE) family, in secondary transporters, according to the computational classification proposed by Saier et al. (http://www.biology.ucsd.edu/~msaier/transport/). We analyzed the topology of AspT biochemically, by using fusion methods in combination with alkaline phosphatase or beta-lactamase. These results suggested that AspT has a unique topology; 8 TMS, a large cytoplasmic loop (183 amino acids) between TMS5 and TMS6, and N- and C-termini that both face the periplasm. These results demonstrated a unique 2D-structure of AspT as the novel AAE family.

  4. Structure-function relationship in the antifreeze activity of synthetic alanine-lysine antifreeze polypeptides.

    PubMed

    Wierzbicki, A; Knight, C A; Rutland, T J; Muccio, D D; Pybus, B S; Sikes, C S

    2000-01-01

    Recently antifreeze proteins (AFP) have been the subject of many structure-function relationship studies regarding their antifreeze activity. Attempts have been made to elucidate the structure-function relationship by various amino acid substitutions, but to our knowledge there has been no successful from first principles design of a polypeptide that would bind to designated ice planes along a specific direction. In this paper we show the results of our first attempt on an entirely de novo design of an alanine-lysine-rich antifreeze polypeptide. This 43 residue alanine-lysine peptide exhibits characteristic nonequilibrium freezing point depression and binds to the designated (210) planes of ice along the [122] vector. The structural and thermodynamic properties of this polypeptide were determined using circular dichroism spectroscopy and its nonequilibrium antifreeze properties were investigated using an ice-etching method and nanoliter osmometry.

  5. Thiophenyl-substituted triazolyl-thione L-alanine: asymmetric synthesis, aggregation and biological properties.

    PubMed

    Saghyan, Ashot S; Simonyan, Hayarpi M; Petrosyan, Satenik G; Geolchanyan, Arpine V; Roviello, Giovanni N; Musumeci, Domenica; Roviello, Valentina

    2014-10-01

    In this work, we report the asymmetric synthesis and characterization of an artificial amino acid based on triazolyl-thione L-alanine, which was modified with a thiophenyl-substituted moiety, as well as in vitro studies of its nucleic acid-binding ability. We found, by dynamic light scattering studies, that the synthetic amino acid was able to form supramolecular aggregates having a hydrodynamic diameter higher than 200 nm. Furthermore, we demonstrated, by UV and CD experiments, that the heteroaromatic amino acid, whose enzymatic stability was demonstrated by HPLC analysis also after 24 h of incubation in human serum, was able to bind a RNA complex, which is a feature of biomedical interest in view of innovative antiviral strategies based on modulation of RNA-RNA molecular recognition.

  6. NQR in Alanine and Lysine Iodates

    NASA Astrophysics Data System (ADS)

    Petrosyan, A. M.; Burbelo, V. M.; Tamazyan, R. A.; Karapetyan, H. A.; Sukiasyan, R. P.

    2000-02-01

    The structure o f iodates of α- and β-alanine ( Ala) (2(β-Ala • HIO3) • H2O , β-Ala-2HIO3 , D L-Ala• HIO3 • 2H2O, L-Ala • HIO3) and L-lysine (L-Lys) (L-Lys • HIO3, L-Lys • 2HIO3,L-Lys • 3HIO3, L-Lys • 6HIO3) have been investigated by means of iodine-127 NQR, IR spectroscopy and X-ray diffraction

  7. Alanine racemase is essential for the growth and interspecies competitiveness of Streptococcus mutans

    PubMed Central

    Wei, Yuan; Qiu, Wei; Zhou, Xue-Dong; Zheng, Xin; Zhang, Ke-Ke; Wang, Shi-Da; Li, Yu-Qing; Cheng, Lei; Li, Ji-Yao; Xu, Xin; Li, Ming-Yun

    2016-01-01

    D-alanine (D-Ala) is an essential amino acid that has a key role in bacterial cell wall synthesis. Alanine racemase (Alr) is a unique enzyme that interconverts L-alanine and D-alanine in most bacteria, making this enzyme a potential target for antimicrobial drug development. Streptococcus mutans is a major causative factor of dental caries. The factors involved in the survival, virulence and interspecies interactions of S. mutans could be exploited as potential targets for caries control. The current study aimed to investigate the physiological role of Alr in S. mutans. We constructed alr mutant strain of S. mutans and evaluated its phenotypic traits and interspecies competitiveness compared with the wild-type strain. We found that alr deletion was lethal to S. mutans. A minimal supplement of D-Ala (150 μg·mL−1) was required for the optimal growth of the alr mutant. The depletion of D-alanine in the growth medium resulted in cell wall perforation and cell lysis in the alr mutant strain. We also determined the compromised competitiveness of the alr mutant strain relative to the wild-type S. mutans against other oral streptococci (S. sanguinis or S. gordonii), demonstrated using either conditioned medium assays or dual-species fluorescent in situ hybridization analysis. Given the importance and necessity of alr to the growth and competitiveness of S. mutans, Alr may represent a promising target to modulate the cariogenicity of oral biofilms and to benefit the management of dental caries. PMID:27740612

  8. Hepatic serine and alanine metabolism during endotoxin-induced fever in sheep.

    PubMed Central

    Southorn, B G; Thompson, J R

    1987-01-01

    Time course changes in plasma amino acid concentrations and the hepatic metabolism of serine and alanine were measured in six mature wethers during endotoxin-induced fever. In separate trials, the animals' responses to injections of saline and endotoxin were measured. The endotoxin was from Escherichia coli serotype 055:B5 and was injected intravenously (4 micrograms/kg body weight). Liver biopsies were obtained from the sheep at 6 h postinjection during both endotoxin and saline injection trials. Rectal temperature in the endotoxin treated animals was increased (P less than 0.05, above that in control animals from 4.25 h to 9 h postinjection, with a maximum rise of 2.43 degrees C at 5.5 h postinjection. Glucose concentration in jugular plasma decreased (P less than 0.05) by 3 h postinjection and remained depressed throughout the 24 h postinjection sampling period. Plasma serine concentration was decreased (P less than 0.05) by 3 h postinjection. Plasma alanine concentration was decreased significantly (P less than 0.05) only at 24 h postinjection. Endotoxin injection increased (P less than 0.05) hepatic oxidation of 14C-serine (162%) and the net incorporation of 14C-serine carbon into hepatic protein (173%) and glycogen (275%). The net incorporation of 14C-alanine carbon into hepatic protein (172%) and glycogen (323%) were increased (P less than 0.05) by endotoxin injection, while alanine oxidation was not affected by endotoxin treatment (P greater than 0.05). The increased hepatic use of serine may explain, in part, the dramatic decrease in plasma concentrations of this amino acid following endotoxin injection into sheep. PMID:3115552

  9. Crystallization of jarosite in the presence of amino acids

    NASA Astrophysics Data System (ADS)

    Crabbe, Harrison; Fernandez, Natalya; Jones, Franca

    2015-04-01

    Jarosite was formed in the presence of five amino acids at two pHs, namely 1.75 and 2.9, to determine what impact amino acids have on its formation. It was found that at the lower pH glycine was the most potent in terms of morphological and yield impacts. XRD analysis showed that incorporation of the amino acid occurs at this low pH for glycine and proline. Dynamic light scattering studies showed that glycine impacts significantly on the jarosite nucleation rate while proline and alanine do not. At the higher pH all of the amino acids had much less impact on morphology or yield. At pH 3 the solids were found to be a 3-phase system consisting of goethite, schwertmannite and jarosite. In this case, alanine appeared to stabilise the presence of schwertmannite more than the other amino acids.

  10. Effect of amino acid intake on brush-border membrane uptake of sulfur amino acids.

    PubMed

    Chesney, R W; Gusowski, N; Padilla, M; Lippincott, S

    1986-07-01

    Alterations in the intake of sulfur amino acids (SAA) changes the rat renal brush-border membrane uptake of the beta-amino acid, taurine. A low-SAA diet enhances and a high-taurine diet reduces uptake (Chesney et al., Kidney Int. 24: 588-594, 1983). Neither the low-SAA diet nor the high-taurine diet alters the time course or concentration-dependent accumulation of the sulfur amino acids methionine and cystine or of inorganic sulfate. By contrast the uptake of beta-alanine, another beta-amino acid that competes with taurine, is greater in animals on the low-SAA diet. The high-taurine diet does not change beta-alanine uptake. The plasma levels of taurine are altered by dietary change, but not the values for methionine and cystine. This study indicates that renal adaptation is expressed for beta-alanine, a nonsulfur-containing beta-amino acid. By contrast, methionine, cystine, and sulfate, which participate in a variety of synthetic and conjugative processes, are not conserved by the renal brush-border surface following ingestion of either a low-methionine and -cystine diet or high-taurine diet.

  11. The Origin of Amino Acids in Lunar Regolith Samples

    NASA Technical Reports Server (NTRS)

    Cook, Jamie E.; Callahan, Michael P.; Dworkin, Jason P.; Glavin, Daniel P.; McLain, Hannah L.; Noble, Sarah K.; Gibson, Everett K., Jr.

    2016-01-01

    We analyzed the amino acid content of seven lunar regolith samples returned by the Apollo 16 and Apollo 17 missions and stored under NASA curation since collection using ultrahigh-performance liquid chromatography with fluorescence detection and time-of-flight mass spectrometry. Consistent with results from initial analyses shortly after collection in the 1970s, we observed amino acids at low concentrations in all of the curated samples, ranging from 0.2 parts-per-billion (ppb) to 42.7 ppb in hot-water extracts and 14.5 ppb to 651.1 ppb in 6M HCl acid-vapor-hydrolyzed, hot-water extracts. Amino acids identified in the Apollo soil extracts include glycine, D- and L-alanine, D- and L-aspartic acid, D- and L-glutamic acid, D- and L-serine, L-threonine, and L-valine, all of which had previously been detected in lunar samples, as well as several compounds not previously identified in lunar regoliths: -aminoisobutyric acid (AIB), D-and L-amino-n-butyric acid (-ABA), DL-amino-n-butyric acid, -amino-n-butyric acid, -alanine, and -amino-n-caproic acid. We observed an excess of the L enantiomer in most of the detected proteinogenic amino acids, but racemic alanine and racemic -ABA were present in some samples.

  12. Amino Acid Transport in Mycobacterium smegmatis

    PubMed Central

    Yabu, Kunihiko

    1970-01-01

    The transport of d-alanine, d-glutamic acid, and d-valine in Mycobacterium smegmatis was compared quantitatively with that of their l-isomers. It appeared that the uptake of d-alanine was mediated by an active process displaying saturation kinetics characteristic of enzyme function, whereas the uptake of d-glutamic acid was accomplished by a passive process showing diffusion kinetics. Both processes were involved in the uptake of l-alanine, l-glutamic acid, d-valine, and l-valine. d-Valine competed with l-valine for entry into the cell through a single active process. d-Alanine and l-alanine also utilized the same active process, but the d-isomer could not enter the cell through the passive process. The passive process exhibited characteristics of diffusion, but was sensitive to sulfhydryl-blocking reagents and showed competition among structurally related amino acids. These last findings suggested that the passive process is a facilitated diffusion. PMID:5437732

  13. Crystallization and preliminary crystallographic analysis of d-alanine-d-alanine ligase from Streptococcus mutans

    SciTech Connect

    Lu, Yong-Zhi; Sheng, Yu; Li, Lan-Fen; Tang, De-Wei; Liu, Xiang-Yu; Zhao, Xiaojun; Liang, Yu-He Su, Xiao-Dong

    2007-09-01

    A potential target for antibiotic drug design, d-alanine-d-alanine ligase from S. mutans, was expressed in E. coli, purified and crystallized. Diffraction data were collected to 2.4 Å resolution. d-Alanine-d-alanine ligase is encoded by the gene ddl (SMU-599) in Streptococcus mutans. This ligase plays a very important role in cell-wall biosynthesis and may be a potential target for drug design. To study the structure and function of this ligase, the gene ddl was amplified from S. mutans genomic DNA and cloned into the expression vector pET28a. The protein was expressed in soluble form in Escherichia coli strain BL21 (DE3). Homogeneous protein was obtained using a two-step procedure consisting of Ni{sup 2+}-chelating and size-exclusion chromatography. Purified protein was crystallized and the cube-shaped crystal diffracted to 2.4 Å. The crystal belongs to space group P3{sub 1}21 or P3{sub 2}21, with unit-cell parameters a = b = 79.50, c = 108.97 Å. There is one molecule per asymmetric unit.

  14. Racemization of alanine by the alanine racemases from Salmonella typhimurium and Bacillus stearothermophilus: energetic reaction profiles

    SciTech Connect

    Faraci, W.S.; Walsh, C.T.

    1988-05-03

    Alanine racemases are bacterial pyridoxal 5'-phosphate (PLP) dependent enzymes providing D-alanine as an essential building block for biosynthesis of the peptidoglycan layer of the cell wall. Two isozymic alanine racemases, encoded by the dadB gene and the alr gene, from the Gram-negative mesophilic Salmonella typhimurium and one from the Gram-positive thermophilic Bacillus stearothermophilus have been examined for the racemization mechanism. Substrate deuterium isotope effects and solvent deuterium isotope effects have been measured in both L ..-->.. D and D..-->.. L directions for all three enzymes to assess the degree to which abstraction of the ..cap alpha..-proton or protonation of substrate PLP carbanion is limiting in catalysis. Additionally, experiments measuring internal return of ..cap alpha..-/sup 3/H from substrate to product and solvent exchange/substrate conversion experiments in /sup 3/H/sub 2/O have been used with each enzyme to examine the partitioning of substrate PLP carbanion intermediates and to obtain the relative heights of kinetically significant energy barriers in alanine racemase catalysis.

  15. β-N-Methylamino-L-alanine (BMAA) perturbs alanine, aspartate and glutamate metabolism pathways in human neuroblastoma cells as determined by metabolic profiling.

    PubMed

    Engskog, Mikael K R; Ersson, Lisa; Haglöf, Jakob; Arvidsson, Torbjörn; Pettersson, Curt; Brittebo, Eva

    2017-02-04

    β-Methylamino-L-alanine (BMAA) is a non-proteinogenic amino acid that induces long-term cognitive deficits, as well as an increased neurodegeneration and intracellular fibril formation in the hippocampus of adult rodents following short-time neonatal exposure and in vervet monkey brain following long-term exposure. It has also been proposed to be involved in the etiology of neurodegenerative disease in humans. The aim of this study was to identify metabolic effects not related to excitotoxicity or oxidative stress in human neuroblastoma SH-SY5Y cells. The effects of BMAA (50, 250, 1000 µM) for 24 h on cells differentiated with retinoic acid were studied. Samples were analyzed using LC-MS and NMR spectroscopy to detect altered intracellular polar metabolites. The analysis performed, followed by multivariate pattern recognition techniques, revealed significant perturbations in protein biosynthesis, amino acid metabolism pathways and citrate cycle. Of specific interest were the BMAA-induced alterations in alanine, aspartate and glutamate metabolism and as well as alterations in various neurotransmitters/neuromodulators such as GABA and taurine. The results indicate that BMAA can interfere with metabolic pathways involved in neurotransmission in human neuroblastoma cells.

  16. Effect of amino acid immobilization on the impedance of graphene oxide

    NASA Astrophysics Data System (ADS)

    Tran, Minh-Hai; Han, Jinwoo; Min, Byeong June; Lee, ChangWoo; Jang, Sei-Heon; Jeong, Hae Kyung

    2015-05-01

    A single residue, dipeptide, or tripeptide of alanine or histidine is covalently attached to graphene oxide (GO), and the effect of the amino acid immobilization on the impedance of GO is investigated using the impedance spectroscopy. The histidine of a tripeptide exhibits the lowest resistance compared to the single or dipeptide histidine in the KCl electrolyte, and the single alanine residue shows the lowest resistance in an acidic electrolyte compared to the dipeptide or tripeptide alanine. The peculiar behavior of the impedance could be explained by different net charges of the amino acids, chain length, and π-π stacking interaction.

  17. Inhibition of glutamine synthesis induces glutamate dehydrogenase-dependent ammonia fixation into alanine in co-cultures of astrocytes and neurons.

    PubMed

    Dadsetan, Sherry; Bak, Lasse K; Sørensen, Michael; Keiding, Susanne; Vilstrup, Hendrik; Ott, Peter; Leke, Renata; Schousboe, Arne; Waagepetersen, Helle S

    2011-09-01

    It has been previously demonstrated that ammonia exposure of neurons and astrocytes in co-culture leads to net synthesis not only of glutamine but also of alanine. The latter process involves the concerted action of glutamate dehydrogenase (GDH) and alanine aminotransferase (ALAT). In the present study it was investigated if the glutamine synthetase (GS) inhibitor methionine sulfoximine (MSO) would enhance alanine synthesis by blocking the GS-dependent ammonia scavenging process. Hence, co-cultures of neurons and astrocytes were incubated for 2.5h with [U-(13)C]glucose to monitor de novo synthesis of alanine and glutamine in the absence and presence of 5.0 mM NH(4)Cl and 10 mM MSO. Ammonia exposure led to increased incorporation of label but not to a significant increase in the amount of these amino acids. However, in the presence of MSO, glutamine synthesis was blocked and synthesis of alanine increased leading to an elevated content intra- as well as extracellularly of this amino acid. Treatment with MSO led to a dramatic decrease in glutamine content and increased the intracellular contents of glutamate and aspartate. The large increase in alanine during exposure to MSO underlines the importance of the GDH and ALAT biosynthetic pathway for ammonia fixation, and it points to the use of a GS inhibitor to ameliorate the brain toxicity and edema induced by hyperammonemia, events likely related to glutamine synthesis.

  18. Crystal Structure of a Thermostable Alanine Racemase from Thermoanaerobacter tengcongensis MB4 Reveals the Role of Gln360 in Substrate Selection.

    PubMed

    Sun, Xiaoliang; He, Guangzheng; Wang, Xiaoyan; Xu, Shujing; Ju, Jiansong; Xu, Xiaoling

    2015-01-01

    Pyridoxal 5'-phosphate (PLP) dependent alanine racemase catalyzes racemization of L-Ala to D-Ala, a key component of the peptidoglycan network in bacterial cell wall. It has been extensively studied as an important antimicrobial drug target due to its restriction in eukaryotes. However, many marketed alanine racemase inhibitors also act on eukaryotic PLP-dependent enzymes and cause side effects. A thermostable alanine racemase (AlrTt) from Thermoanaerobacter tengcongensis MB4 contains an evolutionarily non-conserved residue Gln360 in inner layer of the substrate entryway, which is supposed to be a key determinant in substrate specificity. Here we determined the crystal structure of AlrTt in complex with L-Ala at 2.7 Å resolution, and investigated the role of Gln360 by saturation mutagenesis and kinetic analysis. Compared to typical bacterial alanine racemase, presence of Gln360 and conformational changes of active site residues disrupted the hydrogen bonding interactions necessary for proper PLP immobilization, and decreased both the substrate affinity and turnover number of AlrTt. However, it could be complemented by introduction of hydrophobic amino acids at Gln360, through steric blocking and interactions with a hydrophobic patch near active site pocket. These observations explained the low racemase activity of AlrTt, revealed the essential role of Gln360 in substrate selection, and its preference for hydrophobic amino acids especially Tyr in bacterial alanine racemization. Our work will contribute new insights into the alanine racemization mechanism for antimicrobial drug development.

  19. Detection of non-protein amino acids in the presence of protein amino acids. II.

    NASA Technical Reports Server (NTRS)

    Shapshak, P.; Okaji, M.

    1972-01-01

    Studies conducted with the JEOL 5AH amino acid analyzer are described. This instrument makes possible the programming of the chromatographic process. Data are presented showing the separations of seventeen non-protein amino acids in the presence of eighteen protein amino acids. It is pointed out that distinct separations could be obtained in the case of a number of chemically similar compounds, such as ornithine and lysine, N-amidino alanine and arginine, and iminodiacetic acid and S-carboxymethyl cysteine and aspartic acid.

  20. Purification and characterization of the Oligosaccharyl transferase

    SciTech Connect

    Kapoor, T.M.

    1990-11-01

    Oligosaccharyl transferase was characterized to be a glycoprotein with at least one saccharide unit that had a D-manno or D- glucopyranose configuration with unmodified hydroxy groups at C-3, C-4 and C-6, using a Concanavalin A affinity column. This afforded a 100 fold increase in the transferase purity in the solubilized microsomal sample and also removed over 90% of the microsomal proteins (the cytosolic ones being removed before solubilization). The detergent, N,N-Dimethyldodecylamine N-oxide (LDAO) was used for solubilization and it yielded a system compatible with the assay and the purification steps. An efficient method for detergent extraction without dilution of sample or protein precipitation was also developed.

  1. Neonatal taurine and alanine modulate anxiety-like behavior and decelerate cortical spreading depression in rats previously suckled under different litter sizes.

    PubMed

    Francisco, Elian da Silva; Guedes, Rubem Carlos Araújo

    2015-11-01

    The amino acids taurine and alanine play a role in several physiological processes, including behavior and the electrical activity of the brain. In this study, we investigated the effect of treatment with taurine or alanine on anxiety-like behavior and the excitability-dependent phenomenon known as cortical spreading depression (CSD), using rats suckled in litters with 9 and 15 pups (groups L9 and L15). From postnatal days 7 to 27, the animals received per gavage 300 mg/kg/day of taurine or alanine or both. At 28 days, we tested the animals in the elevated plus maze, and at 33-35 days, we recorded CSD and analyzed its velocity of propagation, amplitude, and duration. Compared with water-treated controls, the L9 groups treated with taurine or alanine displayed anxiolytic behavior (higher number of entries in the open arms; p < 0.05), and reduced CSD velocity (p < 0.001). The effect of both amino acids on CSD was also found in the L15 groups and in five additional L9 groups (naïve, water, taurine, alanine, or both) treated at adulthood (90-110 days). The L15 condition resulted in smaller durations and higher CSD velocities compared with the L9 condition. Besides reinforcing previous evidence of behavioral modulation by taurine and alanine, our data are the first confirmation that treatment with these amino acids decelerates CSD regardless of lactation conditions (normal versus unfavorable lactation) or age at amino acid administration (young versus adult). The results suggest a modulating role for both amino acids on anxiety behavior and neuronal electrical activity.

  2. Identification of a diazinon-metabolizing glutathione S-transferase in the silkworm, Bombyx mori

    PubMed Central

    Yamamoto, Kohji; Yamada, Naotaka

    2016-01-01

    The glutathione S-transferase superfamily play key roles in the metabolism of numerous xenobiotics. We report herein the identification and characterization of a novel glutathione S-transferase in the silkworm, Bombyx mori. The enzyme (bmGSTu2) conjugates glutathione to 1-chloro-2,4-dinitrobenzene, as well as metabolizing diazinon, one of the organophosphate insecticides. Quantitative reverse transcription–polymerase chain reaction analysis of transcripts demonstrated that bmGSTu2 expression was induced 1.7-fold in a resistant strain of B. mori. Mutagenesis of putative amino acid residues in the glutathione-binding site revealed that Ile54, Glu66, Ser67, and Asn68 are crucial for enzymatic function. These results provide insights into the catalysis of glutathione conjugation in silkworm by bmGSTu2 and into the detoxification of organophosphate insecticides. PMID:27440377

  3. Rubber transferase in guayule plants. [Parthenium argentatum

    SciTech Connect

    Rosenfield, C.L.; Foster, M.A.; Benedict, C.R.

    1986-04-01

    Rubber transferase catalyzes the transfer of cis-1,4-polyprenyl-PP to isopentenyl-PP (IPP) with the elimination of PP/sub i/. Rubber transferase activity in guayule (Parthenium argentatum Gray) stems was localized in the lipid fraction of the homogenate following centrifugation in buffer and 0.4M Mannitol. Washed rubber particles were obtained by the chromatography of the lipid fraction on Ultrogel columns with an exclusion limit of 750,000 daltons by the procedure of B.G. Audley (private communication). The rubber particles catalyzed the incorporation of /sup 14/C-IPP into cis-polyisoprene. The radioactive cis-polyisoprene was identified by ozonolysis and chromatography of the resulting /sup 14/C-levulinic acid. The synthesis of cis-polyisoprene in the rubber particles required Mg/sup 2 +/ and IPP and was stimulated 2-fold with the addition of DMAPP. Rubber synthesis in guayule plants growing in the Permian Basin of West Texas does not occur during summer months but is induced by the cold night temperatures of the fall and winter. From August to December individual plants (which were transplanted in May) accumulated from 66mg to 11,800mg or rubber. During this period there was a 4-fold increase in rubber transferase activity in stem homogenates induced by the low temperatures.

  4. International society of sports nutrition position stand: Beta-Alanine.

    PubMed

    Trexler, Eric T; Smith-Ryan, Abbie E; Stout, Jeffrey R; Hoffman, Jay R; Wilborn, Colin D; Sale, Craig; Kreider, Richard B; Jäger, Ralf; Earnest, Conrad P; Bannock, Laurent; Campbell, Bill; Kalman, Douglas; Ziegenfuss, Tim N; Antonio, Jose

    2015-01-01

    The International Society of Sports Nutrition (ISSN) provides an objective and critical review of the mechanisms and use of beta-alanine supplementation. Based on the current available literature, the conclusions of the ISSN are as follows: 1) Four weeks of beta-alanine supplementation (4-6 g daily) significantly augments muscle carnosine concentrations, thereby acting as an intracellular pH buffer; 2) Beta-alanine supplementation currently appears to be safe in healthy populations at recommended doses; 3) The only reported side effect is paraesthesia (tingling), but studies indicate this can be attenuated by using divided lower doses (1.6 g) or using a sustained-release formula; 4) Daily supplementation with 4 to 6 g of beta-alanine for at least 2 to 4 weeks has been shown to improve exercise performance, with more pronounced effects in open end-point tasks/time trials lasting 1 to 4 min in duration; 5) Beta-alanine attenuates neuromuscular fatigue, particularly in older subjects, and preliminary evidence indicates that beta-alanine may improve tactical performance; 6) Combining beta-alanine with other single or multi-ingredient supplements may be advantageous when supplementation of beta-alanine is high enough (4-6 g daily) and long enough (minimum 4 weeks); 7) More research is needed to determine the effects of beta-alanine on strength, endurance performance beyond 25 min in duration, and other health-related benefits associated with carnosine.

  5. Cloning, expression and properties of porcine trachea UDP-galnac: polypeptide N-acetylgalactosaminyl transferase.

    PubMed

    Sangadala, Sreedhara; Swain, Ja Baris; McNear, Adrian; Mendicino, Joseph

    2004-11-01

    A UDP-GalNAc:polypeptide N-acetyl-galactosaminyl transferase which catalyses the transfer of GalNAc from UDP-GalNAc to serine and threonine residues in mucin polypeptide chains was purified to homogeneity from swine trachea epithelium (Mendicino J, Sangadala S: Mol Cell Biochem 185: 135-145, 1998). Peptides obtained by proteolysis of the purified enzyme were isolated, sequenced and used to prepare degenerate oligonucleotide primers. Amplified segments of a gene encoding GalNAc transferase were synthesised using the primers and a swine trachea epithelial cDNA library. Selected cDNA fragments were then used to screen the cDNA library, and a clone containing an open reading frame encoding 559 amino acids was isolated. The predicted amino acid sequence contains type II transmembrane region, three potential N-glycosylation sites as well as all of the isolated peptide sequences. The nucleotide sequence and predicted primary protein structure of the transferase were very similar to those of type T-1 GalNAc transferases. The isolated clone was transiently expressed in COS 7 cells and the recombinant enzyme, which contained an N-terminal hexa-histidine tag, was purified to homogeneity and its enzymatic properties were examined. The Vmax of the recombinant enzyme, 2.08 micromol/(min mg), was nearly the same as the native enzyme, 2.12 micromol/(min mg), when assayed with partially deglycosylated mucins as glycosyl acceptors. Both enzymes showed much higher activities when assayed with peptides prepared by limited acid hydrolysis of incompletely deglycosylated Cowper's gland, swine, and human respiratory mucins and tryptic peptides isolated from deglycosylated mucin polypeptide chains. However, as noted earlier (Mendicino J, Sangadala S: Mol Cell Biochem 185: 135-145, 1998), these enzymes showed very little activity with completely deglycosylated mucin polypeptide chains. When completely deglycosylated polypeptide chains were partially glycosylated by incubation with microsome

  6. Beta-alanine and beta-aminoisobutyric acid levels in two siblings with dihydropyrimidinase deficiency.

    PubMed

    van Kuilenburg, A B P; Stroomer, A E M; Bosch, A M; Duran, M

    2008-06-01

    Dihydropyrimidinase (DHP) deficiency is an inborn error of the pyrimidine degradation pathway, affecting the hydrolytic ring opening of the dihydropyrimidines. In two siblings with a complete DHP deficiency and a variable clinical presentation, a normal concentration of beta-alanine and strongly decreased levels of beta-aminoisobutyric acid were observed in plasma, urine and CSF. No major differences were observed for the concentrations of the beta-amino acids in plasma and urine between the symptomatic and asymptomatic sibling. Thus, the relevance of the shortage of beta-aminoisobutyric acid for the onset of a clinical phenotype in patients with DHP deficiency remains to be established.

  7. Post-translational Introduction of D-Alanine into Ribosomally Synthesized Peptides by the Dehydroalanine Reductase NpnJ.

    PubMed

    Yang, Xiao; van der Donk, Wilfred A

    2015-10-07

    Ribosomally synthesized peptides are generally limited to L-amino acid building blocks. Given the advantageous properties of peptides containing D-amino acids such as stabilization of certain turns and against proteolytic degradation, methods to introduce D-stereocenters are valuable. Here we report the first in vitro reconstitution and characterization of a dehydrogenase that carries out the asymmetric reduction of dehydroalanine. NpnJA reduces dehydroalanine to D-Ala using NAPDH as cosubstrate. The enzyme displays high substrate tolerance allowing introduction of D-Ala into a range of non-native substrates. In addition to the in vitro reactions, we describe five examples of using Escherichia coli as biosynthetic host for D-alanine introduction into ribosomal peptides. A deuterium-label-based coupled-enzyme assay was used to rapidly determine the stereochemistry of the newly installed alanine.

  8. A search for endogenous amino acids in martian meteorite ALH84001

    NASA Technical Reports Server (NTRS)

    Bada, J. L.; Glavin, D. P.; McDonald, G. D.; Becker, L.

    1998-01-01

    Trace amounts of glycine, serine, and alanine were detected in the carbonate component of the martian meteorite ALH84001 by high-performance liquid chromatography. The detected amino acids were not uniformly distributed in the carbonate component and ranged in concentration from 0.1 to 7 parts per million. Although the detected alanine consists primarily of the L enantiomer, low concentrations (<0.1 parts per million) of endogenous D-alanine may be present in the ALH84001 carbonates. The amino acids present in this sample of ALH84001 appear to be terrestrial in origin and similar to those in Allan Hills ice, although the possibility cannot be ruled out that minute amounts of some amino acids such as D-alanine are preserved in the meteorite.

  9. An alternative mechanism for the catalysis of peptide bond formation by L/F transferase: substrate binding and orientation.

    PubMed

    Fung, Angela W; Ebhardt, H Alexander; Abeysundara, Heshani; Moore, Jack; Xu, Zhizhong; Fahlman, Richard P

    2011-06-17

    Eubacterial leucyl/phenylalanyl tRNA protein transferase (L/F transferase) catalyzes the transfer of a leucine or a phenylalanine from an aminoacyl-tRNA to the N-terminus of a protein substrate. This N-terminal addition of an amino acid is analogous to that of peptide synthesis by ribosomes. A previously proposed catalytic mechanism for Escherichia coli L/F transferase identified the conserved aspartate 186 (D186) and glutamine 188 (Q188) as key catalytic residues. We have reassessed the role of D186 and Q188 by investigating the enzymatic reactions and kinetics of enzymes possessing mutations to these active-site residues. Additionally three other amino acids proposed to be involved in aminoacyl-tRNA substrate binding are investigated for comparison. By quantitatively measuring product formation using a quantitative matrix-assisted laser desorption/ionization time-of-flight mass spectrometry-based assay, our results clearly demonstrate that, despite significant reduction in enzymatic activity as a result of different point mutations introduced into the active site of L/F transferase, the formation of product is still observed upon extended incubations. Our kinetic data and existing X-ray crystal structures result in a proposal that the critical roles of D186 and Q188, like the other amino acids in the active site, are for substrate binding and orientation and do not directly participate in the chemistry of peptide bond formation. Overall, we propose that L/F transferase does not directly participate in the chemistry of peptide bond formation but catalyzes the reaction by binding and orientating the substrates for reaction in an analogous mechanism that has been described for ribosomes.

  10. Β-alanine and l-histidine transport across the inner blood-retinal barrier: potential involvement in L-carnosine supply.

    PubMed

    Usui, Takuya; Kubo, Yoshiyuki; Akanuma, Shin-Ichi; Hosoya, Ken-Ichi

    2013-08-01

    The supply of L-carnosine, a bioactive dipeptide of β-alanine and l-histidine, to the retina across the blood-retinal barrier (BRB) was studied. The in vivo and in vitro studies revealed low uptake activities for [(3)H]Gly-Sar, a representative dipeptide, suggesting that l-carnosine transport plays only a minor role at the BRB. The in vivo study using rats showed approximately 18- and 23-fold greater retinal uptake indexes (RUI) for [(3)H]β-alanine and [(3)H]l-histidine compared with that of a paracellular marker, respectively. The RUI of [(3)H]β-alanine was taurine- and γ-aminobutyric acid-sensitive, and the in vitro uptake by TR-iBRB2 cells showed time- concentration- and temperature-dependent [(3)H]β-alanine uptake, suggesting that a carrier-mediated process was involved in β-alanine transport across the inner BRB. [(3)H]β-Alanine uptake was inhibited by taurine and β-guanidinopropionic acid, suggesting that taurine transporter (TAUT/SLC6A6) is responsible for the influx transport of β-alanine across the inner BRB. Regarding l-histidine, the l-leucine-sensitive RUI of [(3)H]l-histidine was identified, and the in vitro [(3)H]l-histidine uptake by TR-iBRB2 cells suggested that a carrier-mediated process was involved in l-histidine transport across the inner BRB. The inhibition profile suggested that L-type amino acid transporter (LAT1/SLC7A5) is responsible for the influx transport of l-histidine across the inner BRB. These results show that the influx transports of β-alanine and l-histidine across the inner BRB is carried out by TAUT and LAT1, respectively, suggesting that the retinal l-carnosine is supplied by enzymatic synthesis from two kinds of amino acids transported across the inner BRB.

  11. Crystal structure of the Apo form of D-Alanine:D-Alanine ligase (DDl) from Streptococcus mutans.

    PubMed

    Lu, Yongzhi; Xu, Hongyan; Zhao, Xiaojun

    2010-08-01

    D-Alanine:D-Alanine ligase (DDl) catalyzes the formation of D-Alanine:D-Alanine dipeptide and is an essential enzyme in bacterial cell wall biosynthesis.. This enzyme does not have a human ortholog, making it an attractive target for developing new antibiotic drugs. We determined the crystal structure at 2.23 A resolution of DDl from Streptococcus mutans (SmDDl), the principal aetiological agent of human dental caries. This structure reveals that SmDDl is a dimer and has a disordered omega-loop region.

  12. Regulation of myocardial amino acid balance in the conscious dog.

    PubMed Central

    Schwartz, R G; Barrett, E J; Francis, C K; Jacob, R; Zaret, B L

    1985-01-01

    The effects in vivo of physiologic increases in insulin and amino acids on myocardial amino acid balance were evaluated in conscious dogs. Arterial and coronary sinus concentrations of amino acids and coronary blood flow were measured during a 30-min basal and a 100-min experimental period employing three protocols: euglycemic insulin clamp (plasma insulin equaled 70 +/- 11 microU/ml, n = 6); euglycemic insulin clamp during amino acid infusion (plasma insulin equaled 89 +/- 12 microU/ml, n = 6); and suppression of insulin with somatostatin during amino acid infusion (plasma insulin equaled 15 +/- 4 microU/ml, n = 6). Basally, only leucine and isoleucine were removed significantly by myocardium (net branched chain amino acid [BCAA] uptake equaled 0.5 +/- 0.2 mumol/min), while glycine, alanine, and glutamine were released. Glutamine demonstrated the highest net myocardial production (1.6 +/- 0.2 mumol/min). No net exchange was seen for valine, phenylalanine, tyrosine, cysteine, methionine, glutamate, asparagine, serine, threonine, taurine, and aspartate. In group I, hyperinsulinemia caused a decline of all plasma amino acids except alanine; alanine balance switched from release to an uptake of 0.6 +/- 0.4 mumol/min (P less than 0.05), while the myocardial balance of other amino acids was unchanged. In group II, amino acid concentrations rose, and were accompanied by a marked rise in myocardial BCAA uptake (0.4 +/- 0.1-2.6 +/- 0.3 mumol/min, P less than 0.001). Uptake of alanine was again stimulated (0.9 +/- 0.3 mumol/min, P less than 0.01), while glutamine production was unchanged (1.3 +/- 0.4 vs. 1.6 +/- 0.3 mumol/min). In group III, there was a 4-5-fold increase in the plasma concentration of the infused amino acids, accompanied by marked stimulation in uptake of only BCAA (6.8 +/- 0.7 mumol/min). Myocardial glutamine production was unchanged (1.9 +/- 0.4-1.3 +/- 0.7 mumol/min). Within the three experimental groups there were highly significant linear correlations

  13. VUV photodynamics and chiral asymmetry in the photoionization of gas phase alanine enantiomers.

    PubMed

    Tia, Maurice; Cunha de Miranda, Barbara; Daly, Steven; Gaie-Levrel, François; Garcia, Gustavo A; Nahon, Laurent; Powis, Ivan

    2014-04-17

    The valence shell photoionization of the simplest proteinaceous chiral amino acid, alanine, is investigated over the vacuum ultraviolet region from its ionization threshold up to 18 eV. Tunable and variable polarization synchrotron radiation was coupled to a double imaging photoelectron/photoion coincidence (i(2)PEPICO) spectrometer to produce mass-selected threshold photoelectron spectra and derive the state-selected fragmentation channels. The photoelectron circular dichroism (PECD), an orbital-sensitive, conformer-dependent chiroptical effect, was also recorded at various photon energies and compared to continuum multiple scattering calculations. Two complementary vaporization methods-aerosol thermodesorption and a resistively heated sample oven coupled to an adiabatic expansion-were applied to promote pure enantiomers of alanine into the gas phase, yielding neutral alanine with different internal energy distributions. A comparison of the photoelectron spectroscopy, fragmentation, and dichroism measured for each of the vaporization methods was rationalized in terms of internal energy and conformer populations and supported by theoretical calculations. The analytical potential of the so-called PECD-PICO detection technique-where the electron spectroscopy and circular dichroism can be obtained as a function of mass and ion translational energy-is underlined and applied to characterize the origin of the various species found in the experimental mass spectra. Finally, the PECD findings are discussed within an astrochemical context, and possible implications regarding the origin of biomolecular asymmetry are identified.

  14. Beta-alanine and taurine as endogenous agonists at glycine receptors in rat hippocampus in vitro.

    PubMed

    Mori, Masahiro; Gähwiler, Beat H; Gerber, Urs

    2002-02-15

    Electrophysiological and pharmacological properties of glycine receptors were characterized in hippocampal organotypic slice cultures. In the presence of ionotropic glutamate and GABA(B) receptor antagonists, pressure-application of glycine onto CA3 pyramidal cells induced a current associated with increased chloride conductance, which was inhibited by strychnine. Similar chloride currents could also be induced with beta-alanine or taurine. Whole-cell glycine responses were significantly greater in CA3 pyramidal cells than in CA1 pyramidal cells and dentate granule cells, while responses to GABA were similar among these three cell types. Although these results demonstrate the presence of functional glycine receptors in the hippocampus, no evidence for their activation during synaptic stimulation was found. Gabazine, a selective GABA(A) receptor antagonist, totally blocked evoked IPSCs in CA3 pyramidal cells. Glycine receptor activation is not dependent on transporter-controlled levels of extracellular glycine, as no chloride current was observed in response to sarcosine, an inhibitor of glycine transporters. In contrast, application of guanidinoethanesulfonic acid, an uptake inhibitor of beta-alanine and taurine, induced strychnine-sensitive chloride current in the presence of gabazine. These data indicate that modulation of transporters for the endogenous amino acids, beta-alanine and taurine, can regulate tonic activation of glycine receptors, which may function in maintenance of inhibitory tone in the hippocampus.

  15. Accumulated analyses of amino acid precursors in returned lunar samples

    NASA Technical Reports Server (NTRS)

    Fox, S. W.; Harada, K.; Hare, P. E.

    1973-01-01

    Six amino acids (glycine, alanine, aspartic acid, glutamic acid, serine, and threonine) obtained by hydrolysis of extracts have been quantitatively determined in ten collections of fines from five Apollo missions. Although the amounts found, 7-45 ng/g, are small, the lunar amino acid/carbon ratios are comparable to those of the carbonaceous chondrites, Murchison and Murray, as analyzed by the same procedures. Since both the ratios of amino acid to carbon, and the four or five most common types of proteinous amino acid found, are comparable for the two extraterrestrial sources despite different cosmophysical histories of the moon and meteorites, common cosmochemical processes are suggested.

  16. Alanine repeats influence protein localization in splicing speckles and paraspeckles.

    PubMed

    Chang, Shuo-Hsiu; Chang, Wei-Lun; Lu, Chia-Chen; Tarn, Woan-Yuh

    2014-12-16

    Mammalian splicing regulatory protein RNA-binding motif protein 4 (RBM4) has an alanine repeat-containing C-terminal domain (CAD) that confers both nuclear- and splicing speckle-targeting activities. Alanine-repeat expansion has pathological potential. Here we show that the alanine-repeat tracts influence the subnuclear targeting properties of the RBM4 CAD in cultured human cells. Notably, truncation of the alanine tracts redistributed a portion of RBM4 to paraspeckles. The alanine-deficient CAD was sufficient for paraspeckle targeting. On the other hand, alanine-repeat expansion reduced the mobility of RBM4 and impaired its splicing activity. We further took advantage of the putative coactivator activator (CoAA)-RBM4 conjoined splicing factor, CoAZ, to investigate the function of the CAD in subnuclear targeting. Transiently expressed CoAZ formed discrete nuclear foci that emerged and subsequently separated-fully or partially-from paraspeckles. Alanine-repeat expansion appeared to prevent CoAZ separation from paraspeckles, resulting in their complete colocalization. CoAZ foci were dynamic but, unlike paraspeckles, were resistant to RNase treatment. Our results indicate that the alanine-rich CAD, in conjunction with its conjoined RNA-binding domain(s), differentially influences the subnuclear localization and biogenesis of RBM4 and CoAZ.

  17. Molecular characterization of a glutathione transferase from Pinus tabulaeformis (Pinaceae).

    PubMed

    Zeng, Qing-Yin; Lu, Hai; Wang, Xiao-Ru

    2005-05-01

    Glutathione transferases (GSTs) play important roles in stress tolerance and detoxification metabolism in plants. To date, studies on GSTs in higher plants have focused largely on agricultural plants. In contrast, there is virtually no information on the molecular characteristics of GSTs in gymnosperms. The present study reports for the first time the cloning, expression and characteristics of a GST gene (PtGSTU1) from a pine, Pinus tabulaeformis, which is widely distributed from northern to central China covering cold temperate and drought regions. The PtGSTU1 gene encodes a protein of 228 amino acid residues with a calculated molecular mass of 26.37 kDa. Reverse transcription PCR revealed that PtGSTU1 was expressed in different tissues, both above and below ground, of P. tabulaeformis. The over-expressed recombinant PtGSTU1 showed high activity towards the substrates 1-chloro-2,4-dinitrobenzene (CDNB) and 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole (NBD-Cl). Kinetic analysis with respect to CDNB as substrate revealed a Km of 0.47 mM and Vmax of 169.1 micromol/min per mg of protein. The recombinant PtGSTU1 retained more than 60% of its maximum enzymatic activity from 15 degrees C to 45 degrees C with a broad optimum Tm range of 25 degrees C - 35 degrees C. The enzyme had a maximum activity at approximately pH 8.5 - 9.0. Site-directed mutagenesis revealed that Ser13 in the N-terminal domain is a critical catalytic residue, responsible for stabilisation of the thiolate anion of enzyme-bound glutathione. Based on comparative analyses of its amino acid sequence, phylogeny and predicted three-dimensional structure, the PtGSTU1 should be classified as a tau class GST.

  18. The structure of alanine racemase from Acinetobacter baumannii.

    PubMed

    Davis, Emily; Scaletti-Hutchinson, Emma; Opel-Reading, Helen; Nakatani, Yoshio; Krause, Kurt L

    2014-09-01

    Acinetobacter baumannii is an opportunistic Gram-negative bacterium which is a common cause of hospital-acquired infections. Numerous antibiotic-resistant strains exist, emphasizing the need for the development of new antimicrobials. Alanine racemase (Alr) is a pyridoxal 5'-phosphate dependent enzyme that is responsible for racemization between enantiomers of alanine. As D-alanine is an essential component of the bacterial cell wall, its inhibition is lethal to prokaryotes, making it an excellent antibiotic drug target. The crystal structure of A. baumannii alanine racemase (AlrAba) from the highly antibiotic-resistant NCTC13302 strain has been solved to 1.9 Å resolution. Comparison of AlrAba with alanine racemases from closely related bacteria demonstrates a conserved overall fold. The substrate entryway and active site of the enzymes were shown to be highly conserved. The structure of AlrAba will provide the template required for future structure-based drug-design studies.

  19. Amino Acid Racemization and the Preservation of Ancient DNA

    NASA Technical Reports Server (NTRS)

    Poinar, Hendrik N.; Hoss, Matthias

    1996-01-01

    The extent of racemization of aspartic acid, alanine, and leucine provides criteria for assessing whether ancient tissue samples contain endogenous DNA. In samples in which the D/L ratio of aspartic acid exceeds 0.08, ancient DNA sequences could not be retrieved. Paleontological finds from which DNA sequences purportedly millions of years old have been reported show extensive racemization, and the amino acids present are mainly contaminates. An exception is the amino acids in some insects preserved in amber.

  20. Characterization of a Thermostable d-Stereospecific Alanine Amidase from Brevibacillus borstelensis BCS-1

    PubMed Central

    Baek, Dae Heoun; Kwon, Seok-Joon; Hong, Seung-Pyo; Kwak, Mi-Sun; Lee, Mi-Hwa; Song, Jae Jun; Lee, Seung-Goo; Yoon, Ki-Hong; Sung, Moon-Hee

    2003-01-01

    A gene encoding a new thermostable d-stereospecific alanine amidase from the thermophile Brevibacillus borstelensis BCS-1 was cloned and sequenced. The molecular mass of the purified enzyme was estimated to be 199 kDa after gel filtration chromatography and about 30 kDa on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, indicating that the enzyme could be composed of a hexamer with identical subunits. The purified enzyme exhibited strong amidase activity towards d-amino acid-containing aromatic, aliphatic, and branched amino acid amides yet exhibited no enzyme activity towards l-amino acid amides, d-amino acid-containing peptides, and NH2-terminally protected amino acid amides. The optimum temperature and pH for the enzyme activity were 85°C and 9.0, respectively. The enzyme remained stable within a broad pH range from 7.0 to 10.0. The enzyme was inhibited by dithiothreitol, 2-mercaptoethanol, and EDTA yet was strongly activated by Co2+ and Mn2+. The kcat/Km for d-alaninamide was measured as 544.4 ± 5.5 mM−1 min−1 at 50°C with 1 mM Co2+. PMID:12571020

  1. Controlled radical polymerization of an acrylamide containing L-alanine moiety via ATRP.

    PubMed

    Rafiee, Zahra

    2016-02-01

    Homopolymerization of an optically active acrylamide having an amino acid moiety in the side chain, N-acryloyl-L-alanine (AAla) was carried out via atom transfer radical polymerization (ATRP) at room temperature using 2-hydroxyethyl-2'-methyl-2'-bromopropionate (HMB) or sodium-4-(bromomethyl)benzoate (SBB) as initiator in pure water, methanol/water mixture and pure methanol solvents. The polymerization reaction resulted in the optically active biocompatible amino acid-based homopolymer in good yield with narrow molecular weight distribution. The number average molecular weight increased with conversion and polydispersity was low. The structure and molecular weight of synthesized polymer were characterized by (1)H NMR, FT-IR spectroscopic techniques and size-exclusion chromatography.

  2. The influence of various cations on the catalytic properties of clays. [polymerization of alanine adenylate

    NASA Technical Reports Server (NTRS)

    Paecht-Horowitz, M.

    1978-01-01

    The polymerization of alanine adenylate in the presence of the sodium form of various clays was studied, and hectorite was found to cause more polymerization than nontronite and montmorillonite (in that order) although the differences were not great. The effect on polymerization of presaturating montmorillonite with different cations was determined. Hectorite, with increased basicity of the interspatial planes, allows polymerization of lysine, which montmorillonite does not. The general trend is that, for the same amino acid, higher degrees of polymerization are obtained when the cation in the octahedral lattice of the clay is divalent rather than trivalent. With the exchangeable cations the order is reversed, for a reason that is explained. The main role of clays in the polymerization mechanism of amino acids is concentration and neutralization of charges.

  3. Comparison of EPR response of alanine and Gd₂O₃-alanine dosimeters exposed to TRIGA Mainz reactor.

    PubMed

    Marrale, M; Schmitz, T; Gallo, S; Hampel, G; Longo, A; Panzeca, S; Tranchina, L

    2015-12-01

    In this work we report some preliminary results regarding the analysis of electron paramagnetic resonance (EPR) response of alanine pellets and alanine pellets added with gadolinium used for dosimetry at the TRIGA research reactor in Mainz, Germany. Two set-ups were evaluated: irradiation inside PMMA phantom and irradiation inside boric acid phantom. We observed that the presence of Gd2O3 inside alanine pellets increases the EPR signal by a factor of 3.45 and 1.24 in case of PMMA and boric acid phantoms, respectively. We can conclude that in the case of neutron beam with a predominant thermal neutron component the addition of gadolinium oxide can significantly improve neutron sensitivity of alanine pellets. Monte Carlo (MC) simulations of both response of alanine and Gd-added alanine pellets with FLUKA code were performed and a good agreement was achieved for pure alanine dosimeters. For Gd2O3-alanine deviations between MC simulations and experimental data were observed and discussed.

  4. Size does matter: 18 amino acids at the N-terminal tip of an amino acid transporter in Leishmania determine substrate specificity

    PubMed Central

    Schlisselberg, Doreen; Mazarib, Eldar; Inbar, Ehud; Rentsch, Doris; Myler, Peter J.; Zilberstein, Dan

    2015-01-01

    Long N-terminal tails of amino acid transporters are known to act as sensors of the internal pool of amino acids and as positive regulators of substrate flux rate. In this study we establish that N-termini of amino acid transporters can also determine substrate specificity. We show that due to alternative trans splicing, the human pathogen Leishmania naturally expresses two variants of the proline/alanine transporter, one 18 amino acid shorter than the other. We demonstrate that the longer variant (LdAAP24) translocates both proline and alanine, whereas the shorter variant (∆18LdAAP24) translocates just proline. Remarkably, co-expressing the hydrophilic N-terminal peptide of the long variant with ∆18LdAAP24 was found to recover alanine transport. This restoration of alanine transport could be mediated by a truncated N-terminal tail, though truncations exceeding half of the tail length were no longer functional. Taken together, the data indicate that the first 18 amino acids of the negatively charged N-terminal LdAAP24 tail are required for alanine transport and may facilitate the electrostatic interactions of the entire negatively charged N-terminal tail with the positively charged internal loops in the transmembrane domain, as this mechanism has been shown to underlie regulation of substrate flux rate for other transporters. PMID:26549185

  5. Active Sites of Spinoxin, a Potassium Channel Scorpion Toxin, Elucidated by Systematic Alanine Scanning.

    PubMed

    Peigneur, Steve; Yamaguchi, Yoko; Kawano, Chihiro; Nose, Takeru; Nirthanan, Selvanayagam; Gopalakrishnakone, Ponnampalam; Tytgat, Jan; Sato, Kazuki

    2016-05-31

    Peptide toxins from scorpion venoms constitute the largest group of toxins that target the voltage-gated potassium channel (Kv). Spinoxin (SPX) isolated from the venom of scorpion Heterometrus spinifer is a 34-residue peptide neurotoxin cross-linked by four disulfide bridges. SPX is a potent inhibitor of Kv1.3 potassium channels (IC50 = 63 nM), which are considered to be valid molecular targets in the diagnostics and therapy of various autoimmune disorders and cancers. Here we synthesized 25 analogues of SPX and analyzed the role of each amino acid in SPX using alanine scanning to study its structure-function relationships. All synthetic analogues showed similar disulfide bond pairings and secondary structures as native SPX. Alanine replacements at Lys(23), Asn(26), and Lys(30) resulted in loss of activity against Kv1.3 potassium channels, whereas replacements at Arg(7), Met(14), Lys(27), and Tyr(32) also largely reduced inhibitory activity. These results suggest that the side chains of these amino acids in SPX play an important role in its interaction with Kv1.3 channels. In particular, Lys(23) appears to be a key residue that underpins Kv1.3 channel inhibition. Of these seven amino acid residues, four are basic amino acids, suggesting that the positive electrostatic potential on the surface of SPX is likely required for high affinity interaction with Kv1.3 channels. This study provides insight into the structure-function relationships of SPX with implications for the rational design of new lead compounds targeting potassium channels with high potency.

  6. The Amino Acid Composition of the Sutter's Mill Carbonaceous Chondrite

    NASA Technical Reports Server (NTRS)

    Glavin, D. P.; Burton, A. S.; Elsila, J. E.; Dworkin, J. P.; Yin, Q. Z.; Cooper, G.; Jenniskens, P.

