Surfactant to dye binding degree based approach for the selective determination of L-glutamate in foodstuffs.
Pedraza, A., Sicilia, M. D., Rubio, S. & Pérez-Bendito, D. (2007). Analytical and Bioanalytical Chemistry, 389(7-8), 2297-2302.
A selective method for the determination of L-glutamate in foodstuffs has been developed. It was based on the competition established between the analyte and the dye Coomassie brilliant blue G (CBBG) to interact with the surfactant didodecyldimethylammonium bromide (DDABr). The measurement parameter was the amount of DDABr required to reach a given dye-to-surfactant binding degree. It was obtained by photometric titration on the basis of the changes observed in the spectral characteristics of the dye when CBBG–DDABr aggregates were formed. The calibration graph obtained was linear in the L-glutamate concentration interval 0.2–5 mM (detection limit 0.05 mM). The high selectivity of the proposed method (other amino acids and food additives did not interfere at the concentrations present in foodstuffs) permitted the direct analysis of food samples after dissolution of the analyte in hot water. The accuracy of the surfactant to the dye binding degree method was demonstrated by determining L-glutamate in different kinds of foodstuffs (liquid and dried soups, seasonings, pasta sauces and dried mushroom creams) and comparing the results obtained with those provided by the commercial Boehringer Mannheim essay.
The rice R2R3-MYB transcription factor OsMYB55 is involved in the tolerance to high temperature and modulates amino acid metabolism.
El-kereamy, A., Bi, Y. M., Ranathunge, K., Beatty, P. H., Good, A. G. & Rothstein, S. J. (2012). PloS one, 7(12), e52030.
Temperatures higher than the optimum negatively affects plant growth and development. Tolerance to high temperature is a complex process that involves several pathways. Understanding this process, especially in crops such as rice, is essential to prepare for predicted climate changes due to global warming. Here, we show that OsMYB55 is induced by high temperature and overexpression of OsMYB55 resulted in improved plant growth under high temperature and decreased the negative effect of high temperature on grain yield. Transcriptome analysis revealed an increase in expression of several genes involved in amino acids metabolism. We demonstrate that OsMYB55 binds to the promoter regions of target genes and directly activates expression of some of those genes including glutamine synthetase (OsGS1;2) glutamine amidotransferase (GAT1) and glutamate decarboxylase 3 (GAD3). OsMYB55 overexpression resulted in an increase in total amino acid content and of the individual amino acids produced by the activation of the above mentioned genes and known for their roles in stress tolerance, namely L-glutamic acid, GABA and arginine especially under high temperature condition. In conclusion, overexpression of OsMYB55 improves rice plant tolerance to high temperature, and this high tolerance is associated with enhanced amino acid metabolism through transcription activation.
Astrocytic glutamate transporter-dependent neuroprotection against glutamate toxicity: An in vitro study of maslinic acid.
Qian, Y., Guan, T., Tang, X., Huang, L., Huang, M., Li, Y., Sun, H., Yu, R. & Zhang, F. (2011). European Journal of Pharmacology, 651(1-3), 59-65.
The astrocytic glutamate transporters GLAST/EAAT1 and GLT-1/EAAT2 are crucial for the removal of glutamate from the synaptic cleft and are essential for maintaining a low concentration of extracellular glutamate in the brain. Enhanced transporter expression is neuroprotective. In the present study, we tested the neuropotective effects of maslinic acid, a natural product from the Olea europaea plant, on cultures of primary neurons from the cerebral cortex. Studies showed that astrocyte-conditioned medium from maslinic acid-treated astrocytes dose-dependently promoted neuron survival during glutamate toxicity by enhancing extracellular glutamate clearance. Real-time PCR and western blot analysis revealed that maslinic acid pre-treatment significantly increased the expression of GLAST and GLT-1 at the protein and mRNA levels. In addition, this neuroprotection was abolished by the glutamate transporter inhibitor, L-Threohydroxy aspartate (THA), in a co-culture of astrocytes and neurons. These findings suggest that maslinic acid regulates the extracellular glutamate concentration by increasing the expression of astrocytic glutamate transporters, which may, in turn, provide neuroprotection.
