scholarly journals Isotopic Tracers Unveil Distinct Fates for Nitrogen Sources during Wine Fermentation with Two Non-Saccharomyces Strains

2020 ◽  
Vol 8 (6) ◽  
pp. 904
Author(s):  
Ying Su ◽  
Pauline Seguinot ◽  
Audrey Bloem ◽  
Anne Ortiz-Julien ◽  
José María Heras ◽  
...  
Keyword(s):  
Amino Acids ◽  
De Novo ◽  
Nitrogen Sources ◽  
Full Potential ◽  
Ehrlich Pathway ◽  
Comprehensive Overview ◽  
Isotopic Tracers ◽  
Yeast Strains ◽  
Nitrogen Redistribution ◽  
Sequential Fermentation

Non-Saccharomyces yeast strains have become increasingly prevalent in the food industry, particularly in winemaking, because of their properties of interest both in biological control and in complexifying flavour profiles in end-products. However, unleashing the full potential of these species would require solid knowledge of their physiology and metabolism, which is, however, very limited to date. In this study, a quantitative analysis using 15N-labelled NH4Cl, arginine, and glutamine, and 13C-labelled leucine and valine revealed the specificities of the nitrogen metabolism pattern of two non-Saccharomyces species, Torulaspora delbrueckii and Metschnikowia pulcherrima. In T. delbrueckii, consumed nitrogen sources were mainly directed towards the de novo synthesis of proteinogenic amino acids, at the expense of volatile compounds production. This redistribution pattern was in line with the high biomass-producer phenotype of this species. Conversely, in M. pulcherrima, which displayed weaker growth capacities, a larger proportion of consumed amino acids was catabolised for the production of higher alcohols through the Ehrlich pathway. Overall, this comprehensive overview of nitrogen redistribution in T. delbrueckii and M. pulcherrima provides valuable information for a better management of co- or sequential fermentation combining these species with Saccharomyces cerevisiae.

scholarly journals Inhibiting Methanogenesis Stimulated de novo Synthesis of Microbial Amino Acids in Mixed Rumen Batch Cultures Growing on Starch but not on Cellulose

2020 ◽  
Vol 8 (6) ◽  
pp. 799
Author(s):  
Emilio M. Ungerfeld ◽  
M. Fernanda Aedo ◽  
Camila Muñoz ◽  
Natalie L. Urrutia ◽  
Emilio D. Martínez ◽  
...  
Keyword(s):  
Amino Acids ◽  
De Novo ◽  
Nitrogen Sources ◽  
Environmental Benefits ◽  
De Novo Synthesis ◽  
Protein Nitrogen ◽  
Protein Supplements ◽  
Batch Cultures ◽  
Sufficient Energy ◽  
Ruminant Diets

Ameliorating methane (CH4) emissions from ruminants would have environmental benefits, but it is necessary to redirect metabolic hydrogen ([H]) toward useful sinks to also benefit animal productivity. We hypothesized that inhibiting rumen methanogenesis would increase de novo synthesis of microbial amino acids (AA) as an alternative [H] sink if sufficient energy and carbon are provided. We examined the effects of inhibiting methanogenesis with 9, 10-anthraquione (AQ) on mixed rumen batch cultures growing on cellulose or starch as sources of energy and carbon contrasting in fermentability, with ammonium (NH4+) or trypticase (Try) as nitrogen (N) sources. Inhibiting methanogenesis with AQ inhibited digestion with cellulose but not with starch, and decreased propionate and increased butyrate molar percentages with both substrates. Inhibiting methanogenesis with 9, 10-anthraquinone increased de novo synthesis of microbial AA with starch but not with cellulose. The decrease in the recovery of [H] caused by the inhibition of methanogenesis was more moderate with starch due to an enhancement of butyrate and AA as [H] sinks. There may be an opportunity to simultaneously decrease the emissions of CH4 and N with some ruminant diets and replace plant protein supplements with less expensive non-protein nitrogen sources such as urea.