    2012-01-01

    In contrast to the Murchison meteorite which had a complex distribution of amino acids with a total C2 to Cs amino acid abundance of approx.14,000 parts-per-billion (ppb) [2], the Sutters Mill meteorite was found to be highly depleted in amino acids. Much lower abundances (approx.30 to 180 ppb) of glycine, beta-alanine, L-alanine and L-serine were detected in SM2 above procedural blank levels indicating that this meteorite sample experienced only minimal terrestrial amino acid contamination after its fall to Earth. Carbon isotope measurements will be necessary to establish the origin of glycine and beta-alanine in SM2. Other non-protein amino acids that are rare on Earth, yet commonly found in other CM meteorites such as aaminoisobutyric acid (alpha-AIB) and isovaline, were not identified in SM2. However, traces of beta-AIB (approx.1 ppb) were detected in SM2 and could be" extraterrestrial in origin. The low abundances of amino acids in the Sutter's Mill meteorite is consistent with mineralogical evidence that at least some parts of the Sutter's Mill meteorite parent body experienced extensive aqueous and/or thermal alteration.

  7. Plasmid-Encoded asp Operon Confers a Proton Motive Metabolic Cycle Catalyzed by an Aspartate-Alanine Exchange Reaction

    PubMed Central

    Abe, Keietsu; Ohnishi, Fumito; Yagi, Kyoko; Nakajima, Tasuku; Higuchi, Takeshi; Sano, Motoaki; Machida, Masayuki; Sarker, Rafiquel I.; Maloney, Peter C.

    2002-01-01

    Tetragenococcus halophila D10 catalyzes the decarboxylation of l-aspartate with nearly stoichiometric release of l-alanine and CO2. This trait is encoded on a 25-kb plasmid, pD1. We found in this plasmid a putative asp operon consisting of two genes, which we designated aspD and aspT, encoding an l-aspartate-β-decarboxylase (AspD) and an aspartate-alanine antiporter (AspT), respectively, and determined the nucleotide sequences. The sequence analysis revealed that the genes of the asp operon in pD1 were in the following order: promoter → aspD → aspT. The deduced amino acid sequence of AspD showed similarity to the sequences of two known l-aspartate-β-decarboxylases from Pseudomonas dacunhae and Alcaligenes faecalis. Hydropathy analyses suggested that the aspT gene product encodes a hydrophobic protein with multiple membrane-spanning regions. The operon was subcloned into the Escherichia coli expression vector pTrc99A, and the two genes were cotranscribed in the resulting plasmid, pTrcAsp. Expression of the asp operon in E. coli coincided with appearance of the capacity to catalyze the decarboxylation of aspartate to alanine. Histidine-tagged AspD (AspDHis) was also expressed in E. coli and purified from cell extracts. The purified AspDHis clearly exhibited activity of l-aspartate-β-decarboxylase. Recombinant AspT was solubilized from E. coli membranes and reconstituted in proteoliposomes. The reconstituted AspT catalyzed self-exchange of aspartate and electrogenic heterologous exchange of aspartate with alanine. Thus, the asp operon confers a proton motive metabolic cycle consisting of the electrogenic aspartate-alanine antiporter and the aspartate decarboxylase, which keeps intracellular levels of alanine, the countersubstrate for aspartate, high. PMID:12003930

  8. Plasmid-encoded asp operon confers a proton motive metabolic cycle catalyzed by an aspartate-alanine exchange reaction.

    PubMed

    Abe, Keietsu; Ohnishi, Fumito; Yagi, Kyoko; Nakajima, Tasuku; Higuchi, Takeshi; Sano, Motoaki; Machida, Masayuki; Sarker, Rafiquel I; Maloney, Peter C

    2002-06-01

    Tetragenococcus halophila D10 catalyzes the decarboxylation of L-aspartate with nearly stoichiometric release of L-alanine and CO(2). This trait is encoded on a 25-kb plasmid, pD1. We found in this plasmid a putative asp operon consisting of two genes, which we designated aspD and aspT, encoding an L-aspartate-beta-decarboxylase (AspD) and an aspartate-alanine antiporter (AspT), respectively, and determined the nucleotide sequences. The sequence analysis revealed that the genes of the asp operon in pD1 were in the following order: promoter --> aspD --> aspT. The deduced amino acid sequence of AspD showed similarity to the sequences of two known L-aspartate-beta-decarboxylases from Pseudomonas dacunhae and Alcaligenes faecalis. Hydropathy analyses suggested that the aspT gene product encodes a hydrophobic protein with multiple membrane-spanning regions. The operon was subcloned into the Escherichia coli expression vector pTrc99A, and the two genes were cotranscribed in the resulting plasmid, pTrcAsp. Expression of the asp operon in E. coli coincided with appearance of the capacity to catalyze the decarboxylation of aspartate to alanine. Histidine-tagged AspD (AspDHis) was also expressed in E. coli and purified from cell extracts. The purified AspDHis clearly exhibited activity of L-aspartate-beta-decarboxylase. Recombinant AspT was solubilized from E. coli membranes and reconstituted in proteoliposomes. The reconstituted AspT catalyzed self-exchange of aspartate and electrogenic heterologous exchange of aspartate with alanine. Thus, the asp operon confers a proton motive metabolic cycle consisting of the electrogenic aspartate-alanine antiporter and the aspartate decarboxylase, which keeps intracellular levels of alanine, the countersubstrate for aspartate, high.

  9. Euphorbia characias latex: micromorphology of rubber particles and rubber transferase activity.

    PubMed

    Spanò, Delia; Pintus, Francesca; Esposito, Francesca; Loche, Danilo; Floris, Giovanni; Medda, Rosaria

    2015-02-01

    We have recently characterized a natural rubber in the latex of Euphorbia characias. Following that study, we here investigated the rubber particles and rubber transferase in that Mediterranean shrub. Rubber particles, observed by scanning electron microscopy, are spherical in shape with diameter ranging from 0.02 to 1.2 μm. Washed rubber particles exhibit rubber transferase activity with a rate of radiolabeled [(14)C]IPP incorporation of 4.5 pmol min(-1)mg(-1). Denaturing electrophoresis profile of washed rubber particles reveals a single protein band of 37 kDa that is recognized in western blot analysis by antibodies raised against the synthetic peptide whose sequence, DVVIRTSGETRLSNF, is included in one of the five regions conserved among cis-prenyl chain elongation enzymes. The cDNA nucleotide sequence of E. characias rubber transferase (GenBank JX564541) and the deduced amino acid sequence appear to be highly homologous to the sequence of several plant cis-prenyltransferases.

  10. Alanine substitutions of noncysteine residues in the cysteine-stabilized αβ motif

    PubMed Central

    Yang, Ying-Fang; Cheng, Kuo-Chang; Tsai, Ping-Hsing; Liu, Chung-Cheng; Lee, Tian-Ren; Ping-Chiang Lyu

    2009-01-01

    The protein scaffold is a peptide framework with a high tolerance of residue modifications. The cysteine-stabilized αβ motif (CSαβ) consists of an α-helix and an antiparallel triple-stranded β-sheet connected by two disulfide bridges. Proteins containing this motif share low sequence identity but high structural similarity and has been suggested as a good scaffold for protein engineering. The Vigna radiate defensin 1 (VrD1), a plant defensin, serves here as a model protein to probe the amino acid tolerance of CSαβ motif. A systematic alanine substitution is performed on the VrD1. The key residues governing the inhibitory function and structure stability are monitored. Thirty-two of 46 residue positions of VrD1 are altered by site-directed mutagenesis techniques. The circular dichroism spectrum, intrinsic fluorescence spectrum, and chemical denaturation are used to analyze the conformation and structural stability of proteins. The secondary structures were highly tolerant to the amino acid substitutions; however, the protein stabilities were varied for each mutant. Many mutants, although they maintained their conformations, altered their inhibitory function significantly. In this study, we reported the first alanine scan on the plant defensin containing the CSαβ motif. The information is valuable to the scaffold with the CSαβ motif and protein engineering. PMID:19533758

  11. High-performance liquid chromatographic determination of beta-alanine, beta-aminoisobutyric acid and gamma-aminobutyric acid in tissue extracts and urine of normal and (aminooxy)acetate-treated rats.

    PubMed

    Abe, T; Kurozumi, Y; Yao, W B; Ubuka, T

    1998-08-07

    A method is described for the simultaneous determination of beta-alanine, beta-aminoisobutyric acid and gamma-aminobutyric acid in biological materials. Amino acids including these beta- and gamma-amino acids were derivatized with 4-dimethylaminoazobenzene-4'-sulfonyl (dabsyl) chloride and dabsyl amino acids formed were separated by reversed-phase high-performance liquid chromatography. Dabsyl derivatives of these beta- and gamma-amino acids were well separated from other dabsyl-amino acids. The method was applied to the determination of these beta- and gamma-amino acids in trichloroacetic acid extracts of various tissues and to the urine of normal rats and those injected with (aminooxy)acetate (AOA). AOA injection (15 mg per kg of body mass) produced remarkable increase in beta-alanine contents in liver, kidney and urine (10.2, 4.6 and 25.7 times, respectively).

  12. AMINO ACID COMPOSITION AND C-TERMINAL RESIDUES OF ALGAL BILIPROTEINS,

    DTIC Science & Technology

    R-phycoerythrin from Ceramium rubrum and C- phycocyanin from Nostoc nuscorum were obtained in purified form by fractional crystallization, followed by...as amino acids. Alanine was identified as the only C-terminal amino acid of R-phycoerythrin, each molecule of which contained about 12 terminal groups. Serine was identified as the only C-terminal group of C- phycocyanin . (Author)

  13. The Formation of Racemic Amino Acids by UV Photolysis of Interstellar Ice Analogs

    NASA Technical Reports Server (NTRS)

    Bernstein, Max P.; Dworkin, Jason P.; Sandford, Scott A.; Cooper, George; Allamandola, Louis J.; DeVincenzi, Donald (Technical Monitor)

    2001-01-01

    Small biologically relevant organic molecules including the amino acids glycine, alanine, and marine were formed in the laboratory by the UV (Ultraviolet) photolysis of realistic interstellar ice analogs, composed primarily of H2O, and including CH3OH, NH3, and HCN, under interstellar conditions. N-formyl glycine, cycloserine (4-amino-3-isoxazolidinone), and glycerol were detected before hydrolysis, and glycine, racemic alanine, racemic marine, glycerol, ethanolamine, and glyceric acid were found after hydrolysis. This suggests that some meteoritic amino acids (and other molecules) may be the direct result of interstellar ice photochemistry, expanding the current paradigm that they formed by reactions in liquid water on meteorite parent bodies.

  14. The influence of manufacture on the free D-amino acid content of Cheddar cheese.

    PubMed

    Csapó, J; Varga-Visi, E; Lóki, K; Albert, Cs

    2006-06-01

    The changes in the concentration and those of composition of alanine, aspartic acid and glutamic acid enantiomers were investigated during manufacture of Cheddar cheese. The amount of D-alanine increased continuously during ripening following the liberation of L-alanine originated from the proteolysis of milk proteins. There was slightly more D-aspartic and D-glutamic acid in the dry matter of curd after pressing than before pressurization. The D-amino acid content and the ratio of the D-enantiomers related to the total amount of free amino acids differed significantly among cheeses produced with different single-strain starters. The D-amino acid composition changed during manufacture, but the influence of the strain selection was not significant on the D-amino acid pattern.

  15. Taurine and beta-alanine act on both GABA and glycine receptors in Xenopus oocyte injected with mouse brain messenger RNA.

    PubMed

    Horikoshi, T; Asanuma, A; Yanagisawa, K; Anzai, K; Goto, S

    1988-09-01

    The responding pathway (process from agonist binding to channel opening) of taurine and beta-alanine was investigated in Xenopus oocytes injected with mouse brain poly(A)+ RNA. Responses to gamma-aminobutyric acid (GABA), glycine, taurine and beta-alanine were induced in oocytes injected with poly(A)+ RNA extracted from 3 regions, cerebrum, cerebellum and brainstem of the mouse brain. From comparison, responses to these 4 inhibitory amino acids in each regional poly(A)+ RNA-injected oocytes were categorized into at least 3 groups: (1) GABA, (2) glycine, and (3) taurine and beta-alanine. No cross-desensitization was observed between GABA response and glycine response, but taurine and beta-alanine responses cross-desensitized both the GABA and glycine responses. Taurine and beta-alanine responses were partially inhibited by the GABA antagonist, bicuculline, and also by the glycine antagonist, strychnine. The results suggest that the taurine or the beta-alanine response in the brain is caused through both the GABA receptor and the glycine receptor.

  16. Human glutathione S-transferases. Characterization of the anionic forms from lung and placenta.

    PubMed Central

    Dao, D D; Partridge, C A; Kurosky, A; Awasthi, Y C

    1984-01-01

    Anionic glutathione S-transferases were purified from human lung and placenta. Chemical and immunochemical characterization, including polyacrylamide-gel electrophoresis, gave strong evidence that the anionic lung and placental enzymes are chemically similar, if not identical, proteins. The electrophoretic mobilities of both proteins were identical in conventional alkaline gels as well as in gels containing sodium dodecyl sulphate. Gel filtration of the intact active enzyme established an Mr value of 45000; however, with sodium dodecyl sulphate/polyacrylamide-gel electrophoresis under dissociating conditions a subunit Mr of 22500 was obtained. Amino acid sequence analysis of the N-terminal region of the placental enzyme revealed a single polypeptide sequence identical with that of lung. Results obtained from immunoelectrophoresis, immunotitration, double immunodiffusion and rocket immunoelectrophoresis also indicated the anionic lung and placental enzymes to be closely similar. The chemical similarity of these two proteins was further supported by protein compositional analysis and fragment analysis after chemical hydrolysis. Immunochemical comparison of the anionic lung and placental enzymes with human liver glutathione S-transferases revealed cross-reactivity with the anionic omega enzyme, but no cross-reactivity was detectable with the cationic enzymes. Comparison of the N-terminal region of the human anionic enzyme with reported sequences of rat liver glutathione S-transferases gave strong evidence of chemical similarity, indicating that these enzymes are evolutionarily related. However, computer analysis of the 30-residue N-terminal sequence did not show any significant chemical similarity to any other reported protein sequence, pointing to the fact that the glutathione S-transferases represent a unique class of proteins. Images Fig. 2. Fig. 5. Fig. 6. Fig. 7. PMID:6466318

  17. On the abiotic formation of amino acids. I - HCN as a precursor of amino acids detected in extracts of lunar samples. II - Formation of HCN and amino acids from simulated mixtures of gases released from lunar samples

    NASA Technical Reports Server (NTRS)

    Yuasa, S.; Flory, D.; Basile, B.; Oro, J.

    1984-01-01

    Two studies on the abiotic formation of amino acids are presented. The first study demonstrates the role of hydrogen cyanide as a precursor of amino acids detected in extracts of lunar samples. The formation of several amino acids, including glycine, alanine, aspartic acid, and glutamic acid, under conditions similar to those used for the analysis of lunar samples is demonstrated. The second study investigates the formation of hydrogen cyanide as well as amino acids from lunar-sample gas mixtures under electrical discharge conditions. These results extend the possibility of synthesis of amino acids to planetary bodies with primordial atmospheres less reducing than a mixture of methane, ammonia, hydrogen and water.

  18. Amino acid uptake by liver of genetically obese Zucker rats.

    PubMed Central

    Ruiz, B; Felipe, A; Casado, J; Pastor-Anglada, M

    1991-01-01

    Alanine and glutamine uptake by the liver of 50-52-day-old genetically obese Zucker rats and their lean littermates has been studied. The net uptake in vivo of L-alanine is 2-fold higher in the obese animals. No significant change in L-glutamine net balance was found. We also studied the Na(+)-dependent uptake of L-alanine and L-glutamine into plasma-membrane vesicles isolated from either obese- or lean-rat livers. Vmax. values of both L-alanine and L-glutamine transport were 2-fold higher in those preparations from obese rats. No change in Km was observed. As suggested by inhibition studies, this seemed to be mediated by an enhancement of the activities of systems A, ASC and N. We conclude that the liver of the obese Zucker rat is extremely efficient in taking up neutral amino acids from the afferent blood, which results in an enhanced net uptake of L-alanine in vivo. The changes in transport activities at the plasma-membrane level might contribute to increase amino acid disposal by liver, probably for lipogenic purposes, as recently reported by Terrettaz & Jeanrenaud [Biochem. J. (1990) 270, 803-807]. PMID:1684102

  19. Glutamine and alanine-induced differential expression of intracellular IL-6, IL-8, and TNF-α in LPS-stimulated monocytes in human whole-blood.

    PubMed

    Raspé, C; Czeslick, E; Weimann, A; Schinke, C; Leimert, A; Kellner, P; Simm, A; Bucher, M; Sablotzki, A

    2013-04-01

    To investigate the effects of the commonly-used immunomodulators l-glutamine, l-alanine, and the combination of both l-alanyl-l-glutamine (Dipeptamin(®)) on intracellular expression of IL-6, IL-8, and TNF-α during endotoxemia, lipopolysaccharide (LPS)-stimulated human monocytes in a whole blood system were investigated by flow cytometry. Whole blood of twenty-seven healthy volunteers was stimulated with LPS and incubated with three different amino acid solutions (1. l-glutamine, 2. l-alanine, 3. l-alanyl-l-glutamine, each concentration 2 mM, 5 mM, incubation time 3 h). CD14(+) monocytes were phenotyped in whole-blood and intracellular expression of cytokines was assessed by flow cytometry. Our investigations showed for the first time in whole blood probes, imitating best physiologically present cellular interactions, that l-glutamine caused a dose-independent inhibitory effect on IL-6 and TNF-α production in human monocytes stimulated with LPS. However, l-alanine had contrary effects on IL-6 expression, significantly upregulating expression of IL-6 in LPS-treated monocytes. The impact of l-alanine on the expression of TNF-α was comparable with glutamine. Neither amino acid was able to affect IL-8 production in LPS-stimulated monocytes. The combination of both did not influence significantly IL-6 and IL-8 expression in monocytes during endotoxemia, however strongly reduced TNF-α production. For the regulation of TNF-α, l-glutamine, l-alanine and the combination of both show a congruent and exponentiated downregulating effect during endotoxemia, for the modulation of IL-6, l-glutamine and l-alanine featured opposite regulation leading to a canceling impact of each other when recombining both amino acids.

  20. Differential distribution of amino acids in plants.

    PubMed

    Kumar, Vinod; Sharma, Anket; Kaur, Ravdeep; Thukral, Ashwani Kumar; Bhardwaj, Renu; Ahmad, Parvaiz

    2017-05-01

    Plants are a rich source of amino acids and their individual abundance in plants is of great significance especially in terms of food. Therefore, it is of utmost necessity to create a database of the relative amino acid contents in plants as reported in literature. Since in most of the cases complete analysis of profiles of amino acids in plants was not reported, the units used and the methods applied and the plant parts used were different, amino acid contents were converted into relative units with respect to lysine for statistical analysis. The most abundant amino acids in plants are glutamic acid and aspartic acid. Pearson's correlation analysis among different amino acids showed that there were no negative correlations between the amino acids. Cluster analysis (CA) applied to relative amino acid contents of different families. Alismataceae, Cyperaceae, Capparaceae and Cactaceae families had close proximity with each other on the basis of their relative amino acid contents. First three components of principal component analysis (PCA) explained 79.5% of the total variance. Factor analysis (FA) explained four main underlying factors for amino acid analysis. Factor-1 accounted for 29.4% of the total variance and had maximum loadings on glycine, isoleucine, leucine, threonine and valine. Factor-2 explained 25.8% of the total variance and had maximum loadings on alanine, aspartic acid, serine and tyrosine. 14.2% of the total variance was explained by factor-3 and had maximum loadings on arginine and histidine. Factor-4 accounted 8.3% of the total variance and had maximum loading on the proline amino acid. The relative content of different amino acids presented in this paper is alanine (1.4), arginine (1.8), asparagine (0.7), aspartic acid (2.4), cysteine (0.5), glutamic acid (2.8), glutamine (0.6), glycine (1.0), histidine (0.5), isoleucine (0.9), leucine (1.7), lysine (1.0), methionine (0.4), phenylalanine (0.9), proline (1.1), serine (1.0), threonine (1

  1. Activity of the lactate-alanine shuttle is independent of glutamate-glutamine cycle activity in cerebellar neuronal-astrocytic cultures.

    PubMed

    Bak, Lasse K; Sickmann, Helle M; Schousboe, Arne; Waagepetersen, Helle S

    The glutamate-glutamine cycle describes the neuronal release of glutamate into the synaptic cleft, astrocytic uptake, and conversion into glutamine, followed by release for use as a neuronal glutamate precursor. This only explains the fate of the carbon atoms, however, and not that of the ammonia. Recently, a role for alanine has been proposed in transfer of ammonia between glutamatergic neurons and astrocytes, denoted the lactate-alanine shuttle (Waagepetersen et al. [ 2000] J. Neurochem. 75:471-479). The role of alanine in this context has been studied further using cerebellar neuronal cultures and corresponding neuronal-astrocytic cocultures. A superfusion paradigm was used to induce repetitively vesicular glutamate release by N-methyl-D-aspartate (NMDA) in the neurons, allowing the relative activity dependency of the lactate-alanine shuttle to be assessed. [(15)N]Alanine (0.2 mM), [2-(15)N]/[5-(15)N]glutamine (0.25 mM), and [(15)N]ammonia (0.3 mM) were used as precursors and cell extracts were analyzed by mass spectrometry. Labeling from [(15)N]alanine in glutamine, aspartate, and glutamate in cerebellar cocultures was independent of depolarization of the neurons. Employing glutamine with the amino group labeled ([2-(15)N]glutamine) as the precursor, an activity-dependent increase in the labeling of both glutamate and aspartate (but not alanine) was observed in the cerebellar neurons. When the amide group of glutamine was labeled ([5-(15)N]glutamine), no labeling could be detected in the analyzed metabolites. Altogether, the results of this study support the existence of the lactate-alanine shuttle and the associated glutamate-glutamine cycle. No direct coupling of the two shuttles was observed, however, and only the glutamate-glutamine cycle seemed activity dependent.

  2. A theoretical study of the XP and NEXAFS spectra of alanine: gas phase molecule, crystal, and adsorbate at the ZnO(10 ̅10) surface.

    PubMed

    Gao, You Kun; Traeger, Franziska; Kotsis, Konstantinos; Staemmler, Volker

    2011-06-14

    The adsorption of alanine on the mixed-terminated ZnO(10 ̅10) surface is studied by means of quantum-chemical ab initio calculations. Using a finite cluster model and the adsorption geometry as obtained both by periodic CPMD and embedded cluster calculations, the C1s, N1s and O1s X-ray photoelectron spectra (XPS) and near-edge X-ray absorption fine structure (NEXAFS) spectra are calculated for single alanine molecules on ZnO(10 ̅10). These spectra are compared with the spectra calculated for alanine in the gas phase and in its crystalline form and with experimental XPS and NEXAFS data for the isolated alanine molecule and for alanine adsorbed on ZnO(10 ̅10) at multilayer and monolayer coverage. The excellent agreement between the experimental and calculated XP and NEXAFS spectra confirms the calculated adsorption geometry: A single alanine molecule is bound to ZnO(10 ̅10) in a dissociated bidentate form with the two O atoms of the acid group bound to two Zn atoms of the surface and the proton transferred to one O atom of the surface. Other possible structures, such as adsorption of alanine in one of its neutral or zwitterionic forms in which the proton of the -COOH group remains at this group or is transferred to the amino group, can be excluded since they would give rise to quite different XP spectra. In the multilayer coverage regime, on the other hand, alanine is in its crystalline form as is also shown by the analysis of the XP spectra.

  3. Microbial Production of Amino Acid-Related Compounds.

    PubMed

    Wendisch, Volker F

    2016-11-22

    Corynebacterium glutamicum is the workhorse of the production of proteinogenic amino acids used in food and feed biotechnology. After more than 50 years of safe amino acid production, C. glutamicum has recently also been engineered for the production of amino acid-derived compounds, which find various applications, e.g., as synthons for the chemical industry in several markets including the polymer market. The amino acid-derived compounds such as non-proteinogenic ω-amino acids, α,ω-diamines, and cyclic or hydroxylated amino acids have similar carbon backbones and functional groups as their amino acid precursors. Decarboxylation of amino acids may yield ω-amino acids such as β-alanine, γ-aminobutyrate, and δ-aminovalerate as well as α,ω-diamines such as putrescine and cadaverine. Since transamination is the final step in several amino acid biosynthesis pathways, 2-keto acids as immediate amino acid precursors are also amenable to production using recombinant C. glutamicum strains. Approaches for metabolic engineering of C. glutamicum for production of amino acid-derived compounds will be described, and where applicable, production from alternative carbon sources or use of genome streamline will be referred to. The excellent large-scale fermentation experience with C. glutamicum offers the possibility that these amino acid-derived speciality products may enter large-volume markets.

  4. Amino acid analyses of R and CK chondrites

    NASA Astrophysics Data System (ADS)

    Burton, Aaron S.; McLain, Hannah; Glavin, Daniel P.; Elsila, Jamie E.; Davidson, Jemma; Miller, Kelly E.; Andronikov, Alexander V.; Lauretta, Dante; Dworkin, Jason P.

    2015-03-01

    Exogenous delivery of amino acids and other organic molecules to planetary surfaces may have played an important role in the origins of life on Earth and other solar system bodies. Previous studies have revealed the presence of indigenous amino acids in a wide range of carbon-rich meteorites, with the abundances and structural distributions differing significantly depending on parent body mineralogy and alteration conditions. Here we report on the amino acid abundances of seven type 3-6 CK chondrites and two Rumuruti (R) chondrites. Amino acid measurements were made on hot water extracts from these meteorites by ultrahigh-performance liquid chromatography with fluorescence detection and time-of-flight mass spectrometry. Of the nine meteorites analyzed, four were depleted in amino acids, and one had experienced significant amino acid contamination by terrestrial biology. The remaining four, comprised of two R and two CK chondrites, contained low levels of amino acids that were predominantly the straight chain, amino-terminal (n-ω-amino) acids β-alanine, and γ-amino-n-butyric acid. This amino acid distribution is similar to what we reported previously for thermally altered ureilites and CV and CO chondrites, and these n-ω-amino acids appear to be indigenous to the meteorites and not the result of terrestrial contamination. The amino acids may have been formed by Fischer-Tropsch-type reactions, although this hypothesis needs further testing.

  5. (2R,1'S,2'R)- and (2S,1'S,2'R)-3-[2-Mono(di,tri)fluoromethylcyclopropyl]alanines and their incorporation into hormaomycin analogues

    PubMed Central

    Kozhushkov, Sergei I; Yufit, Dmitrii S; Grosse, Christian; Kaiser, Marcel

    2014-01-01

    Summary Efficient and scalable syntheses of enantiomerically pure (2R,1'S,2'R)- and (2S,1'S,2'R)-3-[2-mono(di,tri)fluoromethylcyclopropyl]alanines 9a–c, as well as allo-D-threonine (4) and (2S,3R)-β-methylphenylalanine (3), using the Belokon' approach with (S)- and (R)-2-[(N-benzylprolyl)amino]benzophenone [(S)- and (R)-10] as reusable chiral auxiliaries have been developed. Three new fluoromethyl analogues of the naturally occurring octadepsipeptide hormaomycin (1) with (fluoromethylcyclopropyl)alanine moieties have been synthesized and subjected to preliminary tests of their antibiotic activity. PMID:25550751

  6. Function of the D-alanine:D-alanine ligase lid loop: a molecular modeling and bioactivity study.

    PubMed

    Hrast, Martina; Vehar, Blaž; Turk, Samo; Konc, Janez; Gobec, Stanislav; Janežič, Dušanka

    2012-08-09

    D-Alanine:D-alanine ligase (Ddl) is an essential ATP-dependent bacterial enzyme involved in peptidoglycan biosynthesis. Discovery of Ddl inhibitors not competitive with ATP has proven to be difficult because the Ddl bimolecular d-alanine binding pocket is very restricted, as is accessibility to the active site for larger molecules in the catalytically active closed conformation of Ddl. A molecular dynamics study of the opening and closing of the Ddl lid loop informs future structure-based design efforts that allow for the flexibility of Ddl. A virtual screen on generated enzyme conformations yielded some hit inhibitors whose bioactivity was determined.

  7. Enantiomer-specific selection of amino acids

    PubMed Central

    Ren, Xueying; Tellez, Luis A; de Araujo, Ivan E

    2013-01-01

    Dietary intake of L-amino acids impacts on several physiological functions, including the control of gastrointestinal motility, pancreatic secretion, and appetite. However, the biological mechanisms regulating behavioral predilections for certain amino acid types remain poorly understood. We tested the hypothesis that, in mice, the potency with which a given glucogenic amino acid increases glucose utilization reflects its rewarding properties. We have found that: 1. During long-, but not short-, term preference tests, L-alanine and L-serine were preferred over their D-enantiomer counterparts, while no such effect was observed for L-threonine vs. D-threonine; 2. These behavioral patterns were closely associated with the ability of L-amino acids to promote increases in respiratory exchange ratios such that those, and only those, L-amino acids able to promote increases in respiratory exchange ratios were preferred over their D-isomers; 3. These behavioral preferences were independent of gustatory influences, since taste-deficient Trpm5 knockout mice displayed ingestive responses very similar to those of their wild-type counterparts. We conclude that the ability to promote increases in respiratory exchange ratios enhances the reward value of nutritionally relevant amino acids, and suggest a mechanistic link between substrate utilization and amino acid preferences. PMID:24072505

  8. Structural analysis and mutant growth properties reveal distinctive enzymatic and cellular roles for the three major L-alanine transaminases of Escherichia coli.

    PubMed

    Peña-Soler, Esther; Fernandez, Francisco J; López-Estepa, Miguel; Garces, Fernando; Richardson, Andrew J; Quintana, Juan F; Rudd, Kenneth E; Coll, Miquel; Vega, M Cristina

    2014-01-01

    In order to maintain proper cellular function, the metabolism of the bacterial microbiota presents several mechanisms oriented to keep a correctly balanced amino acid pool. Central components of these mechanisms are enzymes with alanine transaminase activity, pyridoxal 5'-phosphate-dependent enzymes that interconvert alanine and pyruvate, thereby allowing the precise control of alanine and glutamate concentrations, two of the most abundant amino acids in the cellular amino acid pool. Here we report the 2.11-Å crystal structure of full-length AlaA from the model organism Escherichia coli, a major bacterial alanine aminotransferase, and compare its overall structure and active site composition with detailed atomic models of two other bacterial enzymes capable of catalyzing this reaction in vivo, AlaC and valine-pyruvate aminotransferase (AvtA). Apart from a narrow entry channel to the active site, a feature of this new crystal structure is the role of an active site loop that closes in upon binding of substrate-mimicking molecules, and which has only been previously reported in a plant enzyme. Comparison of the available structures indicates that beyond superficial differences, alanine aminotransferases of diverse phylogenetic origins share a universal reaction mechanism that depends on an array of highly conserved amino acid residues and is similarly regulated by various unrelated motifs. Despite this unifying mechanism and regulation, growth competition experiments demonstrate that AlaA, AlaC and AvtA are not freely exchangeable in vivo, suggesting that their functional repertoire is not completely redundant thus providing an explanation for their independent evolutionary conservation.

  9. Transfer of Asymmetry between Proteinogenic Amino Acids under Harsh Conditions

    NASA Astrophysics Data System (ADS)

    Tarasevych, Arkadii V.; Vives, Thomas; Snytnikov, Valeriy N.; Guillemin, Jean-Claude

    2017-03-01

    The heating above 400 °C of serine, cysteine, selenocysteine and threonine leads to a complete decomposition of the amino acids and to the formation in low yields of alanine for the three formers and of 2-aminobutyric acid for the latter. At higher temperature, this amino acid is observed only when sublimable α-alkyl-α-amino acids are present, and with an enantiomeric excess dependent on several parameters. Enantiopure or enantioenriched Ser, Cys, Sel or Thr is not able to transmit its enantiomeric excess to the amino acid formed during its decomposition. The presence during the sublimation-decomposition of enantioenriched valine or isoleucine leads to the enantioenrichment of all sublimable amino acids independently of the presence of many decomposition products coming from the unstable derivative. All these studies give information on a potentially prebiotic key-reaction of abiotic transformations between α-amino acids and their evolution to homochirality.

  10. Transfer of Asymmetry between Proteinogenic Amino Acids under Harsh Conditions.

    PubMed

    Tarasevych, Arkadii V; Vives, Thomas; Snytnikov, Valeriy N; Guillemin, Jean-Claude

    2017-03-31

    The heating above 400 °C of serine, cysteine, selenocysteine and threonine leads to a complete decomposition of the amino acids and to the formation in low yields of alanine for the three formers and of 2-aminobutyric acid for the latter. At higher temperature, this amino acid is observed only when sublimable α-alkyl-α-amino acids are present, and with an enantiomeric excess dependent on several parameters. Enantiopure or enantioenriched Ser, Cys, Sel or Thr is not able to transmit its enantiomeric excess to the amino acid formed during its decomposition. The presence during the sublimation-decomposition of enantioenriched valine or isoleucine leads to the enantioenrichment of all sublimable amino acids independently of the presence of many decomposition products coming from the unstable derivative. All these studies give information on a potentially prebiotic key-reaction of abiotic transformations between α-amino acids and their evolution to homochirality.

  11. Surface Propensity of Atmospherically Relevant Amino Acids Studied by XPS.

    PubMed

    Mocellin, Alexandra; Gomes, Anderson Herbert de Abreu; Araújo, Oscar Cardoso; de Brito, Arnaldo Naves; Bjorneholm, Olle

    2017-03-30

    Amino acids constitute an important fraction of the water-soluble organic nitrogen (WSON) compounds in aerosols and are involved in many processes in the atmosphere. In this work, we applied XPS to study aqueous solutions of four amino acids: glycine, alanine, valine and methionine, in their zwitterionic forms. We found that amino acids with hydrophilic side chains and smaller size, GLY and ALA, tend to stay in the bulk of the liquid, while the hydrophobic and bigger amino acids, VAL and MET, are found to concentrate more on the surface. We found experimental evidences that the amino acids have preferential orientation relative to the surface, with the hydrophobic side chain being closer to the surface than the hydrophilic carboxylate group. The observed amino acid surface propensity has implications in atmospheric science as the surface interaction play a central role in cloud droplet formation, and they should be considered in climate models.

  12. Design and Synthesis of Novel Isoxazole Tethered Quinone-Amino Acid Hybrids

    PubMed Central

    Ravi Kumar, P.; Sambaiah, M.; Kandula, Venu; Payili, Nagaraju; Jaya Shree, A.; Yennam, Satyanarayana

    2014-01-01

    A new series of isoxazole tethered quinone-amino acid hybrids has been designed and synthesized involving 1,3-dipolar cycloaddition reaction followed by an oxidation reaction using cerium ammonium nitrate (CAN). Using this method, for the first time various isoxazole tethered quinone-phenyl alanine and quinone-alanine hybrids were synthesized from simple commercially available 4-bromobenzyl bromide, propargyl bromide, and 2,5-dimethoxybenzaldehyde in good yield. PMID:25709839

  13. Prebiotic Synthesis of Hydrophobic and Protein Amino Acids

    PubMed Central

    Ring, David; Wolman, Yecheskel; Friedmann, Nadav; Miller, Stanley L.

    1972-01-01

    The formation of amino acids by the action of electric discharges on a mixture of methane, nitrogen, and water with traces of ammonia was studied in detail. The presence of glycine, alanine, α-amino-n-butyric acid, α-aminoisobutyric acid, valine, norvaline, isovaline, leucine, isoleucine, alloisoleucine, norleucine, proline, aspartic acid, glutamic acid, serine, threonine, allothreonine, α-hydroxy-γ-aminobutyric acid, and α,γ-diaminobutyric acid was confirmed by ion-exchange chromatography and gas chromatography-mass spectrometry. All of the primary α-amino acids found in the Murchison Meteorite have been synthesized by this electric discharge experiment. PMID:4501592

  14. The Path of Carbon in Photosynthesis II. Amino Acids

    DOE R&D Accomplishments Database

    Stepka, W.; Benson, A. A.; Calvin, M.

    1948-05-25

    The radioactive amino acid's synthesized from C{sup 14}O{sub 2} by green algae both in the light and in the dark after CO{sub 2}-free preillumination have been separated and identified using paper chromatography and radioautography. The radioactive amino acids identified were aspartic acid, alanine and smaller amounts of 3- and 4-carbon amino acids. This finding as well as the total absence of radioactive glutamic acid substantiates the mechanism for reduction of CO{sub 2} previously postulated by members of this laboratory.

  15. R76 in transmembrane domain 3 of the aspartate:alanine transporter AspT is involved in substrate transport.

    PubMed

    Suzuki, Satomi; Nanatani, Kei; Abe, Keietsu

    2016-01-01

    The L-aspartate:L-alanine antiporter of Tetragenococcus halophilus (AspT) possesses an arginine residue (R76) within the GxxxG motif in the central part of transmembrane domain 3 (TM3)-a residue that has been estimated to transport function. In this study, we carried out amino acid substitutions of R76 and used proteoliposome reconstitution for analyzing the transport function of each substitution. Both l-aspartate and l-alanine transport assays showed that R76K has higher activity than the AspT-WT (R76), whereas R76D and R76E have lower activity than the AspT-WT. These results suggest that R76 is involved in AspT substrate transport.

  16. Effect of taurine on the concentrations of glutamate, GABA, glutamine and alanine in the rat striatum and hippocampus.

    PubMed

    Molchanova, Svetlana M; Oja, Simos S; Saransaari, Pirjo

    2007-01-01

    Taurine, a non-protein amino acid, acts as an osmoregulator and inhibitory neuromodulator in the brain. Here we studied the effects of intraperitoneal injections of taurine on the concentrations of glutamate and GABA, and their precursors, glutamine and alanine, in the rat striatum and hippocampus. Injections of 0.25, 0.5 and 1 g/kg taurine led to a gradual increase in taurine tissue concentrations in both hippocampus and striatum. Glutamate and GABA also increased in the hippocampus, but not in the striatum. Glutamine increased and alanine decreased markedly in both brain structures. The results corroborate the neuromodulatory role of taurine in the brain. Taurine administration results in an imbalance in inhibitory and excitatory neurotransmission in the glutamatergic (hippocampus) and GABAergic (striatum) brain structures, affecting more markedly the neurotransmitter precursors.

  17. Unprecedented concentrations of indigenous amino acids in primitive CR meteorites

    NASA Astrophysics Data System (ADS)

    Ehrenfreund, Pascale; Martins, Zita; Alexander, Conel; Orzechowska, Grazyna; Fogel, Marylin

    CR meteorites are among the most primitive meteorites. We have performed pioneering work determining the compositional characteristics of amino acids in this type of carbonaceous chondrites. We report the first measurements of amino acids in Antarctic CR meteorites, two of which show the highest amino acid concentrations ever found in a chondrite. We have analyzed the amino acid content of the Antarctic CRs EET92042, GRA95229 and GRO95577 using high performance liquid chromatography with UV fluorescence detection (HPLC-FD) and gas chromatography-mass spectrometry (GC-MS). Additionally, compound-specific carbon isotopic measurements for most of the individual amino acids from the EET92042 and GRA95229 meteorites were achieved by gas chromatography-combustion-isotope ratio mass spectrometry (GC-C-IRMS). Our data show that EET92042 and GRA95229 are the most amino acid-rich chondrites ever analyzed, with total amino acid concentrations of 180 and 249 parts-per-million (ppm), respectively. GRO95577, however, is depleted in amino acids (<1 ppm). The most abundant amino acids present in the EET92042 and GRA95229 meteorites are the α-amino acids glycine, isovaline, α-aminoisobutyric acid (α-AIB), and alanine, with δ 13 C values ranging from +31.6% to +50.5%. The highly enriched carbon isotope results together with racemic enantiomeric ratios determined for most amino acids indicate that primitive organic matter was preserved in these meteorites. In addition, the relative abundances of α-AIB and β-alanine amongst Antarctic CR meteorites appear to correspond to the degree of aqueous alteration on their respective parent body. Investigating the abundances and isotopic composition of amino acids in primitive chondrites helps to understand the role of meteorites as a source of extraterrestrial prebiotic organic compounds to the early Earth.

  18. Noncovalent and covalent functionalization of a (5, 0) single-walled carbon nanotube with alanine and alanine radicals.

    PubMed

    Rajarajeswari, Muthusivarajan; Iyakutti, Kombiah; Kawazoe, Yoshiyuki

    2012-02-01

    We have systematically investigated the noncovalent and covalent adsorption of alanine and alanine radicals, respectively, onto a (5, 0) single-walled carbon nanotube using first-principles calculation. It was found that XH···π (X = N, O, C) interactions play a crucial role in the non-ovalent adsorption and that the functional group close to the carbon nanotube exhibits a significant influence on the binding strength. Noncovalent functionalization of the carbon nanotube with alanine enhances the conductivity of the metallic (5, 0) nanotube. In the covalent adsorption of each alanine radical onto a carbon nanotube, the binding energy depends on the adsorption site on CNT and the electronegative atom that binds with the CNT. The strongest complex is formed when the alanine radical interacts with a (5, 0) carbon nanotube through the amine group. In some cases, the covalent interaction of the alanine radical introduces a half-filled band at the Fermi level due to the local sp (3) hybridization, which modifies the conductivity of the tube.

  19. Structure of D-alanine-D-alanine ligase from Yersinia pestis: nucleotide phosphate recognition by the serine loop.

    PubMed

    Tran, Huyen Thi; Hong, Myoung Ki; Ngo, Ho Phuong Thuy; Huynh, Kim Hung; Ahn, Yeh Jin; Wang, Zhong; Kang, Lin Woo

    2016-01-01

    D-Alanyl-D-alanine is an essential precursor of bacterial peptidoglycan and is synthesized by D-alanine-D-alanine ligase (DDL) with hydrolysis of ATP; this reaction makes DDL an important drug target for the development of antibacterial agents. Five crystal structures of DDL from Yersinia pestis (YpDDL) were determined at 1.7-2.5 Å resolution: apo, AMP-bound, ADP-bound, adenosine 5'-(β,γ-imido)triphosphate-bound, and D-alanyl-D-alanine- and ADP-bound structures. YpDDL consists of three domains, in which four loops, loop 1, loop 2 (the serine loop), loop 3 (the ω-loop) and loop 4, constitute the binding sites for two D-alanine molecules and one ATP molecule. Some of them, especially the serine loop and the ω-loop, show flexible conformations, and the serine loop is mainly responsible for the conformational change in substrate nucleotide phosphates. Enzyme-kinetics assays were carried out for both the D-alanine and ATP substrates and a substrate-binding mechanism was proposed for YpDDL involving conformational changes of the loops.

  20. Reconfiguration of N Metabolism upon Hypoxia Stress and Recovery: Roles of Alanine Aminotransferase (AlaAT) and Glutamate Dehydrogenase (GDH)

    PubMed Central

    Diab, Houssein; Limami, Anis M.

    2016-01-01

    In the context of climatic change, more heavy precipitation and more frequent flooding and waterlogging events threaten the productivity of arable farmland. Furthermore, crops were not selected to cope with flooding- and waterlogging-induced oxygen limitation. In general, low oxygen stress, unlike other abiotic stresses (e.g., cold, high temperature, drought and saline stress), received little interest from the scientific community and less financial support from stakeholders. Accordingly, breeding programs should be developed and agronomical practices should be adapted in order to save plants’ growth and yield—even under conditions of low oxygen availability (e.g., submergence and waterlogging). The prerequisite to the success of such breeding programs and changes in agronomical practices is a good knowledge of how plants adapt to low oxygen stress at the cellular and the whole plant level. In the present paper, we summarized the recent knowledge on metabolic adjustment in general under low oxygen stress and highlighted thereafter the major changes pertaining to the reconfiguration of amino acids syntheses. We propose a model showing (i) how pyruvate derived from active glycolysis upon hypoxia is competitively used by the alanine aminotransferase/glutamate synthase cycle, leading to alanine accumulation and NAD+ regeneration. Carbon is then saved in a nitrogen store instead of being lost through ethanol fermentative pathway. (ii) During the post-hypoxia recovery period, the alanine aminotransferase/glutamate dehydrogenase cycle mobilizes this carbon from alanine store. Pyruvate produced by the reverse reaction of alanine aminotransferase is funneled to the TCA cycle, while deaminating glutamate dehydrogenase regenerates, reducing equivalent (NADH) and 2-oxoglutarate to maintain the cycle function. PMID:27258319

  1. The polyproline II conformation in short alanine peptides is noncooperative.

    PubMed

    Chen, Kang; Liu, Zhigang; Kallenbach, Neville R

    2004-10-26

    The finding that short alanine peptides possess a high fraction of polyproline II (PII) structure (Phi=-75 degrees, Psi=+145 degrees ) at low temperature has broad implications for unfolded states of proteins. An important question concerns whether or not this structure is locally determined or cooperative. We have monitored the conformation of alanine in a series of model peptides AcGGAnGGNH2 (n=1-3) over a temperature range from -10 degrees C to +80 degrees C. Use of 15N-labeled alanine substitutions makes it possible to measure 3JalphaN coupling constants accurately over the full temperature range. Based on a 1D next-neighbor model, the cooperative parameter sigma of PII nucleation is evaluated from the coupling constant data. The finding that sigma is close to unity (1 +/- 0.2) indicates a noncooperative role for alanine in PII structure formation, consistent with statistical surveys of the Protein Data Bank that suggest that most PII structure occurs in isolated residues. Lack of cooperativity in these models implies that hydration effects that influence PII conformation in water are highly localized. Using a nuclear Overhauser effect ratio strategy to define the alanine Psi angle, we estimate that, at 40 degrees C, the time-averaged alanine conformation (Phi=-80 degrees, Psi=+170 degrees ) deviates from canonical PII structure, indicating that PII melts at high temperature. Thus, the high-temperature state of short alanine peptides seems to be an unfolded ensemble with higher distribution in the extended beta structure basin, but not a coil.

  2. EPR/alanine dosimetry for two therapeutic proton beams

    NASA Astrophysics Data System (ADS)

    Marrale, Maurizio; Carlino, Antonio; Gallo, Salvatore; Longo, Anna; Panzeca, Salvatore; Bolsi, Alessandra; Hrbacek, Jan; Lomax, Tony

    2016-02-01

    In this work the analysis of the electron paramagnetic resonance (EPR) response of alanine pellets exposed to two different clinical proton beams employed for radiotherapy is performed. One beam is characterized by a passive delivery technique and is dedicated to the eyes treatment (OPTIS2 beam line). Alanine pellets were irradiated with a 70 MeV proton beam corresponding to 35 mm range in eye tissue. We investigated how collimators with different sizes and shape used to conform the dose to the planned target volume influence the delivered dose. For this purpose we performed measurements with varying the collimator size (Output Factor) and the results were compared with those obtained with other dosimetric techniques (such as Markus chamber and diode detector). This analysis showed that the dosimeter response is independent of collimator diameter if this is larger than or equal to 10 mm. The other beam is characterized by an active spot-scanning technique, the Gantry1 beam line (maximum energy 230 MeV), and is used to treat deep-seated tumors. The dose linearity of alanine response in the clinical dose range was tested and the alanine dose response at selected locations in depth was measured and compared with the TPS planned dose in a quasi-clinical scenario. The alanine response was found to be linear in the dose in the clinical explored range (from 10 to 70 Gy). Furthermore, a depth dose profile in a quasi-clinical scenario was measured and compared to the dose computed by the Treatment Planning System PSIPLAN. The comparison of calibrated proton alanine measurements and TPS dose shows a difference under 1% in the SOBP and a "quenching" effect up to 4% in the distal part of SOBP. The positive dosimetric characteristics of the alanine pellets confirm the feasibility to use these detectors for "in vivo" dosimetry in clinical proton beams.

  3. Concerted modulation of alanine and glutamate metabolism in young Medicago truncatula seedlings under hypoxic stress.

    PubMed

    Limami, Anis M; Glévarec, Gaëlle; Ricoult, Claudie; Cliquet, Jean-Bernard; Planchet, Elisabeth

    2008-01-01

    The modulation of primary nitrogen metabolism by hypoxic stress was studied in young Medicago truncatula seedlings. Hypoxic seedlings were characterized by the up-regulation of glutamate dehydrogenase 1 (GDH1) and mitochondrial alanine aminotransferase (mAlaAT), and down-regulation of glutamine synthetase 1b (GS1b), NADH-glutamate synthase (NADH-GOGAT), glutamate dehydrogenase 3 (GDH3), and isocitrate dehydrogenase (ICDH) gene expression. Hypoxic stress severely inhibited GS activity and stimulated NADH-GOGAT activity. GDH activity was lower in hypoxic seedlings than in the control, however, under either normoxia or hypoxia, the in vivo activity was directed towards glutamate deamination. (15)NH(4) labelling showed for the first time that the adaptive reaction of the plant to hypoxia consisted of a concerted modulation of nitrogen flux through the pathways of both alanine and glutamate synthesis. In hypoxic seedlings, newly synthesized (15)N-alanine increased and accumulated as the major amino acid, asparagine synthesis was inhibited, while (15)N-glutamate was synthesized at a similar rate to that in the control. A discrepancy between the up-regulation of GDH1 expression and the down-regulation of GDH activity by hypoxic stress highlighted for the first time the complex regulation of this enzyme by hypoxia. Higher rates of glycolysis and ethanol fermentation are known to cause the fast depletion of sugar stores and carbon stress. It is proposed that the expression of GDH1 was stimulated by hypoxia-induced carbon stress, while the enzyme protein might be involved during post-hypoxic stress contributing to the regeneration of 2-oxoglutarate via the GDH shunt.

  4. The Genetic Architecture of Murine Glutathione Transferases

    PubMed Central

    Lu, Lu; Pandey, Ashutosh K.; Houseal, M. Trevor; Mulligan, Megan K.

    2016-01-01

    Glutathione S-transferase (GST) genes play a protective role against oxidative stress and may influence disease risk and drug pharmacokinetics. In this study, massive multiscalar trait profiling across a large population of mice derived from a cross between C57BL/6J (B6) and DBA2/J (D2)—the BXD family—was combined with linkage and bioinformatic analyses to characterize mechanisms controlling GST expression and to identify downstream consequences of this variation. Similar to humans, mice show a wide range in expression of GST family members. Variation in the expression of Gsta4, Gstt2, Gstz1, Gsto1, and Mgst3 is modulated by local expression QTLs (eQTLs) in several tissues. Higher expression of Gsto1 in brain and liver of BXD strains is strongly associated (P < 0.01) with inheritance of the B6 parental allele whereas higher expression of Gsta4 and Mgst3 in brain and liver, and Gstt2 and Gstz1 in brain is strongly associated with inheritance of the D2 parental allele. Allele-specific assays confirmed that expression of Gsto1, Gsta4, and Mgst3 are modulated by sequence variants within or near each gene locus. We exploited this endogenous variation to identify coexpression networks and downstream targets in mouse and human. Through a combined systems genetics approach, we provide new insight into the biological role of naturally occurring variants in GST genes. PMID:26829228

  5. Effect of substituting arginine and lysine with alanine on antimicrobial activity and the mechanism of action of a cationic dodecapeptide (CL(14-25)), a partial sequence of cyanate lyase from rice.

    PubMed

    Taniguchi, Masayuki; Takahashi, Nobuteru; Takayanagi, Tomohiro; Ikeda, Atsuo; Ishiyama, Yohei; Saitoh, Eiichi; Kato, Tetsuo; Ochiai, Akihito; Tanaka, Takaaki

    2014-01-01

    The antimicrobial activity of analogs obtained by substituting arginine and lysine in CL(14-25), a cationic α-helical dodecapeptide, with alanine against Porphyromonas gingivalis, a periodontal pathogen, varied significantly depending on the number and position of cationic amino acids. The alanine-substituted analogs had no hemolytic activity, even at a concentration of 1 mM. The antimicrobial activities of CL(K20A) and CL(K20A, K25A) were 3.8-fold and 9.1-fold higher, respectively, than that of CL(14-25). The antimicrobial activity of CL(R15A) was slightly lower than that of CL(14-25), suggesting that arginine at position 15 is not essential but is important for the antimicrobial activity. The experiments in which the alanine-substituted analogs bearing the replacement of arginine at position 24 and/or lysine at position 25 were used showed that arginine at position 24 was crucial for the antimicrobial activity whenever lysine at position 25 was substituted with alanine. Helical wheel projections of the alanine-substituted analogs indicate that the hydrophobicity in the vicinity of leucine at position 16 and alanines at positions 18 and/or 21 increased by substituting lysine at positions 20 and 25 with alanine, respectively. The degrees of diSC3 -5 release from P. gingivalis cells and disruption of GUVs induced by the alanine-substituted analogs with different positive charges were not closely related to their antimicrobial activities. The enhanced antimicrobial activities of the alanine-substituted analogs appear to be mainly attributable to the changes in properties such as hydrophobicity and amphipathic propensity due to alanine substitution and not to their extents of positive charge (cationicity).

  6. Amino acid odorants stimulate microvillar sensory neurons.

    PubMed

    Lipschitz, David L; Michel, William C

    2002-03-01

    The olfactory epithelium (OE) of zebrafish is populated with ciliated and microvillar olfactory sensory neurons (OSNs). Whether distinct classes of odorants specifically activate either of these unique populations of OSNs is unknown. Previously we demonstrated that zebrafish OSNs could be labeled in an activity-dependent fashion by amino acid but not bile acid odorants. To determine which sensory neuron type was stimulated by amino acid odorants, we labeled OSNs using the ion channel permeant probe agmatine (AGB) and analyzed its distribution with conventional light- and electron-microscope immunocytochemical techniques. Approximately 7% of the sensory epithelium was labeled by AGB exposure alone. Following stimulation with one of the eight amino acids tested, the proportion of labeled epithelium increased from 9% for histidine to 19% for alanine; amino acid stimulated increases in labeling of 2-12% over control labeling. Only histidine failed to stimulate a significant increase in the proportion of labeled OSNs compared to control preparations. Most amino acid sensitive OSNs were located superficially in the epithelium and immuno-electron microscopy demonstrated that the labeled OSNs were predominantly microvillar. Large numbers of nanogold particles (20-60 per 1.5 microm(2)) were associated with microvillar olfactory sensory neurons (MSNs), while few such particles (<15 per 1.5 microm(2)) were observed over ciliated olfactory sensory neurons (CSNs), supporting cells (SCs) and areas without tissue, such as the lumen above the OE. Collectively, these findings indicate that microvillar sensory neurons are capable of detecting amino acid odorants.