Production of stable quinine nanodispersions using esterified γ-polyglutamic acid biopolymer.
Hoennscheidt, C., Kreyenschulte, D., Margaritis, A. & Krull, R. (2013). Biochemical Engineering Journal, 79, 259-266.
Novel methods are needed for the development of nanodispersed drug formulations to enhance bioavailability of many hydrophobic pharmaceuticals. The poorly water-soluble quinine is a well-known anti-malaria drug which can be used as a promising model compound for the development of novel nanodispersed formulations. In addition to hydrophobic drug's own affecting properties, surfactants play an important role for the enhancement of their low bioavailability by preparing stable dispersions. Amphiphilic compounds can efficiently be used to stabilize colloidal fragments by preventing the precipitation or crystallization of poorly water-soluble active ingredients during fabrication. A novel biopolymer derivative based on the biotechnologically produced γ-polyglutamic acid (γ-PGA) from Bacillus licheniformis cultivation was developed for encapsulation of the active ingredient. High-molecular γ-PGA is an anionic polyelectrolyte that was optimized and modified with hydrophobic L-phenylalanine ethyl ester (L-PAE) to form an amphiphilic comb polymer P(γ-GA-r-L-PAE) with surfactive properties. The approach of the nanodispersion polymer concentration, molecular weight and grafting degree enables the efficient stabilization of the poorly water-soluble model drug. The research presented in this report indicates the potential benefits of hydrophobically modified γ-PGA and suggests its potential role in forming stable dispersions for future pharmaceutical applications.
Gamma-amino butyric acid, glutamate dehydrogenase and glutamate decarboxylase levels in phylogenetically divergent plants.
Seher, Y., Filiz, O. & Melike, B. (2013). Plant Systematics and Evolution, 299(2), 403-412.
Gamma-amino butyric acid (GABA) is a nonprotein amino acid found in a wide range of organisms including plants. Several studies have shown that GABA plays different roles in plant metabolism including carbon–nitrogen metabolism, energy balance, signaling and development. It has been suggested that the occurrence of GABA and the enzymes related to GABA biosynthesis in prokaryotes and eukaryotes may be important in evolution and diversification. However, studies of GABA biosynthesis and GABA levels in an evolutionary context are restricted to sequenced plant genomes. In this study we aimed to compare the activities of GDH and GAD enzymes and total nitrogen, and the contents of total soluble protein, succinate, glutamate, proline and GABA in plants from different phylogenetic levels including Ulva lactuca, Pseudevernia furfuracea, Nephrolepsis exaltata, Ginkgo biloba, Pinus pinea, Magnolia grandiflora, Nymphaea alba, Urtica dioica, Portulaca oleraceae, Malva sylvestris, Rosa canina, Lavandula stoechas, Washingtonia filifera, Avena barbata and Iris kaempferi. The activities of GAD and GDH enzymes differed according to the species and were not always parallel to GABA levels. The discrepancy in the contents of succinate and GABA between higher and primitive plants was also prominent. Glutamate levels were high with a few exceptions and proline contents were at similar low values as compared to other amino acids. Our results support the hypothesis that the GABA shunt plays a key role in carbon and nitrogen partitioning via linking amino acid metabolism and the tricarboxylic acid cycle which is essential for higher plant species.
Metabolic engineering of the mixed-acid fermentation pathway of Escherichia coli for anaerobic production of glutamate and itaconate.
Vuoristo, K. S., Mars, A. E., Sangra, J. V., Springer, J., Eggink, G., Sanders, J. P. & Weusthuis, R. A. (2015). AMB Express, 5(1), 61.
