Phenylalanine Precursor Research Articles

Whitefly genome expression reveals host-symbiont interaction in amino acid biosynthesis.

BackgroundWhitefly (Bemisia tabaci) complex is a serious insect pest of several crop plants worldwide. It comprises several morphologically indistinguishable species, however very little is known about their genetic divergence and biosynthetic pathways. In the present study, we performed transcriptome sequencing of Asia 1 species of B. tabaci complex and analyzed the interaction of host-symbiont genes in amino acid biosynthetic pathways.Methodology/Principal FindingsWe obtained about 83 million reads using Illumina sequencing that assembled into 72716 unitigs. A total of 21129 unitigs were annotated at stringent parameters. Annotated unitigs were mapped to 52847 gene ontology (GO) terms and 131 Kyoto encyclopedia of genes and genomes (KEGG) pathways. Expression analysis of the genes involved in amino acid biosynthesis pathways revealed the complementation between whitefly and its symbiont partner Candidatus Portiera aleyrodidarum. Most of the non-essential amino acids and intermediates of essential amino acid pathways were supplied by the host insect to its symbiont. The symbiont expressed the pathways for the essential amino acids arginine, threonine and tryptophan and the immediate precursors of valine, leucine, isoleucine and phenyl-alanine. High level expression of the amino acid transporters in the whitefly suggested the molecular mechanisms for the exchange of amino acids between the host and the symbiont.Conclusions/SignificanceOur study provides a comprehensive transcriptome data for Asia 1 species of B. tabaci complex that focusses light on integration of host and symbiont genes in amino acid biosynthesis pathways.

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

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

Plant Cell Biology: The ABC of Monolignol Transport

Lignins are complex aromatic heteropolymers that reinforce the cell walls of terrestrial plants. A new study identifies an ATP-binding cassette ABC transporter that pumps a monolignol lignin precursor across the plasma membrane.

Acute metabolic encephalopathy in two patients treated with asparaginase and ondasetron

L-asparaginase has been an important agent of acute lymphoblastic leukemia (ALL) therapy for more than 40 years. The major adverse events of L-asparaginase are immunoallergic reaction, pancreatitis, liver cytolysis (increased ASAT, ALAT, and bilirubin), and cerebral haemorrhage or thrombosis. Hyperammonemia has already been described with L -asparaginase treatment [1]. In the present article, we report acute metabolic encephalopathy in two young boys receiving asparaginase for B ALL relapse. We discuss the pathophysiology of this metabolic encephalopathy with hyperammonemia and its medical management.

ChemInform Abstract: Chiral 2-Cyanocinnamates in Conjugate Addition Asymmetric Enolate Trapping Reactions.

Chiral 2-cyanocinnamates react with L-selectride® to give an intermediate enolate which can be stereoselectively trapped with different halides to afford α-substituted phenylalanine precursors with excellent chemical yields and good diastereomeric excess.

Synthesis of constrained aromatic β2,3,3-amino alcohol scaffolds — Precursors of non-proteinogenic phenylalanine for peptidomimetics

Novel constrained β2,3,3-amino alcohol scaffolds (11a–11e) with varied substitution patterns have been synthesized by reductive cleavage of the N–O bond of isoxazolidines (10a–10e) derived from intramolecular 1,3-dipolar cycloadditions. A number of methods were probed for reductive cleavage, however, better results were obtained with ammonium formate in the presence of Pd/C or Zn as compared to Zn/acetic acid. The obtained scaffolds (11a–11e) can serve as valuable building blocks for peptidomimetics.Key words: β2,3,3-amino alcohol, peptidomimetics, building blocks, isoxazolidines, reductive cleavage, ammonium formate.