  7. Modifications of the acyl-d-alanyl-d-alanine terminus affecting complex-formation with vancomycin

    PubMed Central

    Nieto, M.; Perkins, H. R.

    1971-01-01

    Vancomycin forms complexes with peptides terminating in d-alanyl-d-alanine that are analogous to the biosynthetic precursors of bacterial mucopeptides. The specificity of complex-formation has been studied by means of many synthetic peptides, prepared by both solid-phase and conventional methods. The following conclusions can be drawn: (a) three amide linkages are required to form a stable complex; (b) the terminal carboxyl group must be free; (c) the carboxyl terminal and subterminal residues must be either glycine or of the d-configuration; (d) the size of the side chain in these residues greatly influences the affinity for vancomycin, a methyl group being the optimum in each case; (e) the nature of the side chain in the third and fourth residues has a smaller effect on complex-formation, but an l-configuration was somewhat better than a d-configuration in the third position. In addition to acyl-d-alanyl-d-alanine, other peptides that occur in bacterial cell walls will combine with vancomycin, although less strongly, e.g. acyl-d-alanyl-d-α-amino acid (where the terminal d-residue may form the cross-link in mucopeptide structure) and acyl-l-alanyl-d-glutamylglycine (a sequence found in the mucopeptide of Micrococcus lysodeikticus and related organisms). These results throw some light on the specificity of the uptake of vancomycin by living bacteria. PMID:5124386

  8. 3-Phosphono-L-alanine as pyrophosphate mimic for DNA synthesis using HIV-1 reverse transcriptase.

    PubMed

    Yang, Shiqiong; Froeyen, Mathy; Lescrinier, Eveline; Marlière, Philippe; Herdewijn, Piet

    2011-01-07

    A series of sulf(on)ate and phosph(on)ate amino acid phosphoramidate analogues of deoxynucleotides were synthesized as potential substrates for HIV-1 reverse transcriptase. Taurine, L-cysteic acid, 3-phosphono-L-alanine, O-sulfonato-L-serine, and O-phospho-L-serine were investigated as leaving groups in an enzyme catalyzed DNA synthesis protocol. Among these analogues, the phosphonate congener performed best and 3-phosphono-L-alanine can be considered as an excellent mimic of the pyrophosphate (PPi) moiety of deoxyadenosine triphosphate, to be used in enzymatic synthesis of nucleic acids. During a single nucleotide incorporation assay the use of 3-phosphono-L-Ala-dAMP as substrate resulted in 95% conversion to a P + 1 strand in 60 min at 50 μM (a concentration 10 times less than found for L-Asp-dAMP) and with improved incorporation kinetics and less stalling. For the sequences investigated, the efficiency of the incorporation is base dependent and decreases in the order (A ≥ T = G > C). In all cases, the incorporation follows Watson-Crick rules.

  9. Tuning electronic transport via hepta-alanine peptides junction by tryptophan doping

    PubMed Central

    Guo, Cunlan; Yu, Xi; Refaely-Abramson, Sivan; Sepunaru, Lior; Bendikov, Tatyana; Pecht, Israel; Kronik, Leeor; Vilan, Ayelet; Sheves, Mordechai; Cahen, David

    2016-01-01

    Charge migration for electron transfer via the polypeptide matrix of proteins is a key process in biological energy conversion and signaling systems. It is sensitive to the sequence of amino acids composing the protein and, therefore, offers a tool for chemical control of charge transport across biomaterial-based devices. We designed a series of linear oligoalanine peptides with a single tryptophan substitution that acts as a “dopant,” introducing an energy level closer to the electrodes’ Fermi level than that of the alanine homopeptide. We investigated the solid-state electron transport (ETp) across a self-assembled monolayer of these peptides between gold contacts. The single tryptophan “doping” markedly increased the conductance of the peptide chain, especially when its location in the sequence is close to the electrodes. Combining inelastic tunneling spectroscopy, UV photoelectron spectroscopy, electronic structure calculations by advanced density-functional theory, and dc current–voltage analysis, the role of tryptophan in ETp is rationalized by charge tunneling across a heterogeneous energy barrier, via electronic states of alanine and tryptophan, and by relatively efficient direct coupling of tryptophan to a Au electrode. These results reveal a controlled way of modulating the electrical properties of molecular junctions by tailor-made “building block” peptides. PMID:27621456

  10. Preferential Treatment: Interaction Between Amino Acids and Minerals

    NASA Astrophysics Data System (ADS)

    Crapster-Pregont, E. J.; Cleaves, H. J.; Hazen, R. M.

    2008-12-01

    Amino acids are the building blocks of proteins and are important for some models of the origin of life. Polymerization of amino acids from dilute solution is unlikely without a scaffold or catalyst. The surfaces of early Earth minerals are the most likely candidates for this role. The surface adsorption behavior of 12 amino acids (L-alanine, L-serine, L-aspartic acid, L-proline, L- phenylalanine, L-valine, L-arginine, d-amino valeric acid, glycine, L-lysine, L-isoleucine, and B-alanine) on 21 minerals (quartz, calcite, enstatite, illite, olivine, pyrrhotite, pyrite, alkali basalt, albite, analcime, chlorite, barite, hydroxyl apatite, hematite, magnetite, aluminum hydroxide, kaolin, silica gel, corundum, rutile, and montmorillonite) was determined via batch adsorption experiments. Absorption was determined for concentrations between 10-4M and 10-6M in the presence of 0.1M NaCl, and between pH values of 3 and 9 at 25 degrees C. The equilibrated solutions were centrifuged, filtered, derivatized using a fluorescent amino group tag (dansyl-chloride) and analyzed by HPLC. Adsorption was standardized using BET surface area measurements for each mineral to give the number of mols of each amino acid adsorbed per square meter for each mineral. The results indicate an enormous difference in the adsorption of amino acids between minerals, along with major differences in the adsorption of individual amino acids on the same mineral surface. There is also a change in the absorbance of amino acids as the pH changes. Many previous studies of amino acid concentration and catalysis by minerals have used clay minerals because of their high surface areas, however, this data suggests that the surfaces of minerals such as calcite, quartz and pyrite have even higher affinities for amino acids. The results suggest mineral surfaces that could be optimal locations for the polymerization of molecules linked to the origin of life.

  11. Combined supplementation of carbohydrate, alanine, and proline is effective in maintaining blood glucose and increasing endurance performance during long-term exercise in mice.

    PubMed

    Nogusa, Yoshihito; Mizugaki, Ami; Hirabayashi-Osada, Yuri; Furuta, Chie; Ohyama, Kana; Suzuki, Katsuya; Kobayashi, Hisamine

    2014-01-01

    Carbohydrate supplementation is extremely important during prolonged exercise because it maintains blood glucose levels during later stages of exercise. In this study, we examined whether maintaining blood glucose levels by carbohydrate supplementation could be enhanced during long-term exercise by combining this supplementation with alanine and proline, which are gluconeogenic amino acids, and whether such a combination would affect exercise endurance performance. Male C57BL/6J mice were orally administered either maltodextrin (1.25 g/kg) or maltodextrin (1.0 g/kg) with alanine (0.225 g/kg) and proline (0.025 g/kg) 15 min before running for 170 min. Combined supplementation of maltodextrin, alanine, and proline induced higher blood glucose levels than isocaloric maltodextrin alone during the late exercise phase (100-170 min). The hepatic glycogen content of mice administered maltodextrin, alanine, and proline was higher than that of mice ingesting maltodextrin alone 60 min after beginning exercise, but the glycogen content of the gastrocnemius muscle showed no difference. We conducted a treadmill running test to determine the effect of alanine and proline on endurance performance. The test showed that running time to exhaustion of mice that were supplemented with maltodextrin (2.0 g/kg) was longer than that of mice that were supplemented with water alone. Maltodextrin supplementation (1.0 g/kg) with alanine (0.9 g/kg) and proline (0.1 g/kg) further increased running time to exhaustion compared to maltodextrin alone (2.0 g/kg). These results indicate that combined supplementation of carbohydrate, alanine, and proline is effective for maintaining blood glucose and hepatic glycogen levels and increasing endurance performance during long-term exercise in mice.

  12. The influence of secretion elicitors and external pH on the kinetics of D-alanine uptake by the trap lobes of Dionaea muscipula Ellis (Venus's Flytrap).

    PubMed

    Rea, P A; Whatley, F R

    1983-08-01

    Simple kinetic techniques were used to examine the mechanism of D-alanine uptake by the adaxial surfaces of the trap lobes of Dionaea muscipula Ellis (Venus's Flytrap.) On the basis of these analyses, the uptake of D-alanine was found to depend on the time during which the trap lobes were inoculated with elicitors of secretion before excision and measurement of uptake. Disks taken from traps that had not been subjected to a preceding period of inoculation with secretion elicitors showed a low basal rate of uptake which was neither pH-dependent nor exhibited saturation with respect to external D-alanine concentration. Disks from preinoculated traps, on the other hand, displayed an enhanced rate of uptake which showed both pH-dependence and saturation with respect to external D-alanine concentration. The capacity for enhanced uptake was lost upon prolonged inoculation or when inoculation was stopped. Of the compounds tested, only elicitors of secretion caused an enhancement of uptake. The enhanced rate of D-alanine uptake is temperature-sensitive with a Q10 characteristic of a mediated process. Uncouplers cause an instantaneous abolition of uptake whereas the effects of terminal-oxidase inhibitors are time-dependent. The pH-dependence of uptake is inferred to result from an increased affinity of the carrier system for D-alanine at low pH values. Although the ionic state of D-alanine is relatively unaffected over the pH range examined, a decrease in the external pH from 6.0 to 3.8 decreases the apparent K m for uptake by four-fold but increases V max by only 30%. It is concluded that the acid secreted by the digestive glands of Dionaea plays a direct role in facilitating the uptake of amino acids from the trap cavity.

  13. Averaged electron collision cross sections for thermal mixtures of \\alpha -alanine conformers in the gas phase

    NASA Astrophysics Data System (ADS)

    Fujimoto, Milton M.; de Lima, Erik V. R.; Tennyson, Jonathan

    2016-11-01

    A theoretical study of elastic electron collisions with 9 conformers of the gas-phase amino acid α-alanine (CH3CH(NH2)COOH) is performed. The eigenphase sums, resonance features, differential and integral cross sections are computed for each individual conformer. Resonance positions for the low-energy {π }* shape resonance are found to vary from 2.6 to 3.1 eV and the resonance widths from 0.3 to 0.5 eV. Averaged cross sections for thermal mixtures of the 9 conformers are presented. Both theoretical and experimental population ratios are considered. Thermally averaged cross sections obtained using the best theoretical estimates give reasonable agreement with the observed thermal cross sections. Excited conformers IIA and IIB make a large contribution to this average due to their large permanent dipole moments.

  14. 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.

  15. On the existence of ``l-threonine formate'', ``l-alanine lithium chloride'' and ``bis l-alanine lithium chloride'' crystals

    NASA Astrophysics Data System (ADS)

    Petrosyan, A. M.; Ghazaryan, V. V.; Fleck, M.

    2013-03-01

    We argue that the recently reported crystals "L-threonine formate" as well as "L-alanine lithium chloride" and "bis L-alanine lithium chloride" actually are the well-known crystals L-threonine and L-alanine, respectively.

  16. First-principles studies of pure and fluorine substituted alanines

    NASA Astrophysics Data System (ADS)

    Ahmad, Sardar; Vaizie, Hamide; Rahnamaye Aliabad, H. A.; Ahmad, Rashid; Khan, Imad; Ali, Zahid; Jalali-Asadabadi, S.; Ahmad, Iftikhar; Khan, Amir Abdullah

    2016-05-01

    This paper communicates the structural, electronic and optical properties of L-alanine, monofluoro and difluoro substituted alanines using density functional calculations. These compounds exist in orthorhombic crystal structure and the calculated structural parameters such as lattice constants, bond angles and bond lengths are in agreement with the experimental results. L-alanine is an indirect band gap insulator, while its fluorine substituted compounds (monofluoroalanine and difluoroalanine) are direct band gap insulators. The substitution causes reduction in the band gap and hence these optically tailored direct wide band gap materials have enhanced optical properties in the ultraviolet (UV) region of electromagnetic spectrum. Therefore, optical properties like dielectric function, refractive index, reflectivity and energy loss function are also investigated. These compounds have almost isotropic nature in the lower frequency range while at higher energies, they have a significant anisotropic nature.

  17. Engineering of alanine dehydrogenase from Bacillus subtilis for novel cofactor specificity.

    PubMed

    Lerchner, Alexandra; Jarasch, Alexander; Skerra, Arne

    2016-09-01

    The l-alanine dehydrogenase of Bacillus subtilis (BasAlaDH), which is strictly dependent on NADH as redox cofactor, efficiently catalyzes the reductive amination of pyruvate to l-alanine using ammonia as amino group donor. To enable application of BasAlaDH as regenerating enzyme in coupled reactions with NADPH-dependent alcohol dehydrogenases, we alterated its cofactor specificity from NADH to NADPH via protein engineering. By introducing two amino acid exchanges, D196A and L197R, high catalytic efficiency for NADPH was achieved, with kcat /KM  = 54.1 µM(-1)  Min(-1) (KM  = 32 ± 3 µM; kcat  = 1,730 ± 39 Min(-1) ), almost the same as the wild-type enzyme for NADH (kcat /KM  = 59.9 µM(-1)  Min(-1) ; KM  = 14 ± 2 µM; kcat  = 838 ± 21 Min(-1) ). Conversely, recognition of NADH was much diminished in the mutated enzyme (kcat /KM  = 3 µM(-1)  Min(-1) ). BasAlaDH(D196A/L197R) was applied in a coupled oxidation/transamination reaction of the chiral dicyclic dialcohol isosorbide to its diamines, catalyzed by Ralstonia sp. alcohol dehydrogenase and Paracoccus denitrificans ω-aminotransferase, thus allowing recycling of the two cosubstrates NADP(+) and l-Ala. An excellent cofactor regeneration with recycling factors of 33 for NADP(+) and 13 for l-Ala was observed with the engineered BasAlaDH in a small-scale biocatalysis experiment. This opens a biocatalytic route to novel building blocks for industrial high-performance polymers.

  18. GMXPBSA 2.1: A GROMACS tool to perform MM/PBSA and computational alanine scanning

    NASA Astrophysics Data System (ADS)

    Paissoni, C.; Spiliotopoulos, D.; Musco, G.; Spitaleri, A.

    2015-01-01

    GMXPBSA 2.1 is a user-friendly suite of Bash/Perl scripts for streamlining MM/PBSA calculations on structural ensembles derived from GROMACS trajectories, to automatically calculate binding free energies for protein-protein or ligand-protein complexes [R.T. Bradshaw et al., Protein Eng. Des. Sel. 24 (2011) 197-207]. GMXPBSA 2.1 is flexible and can easily be customized to specific needs and it is an improvement of the previous GMXPBSA 2.0 [C. Paissoni et al., Comput. Phys. Commun. (2014), 185, 2920-2929]. Additionally, it performs computational alanine scanning (CAS) to study the effects of ligand and/or receptor alanine mutations on the free energy of binding. Calculations require only for protein-protein or protein-ligand MD simulations. GMXPBSA 2.1 performs different comparative analyses, including a posteriori generation of alanine mutants of the wild-type complex, calculation of the binding free energy values of the mutant complexes and comparison of the results with the wild-type system. Moreover, it compares the binding free energy of different complex trajectories, allowing the study of the effects of non-alanine mutations, post-translational modifications or unnatural amino acids on the binding free energy of the system under investigation. Finally, it can calculate and rank relative affinity to the same receptor utilizing MD simulations of proteins in complex with different ligands. In order to dissect the different MM/PBSA energy contributions, including molecular mechanic (MM), electrostatic contribution to solvation (PB) and nonpolar contribution to solvation (SA), the tool combines two freely available programs: the MD simulations software GROMACS [S. Pronk et al., Bioinformatics 29 (2013) 845-854] and the Poisson-Boltzmann equation solver APBS [N.A. Baker et al., Proc. Natl. Acad. Sci. U.S.A 98 (2001) 10037-10041]. All the calculations can be performed in single or distributed automatic fashion on a cluster facility in order to increase the

  19. Specific lysosomal transport of small neutral amino acids

    SciTech Connect

    Pisoni, R.L.; Flickinger, K.S.; Thoene, J.G.; Christensen, H.N.

    1986-05-01

    Studies of amino acid exodus from lysosomes have allowed us previously to describe transport systems specific for cystine and another for cationic amino acids in fibroblast lysosomes. They are now able to study amino acid uptake into highly purified fibroblast lysosomes obtained by separating crude granular fraction on gradients formed by centrifugation in 35% isoosmotic Percoll solutions. Analog inhibition and saturation studies indicate that L-(/sup 14/C)proline (50 ..mu..M) uptake by fibroblast lysosomes at 37/sup 0/C in 50 mM citrate/tris pH 7.0 buffer containing 0.25 M sucrose is mediated by two transport systems, one largely specific for L-proline and the other for which transport is shared with small neutral amino acids such as alanine, serine and threonine. At 7 mM, L-proline inhibits L-(/sup 14/C)proline uptake almost completely, whereas ala, ser, val, thr, gly, N-methylalanine and sarcosine inhibit proline uptake by 50-65%. The system shared by alanine, serine and threonine is further characterized by these amino acids strongly inhibiting the uptakes of each other. Lysosomal proline transport is selective for the L-isomer of the amino acid, and is scarcely inhibited by 7 mM arg, glu, asp, leu, phe, his, met, (methylamino) isobutyrate, betaine or N,N-dimethylglycine. Cis or trans-4-hydroxy-L-proline inhibit proline uptake only slightly. In sharp contrast to the fibroblast plasma membrane in which Na/sup +/ is required for most proline and alanine transport, lysosomal uptake of these amino acids occurs independently of Na/sup +/.

  20. Polymers with complexing properties. Simple poly(amino acids)

    NASA Technical Reports Server (NTRS)

    Roque, J. M.

    1978-01-01

    The free amino (0.3 equiv/residue) and carboxyl (0.5 equiv/residue) groups of thermal polylysine increased dramatically on treatment with distilled water. The total hydrolysis of such a polymer was abnormal in that only about 50% of the expected amino acids were recovered. Poly (lysine-co-alanine-co-glycine) under usual conditions hydrolyzed completely in 8 hours; whereas, when it was pretreated with diazomethane, a normal period of 24 hours was required to give (nearly) the same amounts of each free amino acid as compared with those obtained from the untreated polymer. The amino groups of the basic thermal poly(amino acids) were sterically hindered. The existence of nitrogen atoms linking two or three chains and reactive groups (anhydride, imine) were proposed.

  1. Transport of Amino Acids to the Maize Root 1

    PubMed Central

    Oaks, Ann

    1966-01-01

    When 5-mm maize root tips were excised and placed in an inorganic salts solution for 6 hours, there was a loss of alcohol-insoluble nitrogen. The levels of threonine, proline, valine, isoleucine, leucine, tyrosine, phenylalanine, and lysine in the alcohol soluble fraction were severely reduced, whereas those of glutamate, aspartate, ornithine, and alanine were scarcely affected. There was a 4-fold increase in the level of γ-aminobutyrate. Those amino acids whose synthesis appeared to be deficient in excised root tips also showed poor incorporation of acetate carbon. In addition, the results show that asparagine and the amino acids of the neutral and basic fraction were preferentially transported to the root tip region. The results therefore suggest that the synthesis of certain amino acids in the root tip region is restricted, and that this requirement for amino acids in the growing region could regulate the flow of amino acids to the root tip. PMID:16656225

  2. Polysulfone affinity membranes for the treatment of amino acid mixtures.

    PubMed

    Rodemann, K; Staude, E

    1995-06-20

    Affinity membranes for the treatment of solutions containing amino acids were obtained via lithiating polysulfone that was subsequently converted with glycidylether. From this polymer asymmetric ultrafiltration membranes were cast. The membranes were reacted with iminodiacetic acid yielding membranes fitted out with bidentate chelates. The same reaction path was applied to commercially available symmetric microfiltration membranes. The chelate-bearing membranes were complexed with Cu, Ni, and Zn ions. For the experiments with amino acids only the Cu-complexed membranes were used. The complexation constants for histidine and tryptophan for six different membranes were determined. Because of the affinity of these two amino acids for the complexed Cu ions, they could easily be separated from solutions containing amino acids such as alanine, glycine, and valine. Also, concentrating very dilute amino acid solutions was carried out successfully.

  3. Elevational Variation in Soil Amino Acid and Inorganic Nitrogen Concentrations in Taibai Mountain, China

    PubMed Central

    Yang, Xin; Zhu, Lianfeng; Zhang, Junhua; Jin, Qianyu; Wu, Lianghuan

    2016-01-01

    Amino acids are important sources of soil organic nitrogen (N), which is essential for plant nutrition, but detailed information about which amino acids predominant and whether amino acid composition varies with elevation is lacking. In this study, we hypothesized that the concentrations of amino acids in soil would increase and their composition would vary along the elevational gradient of Taibai Mountain, as plant-derived organic matter accumulated and N mineralization and microbial immobilization of amino acids slowed with reduced soil temperature. Results showed that the concentrations of soil extractable total N, extractable organic N and amino acids significantly increased with elevation due to the accumulation of soil organic matter and the greater N content. Soil extractable organic N concentration was significantly greater than that of the extractable inorganic N (NO3−-N + NH4+-N). On average, soil adsorbed amino acid concentration was approximately 5-fold greater than that of the free amino acids, which indicates that adsorbed amino acids extracted with the strong salt solution likely represent a potential source for the replenishment of free amino acids. We found no appreciable evidence to suggest that amino acids with simple molecular structure were dominant at low elevations, whereas amino acids with high molecular weight and complex aromatic structure dominated the high elevations. Across the elevational gradient, the amino acid pool was dominated by alanine, aspartic acid, glycine, glutamic acid, histidine, serine and threonine. These seven amino acids accounted for approximately 68.9% of the total hydrolyzable amino acid pool. The proportions of isoleucine, tyrosine and methionine varied with elevation, while soil major amino acid composition (including alanine, arginine, aspartic acid, glycine, histidine, leucine, phenylalanine, serine, threonine and valine) did not vary appreciably with elevation (p>0.10). The compositional similarity of many

  4. N-(1-carboxyethyl)alanine (alanopine), a new non-sugar component of lipopolysaccharides of Providencia and Proteus.

    PubMed

    Kocharova, Nina A; Kondakova, Anna N; Ovchinnikova, Olga G; Perepelov, Andrei V; Shashkov, Alexander S; Knirel, Yuriy A

    2009-10-12

    O-Polysaccharides were released by mild acid degradation of lipopolysaccharides of Providencia alcalifaciens O35 and Proteus vulgaris O76 and were studied by 1D and 2D (1)H and (13)C NMR spectroscopies, including HMBC and NOESY (ROESY) experiments. Both polysaccharides were found to contain N-(1-carboxyethyl)alanine (alanopine) that is N-linked to 4-amino-4,6-dideoxyglucose. Analysis of published data [Vinogradov, E.; Perry, M. B. Eur. J. Biochem.2000, 267, 2439-2446] shows that alanopine is present also on the same sugar in the lipopolysaccharide core of Proteus mirabilis O6 and O57.

  5. Growth, spectroscopic, dielectric and nonlinear optical studies of semi organic nonlinear optical crystal - L-Alanine lithium chloride

    NASA Astrophysics Data System (ADS)

    Hanumantharao, Redrothu; Kalainathan, S.

    2012-02-01

    A new and efficient semi organic nonlinear optical crystal (NLO) from the amino acid family L-alanine lithium chloride (LAL) has been grown by slow evaporation technique from aqueous solution. The functional groups were identified from NMR spectral studies. Mass spectral analysis shows the molecular ion mass. Dielectric studies has been done for the grown crystal and relative SHG efficiency is measured by Kurtz and Perry method and found to about 0.43 times that of standard potassium dihydrogen phosphate (KDP) crystals. The compound crystallized in non-centrosymmetric space group Pna21. The results have been discussed in detail.

  6. Growth, spectroscopic, dielectric and nonlinear optical studies of semi organic nonlinear optical crystal--L-alanine lithium chloride.

    PubMed

    Hanumantharao, Redrothu; Kalainathan, S

    2012-02-01

    A new and efficient semi organic nonlinear optical crystal (NLO) from the amino acid family L-alanine lithium chloride (LAL) has been grown by slow evaporation technique from aqueous solution. The functional groups were identified from NMR spectral studies. Mass spectral analysis shows the molecular ion mass. Dielectric studies has been done for the grown crystal and relative SHG efficiency is measured by Kurtz and Perry method and found to about 0.43 times that of standard potassium dihydrogen phosphate (KDP) crystals. The compound crystallized in non-centrosymmetric space group Pna21. The results have been discussed in detail.

  7. Immunocytoma effect upon circadian variation in murine urinary excretion of beta-aminoisobutyric acid, beta-alanine, phenylalanine and tyrosine.

    PubMed

    Halberg, F; Gehrke, C W; Kuo, K; Nelson, W L; Sothern, R B; Cadotte, L M; Haus, E; Scheving, L E

    1978-01-01

    Under the conditions of disynchronization by the manipulation of both the alternation of light and darkness and the availability and unavailability of food, circadian rhythms characterize the excretion of several amino acids by inbred LOU rats bearing an immunocytoma. Large amplitude rhythms can be demonstrated for urinary beta-aminoisobutyric acid, beta-alanine, phenylalanine and tyrosine. Under the same conditions of disynchronization, control animals excrete the same compounds also with a marked circadian variation but at an invariably lower average rate. These excretory rhythms, along with those demonstrated earlier for polyamines and light-chains, are of interest as potential markers for the chronotherapy of cancer.

  8. ANTIGENICITY OF POLYPEPTIDES (POLY ALPHA AMINO ACIDS)

    PubMed Central

    Maurer, Paul H.; Gerulat, Bernard F.; Pinchuck, Paul

    1964-01-01

    A new group of synthetic random polymers of α-L-amino acids has been studied for immunogenicity. With the glutamic acid and alanine copolymers, those consisting of almost equimolar amounts of the two (G60A40 and G40A60) were effective antigens in rabbits whereas those with higher glutamic acid contents (G75A25, G90A10) were poor antigens. The substitution of alanine by valine or leucine (G75V25 and G80Leu20) produced copolymers which were poor antigens in rabbits but effective in guinea pigs. L70A30, although capable of "non-specifically" precipitating serum proteins, was shown not to be antigenic in either rabbits or guinea pigs. The introduction of alanine into glutamic acid and lysine polymers (GLA series) enhanced the immunogenicity of the terpolymers, i.e., GLA30 > GLA20 > GLA10 > GL. The mechanism by which this may be accomplished is discussed as possibly being related to the reduction of the interactions between glutamyl and lysyl residues which allows the carboxyl groups to act as strong immunogenic determinants. PMID:14176288

  9. The origin of amino acids in lunar regolith samples

    NASA Astrophysics Data System (ADS)

    Elsila, Jamie E.; Callahan, Michael P.; Dworkin, Jason P.; Glavin, Daniel P.; McLain, Hannah L.; Noble, Sarah K.; Gibson, Everett K.

    2016-01-01

    We analyzed the amino acid content of seven lunar regolith samples returned by the Apollo 16 and Apollo 17 missions and stored under NASA curation since collection using ultrahigh-performance liquid chromatography with fluorescence detection and time-of-flight mass spectrometry. Consistent with results from initial analyses shortly after collection in the 1970s, we observed amino acids at low concentrations in all of the curated samples, ranging from 0.2 parts-per-billion (ppb) to 42.7 ppb in hot-water extracts and 14.5-651.1 ppb in 6 M HCl acid-vapor-hydrolyzed, hot-water extracts. Amino acids identified in the Apollo soil extracts include glycine, D- and L-alanine, D- and L-aspartic acid, D- and L-glutamic acid, D- and L-serine, L-threonine, and L-valine, all of which had previously been detected in lunar samples, as well as several compounds not previously identified in lunar regoliths: α-aminoisobutyric acid (AIB), D- and L-β-amino-n-butyric acid (β-ABA), DL-α-amino-n-butyric acid, γ-amino-n-butyric acid, β-alanine, and ε-amino-n-caproic acid. We observed an excess of the L enantiomer in most of the detected proteinogenic amino acids, but racemic alanine and racemic β-ABA were present in some samples. We also examined seven samples from Apollo 15, 16, and 17 that had been previously allocated to a non-curation laboratory, as well as two samples of terrestrial dunite from studies of lunar module engine exhaust that had been stored in the same laboratory. The amino acid content of these samples suggested that contamination had occurred during non-curatorial storage. We measured the compound-specific carbon isotopic ratios of glycine, β-alanine, and L-alanine in Apollo regolith sample 70011 and found values of -21‰ to -33‰. These values are consistent with those seen in terrestrial biology and, together with the enantiomeric compositions of the proteinogenic amino acids, suggest that terrestrial biological contamination is a primary source of the

  10. Stereoselective aminoacylation of a dinucleoside monophosphate by the imidazolides of DL-alanine and N-(tert-butoxycarbonyl)-DL-alanine

    NASA Technical Reports Server (NTRS)

    Profy, A. T.; Usher, D. A.

    1984-01-01

    The aminoacylation of diinosine monophosphate was studied experimentally. When the acylating agent was the imidazolide of N-(tert-butoxycarbonyl)-DL-alanine, a 40 percent enantiomeric excess of the isomer was incorporated at the 2' site and the positions of equilibrium for the reversible 2'-3' migration reaction differed for the D and L enantiomers. The reactivity of the nucleoside hydroxyl groups was found to decrease on the order 2'(3') less than internal 2' and less than 5', and the extent of the reaction was affected by the concentration of the imidazole buffer. Reaction of IpI with imidazolide of unprotected DL-alanine, by contrast, led to an excess of the D isomer at the internal 2' site. Finally, reaction with the N-carboxy anhydride of DL-alanine occurred without stereoselection. These results are found to be relevant to the study of the evolution of optical chemical activity and the origin of genetically directed protein synthesis.

  11. D-Amino Acids in the Nervous and Endocrine Systems

    PubMed Central

    Kiriyama, Yoshimitsu

    2016-01-01

    Amino acids are important components for peptides and proteins and act as signal transmitters. Only L-amino acids have been considered necessary in mammals, including humans. However, diverse D-amino acids, such as D-serine, D-aspartate, D-alanine, and D-cysteine, are found in mammals. Physiological roles of these D-amino acids not only in the nervous system but also in the endocrine system are being gradually revealed. N-Methyl-D-aspartate (NMDA) receptors are associated with learning and memory. D-Serine, D-aspartate, and D-alanine can all bind to NMDA receptors. H2S generated from D-cysteine reduces disulfide bonds in receptors and potentiates their activity. Aberrant receptor activity is related to diseases of the central nervous system (CNS), such as Alzheimer's disease, amyotrophic lateral sclerosis, and schizophrenia. Furthermore, D-amino acids are detected in parts of the endocrine system, such as the pineal gland, hypothalamus, pituitary gland, pancreas, adrenal gland, and testis. D-Aspartate is being investigated for the regulation of hormone release from various endocrine organs. Here we focused on recent findings regarding the synthesis and physiological functions of D-amino acids in the nervous and endocrine systems. PMID:28053803

  12. [Regulation of key enzymes of L-alanine biosynthesis by Brevibacterium flavum producer strains].

    PubMed

    Melkonian, L O; Avetisova, G E; Ambartsumian, A A; Chakhalian, A Kh; Sagian, A S

    2013-01-01

    The mechanisms of L-alanine overproduction by Brevibacterium flavum producer strains were studied. It was shown that beta-CI-L-alanine is an inhibitor of some key enzymes involved in the synthesis of L-alanine, including alanine transaminase and valine-pyruvate transaminase. Two highly active B. flavum GL1 and GL1 8 producer strains, which are resistant to the inhibitory effect of beta-Cl-L-alanine, were obtained using a parental B. flavum AA5 producer strain, characterized by a reduced activity of alanine racemase (>or=98%). It was demonstrated that the increased L-alanine synthesis efficiency observed in the producer strains developed in this work is associated with the absence of inhibition of alanine transaminase by the end product of the biosynthesis reaction, as well as with the effect of derepression of both alanine transaminase and valine-pyruvate transaminase synthesis by the studied compound.

  13. Dipeptide Nanotubes Containing Unnatural Fluorine-Substituted β(2,3)-Diarylamino Acid and L-Alanine as Candidates for Biomedical Applications.

    PubMed

    Bonetti, Andrea; Pellegrino, Sara; Das, Priyadip; Yuran, Sivan; Bucci, Raffaella; Ferri, Nicola; Meneghetti, Fiorella; Castellano, Carlo; Reches, Meital; Gelmi, Maria Luisa

    2015-09-18

    The synthesis and the structural characterization of dipeptides composed of unnatural fluorine-substituted β(2,3)-diarylamino acid and L-alanine are reported. Depending on the stereochemistry of the β amino acid, these dipeptides are able to self-assemble into proteolytic stable nanotubes. These architectures were able to enter the cell and locate in the cytoplasmic/perinuclear region and represent interesting candidates for biomedical applications.

  14. STT3, a highly conserved protein required for yeast oligosaccharyl transferase activity in vivo.

    PubMed Central

    Zufferey, R; Knauer, R; Burda, P; Stagljar, I; te Heesen, S; Lehle, L; Aebi, M

    1995-01-01

    N-linked glycosylation is a ubiquitous protein modification, and is essential for viability in eukaryotic cells. A lipid-linked core-oligosaccharide is assembled at the membrane of the endoplasmic reticulum and transferred to selected asparagine residues of nascent polypeptide chains by the oligosaccharyl transferase (OTase) complex. Based on the synthetic lethal phenotype of double mutations affecting the assembly of the lipid-linked core-oligosaccharide and the OTase activity, we have performed a novel screen for mutants in Saccharomyces cerevisiae with altered N-linked glycosylation. Besides novel mutants deficient in the assembly of the lipid-linked oligosaccharide (alg mutants), we identified the STT3 locus as being required for OTase activity in vivo. The essential STT3 protein is approximately 60% identical in amino acid sequence to its human homologue. A mutation in the STT3 locus affects substrate specificity of the OTase complex in vivo and in vitro. In stt3-3 cells very little glycosyl transfer occurs from incomplete lipid-linked oligosaccharide, whereas the transfer of full-length Glc3Man9GlcNAc2 is hardly affected as compared with wild-type cells. Depletion of the STT3 protein results in loss of transferase activity in vivo and a deficiency in the assembly of OTase complex. Images PMID:7588624

  15. Oligomerization of glycine and alanine on metal(II) octacynaomolybdate(IV): role of double metal cyanides in prebiotic chemistry.

    PubMed

    Kumar, Anand; Kamaluddin

    2012-12-01

    Condensation reactions of amino acid (glycine and alanine) on the surface of metal(II) octacyanomolybdate(IV) (MOCMo) complexes are investigated using high-performance liquid chromatography (HPLC) and electron spray ionizations-mass spectroscopy (ESI-MS). The series of MOCMo have been synthesized and the effect of outer sphere metal ions present in the MOCMo on the oligomerization of glycine and alanine at different temperature and time found out. Formation of peptides was observed to start after 7 days at 60 °C. Maximum yield of peptides was found after 35 days at 90 °C. It has been found that zinc(II) octacyanomolybdate(IV) and cobalt(II) were the most effective metal cations present in outer sphere of the MOCMo for the production of high yield of oligomerized products. Surface area of MOCMo seems to play dominating parameter for the oligomerization of alanine and glycine. The results of the present study reveal the role of MOCMo in chemical evolution for the oligomerization of biomolecules.

  16. Amino acids derived from Titan tholins

    NASA Technical Reports Server (NTRS)

    Khare, B. N.; Sagan, C.; Ogino, H.; Nagy, B.; Er, C.; Schram, K. H.; Arakawa, E. T.

    1986-01-01

    An organic heteropolymer (Titan tholin) was produced by continuous dc discharge through a 0.9 N2/0.1 CH4 gas mixture at 0.2 mbar pressure, roughly simulating the cloudtop atmosphere of Titan. Treatment of this tholin with 6N HCl yielded 16 amino acids by gas chromatography after derivatization of N-trifluroacetyl isopropyl esters on two different capillary columns. Identifications were confirmed by GC/MS. Glycine, aspartic acid, and alpha- and beta-alanine were produced in greatest abundance; the total yield of amino acids was approximately 10(-2), approximately equal to the yield of urea. The presence of "nonbiological" amino acids, the absence of serine, and the fact that the amino acids are racemic within experimental error together indicate that these molecules are not due to microbial or other contamination, but are derived from the tholin. In addition to the HCN, HC2CN, and (CN)2 found by Voyager, nitriles and aminonitriles should be sought in the Titanian atmosphere and, eventually, amino acids on the surface. These results suggest that episodes of liquid water in the past or future of Titan might lead to major further steps in prebiological organic chemistry on that body.

  17. Deuterium Fractionation during Amino Acid Formation by Photolysis of Interstellar Ice Analogs Containing Deuterated Methanol

    NASA Astrophysics Data System (ADS)

    Oba, Yasuhiro; Takano, Yoshinori; Watanabe, Naoki; Kouchi, Akira

    2016-08-01

    Deuterium (D) atoms in interstellar deuterated methanol might be distributed into complex organic molecules through molecular evolution by photochemical reactions in interstellar grains. In this study, we use a state-of-the-art high-resolution mass spectrometer coupled with a high-performance liquid chromatography system to quantitatively analyze amino acids and their deuterated isotopologues formed by the photolysis of interstellar ice analogs containing singly deuterated methanol CH2DOH at 10 K. Five amino acids (glycine, α-alanine, β-alanine, sarcosine, and serine) and their deuterated isotopologues whose D atoms are bound to carbon atoms are detected in organic residues formed by photolysis followed by warming up to room temperature. The abundances of singly deuterated amino acids are in the range of 0.3-1.1 relative to each nondeuterated counterpart, and the relative abundances of doubly and triply deuterated species decrease with an increasing number of D atoms in a molecule. The abundances of amino acids increase by a factor of more than five upon the hydrolysis of the organic residues, leading to decreases in the relative abundances of deuterated species for α-alanine and β-alanine. On the other hand, the relative abundances of the deuterated isotopologues of the other three amino acids did not decrease upon hydrolysis, indicating different formation mechanisms of these two groups upon hydrolysis. The present study facilitates both qualitative and quantitative evaluations of D fractionation during molecular evolution in the interstellar medium.

  18. Formation of {gamma}-alumina nanorods in presence of alanine

    SciTech Connect

    Dabbagh, Hossein A.; Rasti, Elham; Yalfani, Mohammad S.; Medina, Francesc

    2011-02-15

    Graphical abstract: Nanorod aluminas with a possible hexagonal symmetry, high surface area and relatively narrow pore size distribution were obtained. Research highlights: {yields} Research highlights {yields} Boehmite was prepared using a green sol-gel process in the presence of alanine. {yields} Nanorod aluminas with a high surface area were obtained. {yields} Addition of alanine would shape the size of the holes and crevices. {yields} The morphologies of the nanorods were revealed by transmission electron microscope. -- Abstract: Boehmite and alumina nanostructures were prepared using a simple green sol-gel process in the presence of alanine in water medium at room temperature. The uncalcined (dried at 200 {sup o}C) and the calcined materials (at 500, 600 and 700 {sup o}C for 4 h) were characterized using XRD, TEM, SEM, N{sub 2} physisorption and TGA. Nanorod aluminas with a possible hexagonal symmetry, high surface area and relatively narrow pore size distribution were obtained. The surface area was enhanced and crystallization was retarded as the alanine content increased. The morphologies of the nanoparticles and nanorods were revealed by a transmission electron microscope (TEM).

  19. A theoretical study of alanine dipeptide and analogs

    SciTech Connect

    Head-Gordon, T.; Head-Gordon, M.; Brooks, C. III; Pople, J. ); Frisch, M.J. )

    1989-01-01

    We Present a preliminary report on the conformational and energetic analysis of the molecule (S)-2-acetylamino-N-methylpropanamide (alanine dipeptide) and an analog molecule, (S)-{alpha}-formylaminopropanamide, using high-quality ab initio methods. Alanine dipeptide and its analogs are of interest since they incorporate many of the structural features found in proteins, such as intramolecular hydrogen bonds, conformational flexibility, and a variety of chemical functionality. One purpose of this study is to provide a useful benchmark calculation, MP2/6-31+G{sup **}//HF/6-31+G{sup *}, for a number of conformations of the alanine system. Based on the comparison of these benchmark calculations with lower-level basis sets, HF/3-21G was chosen to generate a fully relaxed {phi}, {psi} dihedral map. These calculations are the first of their kind on systems of this size. Features of the {phi},{psi} alanine dipeptide map that are discussed include the energetically accessible conformations and possible pathways for their interconversion. In addition, we illustrate the importance of fully optimized geometries and the proper evaluation of correlation energies,

  20. Spectrophotometric readout for an alanine dosimeter for food irradiation applications

    NASA Astrophysics Data System (ADS)

    Ebraheem, S.; Beshir, W. B.; Eid, S.; Sobhy, R.; Kovács, A.

    2003-06-01

    The alanine-electron spin resonance (EPR) readout system is well known as a reference and transfer dosimetry system for the evaluation of high doses in radiation processing. The high cost of an EPR/alanine dosimetry system is a serious handicap for large-scale routine application in irradiation facilities. In this study, the use of a complex produced by dissolving irradiated L-alanine in 1,4-phenyl diammonium dichloride solution was investigated for dosimetry purposes. This complex—having a purple colour—has an increasing absorbance with increasing dose in the range of 1-20 kGy. The applicability of spectrophotometric evaluation was studied by measuring the absorbance intensity of this complex at 360 and 505 nm, respectively. Fluorimetric evaluation was also investigated by measuring the emission of the complex at 435 nm as a function of dose. The present method is easy for routine application. The effect of the dye concentration as well as the suitable amount of irradiated alanine has been studied. With respect to routine application, the stability of the product complex after its formation was also investigated.

  1. Computational alanine scanning with linear scaling semiempirical quantum mechanical methods.

    PubMed

    Diller, David J; Humblet, Christine; Zhang, Xiaohua; Westerhoff, Lance M

    2010-08-01

    Alanine scanning is a powerful experimental tool for understanding the key interactions in protein-protein interfaces. Linear scaling semiempirical quantum mechanical calculations are now sufficiently fast and robust to allow meaningful calculations on large systems such as proteins, RNA and DNA. In particular, they have proven useful in understanding protein-ligand interactions. Here we ask the question: can these linear scaling quantum mechanical methods developed for protein-ligand scoring be useful for computational alanine scanning? To answer this question, we assembled 15 protein-protein complexes with available crystal structures and sufficient alanine scanning data. In all, the data set contains Delta Delta Gs for 400 single point alanine mutations of these 15 complexes. We show that with only one adjusted parameter the quantum mechanics-based methods outperform both buried accessible surface area and a potential of mean force and compare favorably to a variety of published empirical methods. Finally, we closely examined the outliers in the data set and discuss some of the challenges that arise from this examination.

  2. Electrostatic study of Alanine mutational effects on transcription: application to GATA-3:DNA interaction complex.

    PubMed

    El-Assaad, Atlal; Dawy, Zaher; Nemer, Georges

    2015-01-01

    Protein-DNA interaction is of fundamental importance in molecular biology, playing roles in functions as diverse as DNA transcription, DNA structure formation, and DNA repair. Protein-DNA association is also important in medicine; understanding Protein-DNA binding kinetics can assist in identifying disease root causes which can contribute to drug development. In this perspective, this work focuses on the transcription process by the GATA Transcription Factor (TF). GATA TF binds to DNA promoter region represented by `G,A,T,A' nucleotides sequence, and initiates transcription of target genes. When proper regulation fails due to some mutations on the GATA TF protein sequence or on the DNA promoter sequence (weak promoter), deregulation of the target genes might lead to various disorders. In this study, we aim to understand the electrostatic mechanism behind GATA TF and DNA promoter interactions, in order to predict Protein-DNA binding in the presence of mutations, while elaborating on non-covalent binding kinetics. To generate a family of mutants for the GATA:DNA complex, we replaced every charged amino acid, one at a time, with a neutral amino acid like Alanine (Ala). We then applied Poisson-Boltzmann electrostatic calculations feeding into free energy calculations, for each mutation. These calculations delineate the contribution to binding from each Ala-replaced amino acid in the GATA:DNA interaction. After analyzing the obtained data in view of a two-step model, we are able to identify potential key amino acids in binding. Finally, we applied the model to GATA-3:DNA (crystal structure with PDB-ID: 3DFV) binding complex and validated it against experimental results from the literature.

  3. Nutrient uptake by marine invertebrates: cloning and functional analysis of amino acid transporter genes in developing sea urchins (Strongylocentrotus purpuratus).

    PubMed

    Meyer, Eli; Manahan, Donal T

    2009-08-01

    Transport of amino acids from low concentrations in seawater by marine invertebrates has been extensively studied, but few of the genes involved in this physiological process have been identified. We have characterized three amino acid transporter genes cloned from embryos of the sea urchin Strongylocentrotus purpuratus. These genes show phylogenetic proximity to classical amino acid transport systems, including Gly and B0+, and the inebriated gene (INE). Heterologous expression of these genes in frog oocytes induced a 40-fold increase in alanine transport above endogenous levels, demonstrating that these genes mediate alanine transport. Antibodies specific to one of these genes (Sp-AT1) inhibited alanine transport, confirming the physiological activity of this gene in larvae. Whole-mount antibody staining of larvae revealed expression of Sp-AT1 in the ectodermal tissues associated with amino acid transport, as independently demonstrated by autoradiographic localization of radioactive alanine. Maximum rates of alanine transport increased 6-fold during early development, from embryonic to larval stages. Analysis of gene expression during this developmental period revealed that Sp-AT1 transcript abundance remained nearly constant, while that of another transporter gene (Sp-AT2) increased 11-fold. The functional characterization of these genes establishes a molecular biological basis for amino acid transport by developmental stages of marine invertebrates.

  4. Biochemical properties of an omega-class glutathione S-transferase of the silkmoth, Bombyx mori.

    PubMed

    Yamamoto, Kohji; Nagaoka, Sumiharu; Banno, Yutaka; Aso, Yoichi

    2009-05-01

    A cDNA encoding an omega-class glutathione S-transferase of the silkmoth, Bombyx mori (bmGSTO), was cloned by reverse transcriptase-polymerase chain reaction. The resulting clone was sequenced and deduced for amino acid sequence, which revealed 40, 40, and 39% identities to omega-class GSTs from human, pig, and mouse, respectively. A recombinant protein (rbmGSTO) was functionally overexpressed in Escherichia coli cells in a soluble form and purified to homogeneity. rbmGSTO was able to catalyze the biotranslation of glutathione with 1-chloro-2,4-dinitrobenzene, a model substrate for GST, as well as with 4-hydroxynonenal, a product of lipid peroxidation. This enzyme was shown to have high affinity for organophosphorus insecticide and was present abundantly in silkmoth strain exhibiting fenitrothion resistance. These results indicate that bmGSTO could be involved in the increase in level of insecticide resistance for lepidopteran insects.

  5. The unresolved puzzle why alanine extensions cause disease.

    PubMed

    Winter, Reno; Liebold, Jens; Schwarz, Elisabeth

    2013-08-01

    The prospective increase in life expectancy will be accompanied by a rise in the number of elderly people who suffer from ill health caused by old age. Many diseases caused by aging are protein misfolding diseases. The molecular mechanisms underlying these disorders receive constant scientific interest. In addition to old age, mutations also cause congenital protein misfolding disorders. Chorea Huntington, one of the most well-known examples, is caused by triplet extensions that can lead to more than 100 glutamines in the N-terminal region of huntingtin, accompanied by huntingtin aggregation. So far, nine disease-associated triplet extensions have also been described for alanine codons. The extensions lead primarily to skeletal malformations. Eight of these proteins represent transcription factors, while the nuclear poly-adenylate binding protein 1, PABPN1, is an RNA binding protein. Additional alanines in PABPN1 lead to the disease oculopharyngeal muscular dystrophy (OPMD). The alanine extension affects the N-terminal domain of the protein, which has been shown to lack tertiary contacts. Biochemical analyses of the N-terminal domain revealed an alanine-dependent fibril formation. However, fibril formation of full-length protein did not recapitulate the findings of the N-terminal domain. Fibril formation of intact PABPN1 was independent of the alanine segment, and the fibrils displayed biochemical properties that were completely different from those of the N-terminal domain. Although intranuclear inclusions have been shown to represent the histochemical hallmark of OPMD, their role in pathogenesis is currently unclear. Several cell culture and animal models have been generated to study the molecular processes involved in OPMD. These studies revealed a number of promising future therapeutic strategies that could one day improve the quality of life for the patients.

  6. Plasma amino acids

    MedlinePlus

    Amino acids blood test ... types of methods used to determine the individual amino acid levels in the blood. ... test is done to measure the level of amino acids in the blood. An increased level of a ...

  7. Subcritical Water Extraction of Amino Acids from Atacama Desert Soils

    NASA Technical Reports Server (NTRS)

    Amashukeli, Xenia; Pelletier, Christine C.; Kirby, James P.; Grunthaner, Frank J.

    2007-01-01

    Amino acids are considered organic molecular indicators in the search for extant and extinct life in the Solar System. Extraction of these molecules from a particulate solid matrix, such as Martian regolith, will be critical to their in situ detection and analysis. The goals of this study were to optimize a laboratory amino acid extraction protocol by quantitatively measuring the yields of extracted amino acids as a function of liquid water temperature and sample extraction time and to compare the results to the standard HCl vapor- phase hydrolysis yields for the same soil samples. Soil samples from the Yungay region of the Atacama Desert ( Martian regolith analog) were collected during a field study in the summer of 2005. The amino acids ( alanine, aspartic acid, glutamic acid, glycine, serine, and valine) chosen for analysis were present in the samples at concentrations of 1 - 70 parts- per- billion. Subcritical water extraction efficiency was examined over the temperature range of 30 - 325 degrees C, at pressures of 17.2 or 20.0 MPa, and for water- sample contact equilibration times of 0 - 30 min. None of the amino acids were extracted in detectable amounts at 30 degrees C ( at 17.2 MPa), suggesting that amino acids are too strongly bound by the soil matrix to be extracted at such a low temperature. Between 150 degrees C and 250 degrees C ( at 17.2 MPa), the extraction efficiencies of glycine, alanine, and valine were observed to increase with increasing water temperature, consistent with higher solubility at higher temperatures, perhaps due to the decreasing dielectric constant of water. Amino acids were not detected in extracts collected at 325 degrees C ( at 20.0 MPa), probably due to amino acid decomposition at this temperature. The optimal subcritical water extraction conditions for these amino acids from Atacama Desert soils were achieved at 200 degrees C, 17.2 MPa, and a water- sample contact equilibration time of 10 min.

  8. Crystal structures of d-alanine-d-alanine ligase from Xanthomonas oryzae pv. oryzae alone and in complex with nucleotides.

    PubMed

    Doan, Thanh Thi Ngoc; Kim, Jin-Kwang; Ngo, Ho-Phuong-Thuy; Tran, Huyen-Thi; Cha, Sun-Shin; Min Chung, Kyung; Huynh, Kim-Hung; Ahn, Yeh-Jin; Kang, Lin-Woo

    2014-03-01

    D-Alanine-D-alanine ligase (DDL) catalyzes the biosynthesis of d-alanyl-d-alanine, an essential bacterial peptidoglycan precursor, and is an important drug target for the development of antibacterials. We determined four different crystal structures of DDL from Xanthomonas oryzae pv. oryzae (Xoo) causing Bacteria Blight (BB), which include apo, ADP-bound, ATP-bound, and AMPPNP-bound structures at the resolution between 2.3 and 2.0 Å. Similarly with other DDLs, the active site of XoDDL is formed by three loops from three domains at the center of enzyme. Compared with d-alanyl-d-alanine and ATP-bound TtDDL structure, the γ-phosphate of ATP in XoDDL structure was shifted outside toward solution. We swapped the ω-loop (loop3) of XoDDL with those of Escherichia coli and Helicobacter pylori DDLs, and measured the enzymatic kinetics of wild-type XoDDL and two mutant XoDDLs with the swapped ω-loops. Results showed that the direct interactions between ω-loop and other two loops are essential for the active ATP conformation for D-ala-phosphate formation.

  9. Physical reasons for the unusual α-helix stabilization afforded by charged or neutral polar residues in alanine-rich peptides

    PubMed Central

    Vila, Jorge A.; Ripoll, Daniel R.; Scheraga, H. A.