scholarly journals Management of Multiple Nitrogen Sources during Wine Fermentation by Saccharomyces cerevisiae

2017 ◽  
Vol 83 (5) ◽  
Author(s):  
Lucie Crépin ◽  
Nhat My Truong ◽  
Audrey Bloem ◽  
Isabelle Sanchez ◽  
Sylvie Dequin ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Amino Acids ◽  
Fermentation Process ◽  
De Novo ◽  
Nitrogen Sources ◽  
Wine Industry ◽  
Sensory Profile ◽  
Minor Role ◽  
Available Nitrogen ◽  
Intracellular Fate

ABSTRACT During fermentative growth in natural and industrial environments, Saccharomyces cerevisiae must redistribute the available nitrogen from multiple exogenous sources to amino acids in order to suitably fulfill anabolic requirements. To exhaustively explore the management of this complex resource, we developed an advanced strategy based on the reconciliation of data from a set of stable isotope tracer experiments with labeled nitrogen sources. Thus, quantifying the partitioning of the N compounds through the metabolism network during fermentation, we demonstrated that, contrary to the generally accepted view, only a limited fraction of most of the consumed amino acids is directly incorporated into proteins. Moreover, substantial catabolism of these molecules allows for efficient redistribution of nitrogen, supporting the operative de novo synthesis of proteinogenic amino acids. In contrast, catabolism of consumed amino acids plays a minor role in the formation of volatile compounds. Another important feature is that the α-keto acid precursors required for the de novo syntheses originate mainly from the catabolism of sugars, with a limited contribution from the anabolism of consumed amino acids. This work provides a comprehensive view of the intracellular fate of consumed nitrogen sources and the metabolic origin of proteinogenic amino acids, highlighting a strategy of distribution of metabolic fluxes implemented by yeast as a means of adapting to environments with changing and scarce nitrogen resources. IMPORTANCE A current challenge for the wine industry, in view of the extensive competition in the worldwide market, is to meet consumer expectations regarding the sensory profile of the product while ensuring an efficient fermentation process. Understanding the intracellular fate of the nitrogen sources available in grape juice is essential to the achievement of these objectives, since nitrogen utilization affects both the fermentative activity of yeasts and the formation of flavor compounds. However, little is known about how the metabolism operates when nitrogen is provided as a composite mixture, as in grape must. Here we quantitatively describe the distribution through the yeast metabolic network of the N moieties and C backbones of these nitrogen sources. Knowledge about the management of a complex resource, which is devoted to improvement of the use of the scarce N nutrient for growth, will be useful for better control of the fermentation process and the sensory quality of wines.

A Plug-and-Play Approach for the De Novo Generation of Dually Functionalised Bispecifics

2019 ◽  
Author(s):  
Antoine Maruani ◽  
Peter A. Szijj ◽  
Calise Bahou ◽  
João C. F. Nogueira ◽  
Stephen Caddick ◽  
...  
Keyword(s):  
De Novo ◽  
Therapeutic Index ◽  
Antibody Fragments ◽  
Bispecific Antibodies ◽  
Full Potential ◽  
Mechanisms Of Resistance ◽  
Large Excess ◽  
Chemical Methods ◽  
New Class ◽  
Novel Approach

<p>Diseases are multifactorial, with redundancies and synergies between various pathways. However, most of the antibody-based therapeutics in clinical trials and on the market interact with only one target thus limiting their efficacy. The targeting of multiple epitopes could improve the therapeutic index of treatment and counteract mechanisms of resistance. To this effect, a new class of therapeutics emerged: bispecific antibodies.</p><p>Bispecific formation using chemical methods is rare and low yielding and/or requires a large excess of one of the two proteins to avoid homodimerisation. In order for chemically prepared bispecifics to deliver their full potential, high-yielding, modular and reliable cross-linking technologies are required. Herein, we describe a novel approach not only for the rapid and high-yielding chemical generation of bispecific antibodies from native antibody fragments, but also for the site-specific dual functionalisation of the resulting bioconjugates. Based on orthogonal clickable functional groups, this strategy enables the assembly of functionalised bispecifics with controlled loading in a modular and convergent manner.</p>

scholarly journals An enzymatic activation of formaldehyde for nucleotide methylation