Structural studies of prephenate dehydratase from Mycobacterium tuberculosis H37Rv by SAXS, ultracentrifugation, and computational analysis

Tuberculosis (TB) is one of the most common infectious diseases known to man and responsible for millions of human deaths in the world. The increasing incidence of TB in developing countries, the proliferation of multidrug resistant strains, and the absence of resources for treatment have highlighted the need of developing new drugs against TB. The shikimate pathway leads to the biosynthesis of chorismate, a precursor of aromatic amino acids. This pathway is absent from mammals and shown to be essential for the survival of Mycobacterium tuberculosis, the causative agent of TB. Accordingly, enzymes of aromatic amino acid biosynthesis pathway represent promising targets for structure-based drug design. The first reaction in phenylalanine biosynthesis involves the conversion of chorismate to prephenate, catalyzed by chorismate mutase. The second reaction is catalyzed by prephenate dehydratase (PDT) and involves decarboxylation and dehydratation of prephenate to form phenylpyruvate, the precursor of phenylalanine. Here, we describe utilization of different techniques to infer the structure of M. tuberculosis PDT (MtbPDT) in solution. Small angle X-ray scattering and ultracentrifugation analysis showed that the protein oligomeric state is a tetramer and MtbPDT is a flat disk protein. Bioinformatics tools were used to infer the structure of MtbPDT. A molecular model for MtbPDT is presented and molecular dynamics simulations indicate that MtbPDT is stable. Experimental and molecular modeling results were in agreement and provide evidence for a tetrameric state of MtbPDT in solution.

Isotopic Criteria in the Characterization of Aromatic Molecules. 1. Hydrogen Affiliation in Natural Benzenoid/Phenylpropanoid Molecules

The site-specific isotope ratios of several families of aromatic molecules are analyzed in terms of hydrogen affiliation and discriminating potential. Among the aromatic molecules produced by plants, many are biosynthesized by the shikimate pathway, but the terpenic pathway also forms some compounds with a benzenic ring. In compounds of the phenylpropanoid family, specific hydrogen connections are determined with cinnamic acid, a key intermediate in the formation of a large number of aromatic molecules. Then affiliations through the phenylalanine precursor, back to the parent d-erythrose 4-phosphate and phosphoenolpyruvate molecules and finally to glucose, are considered. Typical isotopic profiles of the benzenic ring in natural, as compared to non-natural, molecules are defined. The dispersion observed in the (D/H)i ratios of the lateral chains is illustrative of diverse mechanistic responses and the role of exchange phenomena. The isotopic patterns of aromatic molecules pertaining to the terpenic family are drastically different from those of the shikimate descendants, and they exhibit much less variability. They enable the stereochemical affiliation of individual hydrogen atoms to be traced back first to the parent atoms in the common intermediate, geranyl diphosphate, then to the glyceraldehyde 3-phosphate and pyruvate couple involved in the DOXP pathway, and ultimately to the glucose precursor. The results illustrate the aptitude of the site-specific isotope ratios not only to authenticate natural with respect to chemical molecules but also to characterize different metabolic pathways and to reveal differences associated with the nature of the plant precursor.

Biochemical Characterization and Crystal Structure of Synechocystis Arogenate Dehydrogenase Provide Insights into Catalytic Reaction

The extreme diversity in substrate specificity, and in the regulation mechanism of arogenate/prephenate dehydrogenase enzymes in nature, makes a comparative structural study of these enzymes of great interest. We report here on the biochemical and structural characterization of arogenate dehydrogenase from Synechocystis sp. (TyrAsy). This work paves the way for the understanding of the structural determinants leading to diversity in substrate specificity, and of the regulation mechanisms of arogenate/prephenate dehydrogenases. The overall structure of TyrAsy in complex with NADP was refined to 1.6 A. The asymmetric unit contains two TyrAsy homodimers, with each monomer consisting of a nucleotide binding N-terminal domain and a particularly unique alpha-helical C-terminal dimerization domain. The substrate arogenate was modeled into the active site. The model of the ternary complex enzyme-NADP-arogenate nicely reveals at the atomic level the concerted mechanism of the arogenate/prephenate dehydrogenase reaction.