    2000-01-01

    We have carried out conformational energy calculations on alanine-based copolymers with the sequence Ac-AAAAAXAAAA-NH2 in water, where X stands for lysine or glutamine, to identify the underlying source of stability of alanine-based polypeptides containing charged or highly soluble polar residues in the absence of charge–charge interactions. The results indicate that ionizable or neutral polar residues introduced into the sequence to make them soluble sequester the water away from the CO and NH groups of the backbone, thereby enabling them to form internal hydrogen bonds. This solvation effect dictates the conformational preference and, hence, modifies the conformational propensity of alanine residues. Even though we carried out simulations for specific amino acid sequences, our results provide an understanding of some of the basic principles that govern the process of folding of these short sequences independently of the kind of residues introduced to make them soluble. In addition, we have investigated through simulations the effect of the bulk dielectric constant on the conformational preferences of these peptides. Extensive conformational Monte Carlo searches on terminally blocked 10-mer and 16-mer homopolymers of alanine in the absence of salt were carried out assuming values for the dielectric constant of the solvent ɛ of 80, 40, and 2. Our simulations show a clear tendency of these oligopeptides to augment the α-helix content as the bulk dielectric constant of the solvent is lowered. This behavior is due mainly to a loss of exposure of the CO and NH groups to the aqueous solvent. Experimental evidence indicates that the helical propensity of the amino acids in water shows a dramatic increase on addition of certain alcohols, such us trifluoroethanol. Our results provide a possible explanation of the mechanism by which alcohol/water mixtures affect the free energy of helical alanine oligopeptides relative to nonhelical ones. PMID:11078529

  10. Irritable Bowel Syndrome May Be Associated with Elevated Alanine Aminotransferase and Metabolic Syndrome

    PubMed Central

    Lee, Seung-Hwa; Kim, Kwang-Min; Joo, Nam-Seok

    2016-01-01

    Purpose Recent studies have revealed close relationships between hepatic injury, metabolic pathways, and gut microbiota. The microorganisms in the intestine also cause irritable bowel syndrome (IBS). The aim of this study was to examine whether IBS was associated with elevated hepatic enzyme [alanine aminotransferase (ALT) and aspartate aminotransferase (AST)], gamma-glutamyl transferase (γ-GT) levels, and metabolic syndrome (MS). Materials and Methods This was a retrospective, cross-sectional, case-control study. The case and control groups comprised subjects who visited our health promotion center for general check-ups from June 2010 to December 2010. Of the 1127 initially screened subjects, 83 had IBS according to the Rome III criteria. The control group consisted of 260 age- and sex-matched subjects without IBS who visited our health promotion center during the same period. Results Compared to control subjects, patients with IBS showed significantly higher values of anthropometric parameters (body mass index, waist circumference), liver enzymes, γ-GT, and lipid levels. The prevalences of elevated ALT (16.9% vs. 7.7%; p=0.015) and γ-GT (24.1% vs. 11.5%; p=0.037) levels were significantly higher in patients with IBS than in control subjects. A statistically significant difference was observed in the prevalence of MS between controls and IBS patients (12.7% vs. 32.5%; p<0.001). The relationships between elevated ALT levels, MS, and IBS remained statistically significant after controlling for potential confounding factors. Conclusion On the basis of our study results, IBS may be an important condition in certain patients with elevated ALT levels and MS. PMID:26632395

  11. Kinetic mechanism and inhibition of Mycobacterium tuberculosis D-alanine:D-alanine ligase by the antibiotic D-cycloserine.

    PubMed

    Prosser, Gareth A; de Carvalho, Luiz Pedro S

    2013-02-01

    D-cycloserine (DCS) is an antibiotic that is currently used in second-line treatment of tuberculosis. DCS is a structural analogue of D-alanine, and targets two enzymes involved in the cytosolic stages of peptidoglycan synthesis: alanine racemase (Alr) and D-alanine:D-alanine ligase (Ddl). The mechanisms of inhibition of DCS have been well-assessed using Alr and Ddl enzymes from various bacterial species, but little is known regarding the interactions of DCS with the mycobacterial orthologues of these enzymes. We have over-expressed and purified recombinant Mycobacterium tuberculosis Ddl (MtDdl; Rv2981c), and report a kinetic examination of the enzyme with both its native substrate and DCS. MtDdl is activated by K(+), follows an ordered ter ter mechanism and displays distinct affinities for D-Ala at each D-Ala binding site (K(m,D-Ala1) = 0.075 mm, K(m,D-Ala2) = 3.6 mm). ATP is the first substrate to bind and is necessary for subsequent binding of D-alanine or DCS. The pH dependence of MtDdl kinetic parameters indicate that general base chemistry is involved in the catalytic step. DCS was found to competitively inhibit D-Ala binding at both MtDdl D-Ala sites with equal affinity (K(i,DCS1) = 14 μm, K(i,DCS2) = 25 μm); however, each enzyme active site can only accommodate a single DCS molecule at a given time. The pH dependence of K(i,DCS2) revealed a loss of DCS binding affinity at high pH (pK(a) = 7.5), suggesting that DCS binds optimally in the zwitterionic form. The results of this study may assist in the design and development of novel Ddl-specific inhibitors for use as anti-mycobacterial agents.

  12. Structure of the Mycobacterium tuberculosis D-Alanine:D-Alanine Ligase, a Target of the Antituberculosis Drug D-Cycloserine

    SciTech Connect

    Bruning, John B.; Murillo, Ana C.; Chacon, Ofelia; Barletta, Raúl G.; Sacchettini, James C.

    2011-09-28

    D-Alanine:D-alanine ligase (EC 6.3.2.4; Ddl) catalyzes the ATP-driven ligation of two D-alanine (D-Ala) molecules to form the D-alanyl:D-alanine dipeptide. This molecule is a key building block in peptidoglycan biosynthesis, making Ddl an attractive target for drug development. D-Cycloserine (DCS), an analog of D-Ala and a prototype Ddl inhibitor, has shown promise for the treatment of tuberculosis. Here, we report the crystal structure of Mycobacterium tuberculosis Ddl at a resolution of 2.1 {angstrom}. This structure indicates that Ddl is a dimer and consists of three discrete domains; the ligand binding cavity is at the intersection of all three domains and conjoined by several loop regions. The M. tuberculosis apo Ddl structure shows a novel conformation that has not yet been observed in Ddl enzymes from other species. The nucleotide and D-alanine binding pockets are flexible, requiring significant structural rearrangement of the bordering regions for entry and binding of both ATP and D-Ala molecules. Solution affinity and kinetic studies showed that DCS interacts with Ddl in a manner similar to that observed for D-Ala. Each ligand binds to two binding sites that have significant differences in affinity, with the first binding site exhibiting high affinity. DCS inhibits the enzyme, with a 50% inhibitory concentration (IC{sub 50}) of 0.37 mM under standard assay conditions, implicating a preferential and weak inhibition at the second, lower-affinity binding site. Moreover, DCS binding is tighter at higher ATP concentrations. The crystal structure illustrates potential drugable sites that may result in the development of more-effective Ddl inhibitors.

  13. Beta-N-methylamino-L-alanine (BMAA) in novel South African cyanobacterial isolates.

    PubMed

    Esterhuizen, M; Downing, T G

    2008-10-01

    Beta-N-methylamino-L-alanine (BMAA) is a neurotoxic non-proteinogenic amino acid reportedly produced by the majority of cyanobacterial isolates. A novel method was developed for the detection of BMAA in biological samples. Cultures representing the taxonomic diversity and geographic distribution in Southern Africa were collected and made uni-algal by standard methods before analysis for the presence of both free and protein-associated BMAA. Protein-associated BMAA was released by acid hydrolysis in an inert atmosphere. Samples were analyzed by gas chromatography-mass spectrometry (GC-MS) with pre-derivatization of amino acids using Phenomonex EZ:faast of the tested cultures, 96% were positive for BMAA although several were below the limit for quantification. BMAA presence was not related to the geographic origin or taxonomy of isolates and no correlation between free and bound BMAA concentrations was observed within or between taxonomic groups. These data offer the first confirmation of the taxonomic and geographic ubiquity of BMAA in freshwater cyanobacteria.

  14. Oligomerization of Glycine and Alanine Catalyzed by Iron Oxides: Implications for Prebiotic Chemistry

    NASA Astrophysics Data System (ADS)

    Shanker, Uma; Bhushan, Brij; Bhattacharjee, G.; Kamaluddin

    2012-02-01

    Iron oxide minerals are probable constituents of the sediments present in geothermal regions of the primitive earth. They might have adsorbed different organic monomers (amino acids, nucleotides etc.) and catalyzed polymerization processes leading to the formation of the first living cell. In the present work we tested the catalytic activity of three forms of iron oxides (Goethite, Akaganeite and Hematite) in the intermolecular condensation of each of the amino acids glycine and L-alanine. The effect of zinc oxide and titanium dioxide on the oligomerization has also been studied. Oligomerization studies were performed for 35 days at three different temperatures 50, 90 and 120°C without applying drying/wetting cycling. The products formed were characterized by HPLC and ESI-MS techniques. All three forms of iron oxides catalyzed peptide bond formation (23.2% of gly2 and 10.65% of ala2). The reaction was monitored every 7 days. Formation of peptides was observed to start after 7 days at 50°C. Maximum yield of peptides was found after 35 days at 90°C. Reaction at 120°C favors formation of diketopiperazine derivatives. It is also important to note that after 35 days of reaction, goethite produced dimer and trimer with the highest yield among the oxides tested. We suggest that the activity of goethite could probably be due to its high surface area and surface acidity.

  15. The Cyanobacteria Derived Toxin Beta-N-Methylamino-L-Alanine and Amyotrophic Lateral Sclerosis

    PubMed Central

    Banack, Sandra Anne; Caller, Tracie A.; Stommel, Elijah W.

    2010-01-01

    There is mounting evidence to suggest that environmental factors play a major role in the development of neurodegenerative diseases like ALS (Amyotrophic Lateral Sclerosis). The non-protein amino acid beta-N-methylamino-L-alanine (BMAA) was first associated with the high incidence of Amyotrophic Lateral Sclerosis/Parkinsonism Dementia Complex (ALS/PDC) in Guam, and has been implicated as a potential environmental factor in ALS, Alzheimer’s disease, and other neurodegenerative diseases. BMAA has a number of toxic effects on motor neurons including direct agonist action on NMDA and AMPA receptors, induction of oxidative stress, and depletion of glutathione. As a non-protein amino acid, there is also the strong possibility that BMAA could cause intraneuronal protein misfolding, the hallmark of neurodegeneration. While an animal model for BMAA-induced ALS is lacking, there is substantial evidence to support a link between this toxin and ALS. The ramifications of discovering an environmental trigger for ALS are enormous. In this article, we discuss the history, ecology, pharmacology and clinical ramifications of this ubiquitous, cyanobacteria-derived toxin. PMID:22069578

  16. Computational analysis of binding affinity and neural response at the l-alanine receptor

    NASA Astrophysics Data System (ADS)

    Venanzi, Thomas J.; Bryant, Bruce P.; Venanzi, Carol A.

    1995-10-01

    A model of analogue-receptor binding is developed for the l-alanine receptor in the channel catfish using the AM1-SM2 and ab initio SCRF computational methods. Besides interactions involving the zwitterionic moiety of the amino acid analogue and complementary subsites on the receptor, the model suggests the presence of a hydrophobic pocket with dispersion interactions between the receptor and the residue on the amino acid analogue. Conformational analysis suggests not only a small compact active site on the receptor, but also that the analogues with the highest affinity occupy nearly identical regions of space. Although the binding interaction is dominated by the ionic terms, AM1-SM2 calculations indicate that free energy terms associated with cavity formation, solvent reorganization, and dispersion interactions can be correlated to activation and neural response. From a consideration of this model, molecular features of the analogues that are important for binding and neural response were deduced and other analogues or ligands were developed and tested.

  17. Anisotropy-Guided Enantiomeric Enhancement in AlanineUsing Far-UV Circularly Polarized Light

    NASA Astrophysics Data System (ADS)

    Meinert, Cornelia; Cassam-Chenaï, Patrick; Jones, Nykola C.; Nahon, Laurent; Hoffmann, Søren V.; Meierhenrich, Uwe J.

    2015-06-01

    All life on Earth is characterized by its asymmetry - both the genetic material and proteins are composed of homochiral monomers. Understanding how this molecular asymmetry initially arose is a key question related to the origins of life. Cometary ice simulations, l-enantiomeric enriched amino acids in meteorites and the detection of circularly polarized electromagnetic radiation in star-forming regions point to a possible interstellar/protostellar generation of stereochemical asymmetry. Based upon our recently recorded anisotropy spectra g( λ) of amino acids in the vacuum-UV range, we subjected amorphous films of racemic 13C-alanine to far-UV circularly polarized synchrotron radiation to probe the asymmetric photon-molecule interaction under interstellar conditions. Optical purities of up to 4 % were reached, which correlate with our theoretical predictions. Importantly, we show that chiral symmetry breaking using circularly polarized light is dependent on both the helicity and the wavelength of incident light. In order to predict such stereocontrol, time-dependent density functional theory was used to calculate anisotropy spectra. The calculated anisotropy spectra show good agreement with the experimental ones. The European Space Agency's Rosetta mission, which successfully landed Philae on comet 67P/Churyumov-Gerasimenko on 12 November 2014, will investigate the configuration of chiral compounds and thereby obtain data that are to be interpreted in the context of the results presented here.

  18. Acetate stimulates flux through the tricarboxylic acid cycle in rabbit renal proximal tubules synthesizing glutamine from alanine: a 13C NMR study.

    PubMed Central

    Dugelay, S; Chauvin, M F; Megnin-Chanet, F; Martin, G; Laréal, M C; Lhoste, J M; Baverel, G

    1999-01-01

    Although glutamine synthesis has a major role in the control of acid-base balance and ammonia detoxification in the kidney of herbivorous species, very little is known about the regulation of this process. We therefore studied the influence of acetate, which is readily metabolized by the kidney and whose metabolism is accompanied by the production of bicarbonate, on glutamine synthesis from variously labelled [(13)C]alanine and [(14)C]alanine molecules in isolated rabbit renal proximal tubules. With alanine as sole exogenous substrate, glutamine and, to a smaller extent, glutamate and CO(2), were the only significant products of the metabolism of this amino acid, which was removed at high rates. Absolute fluxes through the enzymes involved in alanine conversion into glutamine were assessed by using a novel model describing the corresponding reactions in conjunction with the (13)C NMR, and to a smaller extent, the radioactive and enzymic data. The presence of acetate (5 mM) led to a large stimulation of fluxes through citrate synthase and alpha-oxoglutarate dehydrogenase. These effects were accompanied by increases in the removal of alanine, in the accumulation of glutamate and in flux through the anaplerotic enzyme pyruvate carboxylase. Acetate did not alter fluxes through glutamate dehydrogenase and glutamine synthetase; as a result, acetate did not change the accumulation of ammonia, which was negligible under both experimental conditions. We conclude that acetate, which seems to be an important energy-provider to the rabbit renal proximal tubule, simultaneously traps as glutamate the extra nitrogen removed as alanine, thus preventing the release of additional ammonia by the glutamate dehydrogenase reaction. PMID:10477267

  19. Topology of AspT, the aspartate:alanine antiporter of Tetragenococcus halophilus, determined by site-directed fluorescence labeling.

    PubMed

    Nanatani, Kei; Fujiki, Takashi; Kanou, Kazuhiko; Takeda-Shitaka, Mayuko; Umeyama, Hideaki; Ye, Liwen; Wang, Xicheng; Nakajima, Tasuku; Uchida, Takafumi; Maloney, Peter C; Abe, Keietsu

    2007-10-01

    The gram-positive lactic acid bacterium Tetragenococcus halophilus catalyzes the decarboxylation of L-aspartate (Asp) with release of L-alanine (Ala) and CO(2). The decarboxylation reaction consists of two steps: electrogenic exchange of Asp for Ala catalyzed by an aspartate:alanine antiporter (AspT) and intracellular decarboxylation of the transported Asp catalyzed by an L-aspartate-beta-decarboxylase (AspD). AspT belongs to the newly classified aspartate:alanine exchanger family (transporter classification no. 2.A.81) of transporters. In this study, we were interested in the relationship between the structure and function of AspT and thus analyzed the topology by means of the substituted-cysteine accessibility method using the impermeant, fluorescent, thiol-specific probe Oregon Green 488 maleimide (OGM) and the impermeant, nonfluorescent, thiol-specific probe [2-(trimethylammonium)ethyl]methanethiosulfonate bromide. We generated 23 single-cysteine variants from a six-histidine-tagged cysteineless AspT template. A cysteine position was assigned an external location if the corresponding single-cysteine variant reacted with OGM added to intact cells, and a position was assigned an internal location if OGM labeling required cell lysis. The topology analyses revealed that AspT has a unique topology; the protein has 10 transmembrane helices (TMs), a large hydrophilic cytoplasmic loop (about 180 amino acids) between TM5 and TM6, N and C termini that face the periplasm, and a positively charged residue (arginine 76) within TM3. Moreover, the three-dimensional structure constructed by means of the full automatic modeling system indicates that the large hydrophilic cytoplasmic loop of AspT possesses a TrkA_C domain and a TrkA_C-like domain and that the three-dimensional structures of these domains are similar to each other even though their amino acid sequences show low similarity.

  20. Incorporation of β-amino acids into dihydrofolate reductase by ribosomes having modifications in the peptidyltransferase center.

    PubMed

    Maini, Rumit; Nguyen, Dan T; Chen, Shengxi; Dedkova, Larisa M; Chowdhury, Sandipan Roy; Alcala-Torano, Rafael; Hecht, Sidney M

    2013-03-01

    Ribosomes containing modifications in three regions of 23S rRNA, all of which are in proximity to the ribosomal peptidyltransferase center (PTC), were utilized previously as a source of S-30 preparations for in vitro protein biosynthesis experiments. When utilized in the presence of mRNAs containing UAG codons at predetermined positions+β-alanyl-tRNA(CUA), the modified ribosomes produced enhanced levels of full length proteins via UAG codon suppression. In the present study, these earlier results have been extended by the use of substituted β-amino acids, and direct evidence for β-amino acid incorporation is provided. Presently, five of the clones having modified ribosomes are used in experiments employing four substituted β-amino acids, including α-methyl-β-alanine, β,β-dimethyl-β-alanine, β-phenylalanine, and β-(p-bromophenyl)alanine. The β-amino acids were incorporated into three different positions (10, 18 and 49) of Escherichia coli dihydrofolate reductase (DHFR) and their efficiencies of suppression of the UAG codons were compared with those of β-alanine and representative α-l-amino acids. The isolated proteins containing the modified β-amino acids were subjected to proteolytic digestion, and the derived fragments were characterized by mass spectrometry, establishing that the β-amino acids had been incorporated into DHFR, and that they were present exclusively in the anticipated peptide fragments. DHFR contains glutamic acid in position 17, and it has been shown previously that Glu-C endoproteinase can hydrolyze DHFR between amino acids residues 17 and 18. The incorporation of β,β-dimethyl-β-alanine into position 18 of DHFR prevented this cleavage, providing further evidence for the position of incorporation of the β-amino acid.

  1. Addition of amino acids and dipeptides to fullerene C{sub 60} giving rise to monoadducts

    SciTech Connect

    Romanova, V.S.; Tsyryapkin, V.A.; Vol`pin, M.E.

    1994-12-01

    The authors have developed a general method for the direct addition of amino acids and dipeptides of various structures to fullerene C{sub 60}. In all cases the addition involves the amino group. The reaction proceeds when the solutions of fullerene and an amino acid (or dipeptide) are mixed at 50-100 {degrees}C. The fullerene derivatives of the following amino acids and dipeptides have been obtained: glycine, p-aminobenzoic acid, {omega}-aminocaproic acid, L-proline, L-alanine, L-alanyl-Lalanine, D,L-alanyl-D,L-alanine, glycyl-L-valine. The adduct of methyl L-ananinate with C{sub 60} was also prepared.

  2. A single amino acid controls the functional switch of human constitutive androstane receptor (CAR) 1 to the xenobiotic-sensitive splicing variant CAR3.

    PubMed

    Chen, Tao; Tompkins, Leslie M; Li, Linhao; Li, Haishan; Kim, Gregory; Zheng, Yuxin; Wang, Hongbing

    2010-01-01

    The constitutive androstane receptor (CAR) is constitutively activated in immortalized cell lines independent of xenobiotic stimuli. This feature of CAR has limited its use as a sensor for xenobiotic-induced expression of drug-metabolizing enzymes. Recent reports, however, reveal that a splicing variant of human CAR (hCAR3), which contains an insertion of five amino acids (APYLT), exhibits low basal but xenobiotic-inducible activities in cell-based reporter assays. Nonetheless, the underlying mechanisms of this functional shift are not well understood. We have now generated chimeric constructs containing various residues of the five amino acids of hCAR3 and examined their response to typical hCAR activators. Our results showed that the retention of alanine (hCAR1+A) alone is sufficient to confer the constitutively activated hCAR1 to the xenobiotic-sensitive hCAR3. It is noteworthy that hCAR1+A was significantly activated by a series of known hCAR activators, and displayed activation superior to that of hCAR3. Moreover, intracellular localization assays revealed that hCAR1+A exhibits nuclear accumulation upon 6-(4-chlorophenyl) imidazo[2,1-b][1,3]thiazole-5-carbaldehyde-O-(3,4-dichlorobenzyl) oxime (CITCO) treatment in COS1 cells, which differs from the spontaneous nuclear distribution of hCAR1 and the nontranslocatable hCAR3. Mammalian two-hybrid and glutathione S-transferase pull-down assays further demonstrated that hCAR1+A interacts with the coactivator SRC-1 and GRIP-1 at low level before activation, while at significantly enhanced level in the presence of CITCO. Thus, the alanine residue in the insertion of hCAR3 seems in charge of the xenobiotic response of hCAR3 through direct and indirect mechanisms. Activation of hCAR1+A may represent a sensitive avenue for the identification of hCAR activators.

  3. Amino Acid Transporters and Release of Hydrophobic Amino Acids in the Heterocyst-Forming Cyanobacterium Anabaena sp. Strain PCC 7120

    PubMed Central

    Pernil, Rafael; Picossi, Silvia; Herrero, Antonia; Flores, Enrique; Mariscal, Vicente

    2015-01-01

    Anabaena sp. strain PCC 7120 is a filamentous cyanobacterium that can use inorganic compounds such as nitrate or ammonium as nitrogen sources. In the absence of combined nitrogen, it can fix N2 in differentiated cells called heterocysts. Anabaena also shows substantial activities of amino acid uptake, and three ABC-type transporters for amino acids have been previously characterized. Seven new loci encoding predicted amino acid transporters were identified in the Anabaena genomic sequence and inactivated. Two of them were involved in amino acid uptake. Locus alr2535-alr2541 encodes the elements of a hydrophobic amino acid ABC-type transporter that is mainly involved in the uptake of glycine. ORF all0342 encodes a putative transporter from the dicarboxylate/amino acid:cation symporter (DAACS) family whose inactivation resulted in an increased uptake of a broad range of amino acids. An assay to study amino acid release from Anabaena filaments to the external medium was set up. Net release of the alanine analogue α-aminoisobutyric acid (AIB) was observed when transport system N-I (a hydrophobic amino acid ABC-type transporter) was engaged in the uptake of a specific substrate. The rate of AIB release was directly proportional to the intracellular AIB concentration, suggesting leakage from the cells by diffusion. PMID:25915115

  4. Amino Acid Transporters and Release of Hydrophobic Amino Acids in the Heterocyst-Forming Cyanobacterium Anabaena sp. Strain PCC 7120.

    PubMed

    Pernil, Rafael; Picossi, Silvia; Herrero, Antonia; Flores, Enrique; Mariscal, Vicente

    2015-04-23

    Anabaena sp. strain PCC 7120 is a filamentous cyanobacterium that can use inorganic compounds such as nitrate or ammonium as nitrogen sources. In the absence of combined nitrogen, it can fix N2 in differentiated cells called heterocysts. Anabaena also shows substantial activities of amino acid uptake, and three ABC-type transporters for amino acids have been previously characterized. Seven new loci encoding predicted amino acid transporters were identified in the Anabaena genomic sequence and inactivated. Two of them were involved in amino acid uptake. Locus alr2535-alr2541 encodes the elements of a hydrophobic amino acid ABC-type transporter that is mainly involved in the uptake of glycine. ORF all0342 encodes a putative transporter from the dicarboxylate/amino acid:cation symporter (DAACS) family whose inactivation resulted in an increased uptake of a broad range of amino acids. An assay to study amino acid release from Anabaena filaments to the external medium was set up. Net release of the alanine analogue α-aminoisobutyric acid (AIB) was observed when transport system N-I (a hydrophobic amino acid ABC-type transporter) was engaged in the uptake of a specific substrate. The rate of AIB release was directly proportional to the intracellular AIB concentration, suggesting leakage from the cells by diffusion.

  5. Amino acid synthesis in a supercritical carbon dioxide - water system.

    PubMed

    Fujioka, Kouki; Futamura, Yasuhiro; Shiohara, Tomoo; Hoshino, Akiyoshi; Kanaya, Fumihide; Manome, Yoshinobu; Yamamoto, Kenji

    2009-06-15

    Mars is a CO(2)-abundant planet, whereas early Earth is thought to be also CO(2)-abundant. In addition, water was also discovered on Mars in 2008. From the facts and theory, we assumed that soda fountains were present on both planets, and this affected amino acid synthesis. Here, using a supercritical CO(2)/liquid H(2)O (10:1) system which mimicked crust soda fountains, we demonstrate production of amino acids from hydroxylamine (nitrogen source) and keto acids (oxylic acid sources). In this research, several amino acids were detected with an amino acid analyzer. Moreover, alanine polymers were detected with LC-MS. Our research lights up a new pathway in the study of life's origin.

  6. Amino Acid Synthesis in a Supercritical Carbon Dioxide - Water System

    PubMed Central

    Fujioka, Kouki; Futamura, Yasuhiro; Shiohara, Tomoo; Hoshino, Akiyoshi; Kanaya, Fumihide; Manome, Yoshinobu; Yamamoto, Kenji

    2009-01-01

    Mars is a CO2-abundant planet, whereas early Earth is thought to be also CO2-abundant. In addition, water was also discovered on Mars in 2008. From the facts and theory, we assumed that soda fountains were present on both planets, and this affected amino acid synthesis. Here, using a supercritical CO2/liquid H2O (10:1) system which mimicked crust soda fountains, we demonstrate production of amino acids from hydroxylamine (nitrogen source) and keto acids (oxylic acid sources). In this research, several amino acids were detected with an amino acid analyzer. Moreover, alanine polymers were detected with LC-MS. Our research lights up a new pathway in the study of life’s origin. PMID:19582225

  7. A 49 kDa microtubule cross-linking protein from Artemia franciscana is a coenzyme A-transferase.

    PubMed

    Oulton, Mindy M; Amons, Reinout; Liang, Ping; MacRae, Thomas H

    2003-12-01

    Embryos and larvae of the brine shrimp, Artemia franciscana, were shown previously to possess a protein, now termed p49, which cross-links microtubules in vitro. Molecular characteristics of p49 were described, but the protein's identity and its role in the cell were not determined. Degenerate oligonucleotide primers designed on the basis of peptide sequence obtained by Edman degradation during this study were used to generate p49 cDNAs by RT-PCR and these were cloned and sequenced. Comparison with archived sequences revealed that the deduced amino acid sequence of p49 resembled the Drosophila gene product CG7920, as well as related proteins encoded in the genomes of Anopheles and Caenorhabditis. Similar proteins exist in several bacteria but no evident homologues were found in vertebrates and plants, and only very distant homologues resided in yeast. When evolutionary relationships were compared, p49 and the homologues from Drosophila, Anopheles and Caenorhabditis formed a distinct subcluster within phylogenetic trees. Additionally, the predicted secondary structures of p49, 4-hydroxybutyrate CoA-transferase from Clostridium aminobutyricum and glutaconate CoA-transferase from Acidaminococcus fermentans were similar and the enzymes may possess related catalytic mechanisms. The purified Artemia protein exhibited 4-hydroxybutyrate CoA-transferase activity, thereby establishing p49 as the first crustacean CoA-transferase to be characterized. Probing of Western blots with an antibody against p49 revealed a cross-reactive protein in Drosophila that associated with microtubules, but to a lesser extent than did p49 from Artemia.

  8. Extraterrestrial amino acids in the Almahata Sitta meteorite

    NASA Astrophysics Data System (ADS)

    Glavin, Daniel P.; Aubrey, Andrew D.; Callahan, Michael P.; Dworkin, Jason P.; Elsila, Jamie E.; Parker, Eric T.; Bada, Jeffrey L.; Jenniskens, Peter; Shaddad, Muawia H.

    2010-10-01

    Amino acid analysis of a meteorite fragment of asteroid 2008 TC3 called Almahata Sitta was carried out using reverse-phase liquid chromatography coupled with UV fluorescence detection and time-of-flight mass spectrometry (LC-FD/ToF-MS) as part of a sample analysis consortium. LC-FD/ToF-MS analyses of hot-water extracts from the meteorite revealed a complex distribution of two- to seven-carbon aliphatic amino acids and one- to three-carbon amines with abundances ranging from 0.5 to 149 parts-per-billion (ppb). The enantiomeric ratios of the amino acids alanine, β-amino-n-butyric acid, 2-amino-2-methylbutanoic acid (isovaline), and 2-aminopentanoic acid (norvaline) in the meteorite were racemic (D/L ˜ 1), indicating that these amino acids are indigenous to the meteorite and not terrestrial contaminants. Several other nonprotein amino acids were also identified in the meteorite above background levels including α-aminoisobutyric acid (α-AIB), 4-amino-2-methylbutanoic acid, 4-amino-3-methylbutanoic acid, and 3-, 4-, and 5-aminopentanoic acid. The total abundances of isovaline and α-AIB in Almahata Sitta are approximately 1000 times lower than the abundances of these amino acids found in the CM carbonaceous chondrite Murchison. The extremely low abundances and unusual distribution of five-carbon amino acids in Almahata Sitta compared to CI, CM, and CR carbonaceous chondrites may reflect extensive thermal alteration of amino acids on the parent asteroid by partial melting during formation or subsequent impact shock heating. It is also possible that amino acids were synthesized by catalytic reactions on the parent body after asteroid 2008 TC3 cooled to lower temperatures, or introduced as a contaminant from unrelated meteorite clasts and chemically altered by α-decarboxylation.

  9. Extraterrestrial Amino Acids in the Almahata Sitta Meteorite

    NASA Technical Reports Server (NTRS)

    Glavin, Daniel P.; Aubrey, Andrew D.; Callahan, Michael P.; Dworkin, Jason P.; Elsila, Jamie E.; Parker, Eric T.; Bada, Jeffrey L.

    2010-01-01

    Amino acid analysis of a meteorite fragment of asteroid 2008 TC3 called Almahata Sitta was carried out using reverse-phase liquid chromatography coupled with UV fluorescence detection and time-of-flight mass spectrometry (LC-FD/ToF-MS) as part of a sample analysis consortium. LC-FD/ToF-MS analyses of hot-water extracts from the meteorite revealed a complex distribution of two- to seven-carbon aliphatic amino acids and one- to three-carbon amines with abundances ranging from 0.5 to 149 parts-per-billion (ppb). The enantiomeric ratios of the amino acids alanine, R-amino-n-butyric acid (beta-ABA), 2-amino-2-methylbutanoic acid (isovaline), and 2-aminopentanoic acid (norvaline) in the meteorite were racemic (D/L approximately 1), indicating that these amino acids are indigenous to the meteorite and not terrestrial contaminants. Several other non-protein amino acids were also identified in the meteorite above background levels including alpha-aminoisobutyric acid (alpha-AIB), 4-amino-2- methylbutanoic acid, 4-amino-3-methylbutanoic acid, and 3-, 4-, and 5-aminopentanoic acid. The total abundances of isovaline and alpha-AIB in Almahata Sitta are 1000 times lower than the abundances of these amino acids found in the CM carbonaceous chondrite Murchison. The extremely low abundances and unusual distribution of five carbon amino acids in Almahata Sitta compared to Cl, CM, and CR carbonaceous chondrites may reflect extensive thermal alteration of amino acids on the parent asteroid by partial melting during formation or subsequent impact shock heating. It is also possible that amino acids were synthesized by catalytic reactions on the parent body after asteroid 2008 TC3 cooled to lower temperatures.

  10. Clinical applications of alanine/electron spin resonance dosimetry.

    PubMed

    Baffa, Oswaldo; Kinoshita, Angela

    2014-05-01

    This paper discusses the clinical applications of electron spin resonance (ESR) dosimetry focusing on the ESR/alanine system. A review of few past studies in this area is presented offering a critical overview of the challenges and opportunities for extending this system into clinical applications. Alanine/ESR dosimetry fulfills many of the required properties for several clinical applications such as water-equivalent composition, independence of the sensitivity for the energy range used in therapy and high precision. Improvements in sensitivity and the development of minidosimeters coupled with the use of a spectrometer of higher microwave frequency expanded the possibilities for clinical applications to the new modalities of radiotherapy (intensity-modulated radiation therapy and radiosurgery) and to the detection of low doses such as those present in some radiological image procedures.

  11. Identification of dietary alanine toxicity and trafficking dysfunction in a Drosophila model of hereditary sensory and autonomic neuropathy type 1.

    PubMed

    Oswald, Matthew C W; West, Ryan J H; Lloyd-Evans, Emyr; Sweeney, Sean T

    2015-12-15

    Hereditary sensory and autonomic neuropathy type 1 (HSAN1) is characterized by a loss of distal peripheral sensory and motorneuronal function, neuropathic pain and tissue necrosis. The most common cause of HSAN1 is due to dominant mutations in serine palmitoyl-transferase subunit 1 (SPT1). SPT catalyses the condensation of serine with palmitoyl-CoA, the initial step in sphingolipid biogenesis. Identified mutations in SPT1 are known to both reduce sphingolipid synthesis and generate catalytic promiscuity, incorporating alanine or glycine into the precursor sphingolipid to generate a deoxysphingoid base (DSB). Why either loss of function in SPT1, or generation of DSBs should generate deficits in distal sensory function remains unclear. To address these questions, we generated a Drosophila model of HSAN1. Expression of dSpt1 bearing a disease-related mutation induced morphological deficits in synapse growth at the larval neuromuscular junction consistent with a dominant-negative action. Expression of mutant dSpt1 globally was found to be mildly toxic, but was completely toxic when the diet was supplemented with alanine, when DSBs were observed in abundance. Expression of mutant dSpt1 in sensory neurons generated developmental deficits in dendritic arborization with concomitant sensory deficits. A membrane trafficking defect was observed in soma of sensory neurons expressing mutant dSpt1, consistent with endoplasmic reticulum (ER) to Golgi block. We found that we could rescue sensory function in neurons expressing mutant dSpt1 by co-expressing an effector of ER-Golgi function, Rab1 suggesting compromised ER function in HSAN1 affected dendritic neurons. Our Drosophila model identifies a novel strategy to explore the pathological mechanisms of HSAN1.

  12. Identification of dietary alanine toxicity and trafficking dysfunction in a Drosophila model of hereditary sensory and autonomic neuropathy type 1

    PubMed Central

    Oswald, Matthew C. W.; West, Ryan J. H.; Lloyd-Evans, Emyr; Sweeney, Sean T.

    2015-01-01

    Hereditary sensory and autonomic neuropathy type 1 (HSAN1) is characterized by a loss of distal peripheral sensory and motorneuronal function, neuropathic pain and tissue necrosis. The most common cause of HSAN1 is due to dominant mutations in serine palmitoyl-transferase subunit 1 (SPT1). SPT catalyses the condensation of serine with palmitoyl-CoA, the initial step in sphingolipid biogenesis. Identified mutations in SPT1 are known to both reduce sphingolipid synthesis and generate catalytic promiscuity, incorporating alanine or glycine into the precursor sphingolipid to generate a deoxysphingoid base (DSB). Why either loss of function in SPT1, or generation of DSBs should generate deficits in distal sensory function remains unclear. To address these questions, we generated a Drosophila model of HSAN1. Expression of dSpt1 bearing a disease-related mutation induced morphological deficits in synapse growth at the larval neuromuscular junction consistent with a dominant-negative action. Expression of mutant dSpt1 globally was found to be mildly toxic, but was completely toxic when the diet was supplemented with alanine, when DSBs were observed in abundance. Expression of mutant dSpt1 in sensory neurons generated developmental deficits in dendritic arborization with concomitant sensory deficits. A membrane trafficking defect was observed in soma of sensory neurons expressing mutant dSpt1, consistent with endoplasmic reticulum (ER) to Golgi block. We found that we could rescue sensory function in neurons expressing mutant dSpt1 by co-expressing an effector of ER–Golgi function, Rab1 suggesting compromised ER function in HSAN1 affected dendritic neurons. Our Drosophila model identifies a novel strategy to explore the pathological mechanisms of HSAN1. PMID:26395456

  13. Cellular and Physiological Effects of Dietary Supplementation with β-Hydroxy-β-Methylbutyrate (HMB) and β-Alanine in Late Middle-Aged Mice

    PubMed Central

    Vallejo, Julian; Spence, Madoka; Cheng, An-Lin; Brotto, Leticia; Edens, Neile K.; Garvey, Sean M.; Brotto, Marco

    2016-01-01

    There is growing evidence that severe decline of skeletal muscle mass and function with age may be mitigated by exercise and dietary supplementation with protein and amino acid ingredient technologies. The purposes of this study were to examine the effects of the leucine catabolite, beta-hydroxy-beta-methylbutyrate (HMB), in C2C12 myoblasts and myotubes, and to investigate the effects of dietary supplementation with HMB, the amino acid β-alanine and the combination thereof, on muscle contractility in a preclinical model of pre-sarcopenia. In C2C12 myotubes, HMB enhanced sarcoplasmic reticulum (SR) calcium release beyond vehicle control in the presence of all SR agonists tested (KCl, P<0.01; caffeine, P = 0.03; ionomycin, P = 0.03). HMB also improved C2C12 myoblast viability (25 μM HMB, P = 0.03) and increased proliferation (25 μM HMB, P = 0.04; 125 μM HMB, P<0.01). Furthermore, an ex vivo muscle contractility study was performed on EDL and soleus muscle from 19 month old, male C57BL/6nTac mice. For 8 weeks, mice were fed control AIN-93M diet, diet with HMB, diet with β-alanine, or diet with HMB and β-alanine. In β-alanine fed mice, EDL muscle showed a 7% increase in maximum absolute force compared to the control diet (202 ± 3vs. 188± 5 mN, P = 0.02). At submaximal frequency of stimulation (20 Hz), EDL from mice fed HMB plus β-alanine showed an 11% increase in absolute force (88.6 ± 2.2 vs. 79.8 ± 2.4 mN, P = 0.025) and a 13% increase in specific force (12.2 ± 0.4 vs. 10.8 ± 0.4 N/cm2, P = 0.021). Also in EDL muscle, β-alanine increased the rate of force development at all frequencies tested (P<0.025), while HMB reduced the time to reach peak contractile force (TTP), with a significant effect at 80 Hz (P = 0.0156). In soleus muscle, all experimental diets were associated with a decrease in TTP, compared to control diet. Our findings highlight beneficial effects of HMB and β-alanine supplementation on skeletal muscle function in aging mice. PMID

  14. The action of acetylcholine antagonists on amino acid responses in the frog spinal cord in vitro.

    PubMed Central

    Nicoll, R A

    1975-01-01

    1 The isolated hemisected frog spinal cord has been used to study the action of acetylcholine antagonists on amino acid responses by means of sucrose gap recording. 2 Primary afferents and motoneurones were shown to contain few, if any, cholinoceptors, since acetylcholine and carbachol responses were essentially abolished when synaptic transmission was blocked with magnesium ions or when action potentials were blocked by tetrodotoxin. 3 Curare antagonized the gamma-aminobutyric acid (GABA) and beta-alanine depolarizations of primary afferents and hyperpolarizing action of these amino acids on motoneurones. Nicotine also antagonized beta-alanine depolarizations and to a small extent GABA depolarizations of primary afferents. These actions are similar to but weaker than those obtained previously with picrotoxin. 4 Atropine selectively antagonized beta-alanine depolarizations of primary afferents and blocked beta-alanine and glycine hyperpolarizations of motoneurones. GABA responses were entirely resistant to the action of atropine. These actions are similar to but 50 times weaker than those obtained previously with strychnine. 5 Dihydro-beta-erythroidine, tetraethylammonium, and gallamine were entirely ineffective in antagonizing amino acid responses. Since these agents are known to block the dorsal root potential elicited by ventral root stimulation but have no effect on the amino acid responses of primary afferents, it is evident that a cholinergic step is involved in this pathway. PMID:1082355

  15. Plasma Amino Acid Levels in Children with Autism and Their Families.

    ERIC Educational Resources Information Center

    Aldred, Sarah; Moore, Kieran M.; Fitzgerald, Michael; Waring, Rosemary H.

    2003-01-01

    Plasma amino acid levels were measured in autistic (n=12), Asperger syndrome (n=11) patients, their parents and siblings. Patients with autism or Asperger syndrome and their siblings and parents all had raised glutamic acid, phenyalanine, asparagine, tyrosine, alanine, and lysine levels than age-matched controls. Results suggest dysregulated amino…

  16. Alanine mutagenesis of the primary antigenic escape residue cluster, c1, of apical membrane antigen 1.

    PubMed

    Dutta, Sheetij; Dlugosz, Lisa S; Clayton, Joshua W; Pool, Christopher D; Haynes, J David; Gasser, Robert A; Batchelor, Adrian H

    2010-02-01

    Antibodies against apical membrane antigen 1 (AMA1) inhibit invasion of Plasmodium merozoites into red cells, and a large number of single nucleotide polymorphisms on AMA1 allow the parasite to escape inhibitory antibodies. The availability of a crystal structure makes it possible to test protein engineering strategies to develop a monovalent broadly reactive vaccine. Previously, we showed that a linear stretch of polymorphic residues (amino acids 187 to 207), localized within the C1 cluster on domain 1, conferred the highest level of escape from inhibitory antibodies, and these were termed antigenic escape residues (AER). Here we test the hypothesis that immunodampening the C1 AER will divert the immune system toward more conserved regions. We substituted seven C1 AER of the FVO strain Plasmodium falciparum AMA1 with alanine residues (ALA). The resulting ALA protein was less immunogenic than the native protein in rabbits. Anti-ALA antibodies contained a higher proportion of cross-reactive domain 2 and domain 3 antibodies and had higher avidity than anti-FVO. No overall enhancement of cross-reactive inhibitory activity was observed when anti-FVO and anti-ALA sera were compared for their ability to inhibit invasion. Alanine mutations at the C1 AER had shifted the immune response toward cross-strain-reactive epitopes that were noninhibitory, refuting the hypothesis but confirming the importance of the C1 cluster as an inhibitory epitope. We further demonstrate that naturally occurring polymorphisms that fall within the C1 cluster can predict escape from cross-strain invasion inhibition, reinforcing the importance of the C1 cluster genotype for antigenic categorization and allelic shift analyses in future phase 2b trials.

  17. Probing alanine transaminase catalysis with hyperpolarized 13CD3-pyruvate

    PubMed Central

    Barb, A.W.; Hekmatyar, S.K.; Glushka, J.N.; Prestegard, J.H.

    2013-01-01

    Hyperpolarized metabolites offer a tremendous sensitivity advantage (>104 fold) when measuring flux and enzyme activity in living tissues by magnetic resonance methods. These sensitivity gains can also be applied to mechanistic studies that impose time and metabolite concentration limitations. Here we explore the use of hyperpolarization by dissolution dynamic nuclear polarization (DNP) in mechanistic studies of alanine transaminase (ALT), a well-established biomarker of liver disease and cancer that converts pyruvate to alanine using glutamate as a nitrogen donor. A specific deuterated, 13C-enriched analog of pyruvic acid, 13C3D3-pyruvic acid, is demonstrated to have advantages in terms of detection by both direct 13C observation and indirect observation through methyl protons introduced by ALT-catalyzed H–D exchange. Exchange on injecting hyperpolarized 13C3D3-pyruvate into ALT dissolved in buffered 1H2O, combined with an experimental approach to measure proton incorporation, provided information on mechanistic details of transaminase action on a 1.5 s timescale. ALT introduced, on average, 0.8 new protons into the methyl group of the alanine produced, indicating the presence of an off-pathway enamine intermediate. The opportunities for exploiting mechanism-dependent molecular signatures as well as indirect detection of hyperpolarized 13C3-pyruvate and products in imaging applications are discussed. PMID:23357427

  18. Pressure-induced phase transitions in L-alanine, revisited.

    PubMed

    Tumanov, N A; Boldyreva, E V; Kolesov, B A; Kurnosov, A V; Quesada Cabrera, R

    2010-08-01

    The effect of pressure on L-alanine has been studied by X-ray powder diffraction (up to 12.3 GPa), single-crystal X-ray diffraction, Raman spectroscopy and optical microscopy (up to approximately 6 GPa). No structural phase transitions have been observed. At approximately 2 GPa the cell parameters a and b become accidentally equal to each other, but without a change in space-group symmetry. Neither of two transitions reported by others (to a tetragonal phase at approximately 2 GPa and to a monoclinic phase at approximately 9 GPa) was observed. The changes in cell parameters were continuous up to the highest measured pressures and the cells remained orthorhombic. Some important changes in the intermolecular interactions occur, which also manifest themselves in the Raman spectra. Two new orthorhombic phases could be crystallized from a MeOH/EtOH/H(2)O pressure-transmitting mixture in the pressure range 0.8-4.7 GPa, but only if the sample was kept at these pressures for at least 1-2 d. The new phases converted back to L-alanine on decompression. Judging from the Raman spectra and cell parameters, the new phases are most probably not L-alanine but its solvates.

  19. Screening for non-protein amino acids in seeds of the Guam cycad, Cycas circinalis, by an improved GC-MS method.

    PubMed

    Oh, C H; Brownson, D M; Mabry, T J

    1995-02-01

    The non-protein amino acid, beta-N-methyl-amino-L-alanine (L-BMAA) from Cycas circinalis seeds, has been implicated as a causative agent of amyotrophic lateral sclerosis-Parkinsonism dementia complex (ALS-PDC), a disease known from Guam and other areas in the western Pacific. We analyzed C. circinalis seeds for additional free non-protein amino acids by a recently developed GC-MS method. The samples were prepared by water extraction of seed flour. The amino acids present in the extract were derivatized by N(O,S)-isobutyloxycarbonylation of the amine functional groups and then tert-butyldimethylsilylation of the carboxyl functional groups. Peaks for a total of seventeen derivatives of non-protein amino acids were detected by GC-MS. In addition to L-BMAA, four other non-protein amino acids were identified as beta-alanine, gamma-amino-butyric acid, pyroglutamic acid, and alpha-aminoadipic acid.

  20. Induction of the d-Amino Acid Oxidase from Trigonopsis variabilis

    PubMed Central

    Horner, R.; Wagner, F.; Fischer, L.

    1996-01-01

    Induction of the d-amino acid oxidase (EC. 1.4.3.3) from the yeast Trigonopsis variabilis was investigated by using a minimal medium containing glucose as the carbon and energy source, (NH(inf4))(inf2)SO(inf4) as the nitrogen source, and various d- and dl-amino acid derivatives as inducers. The best new inducers found were N-carbamoyl-d-alanine, N-acetyl-d-tryptophan, and N-chloroacetyl-d-(alpha)-aminobutyric acid; when the induction effects of these compounds were compared with the effects of d-alanine as the nitrogen source and inducer, the resulting activities of d-amino acid oxidase per gram of dried yeast were 4.2, 2.1, and 1.5 times higher, respectively. The optimum concentration of the best inducer, N-carbamoyl-d-alanine, was 5 mM. This inducer could also be used in its racemic form. The induction was pH dependent. After cultivation of the yeast in a 50-liter bioreactor, d-amino acid oxidase activity of about 3,850 (mu)kat (231,000 U) was obtained. In addition, production of the d-amino acid oxidase was found to be significantly dependent on the metal salt composition of the medium. Addition of zinc ions was required to obtain high d-amino acid oxidase levels in the cells. The optimum concentration of ZnSO(inf4) was about 140 (mu)M. PMID:16535339

  1. Late onset ornithine carbamoyl transferase deficiency in males.

    PubMed Central

    Drogari, E; Leonard, J V

    1988-01-01

    Six boys with ornithine carbamoyl transferase deficiency presenting in infancy or later childhood are described. There was wide variation in both the time of presentation and the symptoms, which may initially suggest a neurological, behavioural, or gastroenterological problem. Two patients died, as did two male siblings who were probably affected, but with early recognition of the hyperammonaemia the outlook is good. PMID:3202644

  2. Histamine N-methyl transferase: inhibition by drugs.

    PubMed Central

    Pacifici, G M; Donatelli, P; Giuliani, L

    1992-01-01

    1. Histamine N-methyl transferase activity was measured in samples of human liver, brain, kidney, lung and intestinal mucosa. The mean (+/- s.d.) rate (nmol min-1 mg-1 protein) of histamine N-methylation was 1.78 +/- 0.59 (liver, n = 60), 1.15 +/- 0.38 (renal cortex, n = 8), 0.79 +/- 0.14 (renal medulla, n = 8), 0.35 +/- 0.08 (lung, n = 20), 0.47 +/- 0.18 (human intestine, n = 30) and 0.29 +/- 0.14 (brain, n = 13). 2. Inhibition of histamine N-methyl transferase by 15 drugs was investigated in human liver. The IC50 for the various drugs ranged over three orders of magnitude; chloroquine was the most potent inhibitor. 3. The average IC50 values for chloroquine were 12.6, 22.0, 19.0, 21.6 microM in liver, renal cortex, brain and colon, respectively. These values are lower than the Michaelis-Menten constant for histamine N-methyltransferase in liver (43.8 microM) and kidney (45.5 microM). Chloroquine carried a mixed non-competitive inhibition of hepatic histamine N-methyl transferase. Some side-effects of chloroquine may be explained by inhibition of histamine N-methyl transferase. PMID:1457266

  3. Rational design of an organometallic glutathione transferase inhibitor

    SciTech Connect

    Ang, W.H.; Parker, L.J.; De Luca, A.; Juillerat-Jeanneret, L.; Morton, C.J.; LoBello, M.; Parker, M.W.; Dyson, P.J.

    2010-08-17

    A hybrid organic-inorganic (organometallic) inhibitor was designed to target glutathione transferases. The metal center is used to direct protein binding, while the organic moiety acts as the active-site inhibitor. The mechanism of inhibition was studied using a range of biophysical and biochemical methods.

  4. Homogentisate solanesyl transferase (HST) cDNA’s in maize

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Maize white seedling 3 (w3) has served as a model albino-seedling mutant since its discovery in 1923. We show that the w3 phenotype is caused by disruptions in homogentisate solanesyl transferase (HST), an enzyme that catalyzes the committed step in plastoquinone-9 (PQ9) biosynthesis. This reaction ...

  5. GLUTATHIONE S-TRANSFERASE-MEDIATED METABOLISM OF BROMODICHLOROMETHANE

    EPA Science Inventory

    GLUTATHIONE s-TRANSFERASE-MEDIATED METABOLISM OF BROMODICHLOROMETHANE. M K Ross1 and R A Pegram2. 1Curriculum in Toxicology, University of North Carolina at Chapel Hill; 2Experimental Toxicology Division, NHEERL/ORD, United States Environmental Protection Agency, Research Triangl...

  6. Isotopic effects in mechanistic studies of biotransformations of fluorine derivatives of L-alanine catalysed by L-alanine dehydrogenase.

    PubMed

    Szymańska-Majchrzak, Jolanta; Pałka, Katarzyna; Kańska, Marianna

    2017-05-01

    Synthesis of 3-fluoro-[2-(2)H]-L-alanine (3-F-[(2)H]-L-Ala) in reductive amination of 3-fluoropyruvic acid catalysed by L-alanine dehydrogenase (AlaDH) was described. Fluorine derivative was used to study oxidative deamination catalysed by AlaDH applied kinetic (for 3-F-L-Ala in H2O - KIE's on Vmax: 1.1; on Vmax/KM: 1.2; for 3-F-L-Ala in (2)H2O - on Vmax: 1.4; on Vmax/KM: 2.1) and solvent isotope effect methods (for 3-F-L-Ala - SIE's on Vmax: 1.0; on Vmax/KM: 0.87; for 3-F-[2-(2)H]-L-Ala - on Vmax: 1.4; on Vmax/KM: 1.5). Studies explain some details of reaction mechanism.

  7. Hydration of amino acids: FTIR spectra and molecular dynamics studies.

    PubMed

    Panuszko, Aneta; Adamczak, Beata; Czub, Jacek; Gojło, Emilia; Stangret, Janusz

    2015-11-01

    The hydration of selected amino acids, alanine, glycine, proline, valine, isoleucine and phenylalanine, has been studied in aqueous solutions by means of FTIR spectra of HDO isotopically diluted in H2O. The difference spectra procedure and the chemometric method have been applied to remove the contribution of bulk water and thus to separate the spectra of solute-affected HDO. To support interpretation of obtained spectral results, molecular dynamics simulations of amino acids were performed. The structural-energetic characteristic of these solute-affected water molecules shows that, on average, water affected by amino acids forms stronger and shorter H-bonds than those in pure water. Differences in the influence of amino acids on water structure have been noticed. The effect of the hydrophobic side chain of an amino acid on the solvent interactions seems to be enhanced because of the specific cooperative coupling of water strong H-bond chain, connecting the carboxyl and amino groups, with the clathrate-like H-bond network surrounding the hydrocarbon side chain. The parameter derived from the spectral data, which corresponds to the contributions of the population of weak hydrogen bonds of water molecules which have been substituted by the stronger ones in the hydration sphere of amino acids, correlated well with the amino acid hydrophobicity indexes.