Itaconic acid, an unsaturated C5-dicarboxylic acid, is a biobased building block for the polymer industry. The purpose of this study was to establish proof of principle for an anaerobic fermentation process for the production of itaconic acid by modification of the mixed acid fermentation pathway of E. coli. E. coli BW25113 (DE3) and the phosphate acetyltransferase (pta) and lactate dehydrogenase (ldhA) deficient strain E. coli BW25113 (DE3) Δpta-ΔldhA were used to study anaerobic itaconate production in E. coli. Heterologous expression of the gene encoding cis-aconitate decarboxylase (cadA) from A. terreus in E. coli BW25113 (DE3) did not result in itaconate production under anaerobic conditions, but 0.08 mM of itaconate was formed when the genes encoding citrate synthase (gltA) and aconitase (acnA) from Corynebacterium glutamicum were also expressed. The same amount was produced when cadA was expressed in E. coli BW25113 (DE3) Δpta-ΔldhA. The titre increased 8 times to 0.66 mM (1.2 % Cmol) when E. coli BW25113 (DE3) Δpta-ΔldhA also expressed gltA and acnA. In addition, this strain produced 8.5 mM (13 % Cmol) of glutamate. The use of a nitrogen-limited growth medium reduced the accumulation of glutamate by nearly 50 % compared to the normal medium, and also resulted in a more than 3-fold increase of the itaconate titre to 2.9 mM. These results demonstrated that E. coli has potential to produce itaconate and glutamate under anaerobic conditions, closing the redox balance by co-production of succinate or ethanol with H2 and CO2.
Genetic, enzymatic and metabolite profiling of the Lactobacillus casei group reveals strain biodiversity and potential applications for flavour diversification.
Stefanovic, E., Kilcawley, K. N., Rea, M. C., Fitzgerald, G. F. & McAuliffe, O. (2017). Journal of Applied Microbiology, 122(5), 1245-1261.
Aims: The Lactobacillus casei group represents a widely explored group of lactic acid bacteria, characterized by a high level of biodiversity. In this study, the genetic and phenotypic diversity of a collection of more than 300 isolates of the Lact. casei group and their potential to produce volatile metabolites important for flavour development in dairy products, was examined. Methods and Results: Following confirmation of species by 16S rRNA PCR, the diversity of the isolates was determined by pulsed-field gel electrophoresis. The activities of enzymes involved in the proteolytic cascade were assessed and significant differences among the strains were observed. Ten strains were chosen based on the results of their enzymes activities and they were analysed for their ability to produce volatiles in media with increased concentrations of a representative aromatic, branched chain and sulphur amino acid. Volatiles were assessed using gas chromatography coupled with mass spectrometry. Strain-dependent differences in the range and type of volatiles produced were evident. Conclusions: Strains of the Lact. casei group are characterized by genetic and metabolic diversity which supports variability in volatile production. Significance and Impact of the Study: This study provides a screening approach for the knowledge-based selection of strains potentially enabling flavour diversification in fermented dairy products.
Development of a production chain from vegetable biowaste to platform chemicals.
Schmidt, A., Sturm, G., Lapp, C. J., Siebert, D., Saravia, F., Horn, H., Ravi, P. P. Lemmer, A. & Gescher, J. (2018). Microbial Cell Factories, 17(1), 90.
Background: A future bioeconomy relies on the development of technologies to convert waste into valuable compounds. We present here an attempt to design a biotechnological cascade for the conversion of vegetable waste into acetoin and electrical energy. Results: A vegetable waste dark fermentation effluent containing mainly acetate, butyrate and propionate was oxidized in a bioelectrochemical system. The achieved average current at a constant anode potential of 0 mV against standard hydrogen electrode was 177.5 ± 52.5 µA/cm2. During this step, acetate and butyrate were removed from the effluent while propionate was the major remaining component of the total organic carbon content comprising on average 75.6%. The key players with regard to carbon oxidation and electrode reduction were revealed using amplicon sequencing and metatranscriptomic analysis. Using nanofiltration, it was possible to concentrate the propionate in the effluent. The effluent was revealed to be a suitable medium for biotechnological production strains. As a proof of principle, the propionate in the effluent of the bioelectrochemical system was converted into the platform chemical acetoin with a carbon recovery of 86%. Conclusions: To the best of our knowledge this is the first report on a full biotechnological production chain leading from vegetable waste to the production of a single valuable platform chemical that integrates carbon elimination steps leading to the production of the valuable side product electrical energy.