Dietary amino acids influence plumage traits and immune responses of male house sparrows, Passer domesticus, but not as expected

Traits that influence social interactions (signals) are widely thought to be honest indicators of some underlying quality of their bearer. One hypothesis is that a signal and the bearer's immunocompetence are linked via a condition-dependent pathway. We tested this idea by measuring the effect of specific dietary components on the development of a melanin-based plumage signal and the production of antibodies in juvenile male house sparrows entering their first moult. We maintained sparrows on one of three artificial diets: high total protein, low total protein, or intermediate protein with the precursors of melanin, phenylalanine and tyrosine (PT), reduced by 50%. Diet did not affect survival or weight gain. Two aspects of male plumage differed between treatments; the white wing bar was significantly smaller in low-protein males, and the black bib feathers had significantly higher reflectance in PT-reduced males. PT reductions had no effect on bib size. PT-reduced birds also produced more antibodies to diphtheria and tetanus antigens than did other subjects. After repeating the experiment using a better control and a different diet formula to compensate for an energy imbalance resulting from reduced PT levels, PT reduction again produced lighter bib feathers, but the effect of diet on antibody production disappeared. We conclude that the amino acid precursors to melanin affect melanin synthesis if scarce in the diet, but in this case, do not affect size, the most conspicuously variable aspect of the signal. We found no evidence of a condition-dependent link between melanin synthesis and immunocompetence.

Synthesis of α,α‐Disubstituted Unnatural Amino Acid Derivatives Using the Aza[2,3]Wittig Sigmatropic Rearrangement.

AbstractFor Abstract see ChemInform Abstract in Full Text.

Synthesis of α,α-disubstituted unnatural amino acid derivatives using the aza-[2,3]-Wittig sigmatropic rearrangement

Aza-[2,3]-Wittig rearrangement precursors derived from alanine, valine, phenylalanine and phenylglycine were synthesised with diethylamide and methylester anion stabilising groups. In the amide series only the alanine derived precursor rearranged upon deprotonation with KH. In the ester series the alanine, valine and phenylalanine precursors rearranged successfully with KH. The phenylalanine ester precursor gave an unselective rearrangement whereas rearrangement of the alanine and valine ester precursors gave levels and sense of diastereoselectivity in line with our transition state model. The products are chiral α,α-disubstituted-α-amino acid derivatives possessing two adjacent stereocentres and a vinyl silane synthetic handle.

Biosynthesis, molecular structure, and domain architecture of potato suberin: a (13)C NMR study using isotopically labeled precursors.

Although suberin in potato wound periderm is known to be a polyester containing long-chain fatty acids and phenolics embedded within the cell wall, many aspects of its molecular structure and polymer-polymer connectivities remain elusive. The present work combines biosynthetic incorporation of site-specifically (13)C-enriched acetates and phenylalanines with one- and two-dimensional solid-state (13)C NMR spectroscopic methods to monitor the developing suberin polymer. Exogenous acetate is found to be incorporated preferentially at the carboxyl end of the aliphatic carbon chains, suggesting addition during the later elongation steps of fatty acid synthesis. Carboxyl-labeled phenylalanine precursors provide evidence for the concurrent development of phenolic esters and of monolignols typical of lignin. Experiments with ring-labeled phenylalanine precursors demonstrate a predominance of sinapyl and guaiacyl structures among suberin's phenolic moieties. Finally, the analysis of spin-exchange (solid-state NOESY) NMR experiments in ring-labeled suberin indicates distances of no more than 0.5 nm between pairs of phenolic and oxymethine carbons, which are attributed to the aromatic-aliphatic polyester and the cell wall polysaccharide matrix, respectively. These results offer direct and detailed molecular information regarding the insoluble intermediates of suberin biosynthesis, indicate probable covalent linkages between moieties of its polyester and polysaccharide domains, and yield a clearer overall picture of this agriculturally important protective material.

Carbon-14 biolabeling of (+)-catechin and proanthocyanidin oligomers in willow tree cuttings.

Proanthocyanidin polymers, oligomers, and the structurally related monomer (+)-catechin were labeled by incorporation of radioactive precursors in shoots of willow tree (Salix caprea L.). [1-(14)C]-Acetate and [U-(14)C]-phenylalanine precursors were fed through the cut stems or petioles of leaves. Optimization of several parameters such as the nature and origin of the plant material, leaf maturity, nature, and quantity of radioactive precursor applied and the duration of metabolism led to incorporation yields of 3.2% and to specific activities of 500 microCi/g. Detailed characterization of the products (polymerization degree, procyanidin/prodelphinidin ratio, specific activities) and purification by chromatography are reported. Some sugars bound to radiolabeled proanthocyanidin polymers were removed by enzymic treatment with a mixture of glycosidases. A radioactive purity close to 100% and specific activities suitable for bioavailability studies were obtained.