  8. Toward amino acid typing for proteins in FFLUX.

    PubMed

    Fletcher, Timothy L; Popelier, Paul L A

    2017-03-05

    Continuing the development of the FFLUX, a multipolar polarizable force field driven by machine learning, we present a modern approach to atom-typing and building transferable models for predicting atomic properties in proteins. Amino acid atomic charges in a peptide chain respond to the substitution of a neighboring residue and this response can be categorized in a manner similar to atom-typing. Using a machine learning method called kriging, we are able to build predictive models for an atom that is defined, not only by its local environment, but also by its neighboring residues, for a minimal additional computational cost. We found that prediction errors were up to 11 times lower when using a model specific to the correct group of neighboring residues, with a mean prediction of ∼0.0015 au. This finding suggests that atoms in a force field should be defined by more than just their immediate atomic neighbors. When comparing an atom in a single alanine to an analogous atom in a deca-alanine helix, the mean difference in charge is 0.026 au. Meanwhile, the same difference between a trialanine and a deca-alanine helix is only 0.012 au. When compared to deca-alanine models, the transferable models are up to 20 times faster to train, and require significantly less ab initio calculation, providing a practical route to modeling large biological systems. © 2016 The Authors. Journal of Computational Chemistry Published by Wiley Periodicals, Inc.

  9. Interaction of metal ions and amino acids - Possible mechanisms for the adsorption of amino acids on homoionic smectite clays

    NASA Technical Reports Server (NTRS)

    Gupta, A.; Loew, G. H.; Lawless, J.

    1983-01-01

    A semiempirical molecular orbital method is used to characterize the binding of amino acids to hexahydrated Cu(2+) and Ni(2+), a process presumed to occur when they are adsorbed in the interlamellar space of homoionic smectite clays. Five alpha-amino acids, beta-alanine, and gamma-aminobutyric acid were used to investigate the metal ion and amino acid specificity in binding. It was assumed that the alpha, beta, and gamma-amino acids would bind as bidentate anionic ligands, forming either 1:1 or 1:2 six-coordinated five, six, and seven-membered-ring chelate complexes, respectively. Energies of complex formation, optimized geometries, and electron and spin distribution were determined; and steric constraints of binding of the amino acids to the ion-exchanged cations in the interlamellar spacing of a clay were examined. Results indicate that hexahydrated Cu(2+) forms more stable complexes than hexahydrated Ni(2+) with all the amino acids studied. However, among these amino acids, complex formation does not favor the adsorption of the biological subset. Calculated energetics of complex formation and steric constraints are shown to predict that 1:1 rather than 1:2 metal-amino acid complexes are generally favored in the clay.

  10. In Vivo d-Serine Hetero-Exchange through Alanine-Serine-Cysteine (ASC) Transporters Detected by Microelectrode Biosensors

    PubMed Central

    2013-01-01

    d-Serine, a co-agonist of N-methyl d-aspartate (NMDA) receptors, has been implicated in neurological and psychiatric disorders such as cerebral ischemia, lateral amyotrophic sclerosis, or schizophrenia. d-Serine signaling represents an important pharmacological target for treating these diseases; however, the biochemical mechanisms controlling extracellular d-serine levels in vivo are still unclear. d-Serine heteroexchange through small neutral amino acid transporters has been shown in cell cultures and brain slices and could provide a biochemical mechanism for the control of d-serine extracellular concentration in vivo. Alternatively, exocytotic d-serine release has also been proposed. In this study, the dynamics of d-serine release and clearance were explored in vivo on a second-by-second time scale using microelectrode biosensors. The rate of d-serine clearance in the rat frontal cortex after a microionophoretic injection revealed a transporter-mediated uptake mechanism. d-Serine uptake was blocked by small neutral l-amino acids, implicating alanine-serine-cysteine (ASC) transporters, in particular high affinity Asc-1 and low affinity ASCT2 transporters. Interestingly, changes in alanine, serine, or threonine levels resulted in d-serine release through ASC transporters. Asc-1, but not ASCT2, appeared to release d-serine in response to changes in amino acid concentrations. Finally, neuronal silencing by tetrodotoxin increased d-serine extracellular concentration by an ASC-transporter-dependent mechanism. Together, these results indicate that d-serine heteroexchange through ASC transporters is present in vivo and may constitute a key component in the regulation of d-serine extracellular concentration. PMID:23581544

  11. Dual-column cation-exchange chromatographic method for beta-aminoisobutyric acid and beta-alanine in biological samples.

    PubMed

    Kuo, K C; Cole, T F; Gehrke, C W; Waalkes, T P; Borek, E

    1978-08-01

    A rapid, automated chromatographic method has been developed for the quantitation of the nucleic acid catabolites beta-aminoisobutyric acid and beta-alanine in urine, serum, and other physiological fluids. The analyses were performed on a modified Beckman 121M amino acid analyzer with dual ion-exchange columns and the use of a single sodium citrate buffer (pH 4.38, 0.20 mol/liter). By carefully matching the elution pattern for the two ion-exchange columns and alternating use of these columns, analyses are completed every 40 min. The chromatography, regeneration, and equilibration of the two columns are precisely programmed, thus the detector sees only the elution of beta-aminoisobutyric acid and beta-alanine alternately from each column. Long-term precision and analytical recovery for the two metabolites in urine were 1.9 and 102%, and 3.3 and 101%, respectively. Their normal physiological values were determined in human serum and urine. Their excretion in the urine was also studied as a function of collection time, to validate a more convenient, less costly method of sampling. This study shows that randomly collected samples are acceptable when the concentration of the two metabolites are expressed in terms of creatinine excretion. In addition, the distribution of the free and conjugated forms of the two metabolites in urine and serum was studied. A preparative method was also developed for the quantitative isolation of beta-amino-isobutyric acid from urine samples. The alternating dual-column technique may be applied to any ion-exchange chromatographic method where many analyses must be performed. This method is currently used in our laboratories for measuring these beta-amino acids in urine and serum of patients with various types of cancers.

  12. Dynamic monitoring of plasma amino acids and carnitine during chemotherapy of patients with alimentary canal malignancies and its clinical value

    PubMed Central

    Wang, Xiaoyu; Wang, Jiaqi; Wang, Zhenghua; Wang, Qingjun; Li, Hua

    2015-01-01

    Objective The aim of this study was to observe the plasma amino acid and carnitine characteristics in patients with metastatic gastrointestinal malignancies during chemotherapy and to identify markers for the early diagnosis and evaluation of adverse reactions and prognosis of the digestive tract malignant tumor patients. Methods Blood samples of 30 patients with metastatic gastrointestinal malignancies were collected at four time points: before chemotherapy, the first day after chemotherapy (+1 day), bone marrow depression period (+14 days), and hematopoietic recovery period (+21 days). The plasma amino acids and carnitine from those 30 patients were determined by high-performance liquid chromatography–tandem mass spectrometry method. Simultaneously, the levels of 21 amino acids were detected in 30 healthy individuals, who were considered as control. Biochemical indexes were also detected at four time points, adverse reactions were recorded during the chemotherapy process, and patients were followed up for 1 year to observe time to progression (TTP) and progression-free survival (PFS). Results Compared to healthy people in the control group, patients with malignancies showed significantly increased levels of plasma amino acids such as Arg, Asp, Cit, Gly, Orn, Tyr, Val, and carnitine (such as C2). The levels of compounds such as C3, Asn, Leu, Lys, Pip, Pro, C0, C5:1 decreased significantly before chemotherapy. The levels of Cit, Cys, Lys, Pro, Tyr, Val, C0, and C2 decreased significantly on the second day of chemotherapy (+1 day), whereas the level of C3 increased significantly. During myelosuppression (+14 days), the levels of Asp, Cit, Met, and Orn were observed to still decrease significantly, whereas the level of Val appeared to increase significantly. The levels of Asp, Glu, and Met were clearly different among patients with gastric carcinoma, rectal cancer, and colon cancer. Compared to the control group, aspartate amino transferase and alanine

  13. 21 CFR 862.1315 - Galactose-1-phosphate uridyl transferase test system.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Galactose-1-phosphate uridyl transferase test... Chemistry Test Systems § 862.1315 Galactose-1-phosphate uridyl transferase test system. (a) Identification. A galactose-1-phosphate uridyl transferase test system is a device intended to measure the...

  14. Loss of Mitochondrial Pyruvate Carrier 2 in the Liver Leads to Defects in Gluconeogenesis and Compensation via Pyruvate-Alanine Cycling.

    PubMed

    McCommis, Kyle S; Chen, Zhouji; Fu, Xiaorong; McDonald, William G; Colca, Jerry R; Kletzien, Rolf F; Burgess, Shawn C; Finck, Brian N

    2015-10-06

    Pyruvate transport across the inner mitochondrial membrane is believed to be a prerequisite for gluconeogenesis in hepatocytes, which is important for the maintenance of normoglycemia during prolonged food deprivation but also contributes to hyperglycemia in diabetes. To determine the requirement for mitochondrial pyruvate import in gluconeogenesis, mice with liver-specific deletion of mitochondrial pyruvate carrier 2 (LS-Mpc2(-/-)) were generated. Loss of MPC2 impaired, but did not completely abolish, hepatocyte conversion of labeled pyruvate to TCA cycle intermediates and glucose. Unbiased metabolomic analyses of livers from fasted LS-Mpc2(-/-) mice suggested that alterations in amino acid metabolism, including pyruvate-alanine cycling, might compensate for the loss of MPC2. Indeed, inhibition of pyruvate-alanine transamination further reduced mitochondrial pyruvate metabolism and glucose production by LS-Mpc2(-/-) hepatocytes. These data demonstrate an important role for MPC2 in controlling hepatic gluconeogenesis and illuminate a compensatory mechanism for circumventing a block in mitochondrial pyruvate import.

  15. Extraterrestrial Amino Acids in the Almahata Sitta Meteorite

    NASA Technical Reports Server (NTRS)

    Glavin, Daniel P.; Aubrey, Andrew D.; Callahan, Michael P.; Dworkin, Jason P.; Elsila, Jamie E.; Parker, Eric T.; Bada, Jeffrey L.

    2009-01-01

    Amino acid analysis of a meteorite fragment of asteroid 2008 TC(sub 3) called Almahata Sitta was carried out using reverse-phase high-perfo rmance liquid chromatography coupled with UV fluorescence detection a nd time-of-flight mass spectrometry (HPLC-FD/ToF-MS) as part of a sam ple analysis consortium. HPLC analyses of hot-water extracts from the meteorite revealed a complex distribution of two- to six-carbon aliph atic amino acids and one- to three carbon amines with abundances rang ing from 0.5 to 149 parts-per-billion (ppb). The enantiomeric ratios of the amino acids alanine, Beta-amino-n-butyric acid (Beta-ABA), 2-amino-2- methylbutanoic acid (isovaline), and 2-aminopentanoic acid (no rvaline) in the meteorite were racemic (D/L approximately 1), indicat ing that these amino acids are indigenous to the meteorite and not te rrestrial contaminants. Several other non-protein amino acids were also identified in the meteorite above background levels including alpha -aminoisobutyric acid (alpha-AIB), 4-amino-2- methybutanoic acid, 4-a mino-3-methylbutanoic acid, and 3-, 4-, and 5-aminopentanoic acid. Th e total abundances of isovaline and AlB in Almahata Sitta are approximately 1000 times lower than the abundances of these amino acids found in the CM carbonaceous meteorite Murchison. The extremely love abund ances and unusual distribution of five carbon amino acids in Almahata Sitta compared to Cl, CM, and CR carbonaceous meteorites and may be due to extensive thermal alteration of amino acids on the parent aster oid by partial melting during formation or impact shock heating.

  16. Modification of amino acids at shock pressures of 3.5 to 32 GPa

    NASA Astrophysics Data System (ADS)

    Peterson, Etta; Horz, Friedrich; Chang, Sherwood

    1997-09-01

    Amino acids were subjected to shock impact over a pressure range of 3.5 to 32 GPa both within and without meteoritic mineral matrices. The extent of amino acid destruction, racemization, and conversion to secondary amino acids was examined. Abundances of parent compounds decreased by a factor of 10 3 over this pressure range. Racemization also occurred, but some residual optical activity remained in the amino acids surviving shocks up to 32 GPa. Secondary amino acids formed in the high peak pressure range; those identified were β-alanine, glycine, alanine, γ-aminobutyric acid, and β-aminoisobutyric acid. At 30 GPa, the abundances of these daughter compounds exceeded those of the remaining initial amino acids. As the concomitant effects of high mechanical stress and temperature accompanying shocks cannot be separated in this work, their relative contribution to the observed transformations cannot be estimated. The survival of amino acids in shock experiments suggests that, after formation or emplacement of amino acids in carbonaceous chondrite parent bodies, these objects never experienced impact velocities greater than 5 km/s, which suffices to generate 30 GPa for typical silicate/silicate impacts. These results also provide guidelines for choosing appropriate capture media for interplanetary dust particles on Earth-orbiting platforms.

  17. Purification and characterization of UDP-GalNAc:polypeptide N-acetylgalactosaminyl transferase from swine trachea epithelium.

    PubMed

    Mendicino, J; Sangadala, S

    1998-08-01

    UDP-GalNac: polypeptide N-acetylgalactosaminyltransferase from swine trachea epithelium was purified to homogeneity by procedures which included affinity chromatography on Sepharose 4B columns containing bound deglycosylated Cowper's gland mucin. The enzyme, purified 12,000-fold from microsomes with a yield of 40%, showed only a single band on dodecyl sulfate polyacrylamide gel electrophoresis. The homogenous enzyme has an apparent molecular mass of 70,000 Da, as determined by gel electrophoresis or gel filtration. The transferase has a broad pH optimum between 6.7-7.8 with maximal activity at pH 7.2, and required Mn2+ for activity with maximal activity at 5-7.5 mM. Higher concentrations of Mn2+, inhibited the enzyme. The purified transferase was specific for UDPGalNAc and glycosylated both threonine and serine residues in tryptic peptides prepared from deglycosylated Cowper's gland and swine and human trachea mucins. The apparent Km of the transferase for UDPGalNAc was 6.3 microM, and the Km values for deglycosylated Cowper's gland and human and swine trachea mucins were 0.83, 1.12 and 0.94 mg/ml, respectively. The Vmax of the purified enzyme was 2.1 micromol/min/mg with deglycosylated Cowper's gland mucin, as the glycosyl acceptor. However, the activities with peptides prepared from deglycosylated mucins by limited acid hydrolysis were 20-fold greater than the intact glycoprotein under identical conditions. The deglycosylated mucins and larger peptides aggregated with time of storage and precipitated from solution. Aggregation was accompanied by a corresponding loss of enzymatic activity even after dispersion of the aggregate by sonication. The deglycosylated mucins which were prepared by chemical treatment and periodate oxidation still contained about 20% of the N-acetylgalactosamine present in the intact mucin. When this residual amino sugar was removed by periodate oxidation the completely deglycosylated mucins became very poor substrates for the purified

  18. Transferability and reproducibility in electron-density studies--bond-topological and atomic properties of tripeptides of the type L-alanyl-X-L-alanine.

    PubMed

    Grabowsky, Simon; Kalinowski, Roman; Weber, Manuela; Förster, Diana; Paulmann, Carsten; Luger, Peter

    2009-08-01

    In the last decade three different data bank approaches have been developed that are intended to make electron-density examinations of large biologically important molecules possible. They rely on Bader's concept of transferability of submolecular fragments with retention of their electronic properties. Therefore, elaborate studies on the quantification of transferability in experiment and theory are still very important. Tripeptides of the type L-alanyl-X-L-alanine (X being any of the 20 naturally encoded amino acids) serve as a model case between amino acids and proteins. The two experimental electron-density determinations (L-alanyl-L-histidinyl-L-alanine and L-alanyl-L-phenylalanyl-L-alanine, highly resolved synchrotron X-ray diffraction data sets) performed in this study and theoretical calculations on all 20 different L-alanyl-X-L-alanine molecules contribute to a better estimation of transferability in the peptide case. As a measure of reproducibility and transferability, standard deviations from averaging over bond-topological and atomic properties of atoms or bonds that are considered equal in their chemical environments were calculated. This way, transferability and reproducibility indices were introduced. It can be shown that experimental transferability indices generally slightly exceed experimental reproducibility indices and that these larger deviations can be attributed to chemical effects such as changes in the geometry (bond lengths and angles), the polarization pattern and the neighboring sphere due to crystal packing. These effects can partly be separated from each other and quantified with the help of gas-phase calculations at optimized and experimental geometries. Thus, the degree of transferability can be quantified in very narrow limits taking into account experimental errors and chemical effects.

  19. Soil Bacteria Take Up D-Amino Acids, Protect Plants

    NASA Astrophysics Data System (ADS)

    Sun, H. J.; Zhang, G.

    2011-12-01

    Recently, many groups reported D-amino acid uptake by plant roots, raising the question of whether soil D-amino acids represent a source of nitrogen or a source of toxicity. The discussion needs to be placed in the context of competition with rhizosphere bacteria. To provide this context, we followed the concentrations of D- and L-enantiomers of alanine, glutamic acid, aspartic acid, and leucine after they were added to soils in the laboratory. In all cases, the uptake of L-enantiomer began immediately and proceeded rapidly until exhausted. In contrast, the uptake of D-enantiomer required induction: an initial period of inactivity followed by rapid consumption comparable in rate to L-enantiomer. The induced nature of the D activity was confirmed by the addition of rifampicin, an mRNA synthesis inhibitor. Preventing the synthesis of new enzymes abolished soil flora's ability to consume D-amino acids, but not L-amino acids. These results suggest that inducible special racemase enzymes, which can convert D-amino acids back to their native L-forms, are widespread among soil microorganisms. This finding does not rule out the possibility that some plants may out-compete microorganisms and be able to access D-amino acids. It does suggest, however, that rhizosphere bacteria can shield plants from the toxic effect of D-amino acids.

  20. New Functions and Potential Applications of Amino Acids.

    PubMed

    Uneyama, Hisayuki; Kobayashi, Hisamine; Tonouchi, Naoto

    2016-11-22

    Currently, several types of amino acids are being produced and used worldwide. Nevertheless, several new functions of amino acids have been recently discovered that could result in other applications. For example, oral stimulation by glutamate triggers the cephalic phase response to prepare for food digestion. Further, the stomach and intestines have specific glutamate-recognizing systems in their epithelial mucosa. Regarding clinical applications, addition of monosodium glutamate to the medicinal diet has been shown to markedly enhance gastric secretion in a vagus-dependent manner. Branched-chain amino acids (BCAAs) are the major components of muscles, and ingestion of BCAAs has been found to be effective for decreasing muscle pain. BCAAs are expected to be a solution for the serious issue of aging. Further, ingestion of specific amino acids could be beneficial. Glycine can be ingested for good night's sleep: glycine ingestion before bedtime significantly improved subjective sleep quality. Ingestion of alanine and glutamine effectively accelerates alcohol metabolism, and ingestion of cystine and theanine effectively prevents colds. Finally, amino acids could be used in a novel clinical diagnostic method: the balance of amino acids in the blood could be an indicator of the risk of diseases such as cancer. These newly discovered functions of amino acids are expected to contribute to the resolution of various issues.

  1. Catabolism of amino acids in livers from cafeteria-fed rats.

    PubMed

    de Castro Ghizoni, Cristiane Vizioli; Gasparin, Fabiana Rodrigues Silva; Júnior, Antonio Sueiti Maeda; Carreño, Fernando Olinto; Constantin, Rodrigo Polimeni; Bracht, Adelar; Ishii Iwamoto, Emy Luiza; Constantin, Jorgete

    2013-01-01

    Most studies using a hypercaloric diet to induce obesity have focused on the metabolism of fat and carbohydrates. Less concern has been given to the metabolism of amino acids, despite evidence of modifications in nitrogen metabolism during obesity. The aim of this study was to evaluate amino acid metabolism in livers from cafeteria diet-induced obese rats. Blood parameters were analysed, and histological sections of livers were stained with Sudan III. The enzymatic activities of some enzymes were determined in liver homogenates. Gluconeogenesis, ureagenesis, and oxygen consumption were evaluated in rat livers perfused with glutamine, alanine, or ammonium chloride. Compared to control rats, cafeteria-fed rats demonstrated higher levels of triacylglycerol and glucose in the blood and greater accumulation of fat in livers. Gluconeogenesis and urea production in livers perfused with glutamine and alanine at higher concentrations showed a substantial reduction in cafeteria-fed rats. However, no significant difference was observed among groups perfused with ammonium chloride. The activities of the enzymes alanine aminotransferase, glutaminase, and aspartate aminotransferase in the livers were reduced in cafeteria-fed rats. Taken together, these data are consistent with the hypothesis that livers from cafeteria diet-induced obese rats exhibit a limitation in their maximal capacity to metabolise glutamine and alanine to glucose, ammonia, and urea, not because of an impairment in gluconeogenesis and/or ureagenesis, but rather due to a depression in the activities of enzymes that catalyse the initial steps of amino acid metabolism.

  2. Interaction of antimicrobial peptide with mycolyl transferase in Mycobacterium tuberculosis.

    PubMed

    Banerjee, Devjani I; Gohil, Tejas P

    2016-03-01

    It is estimated that about 40% of the Indian population are infected with tuberculosis (TB) and that ∼3,000,000 people die as a result of TB annually. TB is caused by Mycobacterium tuberculosis. In 2011, the World Health Organization declared India as having the highest TB burden worldwide. An important criteria for pathogenicity is the presence of mycolic acid linked to the protective outer membrane of bacteria. Mycolyl transferase catalyzes the transfer of mycolic acid and promotes cell wall synthesis. This is also considered as a novel target for drug-mediated intervention strategies. Here, we have attempted to understand the interaction between the antimicrobial peptide (AMP), dermcidin, and mycolyl transferase in M. tuberculosis using a computational approach. The present study was undertaken in order to elucidate the capability of AMPs to treat this bacteria, which is less sensitive to available antibiotics, and to design a novel method for new therapies.

  3. NMR chemical shifts in amino acids: Effects of environments, electric field, and amine group rotation

    SciTech Connect

    Yoon, Young-Gui; Pfrommer, Bernd G.; Louie, Steven G.; Canning, Andrew

    2002-03-03

    The authors present calculations of NMR chemical shifts in crystalline phases of some representative amino acids such as glycine, alanine, and alanyl-alanine. To get an insight on how different environments affect the chemical shifts, they study the transition from the crystalline phase to completely isolated molecules of glycine. In the crystalline limit, the shifts are dominated by intermolecular hydrogen-bonds. In the molecular limit, however, dipole electric field effects dominate the behavior of the chemical shifts. They show that it is necessary to average the chemical shifts in glycine over geometries. Tensor components are analyzed to get the angle dependent proton chemical shifts, which is a more refined characterization method.

  4. Hidden thermodynamic information in protein amino acid mutation tables

    NASA Astrophysics Data System (ADS)

    Phillips, J. C.

    2017-03-01

    We combine the standard 1992 20 × 20 substitution matrix based on block alignment, BLOSUM62, with the standard 1982 amino acid hydropathicity scale KD as well as the modern 2007 hydropathicity scale MZ, and compare the results. The 20-parameter KD and MZ hydropathicity scales have different thermodynamic character, corresponding to first- and second-order transitions. The KD and MZ comparisons show that the mutation rates reflect quantitative iteration of qualitative amino acid-phobic and -philic binary 2 × 10 properties that define quaternary 4 × 5 subgroups (but not quinary 5 × 4 subgroups), with the modern MZ bioinformatic scale giving much better results. The quaternary 5-mer MZ 4 × 5 subgroups are called mutons (Mu5). Among all hydropathicity scales, the MZ scale uniquely exhibits a smooth, deep mutational minimum at its center associated with alanine, glycine, the smallest amino acid, and histidine.

  5. [Glutathione S-transferase of alpha class from pike liver].

    PubMed

    Borvinskaia, E V; Smirnov, L P; Nemova, N N

    2013-01-01

    In this study, glutathione S-transferase (GST) was isolated from the liver of pike Esox lucius, which was homogenous according to SDS-PAGE and isoelectrofocusing. It is a homodimer with subunits mass 25235.36 Da (according to HPLC-MS/MS) and pI about 6.4. Substrate specificity, thermostability, some kinetic characteristics and optimum pH were determined. The enzyme was identified as Alpha class GST.

  6. Enhanced amine and amino acid analysis using Pacific Blue and the Mars Organic Analyzer microchip capillary electrophoresis system.

    PubMed

    Chiesl, Thomas N; Chu, Wai K; Stockton, Amanda M; Amashukeli, Xenia; Grunthaner, Frank; Mathies, Richard A

    2009-04-01

    The fluorescent amine reactive probe Pacific Blue succinimidyl ester (PB) is used for the detection of trace amounts of amines and amino acids by microchip capillary electrophoresis on the Mars Organic Analyzer (MOA). The spectral and chemical properties of PB provide a 200-fold increase in sensitivity and improved resolution compared to fluorescamine derivatization. With the use of cross injection and PB labeling, the MOA detected amino acids at concentrations as low as 75 pM (sub-parts-per-trillion). Micellar electrokinetic chromatography (MEKC) which separates PB-labeled amino acids by their hydrophobicity is also demonstrated. The optimized MEKC conditions (45 mM CHAPSO, pH 6 at 5 degrees C) effectively separated amines and 25 amino acids with enantiomeric resolution of alanine, serine, and citrulline. Samples from the Yungay Hills region in the Atacama Desert, Chile, and from the Murchison meteorite are successfully analyzed using both techniques, and amino acids are found in the parts-per-billion range. Abiotic amino acids such as beta-alanine and epsilon-aminocaprioc acid are detected along with several neutral and acidic amino acids in the Murchison sample. The Atacama Desert sample is found to contain homochiral L-alanine and L-serine indicating the presence of extant or recently extinct life.

  7. Sub-critical water hydrolysis of hog hair for amino acid production.

    PubMed

    Esteban, M B; García, A J; Ramos, P; Márquez, M C

    2010-04-01

    A recycling method using sub-critical water hydrolysis to convert hog hair from slaughterhouses into amino acids was developed. The influence of the reaction parameters such as temperature, time of reaction and initial substrate concentration were investigated in a batch reactor. The quality and quantity of amino acids in hydrolysates were determined and 17 kinds of amino acids were obtained. Under the tested conditions, the highest amino acid yield (325 mg/g protein) was reached at an initial substrate concentration of 10 g/l, a temperature of 250 degrees C and a reaction time of 60 min. A large amount of low-molecular weight amino acids, such alanine and glycine, was observed at these operating conditions. Sub-critical water hydrolysis was confirmed as an effective and practical process to recover amino acids from hog hair waste.

  8. 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.

  9. 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.

  10. Properties of Succinyl-Coenzyme A:l-Malate Coenzyme A Transferase and Its Role in the Autotrophic 3-Hydroxypropionate Cycle of Chloroflexus aurantiacus

    PubMed Central

    Friedmann, Silke; Steindorf, Astrid; Alber, Birgit E.; Fuchs, Georg

    2006-01-01

    The 3-hydroxypropionate cycle has been proposed to operate as the autotrophic CO2 fixation pathway in the phototrophic bacterium Chloroflexus aurantiacus. In this pathway, acetyl coenzyme A (acetyl-CoA) and two bicarbonate molecules are converted to malate. Acetyl-CoA is regenerated from malyl-CoA by l-malyl-CoA lyase. The enzyme forming malyl-CoA, succinyl-CoA:l-malate coenzyme A transferase, was purified. Based on the N-terminal amino acid sequence of its two subunits, the corresponding genes were identified on a gene cluster which also contains the gene for l-malyl-CoA lyase, the subsequent enzyme in the pathway. Both enzymes were severalfold up-regulated under autotrophic conditions, which is in line with their proposed function in CO2 fixation. The two CoA transferase genes were cloned and heterologously expressed in Escherichia coli, and the recombinant enzyme was purified and studied. Succinyl-CoA:l-malate CoA transferase forms a large (αβ)n complex consisting of 46- and 44-kDa subunits and catalyzes the reversible reaction succinyl-CoA + l-malate → succinate + l-malyl-CoA. It is specific for succinyl-CoA as the CoA donor but accepts l-citramalate instead of l-malate as the CoA acceptor; the corresponding d-stereoisomers are not accepted. The enzyme is a member of the class III of the CoA transferase family. The demonstration of the missing CoA transferase closes the last gap in the proposed 3-hydroxypropionate cycle. PMID:16547052

  11. Thermodynamics of Deca-alanine Folding in Water.

    PubMed

    Hazel, Anthony; Chipot, Christophe; Gumbart, James C

    2014-07-08

    The determination of the folding dynamics of polypeptides and proteins is critical in characterizing their functions in biological systems. Numerous computational models and methods have been developed for studying structure formation at the atomic level. Due to its small size and simple structure, deca-alanine is used as a model system in molecular dynamics (MD) simulations. The free energy of unfolding in vacuum has been studied extensively using the end-to-end distance of the peptide as the reaction coordinate. However, few studies have been conducted in the presence of explicit solvent. Previous results show a significant decrease in the free energy of extended conformations in water, but the α-helical state is still notably favored over the extended state. Although sufficient in vacuum, we show that end-to-end distance is incapable of capturing the full complexity of deca-alanine folding in water. Using α-helical content as a second reaction coordinate, we deduce a more descriptive free-energy landscape, one which reveals a second energy minimum in the extended conformations that is of comparable free energy to the α-helical state. Equilibrium simulations demonstrate the relative stability of the extended and α-helical states in water as well as the transition between the two states. This work reveals both the necessity and challenge of determining a proper reaction coordinate to fully characterize a given process.

  12. Formation of chloroform during chlorination of alanine in drinking water.

    PubMed

    Chu, Wen-Hai; Gao, Nai-Yun; Deng, Yang; Dong, Bing-Zhi

    2009-11-01

    Currently, dissolved nitrogenous organic matters in water, important precursors of disinfection by-products (DBPs), are of significant concern. This study was to explore the formation of chloroform (CF) during chlorination of alanine (Ala), an important nitrogenous organic compound commonly present in water sources. Our results indicated that the CF yield reached a maximum value of 0.143% at the molar ratio of chlorine atom to nitrogen atom (Cl/N)=1.0 over a Cl/N range of 0.2-5.0 (pH=7.0, reaction time=5d, and initial Ala=0.1mM). At an acidic-neutral condition (pH 4-7), the formation of CF was suppressed. However, the highest CF yield (0.227%) occurred at weakly alkaline condition (pH 8.0) (initial Ala=0.1mM, and Cl/N=1.0). The increase of Br(-) in water can increase total trihalomethanes (THMs) and bromo-THMs. However, the bromo-THMs level reached a plateau at Br(-)/Cl>0.04. Finally, based on the computation of frontier electron density and identification and measurement of key intermediates during Ala chlorination, we proposed a formation pathway of CF from Ala chlorination: Ala-->monochloro-N-alanine (MC-N-Ala)-->acetaldehyde (AAld)-->monochloroacetaldehyde acetaldehyde (MCAld)-->dichloroacetaldehyde (DCAld)-->trichloroacetaldehyde (TCAld)-->CF.

  13. Formation of Amino Acid Thioesters for Prebiotic Peptide Synthesis: Catalysis By Amino Acid Products

    NASA Technical Reports Server (NTRS)

    Weber, Arthur L.; DeVincenzi, Donald L. (Technical Monitor)

    1999-01-01

    The origin of life can be described as a series of events in which a prebiotic chemical process came increasingly under the control of its catalytic products. In our search for this prebiotic process that yielded catalytic takeover products (such as polypeptides), we have been investigating a reaction system that generates peptide-forming amino acid thioesters from formaldehyde, glycolaldehyde, and ammonia in the presence of thiols. As shown below, this model process begins by aldol condensation of formaldehyde and glycolaldehyde to give trioses and releases. These sugars then undergo beta-dehydration yielding their respective alpha-ketoaldehydes. Addition of ammonia to the alpha-ketoaldehydes yields imines which can either: (a) rearrange in the presence of thesis to give amino acid thioesters or (be react with another molecule of aldehyde to give imidazoles. This 'one-pot' reaction system operates under mild aqueous conditions, and like modem amino acid biosynthesis, uses sugar intermediates which are converted to products by energy-yielding redox reactions. Recently, we discovered that amino acids, such as the alanine reaction product, catalyze the first and second steps of the process. In the presence of ammonia the process also generates other synthetically useful products, like the important biochemical -- pyruvic acid.

  14. A transthyretin variant (alanine 49) associated with familial amyloidotic polyneuropathy in a French family.

    PubMed Central

    Benson, M D; Julien, J; Liepnieks, J; Zeldenrust, S; Benson, M D

    1993-01-01

    A transthyretin mutation was discovered in a French family with familial amyloidotic polyneuropathy originally described in 1983. The syndrome is of early onset (approximate age 35 to 40) with carpal tunnel syndrome. Death is from cardiac disease. By direct genomic DNA sequencing an A-->G mutation was found in the position corresponding to the first base of transthyretin codon 49. The predicted alanine for threonine substitution in the transthyretin protein was proven by amino acid sequencing of transthyretin isolated from the plasma of an affected subject. Since the DNA mutation does not result in the creation or abolition of a restriction endonuclease recognition site, a new DNA analysis technique was used in which site directed mutagenesis is used to create an RFLP when the introduced mutation is in proximity to the natural mutation. Using a 27 nucleotide mutagenesis primer in the PCR reaction, a new Bg1I site was created on amplification of the variant allele. Using this test, termed PCR-IMRA, four affected members of the family were shown to have the mutation. Images PMID:8095301

  15. D-Alanine in the islets of Langerhans of rat pancreas.

    PubMed

    Ota, Nobutoshi; Rubakhin, Stanislav S; Sweedler, Jonathan V

    2014-05-02

    Relatively high levels of D-alanine (D-Ala), an endogenous D-amino acid, have been found in the endocrine systems of several animals, especially in the anterior pituitary; however, its functional importance remains largely unknown. We observed D-Ala in islets of Langerhans isolated from rat pancreas in significantly higher levels than in the anterior/intermediate pituitary; specifically, 180±60 fmol D-Ala per islet (300±100 nmol/gislet), and 10±2.5 nmol/g of wet tissue in pituitary. Additionally, 12±5% of the free Ala in the islets was in the d form, almost an order of magnitude higher than the percentage of D-Ala found in the pituitary. Surprisingly, glucose stimulation of the islets resulted in D-Ala release of 0.6±0.5 fmol per islet. As D-Ala is stored in islets and released in response to changes in extracellular glucose, D-Ala may have a hormonal role.

  16. Membrane topology of aspartate:alanine antiporter AspT from Comamonas testosteroni.

    PubMed

    Fujiki, Takashi; Nanatani, Kei; Nishitani, Kei; Yagi, Kyoko; Ohnishi, Fumito; Yoneyama, Hiroshi; Uchida, Takafumi; Nakajima, Tasuku; Abea, Keietsu

    2007-01-01

    We cloned the aspT gene encoding the L-aspartate:L-alanine antiporter AspTCt in Comamonas testosteroni genomic DNA. Analysis of the nucleotide sequence revealed that C. testosteroni has an asp operon containing aspT upstream of the l-aspartate 4-decarboxylase gene, and that the gene order of the asp operon of C. testosteroni is the inverse of that of Tetragenococcus halophilus. We used proteoliposomes to confirm the transport processes of AspTCt. To elucidate the two-dimensional structure of AspTCt, we analysed its membrane topology by means of alkaline phosphatase (PhoA) and beta-lactamase (BlaM) fusion methods. The fusion analyses revealed that AspTCt has seven transmembrane segments (TMs), a large cytoplasmic loop containing approximately 200 amino acid residues between TM4 and TM5, a cytoplasmic N-terminus, and a periplasmic C-terminus. These results suggest that the orientation of the N-terminus of AspTCt differs from that of tetragenococcal AspT, even though these two AspT orthologues catalyse the same transport reactions.

  17. β-N-methylamino-L-alanine (BMAA) metabolism in the aquatic macrophyte Ceratophyllum demersum.

    PubMed

    Downing, Simoné; Esterhuizen-Londt, Maranda; Grant Downing, Timothy

    2015-10-01

    The cyanobacterial neurotoxin, β-N-methylamino-l-alanine (BMAA) bioaccumulates and biomagnifies within the environment. However, most reports on the environmental presence of BMAA focus on the presence of BMAA in animals rather than in plants. Various laboratory studies have reported that this neurotoxin, implicated in neurodegenerative disease, is rapidly taken up by various aquatic and terrestrial plants, including crop plants. In this study the metabolism of BMAA in the aquatic macrophyte, Ceratophyllum demersum, was investigated using stable isotopically labelled BMAA. Data show that the toxin is rapidly removed from the environment by the plant. However, during depuration cellular BMAA concentrations decrease considerably, without excretion of the toxin back into the environment and without catabolism of BMAA, evidenced by the absence of label transfer to other amino acids. This strongly suggests that BMAA is metabolised via covalent modification and sequestered inside the plant as a BMAA-derivative. This modification may be reversed in humans following consumption of BMAA-containing plant material. These data therefore impact on the assessment of the risk of human exposure to this neurotoxin.

  18. Biotransfer of β-N-methylamino-L-alanine (BMAA) in a eutrophicated freshwater lake.

    PubMed

    Lage, Sandra; Annadotter, Heléne; Rasmussen, Ulla; Rydberg, Sara

    2015-03-02

    β-N-Methylamino-L-alanine (BMAA), a neurotoxic non-protein amino acid, plays a significant role as an environmental risk factor in neurodegenerative diseases, such as amyotrophic lateral sclerosis. BMAA producers occur globally, colonizing almost all habitats and represent species from distinct phytoplanktonic groups, i.e., cyanobacteria, diatoms, and dinoflagellates. Bioaccumulation of BMAA in invertebrate and vertebrate organisms has also been registered around the globe. In the Baltic Sea, BMAA has been detected in several commercial fish species, raising the question of the bioaccumulation of BMAA in Swedish limnic systems. Here we find the presence of BMAA in water samples from Lake Finjasjön and identify its bioaccumulation patterns in both plankti-benthivorous and piscivorous fish, according to fish species, total weight, gender, and season of collection. For the first time, a large number of fish individuals were used in order to draw conclusions on BMAA bioaccumulation in a closed ecological community based on a statistical approach. We may, therefore, conclude that feeding patterns (plankti-benthivorous) and increased age of fish may lead to a higher tissue concentration of BMAA.

  19. Biotransfer of β-N-Methylamino-l-alanine (BMAA) in a Eutrophicated Freshwater Lake

    PubMed Central

    Lage, Sandra; Annadotter, Heléne; Rasmussen, Ulla; Rydberg, Sara

    2015-01-01

    β-N-Methylamino-l-alanine (BMAA), a neurotoxic non-protein amino acid, plays a significant role as an environmental risk factor in neurodegenerative diseases, such as amyotrophic lateral sclerosis. BMAA producers occur globally, colonizing almost all habitats and represent species from distinct phytoplanktonic groups, i.e., cyanobacteria, diatoms, and dinoflagellates. Bioaccumulation of BMAA in invertebrate and vertebrate organisms has also been registered around the globe. In the Baltic Sea, BMAA has been detected in several commercial fish species, raising the question of the bioaccumulation of BMAA in Swedish limnic systems. Here we find the presence of BMAA in water samples from Lake Finjasjön and identify its bioaccumulation patterns in both plankti-benthivorous and piscivorous fish, according to fish species, total weight, gender, and season of collection. For the first time, a large number of fish individuals were used in order to draw conclusions on BMAA bioaccumulation in a closed ecological community based on a statistical approach. We may, therefore, conclude that feeding patterns (plankti-benthivorous) and increased age of fish may lead to a higher tissue concentration of BMAA. PMID:25738330

  20. Lack of behavioral and neuropathological effects of dietary beta-methylamino-L-alanine (BMAA) in mice.

    PubMed

    Cruz-Aguado, Reyniel; Winkler, Daniella; Shaw, Christopher A

    2006-06-01

    Beta-methylamino-L-alanine (BMAA) is an excitotoxin allegedly involved in ALS-parkinsonism-dementia complex (ALS-PDC), a neurological disorder found in Guam and its surrounding islands, in which motor neuron disease symptoms can present alone or can co-occur with parkinsonism and dementia. Although in vitro experiments have shown BMAA's neurotoxic properties, studies using adult animals and systemic administration which better model the case of environmentally-induced human neurodegenerative diseases have not supported the involvement of BMAA in these disorders. In order to better test the hypothesized role of BMAA in neurodegeneration, we fed adult mice BMAA at a dose (28 mg/kg body weight, daily for 30 days) that reproduces the natural levels and tested the animals with a battery of behavioural tests, the latter including the evaluation of motor coordination, motor neuron-mediated reflexes, locomotion, muscular strength and memory. We also assessed whether BMAA exposure triggers cell death in the central nervous system (CNS) of mice by examining neuronal numbers and glial response in the spinal cord and the brain. No motor, cognitive or neuropathological outcome resulted from this feeding paradigm. Our findings support neither the causal role of BMAA in neurodegeneration nor the specific involvement of this amino acid in ALS-PDC.

  1. β-N-Methylamino-L-alanine Induces Neurological Deficits and Shortened Life Span in Drosophila

    PubMed Central

    Zhou, Xianchong; Escala, Wilfredo; Papapetropoulos, Spyridon; Zhai, R. Grace

    2010-01-01

    The neurotoxic non-protein amino acid, β-N-methylamino-L-alanine (BMAA), was first associated with the high incidence of Amyotrophic Lateral Sclerosis/Parkinsonism Dementia Complex (ALS/PDC) in Guam. Recently, BMAA has been implicated as a fierce environmental factor that contributes to the etiology of Alzheimer’s and Parkinson’s diseases, in addition to ALS. However, the toxicity of BMAA in vivo has not been clearly demonstrated. Here we report our investigation of the neurotoxicity of BMAA in Drosophila. We found that dietary intake of BMAA reduced life span, locomotor functions, and learning and memory abilities in flies. The severity of the alterations in phenotype is correlated with the concentration of BMAA detected in flies. Interestingly, developmental exposure to BMAA had limited impact on survival rate, but reduced fertility in females, and caused delayed neurological impairment in aged adults. Our studies indicate that BMAA exposure causes chronic neurotoxicity, and that Drosophila serves as a useful model in dissecting the pathogenesis of ALS/PDC. PMID:22069570

  2. Sequential micellar electrokinetic chromatography analysis of racemization reaction of alanine enantiomers.

    PubMed

    Fu, Rao; Liu, Lina; Guo, Yingna; Guo, Liping; Yang, Li

    2014-02-28

    A novel method for online monitoring racemization reaction of alanine (Ala) enantiomers was developed, by combining sequential sample injection and micellar electrokinetic chromatography (MEKC) technique. Various conditions were investigated to optimize the sequential injection, Ala derivatization and MEKC chiral separation of d-/l-Ala. High reproducibility of the sequential MEKC analysis was demonstrated by analyzing the standard Ala samples, with relative standard deviation values (n=20) of 1.35%, 1.98%, and 1.09% for peak height, peak area and migration time, respectively. Ala racemization was automatically monitored every 40s from the beginning to the end of the reaction, by simultaneous detection of the consumption of the substrate enantiomer and the formation of the product enantiomer. The Michaelis constants of the racemization reaction were obtained by the sequential MEKC method, and were in good agreement with those obtained by traditional off-line enzyme assay. Our study indicated that the present sequential MEKC method can perform fast, efficient, accurate and reproducible analysis of racemization reaction of amino acids, which is of great importance for the determination of the activity of racemase and thus understanding its metabolic functions.

  3. Enantioselective hydrogenation of pyruvic acid oxime to alanine on Pd/Alumina

    SciTech Connect

    Borszeky, K.; Mallat, T.; Aeschiman, R.

    1996-06-01

    The chemo- and enantioselective hydrogenation of pyruvic acid oxime have been studied on Pd/alumina, the latter in the presence of the 1,2-amino alcohol type alkaloids ephedrine, cinchonidine, and cinchonine. High yields of racemic alanine (90-98%) were obtained in the absence of alkaloids in polar solvents at 0-45{degrees}C and 10 bar. Enantioselection increased with higher temperature and alkalid: oxime molar ratio. A 1:1 ephedrine: oxime molar ratio afforded the best enantiomeric excess (26%). The presence of alkaloid resulted in a decrease of reaction rate by a factor of up to 140, compared to the racemic hydrogenation. Based on X-ray crystal structure analysis of the alkaloid-pyruvic acid oxime adduct, a mechanism is proposed for the steric course of the reaction. Extended interactions by multiple H bonds between the adsorbed alkaloid-oxime salt units on the Pd surface is assumed to be at the origin of the moderate enantioselectivity and the very low enantioselective hydrogenation rate. 28 refs., 5 figs., 3 tabs.

  4. Branched-chain amino acid supplementation during bed rest: effect on recovery

    NASA Technical Reports Server (NTRS)

    Stein, T. P.; Donaldson, M. R.; Leskiw, M. J.; Schluter, M. D.; Baggett, D. W.; Boden, G.

    2003-01-01

    Bed rest is associated with a loss of protein from the weight-bearing muscle. The objectives of this study are to determine whether increasing dietary branched-chain amino acids (BCAAs) during bed rest improves the anabolic response after bed rest. The study consisted of a 1-day ambulatory period, 14 days of bed rest, and a 4-day recovery period. During bed rest, dietary intake was supplemented with either 30 mmol/day each of glycine, serine, and alanine (group 1) or with 30 mmol/day each of the three BCAAs (group 2). Whole body protein synthesis was determined with U-(15)N-labeled amino acids, muscle, and selected plasma protein synthesis with l-[(2)H(5)]phenylalanine. Total glucose production and gluconeogenesis from alanine were determined with l-[U-(13)C(3)]alanine and [6,6-(2)H(2)]glucose. During bed rest, nitrogen (N) retention was greater with BCAA feeding (56 +/- 6 vs. 26 +/- 12 mg N. kg(-1). day(-1), P < 0.05). There was no effect of BCAA supplementation on either whole body, muscle, or plasma protein synthesis or the rate of 3-MeH excretion. Muscle tissue free amino acid concentrations were increased during bed rest with BCAA (0.214 +/- 0.066 vs. 0.088 +/- 0.12 nmol/mg protein, P < 0.05). Total glucose production and gluconeogenesis from alanine were unchanged with bed rest but were significantly reduced (P < 0.05) with the BCAA group in the recovery phase. In conclusion, the improved N retention during bed rest is due, at least in part, to accretion of amino acids in the tissue free amino acid pools. The amount accreted is not enough to impact protein kinetics in the recovery phase but does improve N retention by providing additional essential amino acids in the early recovery phase.

  5. Loss of Mitochondrial Pyruvate Carrier 2 in Liver Leads to Defects in Gluconeogenesis and Compensation via Pyruvate-Alanine Cycling

    PubMed Central

    McCommis, Kyle S.; Chen, Zhouji; Fu, Xiaorong; McDonald, William G.; Colca, Jerry R.; Kletzien, Rolf F.; Burgess, Shawn C.; Finck, Brian N.

    2015-01-01

    SUMMARY Pyruvate transport across the inner mitochondrial membrane is believed to be a prerequisite step for gluconeogenesis in hepatocytes, which is important for maintenance of normoglycemia during prolonged food deprivation, but also contributes to hyperglycemia in diabetes. To determine the requirement for mitochondrial pyruvate import in gluconeogenesis, mice with liver-specific deletion of mitochondrial pyruvate carrier 2 (LS-Mpc2−/−) were generated. Loss of MPC2 impaired, but did not completely abolish, hepatocyte pyruvate metabolism, labelled pyruvate conversion to TCA cycle intermediates and glucose, and glucose production from pyruvate. Unbiased metabolomic analyses of livers from fasted LS-Mpc2−/− mice suggested that alterations in amino acid metabolism, including pyruvate-alanine cycling, might compensate for loss of MPC2. Indeed, inhibition of pyruvate-alanine transamination further reduced mitochondrial pyruvate metabolism and glucose production by LS-Mpc2−/− hepatocytes. These data demonstrate an important role for MPC2 in controlling hepatic gluconeogenesis and illuminate a compensatory mechanism for circumventing a block in mitochondrial pyruvate import. PMID:26344101

  6. Plane wave density functional theory studies of the structural and the electronic properties of amino acids attached to graphene oxide via peptide bonding

    NASA Astrophysics Data System (ADS)

    Min, Byeong June; Jeong, Hae Kyung; Lee, ChangWoo

    2015-08-01

    We studied via plane wave pseudopotential total-energy calculations within the local spin density approximation (LSDA) the electronic and the structural properties of amino acids (alanine, glycine, and histidine) attached to graphene oxide (GO) by peptide bonding. The HOMO-LUMO gap, the Hirshfeld charges, and the equilibrium geometrical structures exhibit distinctive variations that depend on the species of the attached amino acid. The GO-amino acid system appears to be a good candidate for a biosensor.

  7. Functional characterization of a member of alanine or glycine: cation symporter family in halotolerant cyanobacterium Aphanothece halophytica.

    PubMed

    Bualuang, Aporn; Kageyama, Hakuto; Tanaka, Yoshito; Incharoensakdi, Aran; Takabe, Teruhiro

    2015-01-01

    Membrane proteins of amino acid-polyamine-organocation (APC) superfamily transport amino acids and amines across membranes and play important roles in the regulation of cellular processes. The alanine or glycine: cation symporter (AGCS) family belongs to APC superfamily and is found in prokaryotes, but its substrate specificity remains to be clarified. In this study, we found that a halotolerant cyanobacterium, Aphanothece halophytica has two putative ApagcS genes. The deduced amino acid sequence of one of genes, ApagcS1, exhibited high homology to Pseudomonas AgcS. The ApagcS1 gene was expressed in Escherichia coli JW4166 which is deficient in glycine uptake. Kinetics studies in JW4166 revealed that ApAgcS1 is a sodium-dependent glycine transporter. Competition experiments showed the significant inhibition by glutamine, asparagine, and glycine. The level of mRNA for ApagcS1 was induced by NaCl and nitrogen-deficient stresses. Uptake of glutamine by ApAgcS1 was also observed. Based on these data, the physiological role of ApAgcS1 was discussed.

  8. Involvement of alanine racemase in germination of Bacillus cereus spores lacking an intact exosporium.

    PubMed

    Venir, Elena; Del Torre, Manuela; Cunsolo, Vincenzo; Saletti, Rosaria; Musetti, Rita; Stecchini, Mara Lucia

    2014-02-01

    The L-alanine mediated germination of food isolated Bacillus cereus DSA 1 spores, which lacked an intact exosporium, increased in the presence of D-cycloserine (DCS), which is an alanine racemase (Alr) inhibitor, reflecting the activity of the Alr enzyme, capable of converting L-alanine to the germination inhibitor D-alanine. Proteomic analysis of the alkaline extracts of the spore proteins, which include exosporium and coat proteins, confirmed that Alr was present in the B. cereus DSA 1 spores and matched to that encoded by B. cereus ATCC 14579, whose spore germination was strongly affected by the block of conversion of L- to D-alanine. Unlike ATCC 14579 spores, L-alanine germination of B. cereus DSA 1 spores was not affected by the preincubation with DCS, suggesting a lack of restriction in the reactant accessibility.

  9. The Helical Alanine Controversy: An (Ala)6 Insertion Dramatically Increases Helicity

    PubMed Central

    Lin, Jasper C.; Barua, Bipasha

    2013-01-01

    Employing chemical shift melts and hydrogen/deuterium exchange NMR techniques, we have determined the stabilization of the Trp-cage miniprotein due to multiple alanine insertions within the N-terminal α-helix. Alanine is shown to be uniquely helix-stabilizing and this stabilization is reflected in the global fold stability of the Trp-cage. The associated free energy change per alanine can be utilized to calculate the alanine propagation value. From the Lifson–Roig formulation, the calculated value (wAla = 1.6) is comparable to those obtained for short, solubilized, alanine-rich helices and is much larger than the values obtained by prior host–guest techniques or in N-terminally templated helices and peptides bearing long contiguous strings of alanines with no capping or solubilizing units present. PMID:15493925

  10. Expression, crystallization and preliminary X-ray crystallographic analysis of Xoo0352, D-alanine-D-alanine ligase A, from Xanthomonas oryzae pv. oryzae.

    PubMed

    Doan, Thanh Thi Ngoc; Kim, Jin-Kwang; Kim, Hyesoon; Ahn, Yeh-Jin; Kim, Jeong-Gu; Lee, Byoung-Moo; Kang, Lin-Woo

    2008-12-01

    Xanthomonas oryzae pv. oryzae (Xoo) causes bacterial blight (BB), which is one of the most devastating diseases of rice in most rice-growing countries. D-Alanine-D-alanine ligase A (DdlA), coded by the Xoo0352 gene, was expressed, purified and crystallized. DdlA is an enzyme that is involved in D-alanine metabolism and the biosynthesis of an essential bacterial peptidoglycan precursor, in which it catalyzes the formation of D-alanyl-D-alanine from two D-alanines, and is thus an attractive antibacterial drug target against Xoo. The DdlA crystals diffracted to 2.3 A resolution and belonged to the primitive tetragonal space group P4(3)2(1)2, with unit-cell parameters a = b = 83.0, c = 97.6 A. There is one molecule in the asymmetric unit, with a corresponding V(M) of 1.88 A(3) Da(-1) and a solvent content of 34.6%. The initial structure was determined by molecular replacement using D-alanine-D-alanine ligase from Staphylococcus aureus (PDB code 2i87) as a template model.

  11. 1H NMR determination of beta-N-methylamino-L-alanine (L-BMAA) in environmental and biological samples.

    PubMed

    Moura, Sidnei; Ultramari, Mariah de Almeida; de Paula, Daniela Mendes Louzada; Yonamine, Mauricio; Pinto, Ernani

    2009-04-01

    A nuclear magnetic resonance (1H NMR) method for the determination of beta-N-methylamino-L-alanine (L-BMAA) in environmental aqueous samples was developed and validated. L-BMAA is a neurotoxic modified amino acid that can be produced by cyanobacteria in aqueous environments. This toxin was extracted from samples by means of solid-phase extraction (SPE) and identified and quantified by 1H NMR without further derivatization steps. The lower limit of quantification (LLOQ) was 5 microg/mL. Good inter and intra-assay precision was also observed (relative standard deviation <8.5%) with the use of 4-nitro-DL-phenylalanine as an internal standard (IS). This method of 1H NMR analysis is not time consuming and can be readily utilized to monitor L-BMAA and confirm its presence in environmental and biological samples.

  12. Stabilization Effect of Amino Acid Side Chains in Peptide Assemblies on Graphite Studied by Scanning Tunneling Microscopy.

    PubMed

    Guo, Yuanyuan; Hou, Jingfei; Zhang, Xuemei; Yang, Yanlian; Wang, Chen

    2017-02-03

    An analysis is presented of the effects of amino acid side chains on peptide assemblies in ambient conditions on a graphite surface. The molecularly resolved assemblies of binary peptides are examined with scanning tunneling microscopy. A comparative analysis of the assembly structures reveals that the lamellae width has an appreciable dependence on the peptide sequence, which could be considered as a manifestation of a stabilizing effect of side-chain moieties of amino acids with high (phenylalanine) and low (alanine, asparagine, histidine and aspartic acid) propensities for aggregation. These amino acids are representative for the chemical structures involving the side chains of charged (histidine and aspartic acid), aromatic (phenylalanine), hydrophobic (alanine), and hydrophilic (asparagine) amino acids. These results might provide useful insight for understanding the effects of sequence on the assembly of surface-bound peptides.

  13. GABA, β-alanine and glycine in the digestive juice of privet-specialist insects: convergent adaptive traits against plant iridoids.

    PubMed

    Konno, Kotaro; Hirayama, Chikara; Yasui, Hiroe; Okada, Sachiko; Sugimura, Masahiro; Yukuhiro, Fumiko; Tamura, Yasumori; Hattori, Makoto; Shinbo, Hiroshi; Nakamura, Masatoshi

    2010-09-01

    The privet tree, Ligustrum obtusifolium (Oleaceae), defends its leaves against insects with a strong lysine-decreasing activity that make proteins non-nutritive. This is caused by oleuropein, an iridoid glycoside. We previously found that some privet-specialist caterpillars adapt by secreting glycine in the digestive juice as a neutralizer that prevents the loss of lysine. Here, we extended the survey into 42 lepidopteran and hymenopteran species. The average concentration of glycine in digestive juice for 11 privet-feeding species (40.396 mM) was higher than that for 32 non-privet-feeding species (2.198 mM). The glycine concentrations exceeded 10 mM in 7 out of 11 privet-feeding species. In Macrophya timida (Hymenoptera), it reached 164.8 mM. Three out of the four remaining privet-feeding species had other amino acids instead. Larvae of a privet-specialist butterfly, Artopoetes pryeri (Lycaenidae), had a high concentration (60.812 mM) of GABA. In two other specialists, β-alanine was found. GABA, β-alanine, and glycine as well as alanine, amines, and ammonium ion inhibited the lysine decrease, indicating that amino residues are responsible for the inhibition. However, the three amino acids found in the specialists were far more effective (20 mM showed 80% inhibition) than the rest (>140 mM was required for 80% inhibition). Our results show a clear and rare case of the apparent convergent evolution of herbivores' molecular adaptations of feeding on a plant with a chemical defense in a manner that minimizes the cost of adaptation. The novel role of GABA in plant-herbivore interactions shown here is probably the first reported non-neuronal role of animal-derived GABA.

  14. GMXPBSA 2.0: A GROMACS tool to perform MM/PBSA and computational alanine scanning

    NASA Astrophysics Data System (ADS)

    Paissoni, C.; Spiliotopoulos, D.; Musco, G.; Spitaleri, A.

    2014-11-01

    GMXPBSA 2.0 is a user-friendly suite of Bash/Perl scripts for streamlining MM/PBSA calculations on structural ensembles derived from GROMACS trajectories, to automatically calculate binding free energies for protein-protein or ligand-protein complexes. GMXPBSA 2.0 is flexible and can easily be customized to specific needs. Additionally, it performs computational alanine scanning (CAS) to study the effects of ligand and/or receptor alanine mutations on the free energy of binding. Calculations require only for protein-protein or protein-ligand MD simulations. GMXPBSA 2.0 performs different comparative analysis, including a posteriori generation of alanine mutants of the wild-type complex, calculation of the binding free energy values of the mutant complexes and comparison of the results with the wild-type system. Moreover, it compares the binding free energy of different complexes trajectories, allowing the study the effects of non-alanine mutations, post-translational modifications or unnatural amino acids on the binding free energy of the system under investigation. Finally, it can calculate and rank relative affinity to the same receptor utilizing MD simulations of proteins in complex with different ligands. In order to dissect the different MM/PBSA energy contributions, including molecular mechanic (MM), electrostatic contribution to solvation (PB) and nonpolar contribution to solvation (SA), the tool combines two freely available programs: the MD simulations software GROMACS and the Poisson-Boltzmann equation solver APBS. All the calculations can be performed in single or distributed automatic fashion on a cluster facility in order to increase the calculation by dividing frames across the available processors. The program is freely available under the GPL license. Catalogue identifier: AETQ_v1_0 Program summary URL:http://cpc.cs.qub.ac.uk/summaries/AETQ_v1_0.html Program obtainable from: CPC Program Library, Queen’s University, Belfast, N. Ireland Licensing

  15. Amino Acid Chemistry as a Link Between Small Solar System Bodies and Carbonaceous Chondrites

    NASA Technical Reports Server (NTRS)

    Glavin, Daniel P.; Ehrenfreund, Pascale; Botta, Oliver; Cooper, George; Bada, Jeffrey L.

    2000-01-01

    Establishing chemical links between meteorites and small solar system bodies, such as comets and asteroids, provides a tool for investigating the processes that occurred during the formation of the solar system. Carbonaceous meteorites are of particular interest, since they may have seeded the early Earth with a variety of prebiotic organic compounds including amino acids, purines and pyrimidines, which are thought to be necessary for the origin of life. Here we report the results of high-performance liquid chromatography (HPLC) based amino acid analyses of the acid-hydrolyzed hot water extracts from pristine interior pieces of the CI carbonaceous chondrites Orgueil and Ivuna and the CM meteorites Murchison and Murray. We found that the CI meteorites Orgueil and Ivuna contained high abundances of beta-alanine and glycine, while only traces of other amino acids like alanine, alpha-amino-n-butryic acid (ABA) and alpha-aminoisobutyric acid (AIB) were detected in these meteorites. Carbon isotopic measurements of beta-alanine and glycine in Orgueil by gas chromatography combustion-isotope ratio mass spectrometry clearly indicate an extraterrestrial origin of these amino acids. The amino acid composition of Orgueil and Ivuna was strikingly different from the CM chondrites Murchison and Murray. The most notable difference was the high relative abundance of B-alanine in Orgueil and Ivuna compared to Murchison and Murray. Furthermore, AIB, which is one of the most abundant amino acids found in Murchison and Murray, was present in only trace amounts in Orgueil and Ivuna. Our amino acid data strongly suggest that the CI meteorites Orgueil and Ivuna came from a different type of parent body than the CM meteorites Murchison and Murray, possibly from an extinct comet. It is generally thought that carbonaceous meteorites are fragments of larger asteroidal bodies delivered via near Earth objects (NEO). Orbital and dynamic studies suggest that both fragments of main belt asteroids

  16. Assessing microbial utilization of free versus sorbed Alanine by using position-specific 13C labeling and 13C-PLFA analysis

    NASA Astrophysics Data System (ADS)

    Herschbach, Jennifer; Apostel, Carolin; Spielvogel, Sandra; Kuzyakov, Yakov; Dippold, Michaela

    2016-04-01

    Microbial utilization is a key transformation process of soil organic matter (SOM). Sorption of low molecular weight organic substances (LMWOS) to soil mineral surfaces blocks or delays microbial uptake and therefore mineralization of LMWOS to CO2, as well as all other biochemical transformations. We used position-specific labeling, a tool of isotope applications novel to soil science, combined with 13C-phospholipid fatty acid (PLFA) analysis, to assess microbial utilization of sorbed and non-sorbed Alanine in soil. Alanine has various functional groups enabling different sorption mechanisms via its positive charge (e.g. to clay minerals by cation exchange), as well as via its negative charge (e.g. to iron oxides by ligand exchange). To assess changes in the transformation pathways caused by sorption, we added uniformly and position-specifically 13C and 14C labeled Alanine to the Ap of a loamy Luvisol in a short-term (10 days) incubation experiment. To allow for sorption of the tracer solution to an aliquot of this soil, microbial activity was minimized in this subsample by sterilizing the soil by γ-radiation. After shaking, the remaining solutions were filtered and the non-sorbed Alanine was removed with Millipore water and then added to non-sterilized soil. For the free Alanine treatment, solutions with Alanine of similar amount and isotopic composition were prepared, added to the soil and incubated as well. The respired CO2 was trapped in NaOH and its 14C-activity was determined at increasing times intervals. Microbial utilization of Alanine's individual C positions was evaluated in distinct microbial groups classified by 13C-PLFA analysis. Sorption to soil minerals delayed respiration to CO2 and reduced initial respiration rate by 80%. Irrespective of sorption, the highest amount was respired from the carboxylic position (C-1), whereas the amino-bound (C-2) and the methylic position (C-3) were preferentially incorporated into PLFA of microorganisms due to the

  17. Carbobenzoxy amino acids: Structural requirements for cholecystokinin receptor antagonist activity

    SciTech Connect

    Maton, P.N.; Sutliff, V.E.; Jensen, R.T.; Gardner, J.D.

    1985-04-01

    The authors used dispersed acini prepared from guinea pig pancreas to examine 28 carbobenzoxy (CBZ) amino acids for their abilities to function as cholecystokinin receptor antagonists. All amino acid derivatives tested, except for CBZ-alanine, CBZ-glycine, and N alpha-CBZ- lysine, were able to inhibit the stimulation of amylase secretion caused by the C-terminal octapeptide of cholecystokinin. In general, there was a good correlation between the ability of a carbobenzoxy amino acid to inhibit stimulated amylase secretion and the ability of the amino acid derivative to inhibit binding of /sup 125/I-cholecystokinin. The inhibition of cholecystokinin-stimulated amylase secretion was competitive, fully reversible, and specific for those secretagogues that interact with the cholecystokinin receptor. The potencies with which the various carbobenzoxy amino acids inhibited the action of cholecystokinin varied 100-fold and CBZ-cystine was the most potent cholecystokinin receptor antagonist. This variation in potency was primarily but not exclusively a function of the hydrophobicity of the amino acid side chain.

  18. Antibacterial Activity of Alanine-Derived Gemini Quaternary Ammonium Compounds.

    PubMed

    Piecuch, Agata; Obłąk, Ewa; Guz-Regner, Katarzyna

    The antibacterial activity of alanine-derived gemini quaternary ammonium salts (chlorides and bromides) with various spacer and alkyl chain lengths was investigated. The studied compounds exhibited a strong bactericidal effect, especially bromides with 10 and 12 carbon alkyl chains and 3 carbon spacer groups (TMPAL-10 Br and TMPAL-12 Br), with a short contact time. Both salts dislodged biofilms of Pseudomonas aeruginosa and Staphylococcus epidermidis, and were lethal to adherent cells of S. epidermidis. Bromide with 2 carbon spacer groups and 12 carbon alkyl chains (TMEAL-12 Br) effectively reduced microbial adhesion by coating polystyrene and silicone surfaces. The results obtained suggest that, after further studies, gemini QAS might be considered as antimicrobial agents in medicine or industry.

  19. Charge dependent photodynamic activity of alanine based zinc phthalocyanines.

    PubMed

    Wang, Ao; Li, Yejing; Zhou, Lin; Yuan, Linxin; Lu, Shan; Lin, Yun; Zhou, Jiahong; Wei, Shaohua

    2014-12-01

    In this paper, to minimize the effects of different structure, three alanine-based zinc phthalocyanines (Pcs) of differing charges were engineered and synthesized with the same basic structure. On this premise, the relationship between nature of charge and photodynamic activity was studied. Besides, further verification and explanation of some inconsistent results were also carried out. The results showed that charge can influence the aggregation state, singlet oxygen generation ability and cellular uptake of Pcs, thereby affecting their photodynamic activity. In addition, the biomolecules inside cells may interact with Pcs of differing charges, which can also influence the aggregation state and singlet oxygen generation of the Pcs, and then influence the relationship between nature of charge and photodynamic activity.

  20. Amino acid analysis

    NASA Technical Reports Server (NTRS)

    Winitz, M.; Graff, J. (Inventor)

    1974-01-01

    The process and apparatus for qualitative and quantitative analysis of the amino acid content of a biological sample are presented. The sample is deposited on a cation exchange resin and then is washed with suitable solvents. The amino acids and various cations and organic material with a basic function remain on the resin. The resin is eluted with an acid eluant, and the eluate containing the amino acids is transferred to a reaction vessel where the eluant is removed. Final analysis of the purified acylated amino acid esters is accomplished by gas-liquid chromatographic techniques.

  1. Parenteral Nutrition: Amino Acids.

    PubMed

    Hoffer, Leonard John

    2017-03-10

    There is growing interest in nutrition therapies that deliver a generous amount of protein, but not a toxic amount of energy, to protein-catabolic critically ill patients. Parenteral amino acids can achieve this goal. This article summarizes the biochemical and nutritional principles that guide parenteral amino acid therapy, explains how parenteral amino acid solutions are formulated, and compares the advantages and disadvantages of different parenteral amino acid products with enterally-delivered whole protein products in the context of protein-catabolic critical illness.

  2. Parenteral Nutrition: Amino Acids

    PubMed Central

    Hoffer, Leonard John

    2017-01-01

    There is growing interest in nutrition therapies that deliver a generous amount of protein, but not a toxic amount of energy, to protein-catabolic critically ill patients. Parenteral amino acids can achieve this goal. This article summarizes the biochemical and nutritional principles that guide parenteral amino acid therapy, explains how parenteral amino acid solutions are formulated, and compares the advantages and disadvantages of different parenteral amino acid products with enterally-delivered whole protein products in the context of protein-catabolic critical illness. PMID:28287411

  3. Efficient L-Alanine Production by a Thermo-Regulated Switch in Escherichia coli.

    PubMed

    Zhou, Li; Deng, Can; Cui, Wen-Jing; Liu, Zhong-Mei; Zhou, Zhe-Min

    2016-01-01

    L-Alanine has important applications in food, pharmaceutical and veterinary and is used as a substrate for production of engineered thermoplastics. Microbial fermentation could reduce the production cost and promote the application of L-alanine. However, the presence of L-alanine significantly inhibit cell growth rate and cause a decrease in the ultimate L-alanine productivity. For efficient L-alanine production, a thermo-regulated genetic switch was designed to dynamically control the expression of L-alanine dehydrogenase (alaD) from Geobacillus stearothermophilus on the Escherichia coli B0016-060BC chromosome. The optimal cultivation conditions for the genetically switched alanine production using B0016-060BC were the following: an aerobic growth phase at 33 °C with a 1-h thermo-induction at 42 °C followed by an oxygen-limited phase at 42 °C. In a bioreactor experiment using the scaled-up conditions optimized in a shake flask, B0016-060BC accumulated 50.3 g biomass/100 g glucose during the aerobic growth phase and 96 g alanine/100 g glucose during the oxygen-limited phase, respectively. The L-alanine titer reached 120.8 g/l with higher overall and oxygen-limited volumetric productivities of 3.09 and 4.18 g/l h, respectively, using glucose as the sole carbon source. Efficient cell growth and L-alanine production were reached separately, by switching cultivation temperature. The results revealed the application of a thermo-regulated strategy for heterologous metabolic production and pointed to strategies for improving L-alanine production.

  4. Amino acid analyses of type 3 chondrites Colony, Ornans, Chainpur, and Bishunpur

    NASA Astrophysics Data System (ADS)

    Chan, H.-S.; Martins, Zita; Sephton, Mark A.

    2012-09-01

    The CO3s Colony and Ornans and LL3s Chainpur and Bishunpur were analyzed for the first time for amino acids using gas chromatography-mass spectrometry (GC-MS). Type 3 chondrites have relatively unaltered metamorphic and petrological histories. Chainpur was the most amino acid rich of the four type 3 chondrites with a total amino acid abundance of 3330 parts per billion (ppb). The other type 3 chondrites had total amino acid abundances that ranged from 660 to 1110 ppb. A D/L ratio of <0.7 for all proteic amino acids suggests at least some amino acid terrestrial contamination. However, a small fraction of indigenous extraterrestrial amino acids cannot be excluded because of the presence of the nonprotein amino acid α-aminoisobutyric acid (α-AIB), and unusually high relative abundances (to glycine) of β-alanine and γ-ABA. The comparisons between the free and total amino acid contents of the samples also indicate a low free/total amino acid ratio (ranging from about 1:4 in CO chondrites to about 1:50 in Chainpur), which indicate that amino acids are present mainly in the bound form and were made detectable after acid hydrolysis.

  5. Branched-chain amino acid administration in surgical patients. Effects on amino acid and fuel substrate profiles.

    PubMed

    Desai, S P; Bistrian, B R; Palombo, J D; Moldawer, L L; Blackburn, G L

    1987-07-01

    During the first five days following gastric bypass surgery, 15 patients received near isotonic amino acid solutions that varied in their branched-chain amino acid (BCAA) content and amino acid profiles (15.6%, 50%, or 100% BCAA solutions). Plasma valine concentrations were elevated in patients receiving 50% and 100% BCAA solutions. Plasma alanine concentrations were highest in patients receiving 50% BCAA. Plasma free fatty acids and blood lactate concentrations were unchanged by either the operation or BCAA administration. Serum glucose concentration was unaffected by the different amino acid administrations and followed the pattern induced by stress initially and later by starvation. beta-Hydroxybutyrate concentrations increased as starvation proceeded and were highest in patients receiving the 15.6% BCAA solution. Branched-chain amino acid-enriched solutions without additional energy may be administered safely to patients recovering from operative trauma. Plasma amino acid concentrations and fuel substrate profiles appear to follow metabolic patterns determined by the physiologic response to stress and starvation and can be affected by large quantities of BCAAs.

  6. Comparison of glycation of glutathione S-transferase by methylglyoxal, glucose or fructose.

    PubMed

    Boušová, Iva; Průchová, Zuzana; Trnková, Lucie; Dršata, Jaroslav

    2011-11-01

    Glycation is a process closely related to the aging and pathogenesis of diabetic complications. In this process, reactive α-dicarbonyl compounds (e.g., methylglyoxal) cause protein modification accompanied with potential loss of their biological activity and persistence of damaged molecules in tissues. We suppose that glutathione S-transferases (GSTs), a group of cytosolic biotransformation enzymes, may be modified by glycation in vivo, which would provide a rationale of its use as a model protein for studying glycation reactions. Glycation of GST by methylglyoxal, fructose, and glucose in vitro was studied. The course of protein glycation was evaluated using the following criteria: enzyme activity, formation of advanced glycation end-products using fluorescence and western blotting, amine content, protein conformation, cross linking and aggregation, and changes in molecular charge of GST. The ongoing glycation by methylglyoxal 2 mM resulted in pronounced decrease in the GST activity. It also led to the loss of 14 primary amino groups, which was accompanied by changes in protein mobility during native polyacrylamide gel electrophoresis. Formation of cross links with molecular weight of 75 kDa was observed. Obtained results can contribute to understanding of changes, which proceed in metabolism of xenobiotics during diabetes mellitus and ageing.

  7. Erythrocyte glutathione transferase: a general probe for chemical contaminations in mammals.

    PubMed

    Bocedi, A; Fabrini, R; Lai, O; Alfieri, L; Roncoroni, C; Noce, A; Pedersen, J Z; Ricci, G

    2016-01-01

    Glutathione transferases (GSTs) are enzymes devoted to the protection of cells against many different toxins. In erythrocytes, the isoenzyme (e-GST) mainly present is GSTP1-1, which is overexpressed in humans in case of increased blood toxicity, as it occurs in nephrophatic patients or in healthy subjects living in polluted areas. The present study explores the possibility that e-GST may be used as an innovative and highly sensitive biomarker of blood toxicity also for other mammals. All distinct e-GSTs from humans, Bos taurus (cow), Sus scrofa (pig), Capra hircus (goat), Equus caballus (horse), Equus asinus (donkey) and Ovis aries (sheep), show very similar amino acid sequences, identical kinetics and stability properties. Reference values for e-GST in all these mammals reared in controlled farms span from 3.5±0.2 U/gHb in the pig to 17.0±0.9 U/gHb in goat; such activity levels can easily be determined with high precision using only a few microliters of whole blood and a simple spectrophotometric assay. Possibly disturbing factors have been examined to avoid artifact determinations. This study provides the basis for future screening studies to verify if animals have been exposed to toxicologic insults. Preliminary data on cows reared in polluted areas show increased expression of e-GST, which parallels the results found for humans.

  8. Erythrocyte glutathione transferase: a general probe for chemical contaminations in mammals

    PubMed Central

    Bocedi, A; Fabrini, R; Lai, O; Alfieri, L; Roncoroni, C; Noce, A; Pedersen, JZ; Ricci, G

    2016-01-01

    Glutathione transferases (GSTs) are enzymes devoted to the protection of cells against many different toxins. In erythrocytes, the isoenzyme (e-GST) mainly present is GSTP1-1, which is overexpressed in humans in case of increased blood toxicity, as it occurs in nephrophatic patients or in healthy subjects living in polluted areas. The present study explores the possibility that e-GST may be used as an innovative and highly sensitive biomarker of blood toxicity also for other mammals. All distinct e-GSTs from humans, Bos taurus (cow), Sus scrofa (pig), Capra hircus (goat), Equus caballus (horse), Equus asinus (donkey) and Ovis aries (sheep), show very similar amino acid sequences, identical kinetics and stability properties. Reference values for e-GST in all these mammals reared in controlled farms span from 3.5±0.2 U/gHb in the pig to 17.0±0.9 U/gHb in goat; such activity levels can easily be determined with high precision using only a few microliters of whole blood and a simple spectrophotometric assay. Possibly disturbing factors have been examined to avoid artifact determinations. This study provides the basis for future screening studies to verify if animals have been exposed to toxicologic insults. Preliminary data on cows reared in polluted areas show increased expression of e-GST, which parallels the results found for humans. PMID:27551520

  9. Ghrelin O-Acyl Transferase in Zebrafish Is an Evolutionarily Conserved Peptide Upregulated During Calorie Restriction

    PubMed Central

    Hatef, Azadeh; Yufa, Roman

    2015-01-01

    Abstract Ghrelin is a multifunctional orexigenic hormone with a unique acyl modification enabled by ghrelin O-acyl transferase (GOAT). Ghrelin is well-characterized in nonmammals, and GOAT sequences of several fishes are available in the GenBank. However, endogenous GOAT in non-mammals remains poorly understood. In this research, GOAT sequence comparison, tissue-specific GOAT expression, and its regulation by nutrient status and exogenous ghrelin were studied. It was found that the bioactive core of zebrafish GOAT amino acid sequence share high identity with that of mammals. GOAT mRNA was most abundant in the gut. GOAT-like immunoreactivity (i.r.) was found colocalized with ghrelin in the gastric mucosa. Food deprivation increased, and feeding decreased GOAT and preproghrelin mRNA expression in the brain and gut. GOAT and ghrelin peptides in the gut and brain showed corresponding decrease in food-deprived state. Intraperitoneal injection of acylated fish ghrelin caused a significant decrease in GOAT mRNA expression, suggesting a feedback mechanism regulating its abundance. Together, these results provide the first in-depth characterization of GOAT in a non-mammal. Our results demonstrate that endogenous GOAT expression is responsive to metabolic status and availability of acylated ghrelin, providing further evidences for GOAT in the regulation of feeding in teleosts. PMID:26226634

  10. A glutathione S-transferase gene associated with antioxidant properties isolated from Apis cerana cerana

    NASA Astrophysics Data System (ADS)

    Liu, Shuchang; Liu, Feng; Jia, Haihong; Yan, Yan; Wang, Hongfang; Guo, Xingqi; Xu, Baohua

    2016-06-01

    Glutathione S-transferases (GSTs) are an important family of multifunctional enzymes in aerobic organisms. They play a crucial role in the detoxification of exogenous compounds, especially insecticides, and protection against oxidative stress. Most previous studies of GSTs in insects have largely focused on their role in insecticide resistance. Here, we isolated a theta class GST gene designated AccGSTT1 from Apis cerana cerana and aimed to explore its antioxidant and antibacterial attributes. Analyses of homology and phylogenetic relationships suggested that the predicted amino acid sequence of AccGSTT1 shares a high level of identity with the other hymenopteran GSTs and that it was conserved during evolution. Quantitative real-time PCR showed that AccGSTT1 is most highly expressed in adult stages and that the expression profile of this gene is significantly altered in response to various abiotic stresses. These results were confirmed using western blot analysis. Additionally, a disc diffusion assay showed that a recombinant AccGSTT1 protein may be roughly capable of inhibiting bacterial growth and that it reduces the resistance of Escherichia coli cells to multiple adverse stresses. Taken together, these data indicate that AccGSTT1 may play an important role in antioxidant processes under adverse stress conditions.

  11. Characterization of the safener-induced glutathione S-transferase isoform II from maize.

    PubMed

    Holt, D C; Lay, V J; Clarke, E D; Dinsmore, A; Jepson, I; Bright, S W; Greenland, A J

    1995-01-01

    The safener-induced maize (Zea mays L.) glutathione S-transferase, GST II (EC 2.5.1.18) and another predominant isoform, GST I, were purified from extracts of maize roots treated with the safeners R-25788 (N,N-diallyl-2-dichloroacetamide) or R-29148 (3-dichloroacetyl-2,2,5-trimethyl-1,3-oxazolidone). The isoforms GST I and GST II are respectively a homodimer of 29-kDa (GST-29) subunits and a heterodimer of 29- and 27-kDa (GST-27) subunits, while GST I is twice as active with 1-chloro-2,4-dinitrobenzene as GST II, GST II is about seven times more active against the herbicide, alachlor. Western blotting using antisera raised against GST-29 and GST-27 showed that GST-29 is present throughout the maize plant prior to safener treatment. In contrast, GST-27 is only present in roots of untreated plants but is induced in all the major aerial organs of maize after root-drenching with safener. The amino-acid sequences of proteolytic fragments of GST-27 show that it is related to GST-29 and identical to the 27-kDa subunit of GST IV.

  12. Identification, genomic organization and expression pattern of glutathione S-transferase in the silkworm, Bombyx mori.

    PubMed

    Yu, Quanyou; Lu, Cheng; Li, Bin; Fang, Shoumin; Zuo, Weidong; Dai, Fangyin; Zhang, Ze; Xiang, Zhonghuai

    2008-12-01

    Glutathione S-transferases (GSTs) are a multifunctional supergene family and some play an important role in insecticide resistance. We have identified 23 putative cytosolic GSTs by searching the new assembly of the Bombyx mori genome sequence. Phylogenetic analyses on the amino acid sequences reveal that 21 of the B. mori GSTs fall into six classes represented in other insects, the other two being unclassified. The majority of the silkworm GSTs belong to the Delta, Epsilon, and Omega classes. Most members of each class are tandemly arranged in the genome, except for the Epsilon GSTs. Expressed sequence tags (ESTs) corresponding to 19 of the 23 GSTs were found in available databases. Furthermore RT-PCR experiments detected expression of all the GSTs in multiple tissues on day 3 of fifth instar larvae. Surprisingly, we found little or no expression of most Delta and Epsilon GSTs in the fat body, which is thought to be the main detoxification organ. This may explain the sensitivity of the silkworm to certain insecticides. Our data provide some insights into the evolution of the B. mori GST family and the functions of individual GST enzymes.

  13. Cloning and functional analysis of a phosphopantetheinyl transferase superfamily gene associated with jadomycin biosynthesis in Streptomyces venezuelae ISP5230.

    PubMed

    Wang, L; McVey, J; Vining, L C

    2001-06-01

    Sequence analysis of a XhoI/SacI fragment of chromosomal DNA downstream of jadL in the Streptomyces venezuelae ISP5230 gene cluster for jadomycin biosynthesis detected a partial ORF similar in its deduced amino acid sequence to the hetI product involved in synthesizing a regulator of heterocyst spacing in ANABAENA: By probing a phage library of S. venezuelae DNA with the XhoI/SacI fragment, the authors identified and isolated a hybridizing clone. The nucleotide sequence of its DNA contained three complete ORFs (jadM, N and X) and one incomplete ORF (jadO). The jadM ORF lay immediately downstream of, and partially overlapped, jadL. It contained 786 nucleotides encoding an amino acid sequence like those of enzymes in the phosphopantetheinyl transferase family. The jadN ORF contained 1794 nucleotides and encoded an amino acid sequence resembling acyl-CoA decarboxylases, thus suggesting a role in polyketide condensation reactions. The jadX ORF was not identified, but the partial jadO showed marked similarities in its deduced amino acid sequence to NDP-hexose-2,3-dehydratases, indicating a role in forming the sugar component of jadomycin B. Expression of jadM in Escherichia coli and examination of the product by SDS-PAGE established that the ORF encoded a 29.1 kDa protein, corresponding in size to the 262 amino acid polypeptide deduced from the jadM sequence. Evidence from a Northern hybridization indicated that jadM expression is correlated with jadomycin B synthesis. Cultures of S. venezuelae ISP5230 disrupted in jadM produced only 2-5% of the wild-type titre of jadomycin B, but grew well and produced chloramphenicol normally. The authors conclude that jadM encodes a holo-ACP synthase needed primarily for jadomycin B biosynthesis.

  14. Identification of the nuclear localisation signal of O-GlcNAc transferase and its nuclear import regulation

    PubMed Central

    Seo, Hyeon Gyu; Kim, Han Byeol; Kang, Min Jueng; Ryum, Joo Hwan; Yi, Eugene C.; Cho, Jin Won

    2016-01-01

    Nucleocytoplasmic O-GlcNAc transferase (OGT) attaches a single GlcNAc to hydroxyl groups of serine and threonine residues. Although the cellular localisation of OGT is important to regulate a variety of cellular processes, the molecular mechanisms regulating the nuclear localisation of OGT is unclear. Here, we characterised three amino acids (DFP; residues 451–453) as the nuclear localisation signal of OGT and demonstrated that this motif mediated the nuclear import of non-diffusible β-galactosidase. OGT bound the importin α5 protein, and this association was abolished when the DFP motif of OGT was mutated or deleted. We also revealed that O-GlcNAcylation of Ser389, which resides in the tetratricopeptide repeats, plays an important role in the nuclear localisation of OGT. Our findings may explain how OGT, which possesses a NLS, exists in the nucleus and cytosol simultaneously. PMID:27713473

  15. 40 CFR 721.520 - Alanine, N-(2-carboxyethyl)-N-alkyl-, salt.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ...-, salt. 721.520 Section 721.520 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED... Specific Chemical Substances § 721.520 Alanine, N-(2-carboxyethyl)-N-alkyl-, salt. (a) Chemical substance... alanine, N-(2-carboxyethyl)-N- alkyl-, salt (P-89-336) is subject to reporting under this section for...

  16. 40 CFR 721.520 - Alanine, N-(2-carboxyethyl)-N-alkyl-, salt.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ...-, salt. 721.520 Section 721.520 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED... Specific Chemical Substances § 721.520 Alanine, N-(2-carboxyethyl)-N-alkyl-, salt. (a) Chemical substance... alanine, N-(2-carboxyethyl)-N- alkyl-, salt (P-89-336) is subject to reporting under this section for...

  17. Polymerization of alanine in the presence of a non-swelling montmorillonite

    NASA Technical Reports Server (NTRS)

    Paecht-Horowitz, M.; Lahav, N.

    1977-01-01

    Alanine, starting from alanine-adenylate, has been polymerized in the presence of non-swelling Al-montmorillonite. The yield of polymerization is much lower than that obtained in the presence of swelling Na-montmorillonite. The possibility that the changing interlayer spacing in Na-montmorillonite might be responsible for its catalytic properties, is discussed.

  18. Regulation of the ald gene encoding alanine dehydrogenase by AldR in Mycobacterium smegmatis.

    PubMed

    Jeong, Ji-A; Baek, Eun-Young; Kim, Si Wouk; Choi, Jong-Soon; Oh, Jeong-Il

    2013-08-01

    The regulatory gene aldR was identified 95 bp upstream of the ald gene encoding L-alanine dehydrogenase in Mycobacterium smegmatis. The AldR protein shows sequence similarity to the regulatory proteins of the Lrp/AsnC family. Using an aldR deletion mutant, we demonstrated that AldR serves as both activator and repressor for the regulation of ald gene expression, depending on the presence or absence of L-alanine. The purified AldR protein exists as a homodimer in the absence of L-alanine, while it adopts the quaternary structure of a homohexamer in the presence of L-alanine. The binding affinity of AldR for the ald control region was shown to be increased significantly by L-alanine. Two AldR binding sites (O1 and O2) with the consensus sequence GA-N₂-ATC-N₂-TC and one putative AldR binding site with the sequence GA-N₂-GTT-N₂-TC were identified upstream of the ald gene. Alanine and cysteine were demonstrated to be the effector molecules directly involved in the induction of ald expression. The cellular level of L-alanine was shown to be increased in M. smegmatis cells grown under hypoxic conditions, and the hypoxic induction of ald expression appears to be mediated by AldR, which senses the intracellular level of alanine.

  19. 40 CFR 721.520 - Alanine, N-(2-carboxyethyl)-N-alkyl-, salt.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ...-, salt. 721.520 Section 721.520 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED... Specific Chemical Substances § 721.520 Alanine, N-(2-carboxyethyl)-N-alkyl-, salt. (a) Chemical substance... alanine, N-(2-carboxyethyl)-N- alkyl-, salt (P-89-336) is subject to reporting under this section for...

  20. 40 CFR 721.520 - Alanine, N-(2-carboxyethyl)-N-alkyl-, salt.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ...-, salt. 721.520 Section 721.520 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED... Specific Chemical Substances § 721.520 Alanine, N-(2-carboxyethyl)-N-alkyl-, salt. (a) Chemical substance... alanine, N-(2-carboxyethyl)-N- alkyl-, salt (P-89-336) is subject to reporting under this section for...

  1. 40 CFR 721.520 - Alanine, N-(2-carboxyethyl)-N-alkyl-, salt.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ...-, salt. 721.520 Section 721.520 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED... Specific Chemical Substances § 721.520 Alanine, N-(2-carboxyethyl)-N-alkyl-, salt. (a) Chemical substance... alanine, N-(2-carboxyethyl)-N- alkyl-, salt (P-89-336) is subject to reporting under this section for...

  2. Glutathione S-transferase from the spruce budworm, Choristoneura fumiferana: identification, characterization, localization, cDNA cloning, and expression.

    PubMed

    Feng, Q L; Davey, K G; Pang, A S; Primavera, M; Ladd, T R; Zheng, S C; Sohi, S S; Retnakaran, A; Palli, S R

    1999-09-01

    A 23-kDa protein that was present at higher levels in diapausing 2nd instar larvae than in feeding 2nd instar larvae of Choristoneura fumiferana was purified, and polyclonal antibodies were raised against this protein. The antibodies were subsequently used to screen a cDNA library that was constructed using RNA from 2nd instar larvae. Eight identical cDNA clones were isolated. The cDNA clone had a 665-bp insert and the longest open reading frame coded for a 203-amino acid protein with a predicted molecular mass of 23.37 kDa. The deduced amino acid sequence showed high similarity to glutathione S-transferases and therefore, the cDNA clone was named C. fumiferana glutathione S-transferase (CfGST). Identity of CfGST was confirmed by using affinity-purification as well as enzyme activity assay. CfGST was closer in similarity to insect GST2 members than GST1 members. The apparent Vmax of the purified CfGST towards the substrates glutathione and 1-chloro-2,4-dinitrobenezene (CDNB) were similar. However, the enzyme had a three-fold higher affinity towards CDNB than glutathione. Analyses using Northern blot, immunoblot and immunocytochemistry demonstrated that the fat body was the major tissue where the enzyme was synthesized and stored. Higher levels of CfGST protein were present in diapausing 2nd instar larvae compared to feeding 2nd and 6th instar larvae, suggesting that besides detoxification CfGST may have other roles during insect development that are not readily apparent at present. The CfGST cDNA was expressed in a recombinant baculovirus expression system and an active enzyme was produced.

  3. Amino acid metabolism by perfused rat hindquarter. Effects of insulin, leucine and 2-chloro-4-methylvalerate.

    PubMed Central

    Davis, E J; Lee, S H

    1985-01-01

    Hindquarters from starved rats were perfused without substrates but in the presence of an O2- and CO2-carrying perfluorocarbon emulsion to evaluate principally the metabolism of individual endogenous and protein-derived amino acids by this muscle preparation. This experimental model was shown, by a battery of metabolite measurements, to maintain cellular homoeostasis for at least 2h. The net appearance of most amino acids closely approximated their frequency of occurrence in muscle proteins, showing that they are not significantly metabolized. Exceptions were the branched-chain amino acids, methionine and those amino acids that are interconvertible with intermediates of the citrate cycle and pyruvate through coupled transaminations. The evidence indicates that only valine, isoleucine, aspartate and probably methionine can be catabolized by skeletal muscle to provide carbon precursors for glutamate/glutamine and alanine that are formed de novo by protein-catabolic muscle. The protein-sparing effects of insulin and leucine were confirmed. Although each decreased proteolysis and the net appearance of free amino acids, they were generally without effect on the ratios of amino acids formed. 2-Chloro-4-methylvalerate selectively stimulated the removal rate for the branched-chain amino acids, confirming the idea that the branched-chain oxo acid dehydrogenase normally limits the rate of their oxidation by muscle. It is also concluded that, since alanine was not formed in excess of that found in muscle proteins when no glucose was added as substrate, the excess of alanine (carbon) released from muscles in other studies is derived to a large extent, but not exclusively, from preformed carbohydrate. PMID:3899101

  4. Unusual nonterrestrial L-proteinogenic amino acid excesses in the Tagish Lake meteorite

    NASA Astrophysics Data System (ADS)

    Glavin, Daniel P.; Elsila, Jamie E.; Burton, Aaron S.; Callahan, Michael P.; Dworkin, Jason P.; Hilts, Robert W.; Herd, Christopher D. K.

    2012-08-01

    The distribution and isotopic and enantiomeric compositions of amino acids found in three distinct fragments of the Tagish Lake C2-type carbonaceous chondrite were investigated via liquid chromatography with fluorescence detection and time-of-flight mass spectrometry and gas chromatography isotope ratio mass spectrometry. Large L-enantiomeric excesses (Lee ˜ 43-59%) of the α-hydrogen aspartic and glutamic amino acids were measured in Tagish Lake, whereas alanine, another α-hydrogen protein amino acid, was found to be nearly racemic (D ≈ L) using both techniques. Carbon isotope measurements of D- and L-aspartic acid and D- and L-alanine in Tagish Lake fall well outside of the terrestrial range and indicate that the measured aspartic acid enantioenrichment is indigenous to the meteorite. Alternate explanations for the L-excesses of aspartic acid such as interference from other compounds present in the sample, analytical biases, or terrestrial amino acid contamination were investigated and rejected. These results can be explained by differences in the solid-solution phase behavior of aspartic acid, which can form conglomerate enantiopure solids during crystallization, and alanine, which can only form racemic crystals. Amplification of a small initial L-enantiomer excess during aqueous alteration on the meteorite parent body could have led to the large L-enrichments observed for aspartic acid and other conglomerate amino acids in Tagish Lake. The detection of nonterrestrial L-proteinogenic amino acid excesses in the Tagish Lake meteorite provides support for the hypothesis that significant enantiomeric enrichments for some amino acids could form by abiotic processes prior to the emergence of life.

  5. Unusual Nonterrestrial L-proteinogenic Amino Acid excesses in the Tagish Lake Meteorite

    NASA Technical Reports Server (NTRS)

    Glavin, Daniel P.; Elsila, Jamie E.; Burton, Aaron S.; Callahan, Michael P.; Dworkin, Jason P.; Hilts, Robert W.; Herd, D. K.

    2012-01-01

    The distribution and isotopic and enantiomeric compositions of amino acids found in three distinct fragments of the Tagish Lake C2-type carbonaceous chondrite were investigated via liquid chromatography with fluorescence detection and time-of-flight mass spectrometry and gas chromatography isotope ratio mass spectrometry. Large L-enantiomeric excesses (L(sub ee) approximately 43-59%) of the alpha-hydrogen aspartic and glutamic amino acids were measured in Tagish Lake, whereas alanine, another alpha hydrogen protein amino acid, was found to be nearly racemic (D much approximately L) using both techniques. Carbon isotope measurements of D- and L-aspartic acid and 1)- and L-alanine in Tagish Lake fall well outside of the terrestrial range and indicate that the measured aspartic acid enantioenrichment is indigenous to the meteorite. Alternate explanations for the L-excesses of aspartic acid such as interference from other compounds present in the sample, analytical biases, or terrestrial amino acid contamination were investigated and rejected. These results can be explained by differences in the solid-solution phase behavior of aspartic acid, which can form conglomerate enantiopure solids during crystallization, and alanine, which can only form racemic crystals. Amplification of a small initial L-enantiomer excess during aqueous alteration on the meteorite parent body could have led to the large L-enrichments observed for aspartic acid and other conglomerate amino acids in Tagish Lake. The detection of non terrestrial L-proteinogenic amino acid excesses in the Tagish Lake meteorite provides support for the hypothesis that significant enantiomeric enrichments for some amino acids could form by abiotic processes prior to the emergence of life.

  6. Nitrogen in dietary glutamate is utilized exclusively for the synthesis of amino acids in the rat intestine.

    PubMed

    Nakamura, Hidehiro; Kawamata, Yasuko; Kuwahara, Tomomi; Torii, Kunio; Sakai, Ryosei

    2013-01-01

    Although previous studies have shown that virtually the entire carbon skeleton of dietary glutamate (glutamate-C) is metabolized in the gut for energy production and amino acid synthesis, little is known regarding the fate of dietary glutamate nitrogen (glutamate-N). In this study, we hypothesized that dietary glutamate-N is an effective nitrogen source for amino acid synthesis and investigated the fate of dietary glutamate-N using [(15)N]glutamate. Fischer male rats were given hourly meals containing [U-(13)C]- or [(15)N]glutamate. The concentration and isotopic enrichment of several amino acids were measured after 0-9 h of feeding, and the net release of each amino acid into the portal vein was calculated. Most of the dietary glutamate-C was metabolized into CO(2), lactate, or alanine (56, 13, and 12% of the dietary input, respectively) in the portal drained viscera (PDV). Most of the glutamate-N was utilized for the synthesis of other amino acids such as alanine and citrulline (75 and 3% of dietary input, respectively) in the PDV, and only minor amounts were released into the portal vein in the form of ammonia and glutamate (2 and 3% of the dietary input, respectively). Substantial incorporation of (15)N into systemic amino acids such as alanine, glutamine, and proline, amino acids of the urea cycle, and branched-chain amino acids was also evident. These results provide quantitative evidence that dietary glutamate-N distributes extensively to amino acids synthesized in the PDV and, consequently, to circulating amino acids.

  7. 21 CFR 862.1315 - Galactose-1-phosphate uridyl transferase test system.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... of the enzyme galactose-1-phosphate uridyl transferase in erythrocytes (red blood cells... hereditary disease galactosemia (disorder of galactose metabolism) in infants. (b) Classification. Class II....

  8. 21 CFR 862.1315 - Galactose-1-phosphate uridyl transferase test system.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... of the enzyme galactose-1-phosphate uridyl transferase in erythrocytes (red blood cells... hereditary disease galactosemia (disorder of galactose metabolism) in infants. (b) Classification. Class II....

  9. 21 CFR 862.1315 - Galactose-1-phosphate uridyl transferase test system.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry Test Systems § 862.1315 Galactose-1-phosphate uridyl transferase test system. (a)...

  10. 21 CFR 862.1315 - Galactose-1-phosphate uridyl transferase test system.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry Test Systems § 862.1315 Galactose-1-phosphate uridyl transferase test system. (a)...

  11. Using Position-Specific 13C and 14C Labeling and 13C-PLFA Analysis to Assess Microbial Transformations of Free Versus Sorbed Alanine

    NASA Astrophysics Data System (ADS)

    Apostel, C.; Herschbach, J.; Bore, E. K.; Kuzyakov, Y.; Dippold, M. A.

    2015-12-01

    Sorption of charged or partially charged low molecular weight organic substances (LMWOS) to soil mineral surfaces delays microbial uptake and therefore mineralization of LMWOS to CO2, as well as all other biochemical transformations. We used position-specific labeling, a tool of isotope applications novel to soil sciences, to compare the transformation mechanisms of sorbed and non-sorbed alanine in soil. Alanine as an amino acid links C- and N-cycles in soil and therefore is a model substance for the pool of LMWOS. To assess transformations of sorbed alanine, we added position-specific and uniformly 13C and 14C labeled alanine tracer to soil that had previously been sterilized by γ-radiation. The labeled soil was added to non-sterilized soil from the same site and incubated. Soil labeled with the same tracers without previous sorption was prepared and incubated as well. We captured the respired CO2 and determined its 14C-activity at increasing time intervals. The incorporation of 14C into microbial biomass was determined by chloroform fumigation extraction (CFE), and utilization of individual C positions by distinct microbial groups was evaluated by 13C-phospholipid fatty acid analysis (PLFA). A dual peak in the respired CO2 revealed two sorption mechanisms. To compare the fate of individual C atoms independent of their concentration and pool size in soil, we applied the divergence index (DI). The DI reveals the convergent or divergent behavior of C from individual molecule positions during microbial utilization. Alanine C-1 position was mainly oxidized to CO2, while its C-2 and C-3 were preferentially incorporated in microbial biomass and PLFA. This indicates that sorption by the COOH group does not protect this group from preferential oxidation. Microbial metabolism was determinative for the preferential oxidation of individual molecule positions. The use of position-specific labeling revealed mechanisms and kinetics of microbial utilization of sorbed and non

  12. Amino Acid Crossword Puzzle

    ERIC Educational Resources Information Center

    Sims, Paul A.

    2011-01-01

    Learning the 20 standard amino acids is an essential component of an introductory course in biochemistry. Later in the course, the students study metabolism and learn about various catabolic and anabolic pathways involving amino acids. Learning new material or concepts often is easier if one can connect the new material to what one already knows;…

  13. The sugar model: catalysis by amines and amino acid products

    NASA Technical Reports Server (NTRS)

    Weber, A. L.

    2001-01-01

    Ammonia and amines (including amino acids) were shown to catalyze the formation of sugars from formaldehyde and glycolaldehyde, and the subsequent conversion of sugars to carbonylcontaining products under the conditions studied (pH 5.5 and 50 degrees C). Sterically unhindered primary amines were better catalysts than ammonia, secondary amines, and sterically hindered primary amines (i.e. alpha-aminoisobutyric acid). Reactions catalyzed by primary amines initially consumed formaldehyde and glycolaldehyde about 15-20 times faster than an uncatalyzed control reaction. The amine-catalyzed reactions yielded aldotriose (glyceraldehyde), ketotriose (dihydroxyacetone), aldotetroses (erythrose and threose), ketotetrose (erythrulose), pyruvaldehyde, acetaldehyde, glyoxal, pyruvate, glyoxylate, and several unindentified carbonyl products. The concentrations of the carbonyl products, except pyruvate and ketotetrose, initially increased and then declined during the reaction, indicating their ultimate conversion to other products (like larger sugars or pyruvate). The uncatalyzed control reaction yielded no pyruvate or glyoxylate, and only trace amounts of pyruvaldehyde, acetaldehyde and glyoxal. In the presence of 15 mM catalytic primary amine, such as alanine, the rates of triose and pyruvaldehyde of synthesis were about 15-times and 1200-times faster, respectively, than the uncatalyzed reaction. Since previous studies established that alanine is synthesized from glycolaldehyde and formaldehyde via pyruvaldehyde as its direct precursor, the demonstration that the alanine catalyzes the conversion of glycolaldehyde and formaldehyde to pyruvaldehyde indicates that this synthetic pathway is capable of autocatalysis. The relevance of this synthetic process, named the Sugar Model, to the origin of life is discussed.

  14. Nucleotidyl transferase assisted DNA labeling with different click chemistries.

    PubMed

    Winz, Marie-Luise; Linder, Eva Christina; André, Timon; Becker, Juliane; Jäschke, Andres

    2015-09-30

    Here, we present a simple, modular and efficient strategy that allows the 3'-terminal labeling of DNA, regardless of whether it has been chemically or enzymatically synthesized or isolated from natural sources. We first incorporate a range of modified nucleotides at the 3'-terminus, using terminal deoxynucleotidyl transferase. In the second step, we convert the incorporated nucleotides, using either of four highly efficient click chemistry-type reactions, namely copper-catalyzed azide-alkyne cycloaddition, strain-promoted azide-alkyne cycloaddition, Staudinger ligation or Diels-Alder reaction with inverse electron demand. Moreover, we create internal modifications, making use of either ligation or primer extension, after the nucleotidyl transferase step, prior to the click reaction. We further study the influence of linker variants on the reactivity of azides in different click reactions. We find that different click reactions exhibit distinct substrate preferences, a fact that is often overlooked, but should be considered when labeling oligonucleotides or other biomolecules with click chemistry. Finally, our findings allowed us to extend our previously published RNA labeling strategy to the use of a different copper-free click chemistry, namely the Staudinger ligation.

  15. Characterization of glutathione S-transferase of Taenia solium.

    PubMed

    Vibanco-Pérez, N; Jiménez, L; Merchant, M T; Landa, A

    1999-06-01

    A Taenia solium glutathione-S-transferase fraction (SGSTF) was isolated from a metacestode crude extract by affinity chromatography on reduced glutathione (GSH)-sepharose. The purified fraction displayed a specific glutathione S-transferase (GST) activity of 2.8 micromol/min/mg and glutathione peroxidase selenium-independent activity of 0.22 micromol/min/mg. Enzymatic characterization of the fraction suggested that the activity was closer to the mammalian mu-class GSTs. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis, gel filtration, and enzyme activity analysis showed that the fraction was composed of a major band of Mr = 26 kd and that the active enzyme was dimeric. Immunohistochemical studies using specific antibodies against the major 26-kd band of the SGSTF indicated that GST protein was present in the tegument, parenchyma, protonephridial, and tegumentary cytons of the T. solium metacestode. Antibodies generated against the SGSTF tested in western blot showed cross-reactivity against GSTs purified from Taenia saginata, T. taeniaeformis, and T. crassiceps, but did not react with GSTs from Schistosoma mansoni, or mice, rabbit, and pig liver tissue. Furthermore, immunization of mice with SGSTF reduced the metacestode burden up to 74.2%. Our findings argue in favor of GST having an important role in the survival of T. solium in its hosts.

  16. Characterization of two Arabidopsis thaliana glutathione S-transferases.

    PubMed

    Nutricati, Eliana; Miceli, Antonio; Blando, Federica; De Bellis, Luigi

    2006-09-01

    Glutathione S-transferases (GST) are multifunctional proteins encoded by a large gene family, divided on the basis of sequence identity into phi, tau, theta, zeta and lambda classes. The phi and tau classes are present only in plants. GSTs appear to be ubiquitous in plants and are involved in herbicide detoxification and stress response, but little is known about the precise role of GSTs in normal plant physiology and during biotic and abiotic stress response. Two cDNAs representing the two plant classes tau and phi, AtGSTF9 and AtGSTU26, were expressed in vitro and the corresponding proteins were analysed. Both GSTs were able to catalyse a glutathione conjugation to 1-chloro-2,4-dinitrobenzene (CDNB), but they were inactive as transferases towards p-nitrobenzylchloride (pNBC). AtGSTF9 showed activity towards benzyl isothiocyanate (BITC) and an activity as glutathione peroxidase with cumene hydroperoxide (CumHPO). AtGSTU26 was not active as glutathione peroxidase and towards BITC. RT-PCR analysis was used to evaluate the expression of the two genes in response to treatment with herbicides and safeners, chemicals, low and high temperature. Our results reveal that AtGSTU26 is induced by the chloroacetanilide herbicides alachlor and metolachlor and the safener benoxacor, and after exposure to low temperatures. In contrast, AtGSTF9 seems not to be influenced by the treatments employed.

  17. Nucleotidyl transferase assisted DNA labeling with different click chemistries

    PubMed Central

    Winz, Marie-Luise; Linder, Eva Christina; André, Timon; Becker, Juliane; Jäschke, Andres

    2015-01-01

    Here, we present a simple, modular and efficient strategy that allows the 3′-terminal labeling of DNA, regardless of whether it has been chemically or enzymatically synthesized or isolated from natural sources. We first incorporate a range of modified nucleotides at the 3′-terminus, using terminal deoxynucleotidyl transferase. In the second step, we convert the incorporated nucleotides, using either of four highly efficient click chemistry-type reactions, namely copper-catalyzed azide-alkyne cycloaddition, strain-promoted azide-alkyne cycloaddition, Staudinger ligation or Diels-Alder reaction with inverse electron demand. Moreover, we create internal modifications, making use of either ligation or primer extension, after the nucleotidyl transferase step, prior to the click reaction. We further study the influence of linker variants on the reactivity of azides in different click reactions. We find that different click reactions exhibit distinct substrate preferences, a fact that is often overlooked, but should be considered when labeling oligonucleotides or other biomolecules with click chemistry. Finally, our findings allowed us to extend our previously published RNA labeling strategy to the use of a different copper-free click chemistry, namely the Staudinger ligation. PMID:26013812

  18. E. coli histidine triad nucleotide binding protein 1 (ecHinT) is a catalytic regulator of D-alanine dehydrogenase (DadA) activity in vivo.