Cloning and characterization of an esophageal-gland-specific chorismate mutase from the phytoparasitic nematode Meloidogyne javanica.

Root-knot nematodes are obligate plant parasites that alter plant cell growth and development by inducing the formation of giant feeder cells. It is thought that nematodes inject secretions from their esophageal glands into plant cells while feeding, and that these secretions cause giant cell formation. To elucidate the mechanisms underlying the formation of giant cells, a strategy was developed to clone esophageal gland genes from the root-knot nematode Meloidogyne javanica. One clone, shown to be expressed in the nematode's esophageal gland, codes for a potentially secreted chorismate mutase (CM). CM is a key branch-point regulatory enzyme in the shikimate pathway and converts chorismate to prephenate, a precursor of phenylalanine and tyrosine. The shikimate pathway is not found in animals, but in plants, where it produces aromatic amino acids and derivative compounds that play critical roles in growth and defense. Therefore, we hypothesize that this CM is involved in allowing nematodes to parasitize plants.

ChemInform Abstract: Concise Synthesis of Arogenate. A Biosynthetic Precursor of Phenylalanine and Tyrosine.

AbstractChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 100 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a “Full Text” option. The original article is trackable via the “References” option.

Pico‐tag analysis of arogenic acid and related free amino acids from plant and fungal extracts

AbstractFree amino acids from various plant tissues and fungal mycelia were extracted with buffered ethanol (75%, v/v) in 0.5% 2‐amino‐2‐methyl‐1‐propanol‐hydrochloride pH 11.0. Following removal of proteins and other high molecular weight components by ultrafiltration, free amino acids in the extracts were dried, derivatized with phenylisothiocyanate and analysed by high performance liquid chromatography using a Waters Pico‐Tag column. Arogenic acid, a non‐aromatic amino acid precursor of phenylalanine and tyrosine, was not detected although both aromatic amino acids were quantitated in all plant and fungal extracts. Nevertheless, arogenic acid in the mycelium of a triply‐blocked mutant of Neurospora crassa was readily detected. It is proposed that the absence of arogenic acid in plant extracts suggests that this intermediate does not accumulate in the metabolic pool thereby negating a possible regulatory role during aromatic amino acid biosynthesis.

Concise synthesis of arogenate. A biosynthetic precursor of phenylalanine and tyrosine

A concise new route to arogenate [L-(8S)-β-(1-carboxy-4-hydroxy-2,5-cyclohexadien-1-yl)alanine diammonium salt]1 in which Michael addition of the anion derived from methyl 1,4-dihydrobenzoate 2 to the dehydroalanine derivative 3 is the key C–C bond-forming step is described.

Effect of the immediate precursors of phenylalanine and tyrosine on growth and protein synthesis in phenylalanine- and tyrosine-deprived HeLa cells

Although Phe is an essential amino acid in mammalian cells, its immediate precursor, β-phenylpyruvic acid (BPP), when present in Phe-deficient medium at 10 −4 and 10 −3 M is converted at a sufficient rate to Phe to sustain growth at 60 and 100% of non-deficient control HeLa S-3 cells, respectively. In contrast, Tyr-deficient cells were unable to convert the immediate precursor of Tyr, OH-β-phenylpyruvic acid (OHBPP), nor could BPP rescue Tyr-deficient cells. The results are considered in terms of the organization of intracellular pathways by which precursors are transaminated and made available for protein synthesis.

Chiral 2-acetamidoacrylates in conjugate addition – asymmetric enolate trapping reactions. Asymmetric synthesis of phenylalanine

A new route to the asymmetric synthesis of α-amino acids based on the reactivity of chiral 2-acetamidoacrylates with nucleophiles through a conjugate addition followed by diastereoselective protonation of the enolate is described. Phenylalanine precursors are obtained in excellent chemical yields (80–95%) with moderate diastereomeric excess (0–44%) through the reaction of chiral 2-acetamidoacrylates with phenylmagnesium bromide in the presence of CuI followed by diastereoselective enolate protonation.