    PubMed

    Bardaweel, Sanaa; Ghosh, Brahma; Chou, Tsui-Fen; Sadowsky, Michael J; Wagner, Carston R

    2011-01-01

    Histidine triad nucleotide binding proteins (Hints) are highly conserved members of the histidine triad (HIT) protein superfamily. Hints comprise the most ancient branch of this superfamily and can be found in Archaea, Bacteria, and Eukaryota. Prokaryotic genomes, including a wide diversity of both gram-negative and gram-positive bacteria, typically have one Hint gene encoded by hinT (ycfF in E. coli). Despite their ubiquity, the foundational reason for the wide-spread conservation of Hints across all kingdoms of life remains a mystery. In this study, we used a combination of phenotypic screening and complementation analyses with wild-type and hinT knock-out Escherichia coli strains to show that catalytically active ecHinT is required in E. coli for growth on D-alanine as a sole carbon source. We demonstrate that the expression of catalytically active ecHinT is essential for the activity of the enzyme D-alanine dehydrogenase (DadA) (equivalent to D-amino acid oxidase in eukaryotes), a necessary component of the D-alanine catabolic pathway. Site-directed mutagenesis studies revealed that catalytically active C-terminal mutants of ecHinT are unable to activate DadA activity. In addition, we have designed and synthesized the first cell-permeable inhibitor of ecHinT and demonstrated that the wild-type E. coli treated with the inhibitor exhibited the same phenotype observed for the hinT knock-out strain. These results reveal that the catalytic activity and structure of ecHinT is essential for DadA function and therefore alanine metabolism in E. coli. Moreover, they provide the first biochemical evidence linking the catalytic activity of this ubiquitous protein to the biological function of Hints in Escherichia coli.

  19. Microbial transformations of free versus sorbed alanine analyzed by position-specific 13C and 14C labeling and 13C-PLFA analysis

    NASA Astrophysics Data System (ADS)

    Apostel, Carolin; Dippold, Michaela; Bore, Ezekiel; Kuzyakov, Yakov

    2015-04-01

    Sorption of charged or partially charged low molecular weight organic substances (LMWOS) to soil mineral surfaces delays microbial uptake and therefore mineralization of LMWOS to CO2, as well as all other biochemical transformations. We used position-specific labeling, a tool of isotope applications novel to soil sciences, to compare the transformation mechanisms of sorbed and non-sorbed alanine in soil. Alanine as an amino acid links C- and N-cycles in soil and therefore is a model representative for the pool of LMWOS. To assess transformations of sorbed alanine, we combined position-specifically and uniformly 13C and 14C labeled alanine tracer solution with a loamy haplic luvisol that had previously been sterilized by γ-radiation. After shaking the mixtures, the supernatant was removed, as was all non-sorbed alanine by repeated shaking with millipore water. The labeled soil was added to non-sterilized soil from the same site. To compare the effect of sorption, soil labeled with the same position-specifically labeled tracers without previous sorption was prepared and incubated as well. We captured the respired CO2 and determined its 14C-activity at increasing time steps. The incorporation of 14C into microbial biomass was determined by CFE, and utilization of individual C positions by distinct microbial groups was evaluated by 13C-PLFA analysis. A dual peak in the respired CO2 revealed the influence of two sorption mechanisms. Microbial uptake and transformation of the sorbed alanine was 3 times slower compared to non-sorbed alanine. To compare the fate of individual C atoms independent of their concentration and pool size in soil, we introduced the divergence index (DI). The DI reveals the convergent or divergent behaviour of C from individual molecule positions during microbial utilization. The DI revealed, that alanines C-1 position was mainly oxidized to CO2, while its C-2 and C-3 were preferentially incorporated in microbial biomass and PLFAs. This indicates

  20. Overview of the potent cyanobacterial neurotoxin β-methylamino-L-alanine (BMAA) and its analytical determination.

    PubMed

    Porojan, Cristina; Mitrovic, Simon M; Yeo, Darren C J; Furey, Ambrose

    2016-10-01

    Blue-green algae are responsible for the production of different types of toxins which can be neurotoxic, hepatotoxic, cytotoxic and dermatotoxic and that can affect both aquatic and terrestrial life. Since its discovery the neurotoxin β-methylamino-L-alanine (BMAA) has been a cause for concern, being associated with the neurodegenerative disease amyotrophic lateral sclerosis/Parkinsonism-dementia complex (ALS/PDC). The initial focus was on Guam where it was observed that a high number of people were affected by the ALS/PDC complex. Subsequently, researchers were surprised to find levels of BMAA in post mortem brains from Canadian patients who also suffered from ALS/PDC. Recent research demonstrates that BMAA has been found at different levels in the aquatic food web in the brackish waters of the Baltic Sea. There is emerging evidence to suggest that sand-borne algae from Qatar can also contain BMAA. Furthermore, there is now concern because BMAA has been found not only in warmer regions of the world but also in temperate regions like Europe. The aim of this review is to focus on the methods of extraction and analysis of the neurotoxic non-protein amino acid BMAA. We also consider the neurotoxicity, aetiology, and diverse sources and routes of exposure to BMAA. In recent years, different methods have been developed for the analysis of BMAA. Some of these use HPLC-FD, UPLC-UV, UPLC-MS and LC-MS/MS using samples that have been derivatised or underivatised. To date the LC-MS/MS approach is the most widely used analytical technique as it is the most selective and sensitive method for BMAA determination.

  1. Alanine-scanning mutagenesis reveals a cytosine deaminase mutant with altered substrate preference.

    PubMed

    Mahan, Sheri D; Ireton, Greg C; Stoddard, Barry L; Black, Margaret E

    2004-07-20

    Suicide gene therapy of cancer is a method whereby cancerous tumors can be selectively eradicated while sparing damage to normal tissue. This is accomplished by delivering a gene, encoding an enzyme capable of specifically converting a nontoxic prodrug into a cytotoxin, to cancer cells followed by prodrug administration. The Escherichia coli gene, codA, encodes cytosine deaminase and is introduced into cancer cells followed by administration of the prodrug 5-fluorocytosine (5-FC). Cytosine deaminase converts 5-FC into cytotoxic 5-fluorouracil, which leads to tumor-cell eradication. One limitation of this enzyme/prodrug combination is that 5-FC is a poor substrate for bacterial cytosine deaminase. The crystal structure of bacterial cytosine deaminase (bCD) reveals that a loop structure in the active site pocket of wild-type bCD comprising residues 310-320 undergoes a conformational change upon cytosine binding, making several contacts to the pyrimidine ring. Alanine-scanning mutagenesis was used to investigate the structure-function relationship of amino acid residues within this region, especially with regard to substrate specificity. Using an E. coli genetic complementation system, seven active mutants were identified (F310A, G311A, H312A, D314A, V315A, F316A, and P318A). Further characterization of these mutants reveals that mutant F316A is 14-fold more efficient than the wild-type at deaminating cytosine to uracil. The mutant D314A enzyme demonstrates a dramatic decrease in cytosine activity (17-fold) as well as a slight increase in activity toward 5-FC (2-fold), indicating that mutant D314A prefers the prodrug over cytosine by almost 20-fold, suggesting that it may be a superior suicide gene.

  2. Instantaneous normal mode analysis of the vibrational relaxation of the amide I mode of alanine dipeptide in water.

    PubMed

    Farag, Marwa H; Zúñiga, José; Requena, Alberto; Bastida, Adolfo

    2013-05-28

    Nonequilibrium Molecular Dynamics (MD) simulations coupled to instantaneous normal modes (INMs) analysis are used to study the vibrational relaxation of the acetyl and amino-end amide I modes of the alanine dipeptide (AlaD) molecule dissolved in water (D2O). The INMs are assigned in terms of the equilibrium normal modes using the Effective Atomic Min-Cost algorithm as adapted to make use of the outputs of standard MD packages, a method which is well suited for the description of flexible molecules. The relaxation energy curves of both amide I modes show multiexponential decays, in good agreement with the experimental findings. It is found that ~85%-90% of the energy relaxes through intramolecular vibrational redistribution. The main relaxation pathways are also identified. The rate at which energy is transferred into the solvent is similar for the acetyl-end and amino-end amide I modes. The conformational changes occurring during relaxation are investigated, showing that the populations of the alpha and beta region conformers are altered by energy transfer in such a way that it takes 15 ps for the equilibrium conformational populations to be recovered after the initial excitation of the AlaD molecule.

  3. Solution nonideality related to solute molecular characteristics of amino acids.

    PubMed Central

    Keener, C R; Fullerton, G D; Cameron, I L; Xiong, J

    1995-01-01

    By measuring the freezing-point depression for dilute, aqueous solutions of all water-soluble amino acids, we test the hypothesis that nonideality in aqueous solutions is due to solute-induced water structuring near hydrophobic surfaces and solute-induced water destructuring in the dipolar electric fields generated by the solute. Nonideality is expressed with a single solute/solvent interaction parameter I, calculated from experimental measure of delta T. A related parameter, I(n), gives a method of directly relating solute characteristics to solute-induced water structuring or destructuring. I(n)-values correlate directly with hydrophobic surface area and inversely with dipolar strength. By comparing the nonideality of amino acids with progressively larger hydrophobic side chains, structuring is shown to increase with hydrophobic surface area at a rate of one perturbed water molecule per 8.8 square angstroms, implying monolayer coverage. Destructuring is attributed to dielectric realignment as described by the Debye-Hückel theory, but with a constant separation of charges in the amino-carboxyl dipole. By using dimers and trimers of glycine and alanine, this destructuring is shown to increase with increasing dipole strength using increased separation of fixed dipolar charges. The capacity to predict nonideal solution behavior on the basis of amino acid characteristics will permit prediction of free energy of transfer to water, which may help predict the energetics of folding and unfolding of proteins based on the characteristics of constituent amino acids. Images FIGURE 6 PMID:7711253

  4. Topology of AspT, the Aspartate:Alanine Antiporter of Tetragenococcus halophilus, Determined by Site-Directed Fluorescence Labeling▿ †

    PubMed Central

    Nanatani, Kei; Fujiki, Takashi; Kanou, Kazuhiko; Takeda-Shitaka, Mayuko; Umeyama, Hideaki; Ye, Liwen; Wang, Xicheng; Nakajima, Tasuku; Uchida, Takafumi; Maloney, Peter C.; Abe, Keietsu

    2007-01-01

    The gram-positive lactic acid bacterium Tetragenococcus halophilus catalyzes the decarboxylation of l-aspartate (Asp) with release of l-alanine (Ala) and CO2. The decarboxylation reaction consists of two steps: electrogenic exchange of Asp for Ala catalyzed by an aspartate:alanine antiporter (AspT) and intracellular decarboxylation of the transported Asp catalyzed by an l-aspartate-β-decarboxylase (AspD). AspT belongs to the newly classified aspartate:alanine exchanger family (transporter classification no. 2.A.81) of transporters. In this study, we were interested in the relationship between the structure and function of AspT and thus analyzed the topology by means of the substituted-cysteine accessibility method using the impermeant, fluorescent, thiol-specific probe Oregon Green 488 maleimide (OGM) and the impermeant, nonfluorescent, thiol-specific probe [2-(trimethylammonium)ethyl]methanethiosulfonate bromide. We generated 23 single-cysteine variants from a six-histidine-tagged cysteineless AspT template. A cysteine position was assigned an external location if the corresponding single-cysteine variant reacted with OGM added to intact cells, and a position was assigned an internal location if OGM labeling required cell lysis. The topology analyses revealed that AspT has a unique topology; the protein has 10 transmembrane helices (TMs), a large hydrophilic cytoplasmic loop (about 180 amino acids) between TM5 and TM6, N and C termini that face the periplasm, and a positively charged residue (arginine 76) within TM3. Moreover, the three-dimensional structure constructed by means of the full automatic modeling system indicates that the large hydrophilic cytoplasmic loop of AspT possesses a TrkA_C domain and a TrkA_C-like domain and that the three-dimensional structures of these domains are similar to each other even though their amino acid sequences show low similarity. PMID:17660287

  5. Formation of [b3 - 1 + cat]+ ions from metal-cationized tetrapeptides containing beta-alanine, gamma-aminobutyric acid or epsilon-aminocaproic acid residues.

    PubMed

    Osburn, Sandra M; Ochola, Sila O; Talaty, Erach R; Van Stipdonk, Michael J

    2008-11-01

    The presence and position of a single beta-alanine (betaA), gamma-aminobutyric acid (gammaABu) or epsilon-aminocaproic acid (Cap) residue has been shown to have a significant influence on the formation of b(n)+ and y(n)+ product ions from a series of model, protonated peptides. In this study, we examined the effect of the same residues on the formation of analogous [b3 - 1 + cat]+ products from metal (Li+, Na+ and Ag+)-cationized peptides. The larger amino acids suppress formation of b3+ from protonated peptides with general sequence AAXG (where X = beta-alanine, gamma-aminobutyric acid or epsilon-aminocaproic acid), presumably because of the prohibitive effect of larger cyclic intermediates in the 'oxazolone' pathway. However, abundant [b3 - 1 + cat]+ products are generated from metal-cationized versions of AAXG. Using a group of deuterium-labeled and exchanged peptides, we found that formation of [b3 - 1 + cat]+ involves transfer of either amide or alpha-carbon position H atoms, and the tendency to transfer the atom from the alpha-carbon position increases with the size of the amino acid in position X. To account for the transfer of the H atom, a mechanism involving formation of a ketene product as [b3 - 1 + cat]+ is proposed.

  6. The Benzyl Ester Group of Amino Acid Monomers Enhances Substrate Affinity and Broadens the Substrate Specificity of the Enzyme Catalyst in Chemoenzymatic Copolymerization.

    PubMed

    Ageitos, Jose Manuel; Yazawa, Kenjiro; Tateishi, Ayaka; Tsuchiya, Kousuke; Numata, Keiji

    2016-01-11

    The chemoenzymatic polymerization of amino acid monomers by proteases involves a two-step reaction: the formation of a covalent acyl-intermediate complex between the protease and the carboxyl ester group of the monomer and the subsequent deacylation of the complex by aminolysis to form a peptide bond. Although the initiation with the ester group of the monomer is an important step, the influence of the ester group on the polymerization has not been studied in detail. Herein, we studied the effect of the ester groups (methyl, ethyl, benzyl, and tert-butyl esters) of alanine and glycine on the synthesis of peptides using papain as the catalyst. Alanine and glycine were selected as monomers because of their substantially different affinities toward papain. The efficiency of the polymerization of alanine and glycine benzyl esters was much greater than that of the other esters. The benzyl ester group therefore allowed papain to equally polymerize alanine and glycine, even though the affinity of alanine toward papain is substantially higher. The characterization of the copolymers of alanine and glycine in terms of the secondary structure and thermal properties revealed that the thermal stability of the peptides depends on the amino acid composition and resultant secondary structure. The current results indicate that the nature of the ester group drastically affects the polymerization efficiency and broadens the substrate specificity of the protease.

  7. Porcine alanine transaminase after liver allo-and xenotransplantation

    PubMed Central

    Ekser, Burcin; Gridelli, Bruno; Cooper, David K.C.

    2013-01-01

    Aspartate transaminase (AST) and alanine transaminase (ALT) are measured following liver transplantation as indicators of hepatocellular injury. During a series of orthotopic liver allo-and xenotransplants, we observed that there was an increase in AST in all cases. The anticipated concomitant rise in ALT did not occur when a wild-type (WT) pig was the source of the liver graft, but did occur when a baboon or a genetically engineered (α1,3-galactosyltransferase gene-knockout [GTKO]) pig was the source of the graft. We hypothesized that the cience of Galα1,3 Gal in GTKO pig livers may render pig hepatocytes similar to human and baboon hepatocytes in their response to hepatocellular injury. Reviewing the literature, after WT pig liver allotransplantation or xenotransplantation, in the majority of reports, although changes in AST were reported, no mention was made of changes in ALT, suggesting that there was no change in ALT. However, Ramirez et al. reported two cases of liver xenotransplants from hCD55 pigs, following which there were increases in both AST and ALT, suggesting that it is not simply the cience of expression of Galα1,3 Gal that is the cause. We acknowledge that our observation is based on a small number of experiments, but we believe it is worth recording. PMID:22360753

  8. Porcine alanine transaminase after liver allo-and xenotransplantation.

    PubMed

    Ekser, Burcin; Gridelli, Bruno; Cooper, David K C

    2012-01-01

    Aspartate transaminase (AST) and alanine transaminase (ALT) are measured following liver transplantation as indicators of hepatocellular injury. During a series of orthotopic liver allo-and xenotransplants, we observed that there was an increase in AST in all cases. The anticipated concomitant rise in ALT did not occur when a wild-type (WT) pig was the source of the liver graft, but did occur when a baboon or a genetically engineered (α1,3-galactosyltransferase gene-knockout [GTKO]) pig was the source of the graft. We hypothesized that the cience of Galα1,3Gal in GTKO pig livers may render pig hepatocytes similar to human and baboon hepatocytes in their response to hepatocellular injury. Reviewing the literature, after WT pig liver allotransplantation or xenotransplantation, in the majority of reports, although changes in AST were reported, no mention was made of changes in ALT, suggesting that there was no change in ALT. However, Ramirez et al. reported two cases of liver xenotransplants from hCD55 pigs, following which there were increases in both AST and ALT, suggesting that it is not simply the cience of expression of Galα1,3Gal that is the cause. We acknowledge that our observation is based on a small number of experiments, but we believe it is worth recording.

  9. Alanine synthesis from glyceraldehyde and ammonium ion in aqueous solution

    NASA Technical Reports Server (NTRS)

    Weber, A. L.

    1985-01-01

    The formation of alanine (ala) form C(14)-glyceraldehyde and ammonium phosphate in the presence or absence of a thiol is reported. At ambient temperature, ala synthesis was six times more rapid in the presence of 3-mercaptopropionic acid than in its absence (0.6 and 0.1 percent, respectively, after 60 days). Similarly, the presence of another thiol, N-acetylcysteinate, increased the production of ala, as well as of lactate. The reaction pathway of thiol-catalyzed synthesis of ala, with the lactic acid formed in a bypath, is suggested. In this, dehydration of glyceraldehyde is followed by the formation of hemithioacetal. In the presence of ammonia, an imine is formed, which eventually yields ala. This pathway is consistent with the observation that the rate ratio of ala/lactate remains constant throughout the process. The fact that the reaction takes place under anaerobic conditions in the presence of H2O and with the low concentrations of simple substrates and catalysts makes it an attractive model prebiotic reaction in the process of molecular evolution.

  10. Free amino acids in fibromyalgia syndrome: relationship with clinical picture.

    PubMed

    Ruggiero, Valeria; Mura, Massimiliano; Cacace, Enrico; Era, Benedetta; Peri, Marcella; Sanna, Giuseppina; Fais, Antonella

    2017-04-01

    The objectives of our study were to evaluate free amino acid (FAA) concentrations in the serum of patients affected by fibromyalgia syndrome (FMS) and to determine the relationships between FAA levels and FMS clinical parameters. Thus, serum amino acid concentrations were quantified (HPLC analysis) in 23 females with fibromyalgia (according to the American College of Rheumatology classification criteria) and 20 healthy females. The results showed significantly higher serum concentrations of aspartate, cysteine, glutamate, glycine, isoleucine, leucine, methionine, ornithine, phenylalanine, sarcosine, serine, taurine, tyrosine and valine in FMS patients vs. healthy controls. Patients with higher Fibromyalgia Impact Questionnaire (FIQ) scores showed increased levels of alanine, glutamine, isoleucine, leucine, phenylalanine, proline and valine. In conclusion, our results indicate an imbalance in some FAAs in FMS patients. Increased Glu is particularly interesting, as it could explain the deficit in monoaminergic transmission involved in pain.

  11. Enzymatic characterization and crystal structure analysis of the D-alanine-D-alanine ligase from Helicobacter pylori.

    PubMed

    Wu, Dalei; Zhang, Liang; Kong, Yunhua; Du, Jiamu; Chen, Shuai; Chen, Jing; Ding, Jianping; Jiang, Hualiang; Shen, Xu

    2008-09-01

    D-Alanine-D-alanine ligase is the second enzyme in the D-Ala branch of bacterial cell wall peptidoglycan assembly, and recognized as an attractive antimicrobial target. In this work, the D-Ala-D-Ala ligase of Helicobacter pylori strain SS1 (HpDdl) was kinetically and structurally characterized. The determined apparent K(m) of ATP (0.87 microM), the K(m1) (1.89 mM) and K(m2) of D-Ala (627 mM), and the k(cat) (115 min(-1)) at pH 8.0 indicated its relatively weak binding affinity and poor catalytic activity against the substrate D-Ala in vitro. However, by complementary assay of expressing HpDdl in Escherichia coli Delta ddl mutant, HpDdl was confirmed to be capable of D-Ala-D-Ala ligating in vivo. Through sequence alignment with other members of the D-Ala-D-X ligase superfamily, HpDdl keeps two conservatively substituted residues (Ile16 and Leu241) and two nonconserved residues (Leu308 and Tyr311) broadly located in the active region of the enzyme. Kinetic analyses against the corresponding HpDdl mutants (I16V, L241Y, L241F, L308T, and Y311S) suggested that these residues, especially Leu308 and Tyr311, might partly contribute to the unique catalytic properties of the enzyme. This was fairly proved by the crystal structure of HpDdl, which revealed that there is a 3(10)-helix (including residues from Gly306 to Leu312) near the D-Ala binding region in the C-terminal domain, where HpDdl has two sequence deletions compared with other homologs. Such 3(10)-helix may participate in D-Ala binding and conformational change of the enzyme. Our present work hopefully provides useful information for understanding the D-Ala-D-Ala ligase of Helicobacter pylori.

  12. Amino acid racemization dating of fossil bones, I. inter-laboratory comparison of racemization measurements

    USGS Publications Warehouse

    Bada, J.L.; Hoopes, E.; Darling, D.; Dungworth, G.; Kessels, H.J.; Kvenvolden, K.A.; Blunt, D.J.

    1979-01-01

    Enantiomeric measurements for aspartic acid, glutamic acid, and alanine in twenty-one different fossil bone samples have been carried out by three different laboratories using different analytical methods. These inter-laboratory comparisons demonstrate that D/L aspartic acid measurements are highly reproducible, whereas the enantiomeric measurements for the other amino acids show a wide variation between the three laboratories. At present, aspartic acid measurements are the most suitable for racemization dating of bone because of their superior analytical precision. ?? 1979.

  13. Photon Atomic Parameters of Nonessential Amino Acids for Radiotherapy and Diagnostics

    PubMed Central

    Bursalıoğlu, Ertuğrul O.; İçelli, Orhan; Balkan, Begüm; Kavanoz, H. Birtan; Okutan, Mustafa

    2014-01-01

    The total mass attenuation coefficients (μt) (cm2/g) and atomic, molecular, and electronic effective cross sections have been calculated for nonessential amino acids that contain H, C, N, and O such as tyrosine, aspartate, glutamine, alanine, asparagine, aspartic acid, cysteine, and glycine in the wide energy region 0.015–15 MeV. The variations with energy of total mass attenuation coefficients and atomic, molecular, and electronic cross sections are shown for all photon interactions. PMID:25548658

  14. Differentiation of green, white, black, Oolong, and Pu-erh teas according to their free amino acids content.

    PubMed

    Alcázar, A; Ballesteros, O; Jurado, J M; Pablos, F; Martín, M J; Vilches, J L; Navalón, A

    2007-07-25

    In this paper, the differentiation of green, black, Oolong, white, and Pu-erh teas has been carried out according to their free amino acid contents. Alanine, arginine, asparagine, aspartic acid, glutamic acid, isoleucine, histidine, leucine, phenylalanine, serine, theanine, threonine, and tyrosine have been determined by liquid chromatography with derivatization with o-phthalaldehyde and fluorescence detection. The chromatographic separation was achieved with a Hypersil ODS column and gradient elution. The amino acid contents were used as chemometric descriptors for classification purposes of different tea varieties. Principal component analysis, k-nearest neighbors, linear discriminant analysis, and artificial neural networks were applied to differentiate tea varieties. Using back-propagation multilayer perceptron artificial neural networks, 100% success in the classification was obtained. The most differentiating amino acids were glutamic acid, asparagine, serine, alanine, leucine, and isoleucine.

  15. Hydrogen production using amino acids obtained by protein degradation in waste biomass by combined dark- and photo-fermentation.

    PubMed

    Cheng, Jun; Ding, Lingkan; Xia, Ao; Lin, Richen; Li, Yuyou; Zhou, Junhu; Cen, Kefa

    2015-03-01

    The biological hydrogen production from amino acids obtained by protein degradation was comprehensively investigated to increase heating value conversion efficiency. The five amino acids (i.e., alanine, serine, aspartic acid, arginine, and leucine) produced limited hydrogen (0.2-16.2 mL/g) but abundant soluble metabolic products (40.1-84.0 mM) during dark-fermentation. The carbon conversion efficiencies of alanine (85.3%) and serine (94.1%) during dark-fermentation were significantly higher than those of other amino acids. Residual dark-fermentation solutions treated with zeolite for NH4(+) removal were inoculated with photosynthetic bacteria to further produce hydrogen during photo-fermentation. The hydrogen yields of alanine and serine through combined dark- and photo-fermentation were 418.6 and 270.2 mL/g, respectively. The heating value conversion efficiency of alanine to hydrogen was 25.1%, which was higher than that of serine (21.2%).

  16. Effects of Monovalent Cations on the Sodium-Alanine Interaction in Rabbit Ileum

    PubMed Central

    Frizzell, Raymond A.; Schultz, Stanley G.

    1970-01-01

    H, K, Rb, and Li inhibit Na-dependent alanine influx across the brush border of rabbit ileum. Kinetic analysis indicates that H and K behave as competitive inhibitors of influx so that increasing the concentration of H or K in the mucosal solution is kinetically indistinguishable from decreasing the Na concentration. In addition the coupling between alanine and Na influxes is markedly reduced at pH 2.5. With the exception of H and Li, none of these monovalent cations significantly affects carrier-mediated alanine influx in the absence of Na indicating that their inhibitory effects are largely restricted to the Na-dependent fraction of influx. Increasing H concentration from 0.03 to 3 mM does not affect influx in the absence of Na but markedly inhibits influx in the presence of Na. Li significantly enhances alanine influx in the absence of Na. Ag, UO2, and La also inhibit the Na-dependent fraction of alanine influx. These findings suggest that anionic groups having a pKa of approximately 4 are involved in the interaction between Na and the alanine-carrier complex; present evidence implicates carboxylate groups however, phosphoryl residues cannot be ruled out. The previously proposed kinetic model for the Na-alanine interaction has been extended to accommodate these effects of H and other monovalent cations. The mechanistic and physiological implications of these findings are discussed. PMID:5507092

  17. Revised mechanism of D-alanine incorporation into cell wall polymers in Gram-positive bacteria.

    PubMed

    Reichmann, Nathalie T; Cassona, Carolina Picarra; Gründling, Angelika

    2013-09-01

    Teichoic acids (TAs) are important for growth, biofilm formation, adhesion and virulence of Gram-positive bacterial pathogens. The chemical structures of the TAs vary between bacteria, though they typically consist of zwitterionic polymers that are anchored to either the peptidoglycan layer as in the case of wall teichoic acid (WTA) or the cell membrane and named lipoteichoic acid (LTA). The polymers are modified with D-alanines and a lack of this decoration leads to increased susceptibility to cationic antimicrobial peptides. Four proteins, DltA-D, are essential for the incorporation of d-alanines into cell wall polymers and it has been established that DltA transfers D-alanines in the cytoplasm of the cell onto the carrier protein DltC. However, two conflicting models have been proposed for the remainder of the mechanism. Using a cellular protein localization and membrane topology analysis, we show here that DltC does not traverse the membrane and that DltD is anchored to the outside of the cell. These data are in agreement with the originally proposed model for D-alanine incorporation through a process that has been proposed to proceed via a D-alanine undecaprenyl phosphate membrane intermediate. Furthermore, we found that WTA isolated from a Staphylococcus aureus strain lacking LTA contains only a small amount of D-alanine, indicating that LTA has a role, either direct or indirect, in the efficient D-alanine incorporation into WTA in living cells.

  18. Alanine-EPR as a transfer standard dosimetry system for low energy X radiation

    NASA Astrophysics Data System (ADS)

    Khoury, H. J.; da Silva, E. J.; Mehta, K.; de Barros, V. S.; Asfora, V. K.; Guzzo, P. L.; Parker, A. G.

    2015-11-01

    The purpose of this paper is to evaluate the use of alanine-EPR as a transfer standard dosimetry system for low energy X radiation, such as that in RS-2400, which operates in the range from 25 to 150 kV and 2 to 45 mA. Two types of alanine dosimeters were investigated. One is a commercial alanine pellets from Aérial-Centre de Ressources Technologiques, France and one was prepared in our laboratory (LMRI-DEN/UFPE). The EPR spectra of the irradiated dosimeters were recorded in the Nuclear Energy Department of UFPE, using a Bruker EMX10 EPR spectrometer operating in the X-band. The alanine-EPR dosimetry system was calibrated in the range of 20-220 Gy in this X-ray field, against an ionization chamber calibrated at the relevant X-ray energy with traceability to PTB. The results showed that both alanine dosimeters presented a linear dose response the same sensitivity, when the EPR signal was normalized to alanine mass. The total uncertainty in the measured dose was estimated to be about 3%. The results indicate that it is possible to use the alanine-EPR dosimetry system for validation of a low-energy X ray irradiator, such as RS-2400.

  19. Characterization of alanine catabolism in Pseudomonas aeruginosa and its importance for proliferation in vivo.

    PubMed

    Boulette, Megan L; Baynham, Patricia J; Jorth, Peter A; Kukavica-Ibrulj, Irena; Longoria, Aissa; Barrera, Karla; Levesque, Roger C; Whiteley, Marvin

    2009-10-01

    The opportunistic pathogen Pseudomonas aeruginosa causes a variety of infections in immunocompromised individuals, including individuals with the heritable disease cystic fibrosis. Like the carbon sources metabolized by many disease-causing bacteria, the carbon sources metabolized by P. aeruginosa at the host infection site are unknown. We recently reported that l-alanine is a preferred carbon source for P. aeruginosa and that two genes potentially involved in alanine catabolism (dadA and dadX) are induced during in vivo growth in the rat peritoneum and during in vitro growth in sputum (mucus) collected from the lungs of individuals with cystic fibrosis. The goals of this study were to characterize factors required for alanine catabolism in P. aeruginosa and to assess the importance of these factors for in vivo growth. Our results reveal that dadA and dadX are arranged in an operon and are required for catabolism of l-alanine. The dad operon is inducible by l-alanine, d-alanine, and l-valine, and induction is dependent on the transcriptional regulator Lrp. Finally, we show that a mutant unable to catabolize dl-alanine displays decreased competitiveness in a rat lung model of infection.

  20. Amino Acid Analyses of Acid Hydrolysates in Desert Varnish

    NASA Technical Reports Server (NTRS)

    Perry, Randall S.; Staley, James T.; Dworkin, Jason P.; Engel, Mike

    2001-01-01

    There has long been a debate as to whether rock varnish deposits are microbially mediated or are deposited by inorganic processes. Varnished rocks are found throughout the world primarily in arid and semi-arid regions. The varnish coats are typically up to 200 microns thick and are composed of clays and alternating layers enriched in manganese and iron oxides. The individual layers range in thickness from 1 micron to greater than 10 microns and may continue laterally for more than a 100 microns. Overlapping botryoidal structures are visible in thin section and scanning electron micrographs. The coatings also include small amounts of organic mater and detrital grains. Amino-acid hydrolysates offer a means of assessing the organic composition of rock varnish collected from the Sonoran Desert, near Phoenix, AZ. Chromatographic analyses of hydrolysates from powdered samples of rock varnish suggest that the interior of rock varnish is relatively enriched in amino acids and specifically in d-alanine and glutamic acid. Peptidoglycan (murein) is the main structural component of gram-positive bacterial cell walls. The d-enantiomer of alanine and glutamic acid are specific to peptidoglycan and are consequently an indicator for the presence of bacteria. D-alanine is also found in teichoic acid which is only found in gram-positive bacteria. Several researchers have cultured bacteria from the surface of rock varnish and most have been gram-positive, suggesting that gram-positive bacteria are intimately associated with varnish coatings and may play a role in the formation of varnish coatings.

  1. Inhibition of small-intestinal sugar and amino acid transport by the enterotoxin of Shigella dysenteriae I.

    PubMed

    Binder, H J; Whiting, D S

    1977-05-01

    The enterotoxin of Shigella dysenteriae I produces fluid and electrolyte secretion in the rabbit ileum. These present studies were designed to evaluate nonelectrolyte transport in rabbit ileal mucosa exposed to Shigella enterotoxin. Both 10 mM galactose and 5 mM L-alanine absorptions were significantly impaired in enterotoxin-exposed ileal mucosa compared with control mucosa. L-Alanine influx was not imparied in two other secretory processes: that induced by cholera enterotoxin and hyperosmolarity. These studies provide evidence that both surgar and amino acid absorptions are diminished in the small intestine by the enterotoxin of S. dysenteriae I.

  2. Inhibition of small-intestinal sugar and amino acid transport by the enterotoxin of Shigella dysenteriae I.

    PubMed Central

    Binder, H J; Whiting, D S

    1977-01-01

    The enterotoxin of Shigella dysenteriae I produces fluid and electrolyte secretion in the rabbit ileum. These present studies were designed to evaluate nonelectrolyte transport in rabbit ileal mucosa exposed to Shigella enterotoxin. Both 10 mM galactose and 5 mM L-alanine absorptions were significantly impaired in enterotoxin-exposed ileal mucosa compared with control mucosa. L-Alanine influx was not imparied in two other secretory processes: that induced by cholera enterotoxin and hyperosmolarity. These studies provide evidence that both surgar and amino acid absorptions are diminished in the small intestine by the enterotoxin of S. dysenteriae I. PMID:324910

  3. Amino Acids and Chirality

    NASA Technical Reports Server (NTRS)

    Cook, Jamie E.

    2012-01-01

    Amino acids are among the most heavily studied organic compound class in carbonaceous chondrites. The abundance, distributions, enantiomeric compositions, and stable isotopic ratios of amino acids have been determined in carbonaceous chondrites fi'om a range of classes and petrographic types, with interesting correlations observed between these properties and the class and typc of the chondritcs. In particular, isomeric distributions appear to correlate with parent bodies (chondrite class). In addition, certain chiral amino acids are found in enantiomeric excess in some chondrites. The delivery of these enantiomeric excesses to the early Earth may have contributed to the origin of the homochirality that is central to life on Earth today. This talk will explore the amino acids in carbonaceous chondritcs and their relevance to the origin of life.

  4. Amino Acid Metabolism Disorders

    MedlinePlus

    ... breaks the food parts down into sugars and acids, your body's fuel. Your body can use this ... process. One group of these disorders is amino acid metabolism disorders. They include phenylketonuria (PKU) and maple ...

  5. Serum Alanine Aminotransferase Levels, Hematocrit Rate and Body Weight Correlations Before and After Hemodialysis Session

    PubMed Central

    Lopes, Edmundo Pessoa; Sette, Luis Henrique B. C.; Sette, Jorge Bezerra C.; Luna, Carlos F.; Andrade, Amaro M.; Moraes, Maviael; Sette, Paulo C. A.; Menezes, Roberto; Cavalcanti, Rui L.; Conceição, Sergio C.

    2009-01-01

    PURPOSE To evaluate alanine aminotransferase levels before and after a hemodialysis session and to correlate these values with the hematocrit rate and weight loss during hemodialysis. PATIENTS AND METHODS The serum alanine aminotransferase levels, hematocrit rate and body weight were measured and correlated before and after a single hemodialysis session for 146 patients with chronic renal failure. An receiver operating characteristic (ROC) curve for the serum alanine aminotransferase levels collected before and after hemodialysis was plotted to identify hepatitis C virus-infected patients. RESULTS The mean weight loss of the 146 patients during hemodialysis was 5.3% (p < 0.001). The mean alanine aminotransferase levels before and after hemodialysis were 18.8 and 23.9 IU/, respectively, denoting a significant 28.1% increase. An equally significant increase of 16.4% in the hematocrit rate also occurred after hemodialysis. The weight loss was inversely correlated with the rise in both the alanine aminotransferase level (r = 0.3; p < 0.001) and hematocrit rate (r = 0.5; p < 0.001). A direct correlation was found between the rise in alanine aminotransferase levels and the hematocrit during the hemodialysis session (r = 0.4; p < 0.001). Based on the ROC curve, the upper limit of the normal alanine aminotransferase level should be reduced by 40% relative to the upper limit of normal if the blood samples are collected before the hemodialysis session or by 60% if blood samples are collected after the session. CONCLUSION In the present study, significant elevations in the serum alanine aminotransferase levels and hematocrit rates occurred in parallel to a reduction in body weight after the hemodialysis session. These findings suggest that one of the factors for low alanine aminotransferase levels prior to hemodialysis could be hemodilution in patients with chronic renal failure. PMID:19841699

  6. Effect of environmental salinity manipulation on uptake rates and distribution patterns of waterborne amino acids in the Pacific hagfish.

    PubMed

    Glover, Chris N; Blewett, Tamzin A; Wood, Chris M

    2017-02-01

    Among vertebrates, hagfish are the only known iono- and osmoconformers, and the only species thus far documented to absorb amino acids directly across the skin. In the current study, short-term (6h) manipulations of exposure salinities (75-125% seawater) were conducted to determine whether changes in osmotic demands influenced the uptake and tissue distribution of waterborne amino acids (alanine, glycine and phenylalanine), in the Pacific hagfish, Eptatretus stoutii. No changes in erythrocyte or muscle amino acid accumulation rates were noted, but the patterns of plasma amino acid accumulation were suggestive of regulation. Contrary to expectations, glycine transport across the skin in vitro was enhanced in the lowest exposure salinity, but no other salinity-dependent changes were demonstrated. Overall, this study indicates that uptake and distribution of amino acids varies with salinity, but not in a manner that is consistent with a role for the studied amino acids in maintaining osmotic balance in hagfish.

  7. Manageable cytotoxicity of nanocapsules immobilizing D-amino acid oxidase via exogenous administration of nontoxic prodrug

    NASA Astrophysics Data System (ADS)

    Zhao, Yang; Zhu, Yingchun; Fu, Jingke

    2014-02-01

    D-Amino acid oxidase (DAO), which could catalyze generation of hydrogen peroxide with strong oxidbility and cytotoxicity, has become of interest as a biocatalyst for therapeutic treatments. Herein we report that amino-functional hollow mesoporous silica with large pore size (10.27 nm) and positively charged surface effectively immobilize DAO with negative charge. The adsorption, activity and stability of DAO are demonstrated to depend mainly on the amino-functionalization of surface. Significant cancer cell killing effect is observed when the cells are treated by the nanocapsules entrapping DAO together with D-alanine, showing distinct dose-dependency on concentration of the nanocapsules entrapping DAO or D-alanine. Nevertheless, the toxicity is completely neutralized by the addition of catalase, and anti-tumor effect is not observed when either the nanocapsules entrapping DAO or D-alanine is applied alone. The results indicate that cytotoxicity of the nanocapsules entrapping DAO could be managed by exogenous administration of nontoxic prodrug to tumor tissue, due to the stereoselectivity of DAO and the scarcity of its substrates in mammalian organisms. Thus, the method might be exploited as a potential treatment for cancer therapy.

  8. Glutathione S-transferase class {pi} polymorphism in baboons

    SciTech Connect

    Aivaliotis, M.J.; Cantu, T.; Gilligan, R.

    1995-02-01

    Glutathione S-transferase (GST) comprises a family of isozymes with broad substrate specificities. One or more GST isozymes are present in most animal tissues and function in several detoxification pathways through the conjugation of reduced glutathione with various electrophiles, thereby reducing their potential toxicity. Four soluble GST isozymes encoded by genes on different chromosomes have been identified in humans. The acidic class pi GST, GSTP (previously designated GST-3), is widely distributed in adult tissues and appears to be the only GST isozyme present in leukocytes and placenta. Previously reported electrophoretic analyses of erythrocyte and leukocyte extracts revealed single bands of activity, which differed slightly in mobility between the two cell types, or under other conditions, a two-banded pattern. To our knowledge, no genetically determined polymorphisms have previously been reported in GSTP from any species. We now report a polymorphism of GSTP in baboon leukocytes, and present family data that verifies autosomal codominant inheritance. 14 refs., 2 figs., 1 tab.

  9. MOF Acetyl Transferase Regulates Transcription and Respiration in Mitochondria.

    PubMed

    Chatterjee, Aindrila; Seyfferth, Janine; Lucci, Jacopo; Gilsbach, Ralf; Preissl, Sebastian; Böttinger, Lena; Mårtensson, Christoph U; Panhale, Amol; Stehle, Thomas; Kretz, Oliver; Sahyoun, Abdullah H; Avilov, Sergiy; Eimer, Stefan; Hein, Lutz; Pfanner, Nikolaus; Becker, Thomas; Akhtar, Asifa

    2016-10-20

    A functional crosstalk between epigenetic regulators and metabolic control could provide a mechanism to adapt cellular responses to environmental cues. We report that the well-known nuclear MYST family acetyl transferase MOF and a subset of its non-specific lethal complex partners reside in mitochondria. MOF regulates oxidative phosphorylation by controlling expression of respiratory genes from both nuclear and mtDNA in aerobically respiring cells. MOF binds mtDNA, and this binding is dependent on KANSL3. The mitochondrial pool of MOF, but not a catalytically deficient mutant, rescues respiratory and mtDNA transcriptional defects triggered by the absence of MOF. Mof conditional knockout has catastrophic consequences for tissues with high-energy consumption, triggering hypertrophic cardiomyopathy and cardiac failure in murine hearts; cardiomyocytes show severe mitochondrial degeneration and deregulation of mitochondrial nutrient metabolism and oxidative phosphorylation pathways. Thus, MOF is a dual-transcriptional regulator of nuclear and mitochondrial genomes connecting epigenetics and metabolism.

  10. Thermodynamics of Enzyme-Catalyzed Reactions: Part 2. Transferases

    NASA Astrophysics Data System (ADS)

    Goldberg, Robert N.; Tewari, Yadu B.

    1994-07-01

    Equilibrium constants and enthalpy changes for reactions catalyzed by the transferase class of enzymes have been compiled. For each reaction the following information is given: the reference for the data; the reaction studied; the name of the enzyme used and its Enzyme Commission number; the method of measurement; the conditions of measurement [temperature, pH, ionic strength, and the buffer(s) and cofactor(s) used]; the data and an evaluation of it; and, sometimes, commentary on the data and on any corrections which have been applied to it or any calculations for which the data have been used. The data from 285 references have been examined and evaluated. Chemical Abstract Service registry numbers are given for the substances involved in these various reactions. There is a cross reference between the substances and the Enzyme Commission numbers of the enzymes used to catalyze the reactions in which the substances participate.

  11. Characterization of the l-alanine exporter AlaE of Escherichia coli and its potential role in protecting cells from a toxic-level accumulation of l-alanine and its derivatives.

    PubMed

    Kim, Seryoung; Ihara, Kohei; Katsube, Satoshi; Hori, Hatsuhiro; Ando, Tasuke; Isogai, Emiko; Yoneyama, Hiroshi

    2015-08-01

    We previously reported that the alaE gene of Escherichia coli encodes the l-alanine exporter AlaE. The objective of this study was to elucidate the mechanism of the AlaE exporter. The minimum inhibitory concentration of l-alanine and l-alanyl-l-alanine in alaE-deficient l-alanine-nonmetabolizing cells MLA301ΔalaE was 4- and >4000-fold lower, respectively, than in the alaE-positive parent cells MLA301, suggesting that AlaE functions as an efflux pump to avoid a toxic-level accumulation of intracellular l-alanine and its derivatives. Furthermore, the growth of the alaE-deficient mutant derived from the l-alanine-metabolizing strain was strongly inhibited in the presence of a physiological level of l-alanyl-l-alanine. Intact MLA301ΔalaE and MLA301ΔalaE/pAlaE cells producing plasmid-borne AlaE, accumulated approximately 200% and 50%, respectively, of the [(3) H]l-alanine detected in MLA301 cells, suggesting that AlaE exports l-alanine. When 200 mmol/L l-alanine-loaded inverted membrane vesicles prepared from MLA301ΔalaE/pAlaE were placed in a solution containing 200 mmol/L or 0.34 μmol/L l-alanine, energy-dependent [(3) H]l-alanine accumulation occurred under either condition. This energy-dependent uphill accumulation of [(3) H]l-alanine was strongly inhibited in the presence of carbonyl cyanide m-chlorophenylhydrazone but not by dicyclohexylcarbodiimide, suggesting that the AlaE-mediated l-alanine extrusion was driven by proton motive force. Based on these results, physiological roles of the l-alanine exporter are discussed.

  12. Progress towards an alanine/ESR therapy level reference dosimetry service at NPL.

    PubMed

    Sharpe, P H; Rajendran, K; Sephton, J P

    1996-01-01

    This paper describes work being carried out at the National Physical Laboratory towards the establishment of an alanine reference dosimetry service for radiotherapy applications. A precision fused quartz holder has been constructed to allow precise positioning of alanine dosimeters in the ESR cavity. A novel method of signal analysis based on spectrum fitting has been developed to minimize the effect of baseline distortions. Data are also presented on the relative response of alanine to 60Co gamma rays and high energy photons (4-12 MeV).

  13. Applicability of EPR/alanine dosimetry for quality assurance in proton eye radiotherapy.

    PubMed

    Michalec, B; Mierzwinska, G; Ptaszkiewicz, M; Sowa, U; Stolarczyk, L; Weber, A

    2014-06-01

    A new quality assurance and quality control method for proton eye radiotherapy based on electron paramagnetic resonance (EPR)/alanine dosimetry has been developed. It is based on Spread-Out Bragg Peak entrance dose measurement with alanine detectors. The entrance dose is well correlated with the dose at the facility isocenter, where, during the therapeutic irradiation, the tumour is placed. The unique alanine detector features namely keeping the dose record in a form of stable radiation-induced free radicals trapped in the material structure, and the non-destructive read-out makes this type of detector a good candidate for additional documentation of the patient's exposure over the therapy course.

  14. Temperature dependences of piezoelectric, elastic and dielectric constants of L-alanine crystal

    NASA Astrophysics Data System (ADS)

    Tylczyński, Z.; Sterczyńska, A.; Wiesner, M.

    2011-09-01

    Temperature changes in the components of piezoelectric, elastic and dielectric tensors were studied in L-alanine crystals in the range 100-300 K. A jumpwise increase in the c55 component of the elastic stiffness accompanied by maxima in damping of all face-shear modes observed at 199 K in L-alanine crystal were interpreted as a result of changes in the NH3+ vibrations occurring through electron-phonon coupling. All components of the piezoelectric tensor show small anomalies in this temperature range. The components of the electromechanical coupling coefficient determined indicate that L-alanine is a weak piezoelectric.

  15. Temperature dependences of piezoelectric, elastic and dielectric constants of L-alanine crystal.

    PubMed

    Tylczyński, Z; Sterczyńska, A; Wiesner, M

    2011-09-07

    Temperature changes in the components of piezoelectric, elastic and dielectric tensors were studied in L-alanine crystals in the range 100-300 K. A jumpwise increase in the c(55) component of the elastic stiffness accompanied by maxima in damping of all face-shear modes observed at 199 K in L-alanine crystal were interpreted as a result of changes in the NH(3)(+) vibrations occurring through electron-phonon coupling. All components of the piezoelectric tensor show small anomalies in this temperature range. The components of the electromechanical coupling coefficient determined indicate that L-alanine is a weak piezoelectric.

  16. Molecular dynamics simulations of mutated Mycobacterium tuberculosis L-alanine dehydrogenase to illuminate the role of key residues.

    PubMed

    Ling, Baoping; Bi, Siwei; Sun, Min; Jing, Zhihong; Li, Xiaoping; Zhang, Rui

    2014-05-01

    L-Alanine dehydrogenase from Mycobacterium tuberculosis (L-MtAlaDH) catalyzes the NADH-dependent interconversion of l-alanine and pyruvate, and it is considered to be a potential target for the treatment of tuberculosis. The experiment has verified that amino acid replacement of the conserved active-site residues which have strong stability and no great changes in biological evolutionary process, such as His96 and Asp270, could lead to inactive mutants [Ågren et al., J. Mol. Biol. 377 (2008) 1161-1173]. However, the role of these conserved residues in catalytic reaction still remains unclear. Based on the crystal structures, a series of mutant structures were constructed to investigate the role of the conserved residues in enzymatic reaction by using molecular dynamics simulations. The results show that whatever the conserved residues were mutated, the protein can still convert its conformation from open state to closed state as long as NADH is present in active site. Asp270 maintains the stability of nicotinamide ring and ribose of NADH through hydrogen bond interactions, and His96 is helpful to convert the protein conformation by interactions with Gln271, whereas, they would lead to the structural rearrangement in active site and lose the catalytic activity when they were mutated. Additionally, we deduce that Met301 plays a major role in catalytic reaction due to fixing the nicotinamide ring of NADH to prevent its rotation, and we propose that Met301 would be mutated to the hydrophobic residue with large steric hindrance in side chain to test the activity of the protein in future experiment.

  17. Kinetic energy releases of small amino acids upon interaction with keV ions

    NASA Astrophysics Data System (ADS)

    Bari, S.; Alvarado, F.; Postma, J.; Sobocinski, P.; Hoekstra, R.; Schlathölter, T.

    2009-01-01

    In chromatin, DNA is tightly packed into one complex together with histone and non-histone proteins. These proteins are known to protect the DNA against indirect and to some extent even direct radiation damage. Radiation action upon amino acids is thus one of the primary steps in biological radiation action. In this paper we investigate the ionization and fragmentation of the gas-phase amino acids glycine, alanine and valine upon interaction with keV α-particles. High resolution coincidence time-of-flight mass spectrometry is used to determine the dominant fragmentation channels as well as fragment kinetic energies.

  18. Deuterium isotope effect on 13C chemical shifts of tetrabutylammonium salts of Schiff bases amino acids.

    PubMed

    Rozwadowski, Z

    2006-09-01

    Deuterium isotope effects on 13C chemical shift of tetrabutylammonium salts of Schiff bases, derivatives of amino acids (glycine, L-alanine, L-phenylalanine, L-valine, L-leucine, L-isoleucine and L-methionine) and various ortho-hydroxyaldehydes in CDCl3 have been measured. The results have shown that the tetrabutylammonium salts of the Schiff bases amino acids, being derivatives of 2-hydroxynaphthaldehyde and 3,5-dibromosalicylaldehyde, exist in the NH-form, while in the derivatives of salicylaldehyde and 5-bromosalicylaldehyde a proton transfer takes place. The interactions between COO- and NH groups stabilize the proton-transferred form through a bifurcated intramolecular hydrogen bond.

  19. Production of amino acids by analog-resistant mutants of the cyanobacterium Spirulina platensis.

    PubMed Central

    Riccardi, G; Sora, S; Ciferri, O

    1981-01-01

    Mutants of Spirulina platensis resistant to 5-fluorotryptophan, beta-3-thienyl-alanine, ethionine, p-fluorophenylalanine, or azetidine-2-carboxylic acid were isolated. Some of these mutants appeared to be resistant to more than one analog and to overproduce the corresponding amino acids. A second group was composed of mutants that were resistant to one analog only. Of the latter mutants, one resistant to azetidine-2-carboxylic acid was found to overproduce proline only, whereas one resistant to fluorotryptophan and one resistant to ethionine did not overproduce any of the tested amino acids. PMID:6792182

  20. Synthesis and characterization of bis-thiourea having amino acid derivatives

    NASA Astrophysics Data System (ADS)

    Fakhar, Imran; Yamin, Bohari M.; Hasbullah, Siti Aishah

    2016-11-01

    In this article four new symmetric bis-thiourea derivatives having amino acid linkers were reported with good yield. Isophthaloyl dichloride was used as spacer and L-alanine, L-aspartic acid, L-phenylalanine and L-glutamic acid were used as linkers. Bis-thiourea derivatives were prepared from relatively stable isophthaloyl isothiocyanate intermediate. Newly synthesized bis-thiourea derivatives were characterized by FTIR, H-NMR, 13C-NMR and CHNS-O elemental analysis techniques. Characterization data was in good agreement with the expected derivatives, hence confirmed the synthesis of four new derivatives of bis-thiourea having amino acids.

  1. Solvation Free Energies of Alanine Peptides: The Effect of Flexibility

    SciTech Connect

    Kokubo, Hironori; Harris, Robert C.; Asthagiri, Dilip; Pettitt, Bernard M.

    2013-12-03

    The electrostatic (?Gel), cavity-formation (?Gvdw), and total (?G) solvation free energies for 10 alanine peptides ranging in length (n) from 1 to 10 monomers were calculated. The free energies were computed both with xed, extended conformations of the peptides and again for some of the peptides without constraints. The solvation free energies, ?Gel, ?Gvdw, and ?G, were found to be linear in n, with the slopes of the best-fit lines being gamma_el, gamma_vdw, and gamma, respectively. Both gamma_el and gamma were negative for fixed and flexible peptides, and gamma_vdw was negative for fixed peptides. That gamma_vdw was negative was surprising, as experimental data on alkanes, theoretical models, and MD computations on small molecules and model systems generally suggest that gamma_vdw should be positive. A negative gamma_vdw seemingly contradicts the notion that ?Gvdw drives the initial collapse of the protein when it folds by favoring conformations with small surface areas, but when we computed ?Gvdw for the flexible peptides, thereby allowing the peptides to assume natural ensembles of more compact conformations, gamma-vdw was positive. Because most proteins do not assume extended conformations, a ?Gvdw that increases with increasing surface area may be typical for globular proteins. An alternative hypothesis is that the collapse is driven by intramolecular interactions. We show that the intramolecular van der Waal's interaction energy is more favorable for the flexible than for the extended peptides, seemingly favoring this hypothesis, but the large fluctuations in this energy may make attributing the collapse of the peptide to this intramolecular energy difficult.

  2. Responses of single facial taste fibers in the sea catfish, Arius felis, to amino acids.

    PubMed

    Michel, W; Caprio, J

    1991-07-01

    1. Taste buds in catfish are found not only within the oropharyngeal cavity, as in mammals, but are also located along the external body surface of the animal from the barbels and lips to the caudal fin. Because these taste buds are innervated by the facial (cranial VII) nerve, the extraoral taste system of catfish is analogous to the mammalian taste system of the anterior two-thirds of the tongue, which contains taste buds innervated by the chorda tympani nerve, and of the soft palate and nasoincisor ducts, which contain taste buds innervated by the greater superficial petrosal nerve. 2. The majority of information concerning the specificity of individual taste fibers in vertebrates has been obtained primarily in mammals to stimuli representing the four basic human taste qualities (i.e., salty, sweet, sour, and bitter). In the present report, we examine the evidence for gustatory fiber types within the stimulus class of amino acids, compounds known to be especially relevant gustatory stimuli for catfish and other teleosts. 3. Action potentials were recorded from 60 individual facial taste neurons obtained from 28 sea catfish (Arius felis). Stimuli were 10(-4) M concentrations of L-alanine, D-alanine, glycine, L-proline, L-histidine, and L-arginine, compounds selected from an original stimulus list of 28 amino acids. Responses were quantified as the number of action potentials evoked at various time intervals from the first 0.5 s up to 10 s of response time. 4. The spontaneous activity of 42 fully characterized neurons was 0.8 +/- 2.1 SD spikes/3 s. The average rate of spike discharge increased 50-fold during stimulation with the most effective amino acid (42 +/- 31 spikes/3 s, mean +/- SD). The majority of the sampled neurons were not narrowly tuned to the amino acid stimulants tested (mean breadth of responsiveness, H = 0.60; range 0-0.95). 5. Hierarchical cluster analysis of the fully characterized neurons identified two large and two small groups of cells. The

  3. Rapid Ti(III) reduction of perchlorate in the presence of beta-alanine: kinetics, pH effect, complex formation, and beta-alanine effect.

    PubMed

    Wang, Chao; Huang, Zhengdao; Lippincott, Lee; Meng, Xiaoguang

    2010-03-15

    Ti(III) reduction of perchlorate might be a useful method for the treatment of highly perchlorate-contaminated water. Though the reaction rate was usually low, we observed that beta-alanine (HOOCCH(2)CH(2)NH(2)) could significantly promote the reaction. A complete (>99.9%) perchlorate removal was obtained in a solution containing [ClO(4)(-)]=1.0mM, [Ti(III)]=40 mM, and [beta-alanine]=120 mM after 2.5h of reaction under 50 degrees C. The effects of both pH and complex formation on the reaction were then studied. The results showed that without beta-alanine the optimal pH was 2.3. When pH increased from 1.6 to 2.3, the reduction rate increased remarkably. In the pH range >2.3, however, the reduction was significantly inhibited, attributed to the formation of Ti(III) precipitate. The presence of beta-alanine at a molar ratio of [beta-alanine]:[Ti(III)]=3:1 significantly increased the reduction rate of perchlorate even at near neutral pH. This is because beta-alanine formed complexes with Ti(III), which greatly improved the total soluble [Ti(III)] in the pH range between 3.5 and 6. The findings may lead to the development of rapid treatment methods for intermittent and small stream of highly perchlorate-contaminated water, which are resulted from the manufacturing, storage, handling, use and/or disposal of large quantities of perchlorate salts.

  4. Monoclonal antibodies recognizing single amino acid substitutions in hemoglobin

    SciTech Connect

    Stanker, L.H.; Branscomb, E.; Vanderlaan, M.; Jensen, R.H.

    1986-06-01

    Four monoclonal antibodies (mAb) to non-human primate hemoglobin referred to as Cap-4, Cap-5, Rh-2, and Rh-4, and two mAb to human hemoglobin, referred to as H-1 and H-3 were isolated and were partially characterized. Binding studies with these mAb on a panel of hemoglobins and isolated ..cap alpha.. and ..beta.. globin chains revealed a unique reactivity pattern for each mAb. Amino acid sequence analysis of the antigens used to generate the binding data suggests that the specific recognition of certain hemoglobin antigens by each mAb is controlled by the presence of a particular amino acid at a specific position within the epitope. The use of synthetic peptides as antigens confirmed this observation for five of the mAb. No synthetic peptides were tested with the sixth mAb, Rh-2. The amino acids required for binding of mAb Cap-4, Cap-5, Rh-4, and Rh-2 to hemoglobin are alanine at ..beta..5, threonine at ..beta..13, glutamine at ..beta..125, and leucine at ..cap alpha..68. The non-human primate hemoglobin antibodies require a specific amino acid that is not present in human hemoglobin. The amino acid required for binding of Cap-4, Cap-5, and Rh-4 could arise by a single base change in the ..beta.. globin gene, whereas the amino acid required for Rh-2 binding could only occur if two base changes occurred. Thus these mAb are candidate probes for a somatic cell mutation assay on the basis of the detection of peripheral blood red cells that possess single amino acid substituted hemoglobin as a result of single base substitutions in the globin genes of precursor cells.

  5. Inducible l-Alanine Exporter Encoded by the Novel Gene ygaW (alaE) in Escherichia coli ▿

    PubMed Central

    Hori, Hatsuhiro; Yoneyama, Hiroshi; Tobe, Ryuta; Ando, Tasuke; Isogai, Emiko; Katsumata, Ryoichi

    2011-01-01

    We previously isolated a mutant hypersensitive to l-alanyl-l-alanine from a non-l-alanine-metabolizing Escherichia coli strain and found that it lacked an inducible l-alanine export system. Consequently, this mutant showed a significant accumulation of intracellular l-alanine and a reduction in the l-alanine export rate compared to the parent strain. When the mutant was used as a host to clone a gene(s) that complements the dipeptide-hypersensitive phenotype, two uncharacterized genes, ygaW and ytfF, and two characterized genes, yddG and yeaS, were identified. Overexpression of each gene in the mutant resulted in a decrease in the intracellular l-alanine level and enhancement of the l-alanine export rate in the presence of the dipeptide, suggesting that their products function as exporters of l-alanine. Since ygaW exhibited the most striking impact on both the intra- and the extracellular l-alanine levels among the four genes identified, we disrupted the ygaW gene in the non-l-alanine-metabolizing strain. The resulting isogenic mutant showed the same intra- and extracellular l-alanine levels as observed in the dipeptide-hypersensitive mutant obtained by chemical mutagenesis. When each gene was overexpressed in the wild-type strain, which does not intrinsically excrete alanine, only the ygaW gene conferred on the cells the ability to excrete alanine. In addition, expression of the ygaW gene was induced in the presence of the dipeptide. On the basis of these results, we concluded that YgaW is likely to be the physiologically most relevant exporter for l-alanine in E. coli and proposed that the gene be redesignated alaE for alanine export. PMID:21531828

  6. Changes in plasma amino acid concentrations with increasing age in patients with propionic acidemia.

    PubMed

    Scholl-Bürgi, Sabine; Sass, Jörn Oliver; Heinz-Erian, Peter; Amann, Edda; Haberlandt, Edda; Albrecht, Ursula; Ertl, Claudia; Sigl, Sara Baumgartner; Lagler, Florian; Rostasy, Kevin; Karall, Daniela

    2010-05-01

    The objective of the study is to analyze plasma amino acid concentrations in propionic acidemia (PA) for the purpose of elucidating possible correlations between propionyl-CoA carboxylase deficiency and distinct amino acid behavior. Plasma concentrations of 19 amino acids were measured in 240 random samples from 11 patients (6 families) with enzymatically and/or genetically proven propionic acidemia (sampling period, January 2001-December 2007). They were compared with reference values from the literature and correlated with age using the Pearson correlation coefficient test. Decreased plasma concentrations were observed for glutamine, histidine, threonine, valine, isoleucine, leucine, phenylalanine and arginine. Levels of glycine, alanine and aspartate were elevated, while values of serine, asparagine, ornithine and glutamate were normal. For lysine, proline and methionine a clear association was not possible. Significant correlations with age were observed for 13 amino acids (positive correlation: asparagine, glutamine, proline, alanine, histidine, threonine, methionine, arginine; negative correlation: leucine, phenylalanine, ornithine, glutamate and aspartate). This study gives new insight over long-term changes in plasma amino acid concentrations and may provide options for future therapies (e.g., substitution of anaplerotic substances) in PA patients.

  7. Determination of the D and L isomers of some protein amino acids present in soils

    NASA Technical Reports Server (NTRS)

    Pollock, G. E.; Cheng, C.-N.; Cronin, S. E.

    1977-01-01

    The D and L isomers of some protein amino acids present in soils were measured by using a gas chromatographic technique. The results of two processing procedures were compared to determine the better method. Results of the comparison indicated that the determination of D and L percentages requires amino acid purification if one is to obtain accurate data. It was found that very significant amounts of D-alanine, D-aspartic acid, and D-glutamic acid were present in the contemporary soils studied. Valine, isoleucine, leucine, proline, and phenylalanine generally contained only a trace to very small amounts of the D isomer. It is probable that the D-amino acids from the alanine, aspartic, and glutamic acids are contributed to the soil primarily via microorganisms. The finding of very significant quantities of some D-amino acids (about 5-16%) in present-day soils may alert some investigators of geological sediments to a possible problem in using amino acid racemization as an age-dating technique.

  8. Determination of the carbon, hydrogen and nitrogen contents of alanine and their uncertainties using the certified reference material L-alanine (NMIJ CRM 6011-a).

    PubMed

    Itoh, Nobuyasu; Sato, Ayako; Yamazaki, Taichi; Numata, Masahiko; Takatsu, Akiko

    2013-01-01

    The carbon, hydrogen, and nitrogen (CHN) contents of alanine and their uncertainties were estimated using a CHN analyzer and the certified reference material (CRM) L-alanine. The CHN contents and their uncertainties, as measured using the single-point calibration method, were 40.36 ± 0.20% for C, 7.86 ± 0.13% for H, and 15.66 ± 0.09% for N; the results obtained using the bracket calibration method were also comparable. The method described in this study is reasonable, convenient, and meets the general requirement of having uncertainties ≤ 0.4%.

  9. In silico characterization of the family of PARP-like poly(ADP-ribosyl)transferases (pARTs)

    PubMed Central

    Otto, Helge; Reche, Pedro A; Bazan, Fernando; Dittmar, Katharina; Haag, Friedrich; Koch-Nolte, Friedrich

    2005-01-01

    Background ADP-ribosylation is an enzyme-catalyzed posttranslational protein modification in which mono(ADP-ribosyl)transferases (mARTs) and poly(ADP-ribosyl)transferases (pARTs) transfer the ADP-ribose moiety from NAD onto specific amino acid side chains and/or ADP-ribose units on target proteins. Results Using a combination of database search tools we identified the genes encoding recognizable pART domains in the public genome databases. In humans, the pART family encompasses 17 members. For 16 of these genes, an orthologue exists also in the mouse, rat, and pufferfish. Based on the degree of amino acid sequence similarity in the catalytic domain, conserved intron positions, and fused protein domains, pARTs can be divided into five major subgroups. All six members of groups 1 and 2 contain the H-Y-E trias of amino acid residues found also in the active sites of Diphtheria toxin and Pseudomonas exotoxin A, while the eleven members of groups 3 – 5 carry variations of this motif. The pART catalytic domain is found associated in Lego-like fashion with a variety of domains, including nucleic acid-binding, protein-protein interaction, and ubiquitylation domains. Some of these domain associations appear to be very ancient since they are observed also in insects, fungi, amoebae, and plants. The recently completed genome of the pufferfish T. nigroviridis contains recognizable orthologues for all pARTs except for pART7. The nearly completed albeit still fragmentary chicken genome contains recognizable orthologues for twelve pARTs. Simpler eucaryotes generally contain fewer pARTs: two in the fly D. melanogaster, three each in the mosquito A. gambiae, the nematode C. elegans, and the ascomycete microfungus G. zeae, six in the amoeba E. histolytica, nine in the slime mold D. discoideum, and ten in the cress plant A. thaliana. GenBank contains two pART homologues from the large double stranded DNA viruses Chilo iridescent virus and Bacteriophage Aeh1 and only a single entry

  10. Degradation of aromatics and chloroaromatics by Pseudomonas sp. strain B13: purification and characterization of 3-oxoadipate:succinyl-coenzyme A (CoA) transferase and 3-oxoadipyl-CoA thiolase.

    PubMed

    Kaschabek, Stefan R; Kuhn, Bernd; Müller, Dagmar; Schmidt, Eberhard; Reineke, Walter

    2002-01-01

    The degradation of 3-oxoadipate in Pseudomonas sp. strain B13 was investigated and was shown to proceed through 3-oxoadipyl-coenzyme A (CoA) to give acetyl-CoA and succinyl-CoA. 3-Oxoadipate:succinyl-CoA transferase of strain B13 was purified by heat treatment and chromatography on phenyl-Sepharose, Mono-Q, and Superose 6 gels. Estimation of the native molecular mass gave a value of 115,000 +/- 5,000 Da with a Superose 12 column. Polyacrylamide gel electrophoresis under denaturing conditions resulted in two distinct bands of equal intensities. The subunit A and B values were 32,900 and 27,000 Da. Therefore it can be assumed that the enzyme is a heterotetramer of the type A2B2 with a molecular mass of 120,000 Da. The N-terminal amino acid sequences of both subunits are as follows: subunit A, AELLTLREAVERFVNDGTVALEGFTHLIPT; subunit B, SAYSTNEMMTVAAARRLKNGAVVFV. The pH optimum was 8.4. Km values were 0.4 and 0.2 mM for 3-oxoadipate and succinyl-CoA, respectively. Reversibility of the reaction with succinate was shown. The transferase of strain B13 failed to convert 2-chloro- and 2-methyl-3-oxoadipate. Some activity was observed with 4-methyl-3-oxoadipate. Even 2-oxoadipate and 3-oxoglutarate were shown to function as poor substrates of the transferase. 3-oxoadipyl-CoA thiolase was purified by chromatography on DEAE-Sepharose, blue 3GA, and reactive brown-agarose. Estimation of the native molecular mass gave 162,000 +/- 5,000 Da with a Superose 6 column. The molecular mass of the subunit of the denatured protein, as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, was 42 kDa. On the basis of these results, 3-oxoadipyl-CoA thiolase should be a tetramer of the type A4. The N-terminal amino acid sequence of 3-oxoadipyl-CoA thiolase was determined to be SREVYI-DAVRTPIGRFG. The pH optimum was 7.8. Km values were 0.15 and 0.01 mM for 3-oxoadipyl-CoA and CoA, respectively. Sequence analysis of the thiolase terminus revealed high percentages of identity

  11. Titration of Alanine Monitored by NMR Spectroscopy: A Biochemistry Laboratory Experiment

    ERIC Educational Resources Information Center

    Waller, Francis J.; And Others

    1977-01-01

    The experiment described here involves simultaneous monitoring of pH and NMR chemical shifts during an aqueous titration of alpha- and beta-alanine. This experiment is designed for use in an undergraduate biochemistry course. (MR)

  12. Second harmonic generation studies in L-alanine single crystals grown from solution

    NASA Astrophysics Data System (ADS)

    Boomadevi, Shanmugam; Pandiyan, Krishnamoorthy

    2014-01-01

    Single crystals of L-alanine of dimensions 2×1.1×0.5 cm3 were grown by evaporation method using deionised water as a solvent. The morphology of the grown crystals had (1 2 0) and (0 1 1) as their prominent faces. UV-vis-near IR spectrum shows the transparency range of L-alanine crystal available for frequency doubling from 250 to 1400 nm. Phase-matched second harmonic generation was observed in L-alanine sample by using 7 ns Q-switched Nd:YAG laser with OPO set up. In the present work, phase matching was achieved by angle and wavelength tuning. The angular and spectral phase-matching bandwidths were determined experimentally for a 1.5 mm thick L-alanine crystal and the results have been compared with their theoretical results. Further the possible reasons for the broadening of SHG spectrum have been discussed.

  13. Effects of preservation methods on amino acids and 5'-nucleotides of Agaricus bisporus mushrooms.

    PubMed

    Liu, Ying; Huang, Fan; Yang, Hong; Ibrahim, S A; Wang, Yan-Feng; Huang, Wen

    2014-04-15

    In this study, the proximate composition, free amino acids content and 5'-nucleotides in frozen, canned and salted Agaricus bisporus (A. bisporus) were investigated. We found that the three kinds of A. bisporus products were good sources of protein, with amount varying in the ranges of 16.54-24.35g/100g (dry weight). Freezing, canning and salting process, followed by 6months of storage led to a significant reduction in free amino acids, especially tyrosine, alanine, glutamine and cysteine. There were medium levels of MSG-like amino acids in frozen A. bisporus and canned A. bisporus, and low levels of MSG-like amino acids in salted A. bisporus. The mount of flavor 5'-nucleotides in frozen A. bisporus was higher than that of canned and salted A. bisporus. The present study thus suggests that freezing is beneficial for the preservation of A. bisporus.

  14. Study of stationary phase metabolism via isotopomer analysis of amino acids from an isolated protein.

    PubMed

    Shaikh, Afshan S; Tang, Yinjie J; Mukhopadhyay, Aindrila; Martín, Héctor García; Gin, Jennifer; Benke, Peter I; Keasling, Jay D

    2010-01-01

    Microbial production of many commercially important secondary metabolites occurs during stationary phase, and methods to measure metabolic flux during this growth phase would be valuable. Metabolic flux analysis is often based on isotopomer information from proteinogenic amino acids. As such, flux analysis primarily reflects the metabolism pertinent to the growth phase during which most proteins are synthesized. To investigate central metabolism and amino acids synthesis activity during stationary phase, addition of fully (13)C-labeled glucose followed by induction of green fluorescent protein (GFP) expression during stationary phase was used. Our results indicate that Escherichia coli was able to produce new proteins (i.e., GFP) in the stationary phase, and the amino acids in GFP were mostly from degraded proteins synthesized during the exponential growth phase. Among amino acid biosynthetic pathways, only those for serine, alanine, glutamate/glutamine, and aspartate/asparagine had significant activity during the stationary phase.

  15. Study of Stationary Phase Metabolism Via Isotopomer Analysis of Amino Acids from an Isolated Protein

    SciTech Connect

    Shaikh, AfshanS.; Tang, YinjieJ.; Mukhopadhyay, Aindrila; Martin, Hector Garcia; Gin, Jennifer; Benke, Peter; Keasling, Jay D.

    2009-09-14

    Microbial production of many commercially important secondary metabolites occurs during stationary phase, and methods to measure metabolic flux during this growth phase would be valuable. Metabolic flux analysis is often based on isotopomer information from proteinogenic amino acids. As such, flux analysis primarily reflects the metabolism pertinent to the growth phase during which most proteins are synthesized. To investigate central metabolism and amino acids synthesis activity during stationary phase, addition of fully 13C-labeled glucose followed by induction of green fluorescent protein (GFP) expression during stationary phase was used. Our results indicate that Escherichia coli was able to produce new proteins (i.e., GFP) in the stationary phase, and the amino acids in GFP were mostly from degraded proteins synthesized during the exponential growth phase. Among amino acid biosynthetic pathways, only those for serine, alanine, glutamate/glutamine, and aspartate/asparagine had significant activity during the stationary phase.

  16. Synthesis of amino acyl adenylates using the tert-butoxycarbonyl protecting group

    NASA Technical Reports Server (NTRS)

    Armstrong, D. W.; Seguin, R.; Saburi, M.; Fendler, J. H.

    1979-01-01

    The synthesis of amino acyl adenylates using N-tert-butoxycarbonyl-protected amino acids is reported. Anhydrous solutions containing N-tert-butoxycarbonyl alanine, phenylalanine, and methionine were combined with the anhydrous mono (tri-n-octylammonium) salt of adenosine 5'-phosphate and the resultant amino acyl adenylates were characterized by means of elemental analysis, and infrared and proton NMR spectroscopy. Amino acyl adenylate yields of up to 60% were obtained with high purity at room temperatures. The reported synthesis is considered to represent a large improvement over previous methods due to the purity of the products, normal temperature requirements, and the stability of the starting compounds, which suggests its use in investigations of prebiotic oligo- and polypeptide synthesis.

  17. 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.

  18. Amino Acid and Protein Metabolism in Bermuda Grass During Water Stress 12

    PubMed Central

    Barnett, N. M.; Naylor, A. W.

    1966-01-01

    The ability of Arizona Common and Coastal Bermuda grass [Cynodon dactylon (L.) Pers.] to synthesize amino acids and proteins during water stress was investigated. Amino acids were continually synthesized during the water stress treatments, but protein synthesis was inhibited and protein levels decreased. Water stress induced a 10- to 100-fold accumulation of free proline in shoots and a 2- to 6-fold accumulation of free asparagine, both of which are characteristic responses of water-stressed plants. Valine levels increased, and glutamic acid and alanine levels decreased. 14C labeling experiments showed that free proline turns over more slowly than any other free amino acid during water stress. This proline is readily synthesized and accumulated from glutamic acid. It is suggested that during water stress free proline functions as a storage compound. No significant differences were found in the amino acid and protein metabolism of the 2 varieties of Bermuda grass. PMID:16656387

  19. Formation of amino acids by cobalt-60 irradiation of hydrogen cyanide solutions

    NASA Technical Reports Server (NTRS)

    Sweeney, M. A.; Toste, A. P.; Ponnamperuma, C.

    1976-01-01

    Experiments were conducted to study the pathway for the prebiotic origin of amino acids from hydrogen cyanide (HCN) under the action of ionizing radiation considered as an effective source of energy on the primitive earth. The irradiations were performed in a cobalt-60 source with a dose rate of 200,000 rad/hr. Seven naturally occurring amino acids are identified among the products formed by the hydrolysis of gamma-irradiated solutions of HCN: glycine, alanine, valine, serine, threonine, aspartic acid, and glutamic acid. The identity of these amino acids is established by gas chromatography and mass spectrometry. Control experiments provided evidence that the amino acids are not the result of contamination.

  20. In-Situ Measurements of the Radiation Stability of Amino Acids at 15-140 K

    NASA Technical Reports Server (NTRS)

    Gerakines, Perry A.; Hudson, Reggie L.; Moore, Marla H.; Bell, Jan-Luca

    2012-01-01

    We present new kinetics data on the radiolytic destruction of amino acids measured in situ with infrared spectroscopy. Samples were irradiated at 15, 100, and 140 K with D.8-MeV protons, and amino-acid decay was followed at each temperature with and without H2O present. Observed radiation products included CO2 and amines, consistent with amino-acid decarboxylation. The half-lives of glycine, alanine, and phenylalanine were estimated for various extraterrestrial environments. Infrared spectral changes demonstrated the conversion from the non-zwitterion structure NH2-CH2(R)-COOH at 15 K to the zwitterion structure +NH3-CH2(R)-COO- at 140 K for each amino acid studied.

  1. Extraterrestrial Amino Acids in the Almahata Sitta Meteorite

    NASA Astrophysics Data System (ADS)

    Callahan, Michael; Aubrey, A.; Bada, J. L.; Dworkin, J. P.; Elsila, J. E.; Glavin, D. P.; Parker, E.; Jenniskens, P.

    2009-09-01

    The recovery of meteorite fragments from the 2008 TC3 asteroid impact, collectively named Almahata Sitta, revealed a rare, anomalous polymict ureilite containing large carbonaceous grains (Jenniskens et al. 2009). Here we report the first amino acid analysis of a meteorite from an F-type asteroid as part of the Almahata Sitta meteorite sample analysis consortium. A single fragment (piece #4, 1.2 grams) was crushed to a powder, and separate 0.1 g aliquots of the same meteorite were carried through identical hot-water extraction, acid hydrolysis and desalting procedures at NASA Goddard Space Flight Center and the Scripps Institution of Oceanography. The o-phthaldialdehyde/N-acetyl-L-cysteine amino acid derivatives in the extracts were analyzed by high performance liquid chromatography with UV fluorescence detection and time-of-flight mass spectrometry. Analyses of the meteorite extracts revealed a complex distribution of two- to six-carbon aliphatic amino acids with abundances ranging from 0.5 to 69 parts-per-billion (ppb). Glycine was the most abundant amino acid detected, however, since this protein amino acid is a common terrestrial contaminant, we are currently unable to rule out at least a partial terrestrial source. However, the D/L ratio of alanine in the meteorite was racemic, suggesting that very little terrestrial amino acid contamination. Several non-protein amino acids that are rare in the biosphere were also identified in the meteorite above background levels including D,L-4-amino-2-methybutyric acid (65 ± 8 ppb), D-isovaline (1.3 ± 0.1 ppb), L-isovaline (1.4 ± 0.1 ppb), and α-aminoisobutryic acid (7.1 ± 5.8 ppb). The abundance of isovaline and AIB are 1000 times lower than the abundances found in the CM2 meteorite Murchison while D,L-4-amino-2-methybutyric acid is similar. The very low amino acid abundances and the presence of several amino acid decomposition products including methylamine, ethylamine, and isopropylamine are consistent with

  2. Quantification of amino acids in fermentation media by isocratic HPLC analysis of their α-hydroxy acid derivatives.

    PubMed

    Pleissner, Daniel; Wimmer, Reinhard; Eriksen, Niels T

    2011-01-01

    In this paper we describe a novel method for quantification of amino acids. First, α-hydroxy acid derivatives of amino acids were formed after reaction with dinitrogen trioxide by the van Slyke reaction. Second, the α-hydroxy acid derivatives were separated on an Aminex HPX-87H column (Bio-Rad) eluted isocratically with 5 mM H(2)SO(4) and quantified by refractive index detection. We were able to measure the reaction products of 13 of the 20 classical amino acids: glycine, l-alanine, l-valine, l-leucine, l-isoleucine, l-methionine, l-serine, l-threonine, l-asparagine, l-glutamine, l-aspartic acid, l-glutamic acid, and l-proline. We obtained linear relationships between the product peak areas and initial amino acid concentration, whereby the concentrations of these amino acids could be quantified on the basis of the quantification of their products. The method can be used to analyze amino acids in parallel with other small molecules, such as sugars or short chain fatty acids, and was used for parallel quantification of glycine, l-alanine, or l-glutamic acid, and glucose uptake in cultures of the heterotrophic dinoflagellate Crypthecodinium cohnii . The method can also be used to quantify other amines, as demonstrated by detection of Tris (2-amino-2-(hydroxymethyl)propane-1,3-diol).

  3. Rational design of mirror-like peptides with alanine regulation.

    PubMed

    Li, Weizhong; Tan, Tingting; Xu, Wei; Xu, Lin; Dong, Na; Ma, Deying; Shan, Anshan

    2016-02-01

    To generate effective antimicrobial peptides (AMPs) with good antimicrobial activities and cell selectivity, many synthetic strategies have been implemented to facilitate the development of AMPs. However, these synthetic strategies represent only a small proportion of the methods used for the development of AMPs and are not optimal with the requirements needed for the design of AMPs. In this investigation, we designed a mirror-like structure with a lower charge and a higher number of hydrophobic amino acids. The amino acid sequence of the designed mirror-like peptides was XXYXXXYXXXYXX [X represents L (Leu) and/or A (Ala); Y represents K (Lys)]. These mirror-like peptides displayed antimicrobial activity against both Gram-positive and Gram-negative bacteria. Hemolysis activity and cytotoxicity, detected by using human red blood cells (hRBCs) and human embryonic kidney cells (HEK293), respectively, demonstrated that the frequency of Ala residues in this structure had a regulatory effect on the high hydrophobic region. In particular, KL4A6 showed a greater antimicrobial potency than the other three mirror-like peptides, folded into an α-helical structure, and displayed the highest therapeutic index, suggesting its good cell selectivity. Observations from fluorescence spectroscopy, flow cytometry, and electron microscopy experiments indicated that KL4A6 exhibited good membrane penetration potential by inducing membrane blebbing, disruption and lysis. Therefore, generating mirror-like peptides is a promising strategy for designing effective AMPs with regions of high hydrophobicity.

  4. SU-E-T-643: Pure Alanine Dosimeter for Verification Dosimetry in IMRT

    SciTech Connect

    Al-Karmi, Anan M.; Zraiqat, Fadi

    2015-06-15

    Purpose: The objective of this study was evaluation of accuracy of pure alanine dosimeters measuring intensity-modulated radiation therapy (IMRT) dose distributions in a thorax phantom. Methods: Alanine dosimeters were prepared in the form of 110 mg pure L-α-alanine powder filled into clear tissue-equivalent polymethylmethacrylate (PMMA) plastic tubes with the dimensions 25 mm length, 3 mm inner diameter, and 1 mm wall thickness. A dose-response calibration curve was established for the alanine by placing the dosimeters at 1.5 cm depth in a 30×30×30 cm{sup 3} solid water phantom and then irradiating on a linac with 6 MV photon beam at 10×10 cm{sup 2} field size to doses ranging from 1 to 5 Gy. Electron paramagnetic resonance (EPR) spectroscopy was used to determine the absorbed dose in alanine. An IMRT treatment plan was designed for a commercial heterogeneous CIRS thorax phantom and the dose values were calculated at three different points located in tissue, lung, and bone equivalent materials. A set of dose measurements was carried out to compare measured and calculated dose values by placing the alanine dosimeters at those selected locations inside the thorax phantom and delivering the IMRT to the phantom. Results: The alanine dose measurements and the IMRT plan dose calculations were found to be in agreement within ±2%. Specifically, the deviations were −0.5%, 1.3%, and −1.7% for tissue, lung, and bone; respectively. The slightly large deviations observed for lung and bone may be attributed to tissue inhomogeneity, steep dose gradients in these regions, and uncontrollable changes in spectrometer conditions. Conclusion: The results described herein confirmed that pure alanine dosimeter was suitable for in-phantom dosimetry of IMRT beams because of its high sensitivity and acceptable accuracy. This makes the dosimeter a promising option for quality control of the therapeutic beams, complementing the commonly used ionization chambers, TLDs, and films.

  5. Identification of the Fluvirucin B2 (Sch 38518) Biosynthetic Gene Cluster from Actinomadura fulva subsp. indica ATCC 53714: substrate Specificity of the β-Amino Acid Selective Adenylating Enzyme FlvN.

    PubMed

    Miyanaga, Akimasa; Hayakawa, Yuki; Numakura, Mario; Hashimoto, Junko; Teruya, Kuniko; Hirano, Takashi; Shin-Ya, Kazuo; Kudo, Fumitaka; Eguchi, Tadashi

    2016-05-01

    Fluvirucins are 14-membered macrolactam polyketides that show antifungal and antivirus activities. Fluvirucins have the β-alanine starter unit at their polyketide skeletons. To understand the construction mechanism of the β-alanine moiety in fluvirucin biosyntheses, we have identified the biosynthetic cluster of fluvirucin B2 produced from Actinomadura fulva subsp. indica ATCC 53714. The identified gene cluster contains three polyketide synthases, four characteristic β-amino acid-carrying enzymes, one decarboxylase, and one amidohydrolase. We next investigated the activity of the adenylation enzyme FlvN, which is a key enzyme for the selective incorporation of a β-amino acid substrate. FlvN showed strong preference for l-aspartate over other amino acids such as β-alanine. Based on these results, we propose a biosynthetic pathway for fluvirucin B2.

  6. PARTICIPATION OF Y89 AND Y97 IN THE CONJUGATING ACTIVITY OF Drosophila melanogaster GLUTATHIONE S-TRANSFERASE D3 (DmGSTD3).

    PubMed

    Vignesvaran, Kithalakshmi; Alias, Zazali

    2016-07-01

    Drosophila melanogaster glutathione S-transferase D3 (DmGSTD3) has a shorter amino acid sequence as compared to other GSTs known in the fruit flies. This is due to the 15 amino acid N-terminal truncation in which normally active amino acid residue is located. The work has made use of homology modeling to visualize the arrangement of amino acid side chains in the glutathione (GSH) substrate cavity. The identified amino acids were then replaced with amino acids without functional groups in the side chains and the mutants were analyzed kinetically. Homology modeling revealed that the side chains of Y89 and Y97 were shown facing toward the substrate cavity proposing their possible role in catalyzing the conjugation. Y97A and Y89A GSH gave large changes in Km (twofold increase), Vmax (fivefold reduction), and Kcat /Km values for GSH suggesting their significant role in the conjugation reaction. The replacement at either positions has not affected the affinity of the enzyme toward 1-chloro-2,4-dinitrobenzene as no significant change in values of Kmax was observed. The replacement, however, had significantly reduced the catalytic efficiency of both mutants with (Kcat /Km )(GSH) and (Kcat /Km )(CDNB) of eight- and twofold reduction. The recombinant DmGSTD3 has shown no activity toward 1,2-dichloro-4-nitrobenzene, 2,4-hexadienal, 2,4-heptadienal, p-nitrobenzyl chloride, ethacrynic acid, and sulfobromophthalein. Therefore, it was evident that DmGSTD3 has made use of distal amino acids Y97 and Y89 for GSH conjugation.

  7. [Alanine solution as enzyme reaction buffer used in A to O blood group conversion].

    PubMed

    Li, Su-Bo; Zhang, Xue; Zhang, Yin-Ze; Tan, Ying-Xia; Bao, Guo-Qiang; Wang, Ying-Li; Ji, Shou-Ping; Gong, Feng; Gao, Hong-Wei

    2014-06-01

    The aim of this study was to investigate the effect of alanine solution as α-N-acetylgalactosaminidase enzyme reaction buffer on the enzymatic activity of A antigen. The binding ability of α-N-acetylgalactosaminidase with RBC in different reaction buffer such as alanine solution, glycine solution, normal saline (0.9% NaCl), PBS, PCS was detected by Western blot. The results showed that the efficiency of A to O conversion in alanine solution was similar to that in glycine solution, and Western blot confirmed that most of enzymes blinded with RBC in glycine or alanine solution, but few enzymes blinded with RBC in PBS, PCS or normal saline. The evidences indicated that binding of enzyme with RBC was a key element for A to O blood group conversion, while the binding ability of α-N-acetylgalactosaminidase with RBC in alanine or glycine solution was similar. It is concluded that alanine solution can be used as enzyme reaction buffer in A to O blood group conversion. In this buffer, the α-N-acetylgalactosaminidase is closely blinded with RBC and α-N-acetylgalactosaminidase plays efficient enzymatic activity of A antigen.

  8. The Synthesis and Evaluation of Arctigenin Amino Acid Ester Derivatives.

    PubMed

    Cai, En-Bo; Yang, Li-Min; Jia, Cai-Xia; Zhang, Wei-Yuan; Zhao, Yan; Li, Wei; Song, Xing-Zhuo; Zheng, Man-Ling

    2016-10-01

    The use of arctigenin (ARG), a traditional medicine with many pharmacological activities, has been restricted due to its poor solubility in water. Five amino acid derivatives of ARG have been synthesized using glycine, o-alanine, valine, leucine, and isoleucine, which have t-butyloxy carbonyl (BOC) as a protective group. In this study, we examined the effects of removing these protective groups. The results showed that the amino acid derivatives have better solubility and nitrite-clearing ability than ARG. Among the compounds tested, the amino acid derivatives without protective group were the best. Based on these results, ARG and its two amino acid derivatives without protective group (ARG8, ARG10) were selected to evaluate their anti-tumor activity in vivo at a dosage of 40 mg/kg. The results indicated that ARG8 and ARG10 both exhibit more anti-tumor activity than ARG in H22 tumor-bearing mice. The tumor inhibition rates of ARG8 and ARG10 were 69.27 and 43.58%, which was much higher than ARG. Furthermore, the mice treated with these compounds exhibited less damage to the liver, kidney and immune organs compared with the positive group. Furthermore, ARG8 and ARG10 improved the serum cytokine levels significantly compared to ARG. In brief, this study provides a method to improve the water solubility of drugs, and we also provide a reference basis for new drug development.

  9. Industrial production of amino acids by coryneform bacteria.

    PubMed

    Hermann, Thomas

    2003-09-04

    In the 1950s Corynebacterium glutamicum was found to be a very efficient producer of L-glutamic acid. Since this time biotechnological processes with bacteria of the species Corynebacterium developed to be among the most important in terms of tonnage and economical value. L-Glutamic acid and L-lysine are bulk products nowadays. L-Valine, L-isoleucine, L-threonine, L-aspartic acid and L-alanine are among other amino acids produced by Corynebacteria. Applications range from feed to food and pharmaceutical products. The growing market for amino acids produced with Corynebacteria led to significant improvements in bioprocess and downstream technology as well as in molecular biology. During the last decade big efforts were made to increase the productivity and to decrease the production costs. This review gives an overview of the world market for amino acids produced by Corynebacteria. Significant improvements in bioprocess technology, i.e. repeated fed batch or continuous production are summarised. Bioprocess technology itself was improved furthermore by application of more sophisticated feeding and automatisation strategies. Even though several amino acids developed towards commodities in the last decade, side aspects of the production process like sterility or detection of contaminants still have increasing relevance. Finally one focus of this review is on recent developments in downstream technology.

  10. Comparison of Free Total Amino Acid Compositions and Their Functional Classifications in 13 Wild Edible Mushrooms.

    PubMed

    Sun, Liping; Liu, Qiuming; Bao, Changjun; Fan, Jian

    2017-02-24

    Thirteen popular wild edible mushroom species in Yunnan Province, Boletus bicolor, Boletus speciosus, Boletus sinicus, Boletus craspedius, Boletus griseus, Boletus ornatipes, Xerocomus, Suillus placidus, Boletinus pinetorus, Tricholoma terreum, Tricholomopsis lividipileata, Termitomyces microcarpus, and Amanita hemibapha, were analyzed for their free amino acid compositions by online pre-column derivazation reversed phase high-performance liquid chromatography (RP-HPLC) analysis. Twenty free amino acids, aspartic acid, glutamic acid, serine, glycine, alanine, praline, cysteine, valine, methionine, phenylalanine, isoleucine, leucine, lysine, histidine, threonine, asparagines, glutamine, arginine, tyrosine, and tryptophan, were determined. The total free amino acid (TAA) contents ranged from 1462.6 mg/100 g in B. craspedius to 13,106.2 mg/100 g in T. microcarpus. The different species showed distinct free amino acid profiles. The ratio of total essential amino acids (EAA) to TAA was 0.13-0.41. All of the analyzed species showed high contents of hydrophobic amino acids, at 33%-54% of TAA. Alanine, cysteine, glutamine, and glutamic acid were among the most abundant amino acids present in all species. The results showed that the analyzed mushrooms possessed significant free amino acid contents, which may be important compounds contributing to the typical mushroom taste, nutritional value, and potent antioxidant properties of these wild edible mushrooms. Furthermore, the principal component analysis (PCA) showed that the accumulative variance contribution rate of the first four principal components reached 94.39%. Cluster analysis revealed EAA composition and content might be an important parameter to separate the mushroom species, and T. microcarpus and A. hemibapha showed remarkable EAA content among the 13 species.

  11. Plant and Soil Emissions of Amines and Amino Acids: A Source of Secondary Aerosol Precursors

    NASA Astrophysics Data System (ADS)

    Jackson, M. L.; Doskey, P. V.; Pypker, T. G.

    2011-12-01

    Ammonia (NH3) is the most abundant alkaline gas in the atmosphere and forms secondary aerosol by neutralizing sulfuric and nitric acids that are released during combustion of fossil fuels. Ammonia is primarily emitted by cropping and livestock operations. However, C2 and C3 amines (pKb 3.3-3.4), which are stronger bases than NH3 (pKb 4.7) have been observed in nuclei mode aerosol that is the precursor to secondary aerosol. Mixtures of amines and amino acids have been identified in diverse environments in aerosol, fog water, cloud water, the soluble fraction of precipitation, and in dew. Glycine (pKb 4.2), serine (pKb 4.8) and alanine (pKb 3.7 and 4.1 for the D and L forms, respectively) are typically the most abundant species. The only reported values of gas-phase glycine, serine and alanine were in marine air and ranged from 6-14 pptv. The origin of atmospheric amines and amino acids has not been fully identified, although sources are likely similar to NH3. Nitrate assimilation in plants forms glycine, serine, and L-alanine, while D-alanine is present in bacterial cell walls. Glycine is converted to serine during C3 plant photorespiration, producing CO2 and NH3. Bacteria metabolize glycine and alanine to methylamine and ethylamine via decarboxylation. Likely sources of amino acids are plants and bacteria, thus concentrations near continental sources are likely greater than those measured in marine air. The overall goal of the research is to examine seasonal variations and relationships between the exchange of CO2, NH3, amines, and amino acids with a corn/soybean rotation in the Midwest Corn Belt. The study presents gaseous profiles of organic amine compounds from various species of vegetation using a mist chamber trapping technique and analysis of the derivatized species by high pressure liquid chromatography with fluorescence detection. Amino acid and amine profiles were obtained for red oak (Quercus rubra), sugar maple (Acer saccharinum), white pine (Pinus

  12. Role of Host Cell-Derived Amino Acids in Nutrition of Intracellular Salmonella enterica

    PubMed Central

    Popp, Jasmin; Noster, Janina; Busch, Kim; Kehl, Alexander; zur Hellen, Gero

    2015-01-01

    The facultative intracellular pathogen Salmonella enterica resides in a specific membrane-bound compartment termed the Salmonella-containing vacuole (SCV). Despite being segregated from access to metabolites in the host cell cytosol, Salmonella is able to efficiently proliferate within the SCV. We set out to unravel the nutritional supply of Salmonella in the SCV with focus on amino acids. We studied the availability of amino acids by the generation of auxotrophic strains for alanine, asparagine, aspartate, glutamine, and proline in a macrophage cell line (RAW264.7) and an epithelial cell line (HeLa) and examined access to extracellular nutrients for nutrition. Auxotrophies for alanine, asparagine, or proline attenuated intracellular replication in HeLa cells, while aspartate, asparagine, or proline auxotrophies attenuated intracellular replication in RAW264.7 macrophages. The different patterns of intracellular attenuation of alanine- or aspartate-auxotrophic strains support distinct nutritional conditions in HeLa cells and RAW264.7 macrophages. Supplementation of medium with individual amino acids restored the intracellular replication of mutant strains auxotrophic for asparagine, proline, or glutamine. Similarly, a mutant strain deficient in succinate dehydrogenase was complemented by the extracellular addition of succinate. Complementation of the intracellular replication of auxotrophic Salmonella by external amino acids was possible if bacteria were proficient in the induction of Salmonella-induced filaments (SIFs) but failed in a SIF-deficient background. We propose that the ability of intracellular Salmonella to redirect host cell vesicular transport provides access of amino acids to auxotrophic strains and, more generally, is essential to continuously supply bacteria within the SCV with nutrients. PMID:26351287

  13. The secretory omega-class glutathione transferase OvGST3 from the human pathogenic parasite Onchocerca volvulus.

    PubMed

    Liebau, Eva; Höppner, Jana; Mühlmeister, Mareike; Burmeister, Cora; Lüersen, Kai; Perbandt, Markus; Schmetz, Christel; Büttner, Dietrich; Brattig, Norbert

    2008-07-01

    Onchocerciasis or river blindness, caused by the filarial nematode Onchocerca volvulus, is the second leading cause of blindness due to infectious diseases. The protective role of the omega-class glutathione transferase 3 from O. volvulus (OvGST3) against intracellular and environmental reactive oxygen species has been described previously. In the present study, we continue our investigation of the highly stress-responsive OvGST3. Alternative splicing of two exons and one intron retention generates five different transcript isoforms that possess a spliced leader at their 5'-end, indicating that the mechanism of mature mRNA production involves alternative-, cis- and trans-splicing processes. Interestingly, the first two exons of the ovgst3 gene encode a signal peptide before sequence identity to other omega-class glutathione transferases begins. Only the recombinant expression of the isoform that encodes the longest deduced amino acid sequence (OvGST3/5) was successful, with the purified enzyme displaying modest thiol oxidoreductase activity. Significant IgG1 and IgG4 responses against recombinantly expressed OvGST3/5 were detected in sera from patients with the generalized as well as the chronic hyperreactive form of onchocerciasis, indicating exposure of the secreted protein to the human host's immune system and its immunogenicity. Immunohistological localization studies performed at light and electron microscopy levels support the extracellular localization of the protein. Intensive labeling of the OvGST3 was observed in the egg shell at the morula stage of the embryo, indicating extremely defined, stage-specific expression for a short transient period only.

  14. Expression, crystallization and preliminary X-ray crystallographic analysis of D-alanine-D-alanine ligase from OXA-23-producing Acinetobacter baumannii K0420859.

    PubMed

    Huynh, Kim-Hung; Tran, Huyen-Thi; Pham, Tan-Viet; Ngo, Ho-Phuong-Thuy; Cha, Sun-Shin; Chung, Kyung Min; Lee, Sang Hee; Kang, Lin-Woo

    2014-04-01

    Acinetobacter baumannii causes bacteraemia, pneumonia, other respiratory-tract and urinary-tract infections in humans. OXA-23 carbapenemase-producing A. baumannii K0420859 (A. baumannii OXA-23) is resistant to carbapenem, a common antibacterial drug. To develop an efficient and novel antibacterial drug against A. baumannii OXA-23, D-alanine-D-alanine ligase, which is essential in bacterial cell-wall synthesis, is of interest. Here, the D-alanine-D-alanine ligase (AbDdl) gene from A. baumannii OXA-23 was cloned and expressed, and the AbDdl protein was purified and crystallized; this enzyme can be used as a novel target for an antibacterial drug against A. baumannii OXA-23. The AbDdl crystal diffracted to a resolution of 2.8 Å and belonged to the orthorhombic space group P212121, with unit-cell parameters a = 113.4, b = 116.7, c = 176.5 Å, a corresponding VM of 2.8 Å(3) Da(-1) and a solvent content of 56.3%, and six protomers in the asymmetric unit.

  15. Enrichment of Non-Terrestrial L-Proteinogenic Amino Acids by Aqueous Alteration on the Tagish Lake Meteorite Parent Body

    NASA Technical Reports Server (NTRS)

    Glavin, Daniel P.; Elsila, Jamie E.; Burton, Aaron S.; Callahan, Michael P.; Dworkin, Jason P.; Herd, Christopher D. K.

    2012-01-01

    The distribution and isotopic and enantiomeric compositions of amino acids found in three distinct fragments of the Tagish Lake C2-type carbonaceous chondrite were investigated via liquid chromatography fluorescence detection time-of-flight mass spectrometry and gas chromatography isotope ratio mass spectrometry. Large L-enantiomeric excesses (L(sub ee) approx. 43 to 59%) of the a-hydrogen aspartic and glutamic amino acids were measured in Tagish Lake, whereas alanine, another alpha-hydrogen protein amino acid, was found to be nearly racemic (D approx. L) using both techniques. Carbon isotope measurements of D- and L-aspartic acid and D- and L-alanine in Tagish Lake fall well outside of the terrestrial range and indicate that the measured aspartic acid enantioenrichment is indigenous to the meteorite. Alternate explanations for the Lexcesses of aspartic acid such as interference from other compounds present in the sample, analytical biases, or terrestrial amino acid contamination were investigated and rejected. These results can be explained by differences in the solid-solution phase behavior of aspartic acid, which can form conglomerate enantiopure solids during crystallization, and alanine, which can only form racemic crystals.

  16. Methyl jasmonate differentially affects tocopherol content and tyrosine amino transferase activity in cultured cells of Amaranthus caudatus and Chenopodium quinoa.

    PubMed

    Antognoni, F; Faudale, M; Poli, F; Biondi, S

    2009-03-01

    Tocopherols are lipid-soluble compounds synthesised exclusively by photosynthetic organisms. In this study, in vitro callus cultures were established from two plants that are naturally rich in tocopherols, Amaranthus caudatus and Chenopodium quinoa, in order to examine whether callus cultures were able to produce these compounds at levels comparable to those observed in planta. In both species, cotyledon explants produced the best callus induction and, once established, callus cultures were grown under two different hormonal treatments to check for effects of growth and to induce chloroplast differentiation in the cells. A rapid differentiation of chloroplasts occurred only in C. quinoa cell aggregates grown in the presence of benzyladenine, leading to the production of a homogeneous green callus. In both species, only alpha-tocopherol was produced by callus cultures, although levels were much lower than in planta, and the production was not influenced by the hormonal conditions. Interestingly, cell cultures of the two species responded in different ways to methyl jasmonate (MJ). In A. caudatus cultures, treatment with 100 mum MJ increased the production of alpha-tocopherol up to fivefold, and the inductive effect was influenced by the hormonal composition of the medium. This increase in alpha-tocopherol was associated with a proportional increase in tyrosine aminotransferase (TAT) activity, one of the key enzymes involved in tocopherol biosynthesis. By contrast, in C. quinoa cultures, elicitation with MJ did not have any effect, neither on tocopherol production, nor on TAT activity. These results are discussed in relation to chloroplast differentiation and the interplay between jasmonates and phytohormones.

  17. Nuclear translocation of glutathione S-transferase {pi} is mediated by a non-classical localization signal

    SciTech Connect

    Kawakatsu, Miho; Goto, Shinji; Yoshida, Takako; Urata, Yoshishige; Li, Tao-Sheng

    2011-08-12

    Highlights: {yields} Nuclear translocation of GST{pi} is abrogated by the deletion of the last 16 amino acid residues in the carboxy-terminal region, indicating that residues 195-208 of GST{pi} are required for nuclear translocation. {yields} The lack of a contiguous stretch of positively charged amino acid residues within the carboxy-terminal region of GST{pi}, suggests that the nuclear translocation of GST{pi} is mediated by a non-classical nuclear localization signal. {yields} An in vitro transport assay shows that the nuclear translocation of GST{pi} is dependent on cytosolic factors and ATP. -- Abstract: Glutathione S-transferase {pi} (GST{pi}), a member of the GST family of multifunctional enzymes, is highly expressed in human placenta and involved in the protection of cellular components against electrophilic compounds or oxidative stress. We have recently found that GST{pi} is expressed in the cytoplasm, mitochondria, and nucleus in some cancer cells, and that the nuclear expression of GST{pi} appears to correlate with resistance to anti-cancer drugs. Although the mitochondrial targeting signal of GST{pi} was previously identified in the amino-terminal region, the mechanism of nuclear translocation remains completely unknown. In this study, we find that the region of GST{pi}195-208 is critical for nuclear translocation, which is mediated by a novel and non-classical nuclear localization signal. In addition, using an in vitro transport assay, we demonstrate that the nuclear translocation of GST{pi} depends on the cytosolic extract and ATP. Although further experiments are needed to understand in depth the precise mechanism of nuclear translocation of GST{pi}, our results may help to establish more efficient anti-cancer therapy, especially with respect to resistance to anti-cancer drugs.

  18. Free amino acid composition in hemolymph and muscle of the ghost crab, Ocypode platytarsis.

    PubMed

    Sankar, R Siva; Yogamoorthi, A

    2012-05-15

    Free amino acid plays an important role in physiological functions. The ghost crab, Ocypode platytarsis caught off the Puducherry sandy beaches, south east coast of India were investigated for the pattern of distribution of free amino acids. The hemolymph and muscle were sampled from male and female crabs and then subjected to free amino acid analysis quantitatively by HPLC. Fourteen free amino acids have been determined in hemolymph and muscle of both sexes in Ocypode platytarsis. The essential amino acids were leucine, lysine, methionine, phenylalanine, threonine, valine and histidine and non-essential amino acids were alanine, arginine, glutamic acid, glycine, serine and tyrosine. The total concentration of free amino acid found in the male and female hemolymph was lower than that in the muscle. Glutamic acid is of higher concentration in the hemolymph whereas histidine found was higher in the muscle of both sexes. The male crab contains a strikingly higher concentration of histidine of about eight times than that of female in muscles. The study revealed that distribution pattern of free amino acids differ in relation to hemolymph and muscle of male and female crabs.

  19. Composition of antioxidants and amino acids in Stevia leaf infusions.

    PubMed

    Periche, Angela; Koutsidis, Georgios; Escriche, Isabel

    2014-03-01

    Stevia, a non-caloric natural sweetener with beneficial properties and considerable antioxidants and amino acids, is increasingly consumed as an infusion. This work evaluates the influence of the conditions (temperature: 50, 70 or 90 °C and time: 1, 5, 20 or 40 min) applied to obtain Stevia infusions, on antioxidants (total phenols, flavonoids and antioxidant activity) and amino acids. The total concentration of the eleven amino acids found was 11.70 mg/g in dried leaves and from 6.84 to 9.11 mg/g per gram of Stevia in infusions. However, infusions showed higher levels of certain amino acids (alanine, asparagine, leucine and proline), and greater values of the three antioxidant parameters in comparison with dry leaves. Temperature had more influence (minimum values at 50 °C and maximum at 90 °C) than time in the case of antioxidants. At 90 °C there were no important increases in the extraction of antioxidant compounds after 5 min; each gram of Stevia had 117 mg trolox (total antioxidant activity), 90 mg gallic acid (total phenols) and 56 mg catechin equivalents (flavonoids). Varying the temperature and time conditions no notable differences were observed in the concentrations of the majority of amino acids. However, the infusion treatment at 90 °C for 5 min was the best, as it gave the highest yield of 8 of the 11 amino acids. Therefore, with respect to the compounds analyzed in this study, the best way to obtain Stevia leaf infusions is the same as the domestic process, almost boiling water for a short time.

  20. Energetics of the molecular interactions of L-alanine and L-serine with xylitol, D-sorbitol, and D-mannitol in aqueous solutions at 298.15 K

    NASA Astrophysics Data System (ADS)

    Mezhevoi, I. N.; Badelin, V. G.

    2013-04-01

    Integral enthalpies of dissolution Δsol H m of L-alanine and L-serine are measured via the calorimetry of dissolution in aqueous solutions of xylitol, D-sorbitol, and D-mannitol. Standard enthalpies of dissolution (Δsol H ○) and the transfer (Δtr H ○) of amino acids from water to binary solvent are calculated from the experimental data. Using the McMillan-Mayer theory, enthalpy coefficients of pairwise interactions h xy of amino acids with molecules of polyols are calculated that are negative. The obtained results are discussed within the theory of the prevalence of different types of interactions in mixed solutions and the effect of the structural features of interacting biomolecules on the thermochemical parameters of dissolution of amino acids.