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Charles Bou-Nader ◽  
Frederick W. Stull ◽  
Ludovic Pecqueur ◽  
Philippe Simon ◽  
Vincent Guérineau ◽  
...  
Keyword(s):  
Amino Acids ◽  
Thymidylate Synthase ◽  
Active Site ◽  
De Novo ◽  
Crystallographic Structure ◽  
De Novo Synthesis ◽  
Active Site Residues ◽  
Enzymatic Activation ◽  
Enzyme Cofactors ◽  
Unique Ability

AbstractFolate enzyme cofactors and their derivatives have the unique ability to provide a single carbon unit at different oxidation levels for the de novo synthesis of amino-acids, purines, or thymidylate, an essential DNA nucleotide. How these cofactors mediate methylene transfer is not fully settled yet, particularly with regard to how the methylene is transferred to the methylene acceptor. Here, we uncovered that the bacterial thymidylate synthase ThyX, which relies on both folate and flavin for activity, can also use a formaldehyde-shunt to directly synthesize thymidylate. Combining biochemical, spectroscopic and anaerobic crystallographic analyses, we showed that formaldehyde reacts with the reduced flavin coenzyme to form a carbinolamine intermediate used by ThyX for dUMP methylation. The crystallographic structure of this intermediate reveals how ThyX activates formaldehyde and uses it, with the assistance of active site residues, to methylate dUMP. Our results reveal that carbinolamine species promote methylene transfer and suggest that the use of a CH2O-shunt may be relevant in several other important folate-dependent reactions.

scholarly journals Intracellular Fate of Universally Labelled 13C Isotopic Tracers of Glucose and Xylose in Central Metabolic Pathways of Xanthomonas oryzae

Metabolites ◽  
2018 ◽  
Vol 8 (4) ◽  
pp. 66 ◽  
Author(s):  
Manu Shree ◽  
Shyam K. Masakapalli
Keyword(s):  
Amino Acids ◽  
Metabolic Pathways ◽  
Metabolic Networks ◽  
Tca Cycle ◽  
Xanthomonas Oryzae ◽  
Flux Analysis ◽  
Isotopic Tracers ◽  
Feeding Experiments ◽  
The Tca Cycle ◽  
Metabolic Route

The goal of this study is to map the metabolic pathways of poorly understood bacterial phytopathogen, Xanthomonas oryzae (Xoo) BXO43 fed with plant mimicking media XOM2 containing glutamate, methionine and either 40% [13C5] xylose or 40% [13C6] glucose. The metabolic networks mapped using the KEGG mapper and the mass isotopomer fragments of proteinogenic amino acids derived from GC-MS provided insights into the activities of Xoo central metabolic pathways. The average 13C in histidine, aspartate and other amino acids confirmed the activities of PPP, the TCA cycle and amino acid biosynthetic routes, respectively. The similar labelling patterns of amino acids (His, Ala, Ser, Val and Gly) from glucose and xylose feeding experiments suggests that PPP would be the main metabolic route in Xoo. Owing to the lack of annotated gene phosphoglucoisomerase in BXO43, the 13C incorporation in alanine could not be attributed to the competing pathways and hence warrants additional positional labelling experiments. The negligible presence of 13C incorporation in methionine brings into question its potential role in metabolism and pathogenicity. The extent of the average 13C labelling in several amino acids highlighted the contribution of pre-existing pools that need to be accounted for in 13C-flux analysis studies. This study provided the first qualitative insights into central carbon metabolic pathway activities in Xoo.

scholarly journals Metabolic and endocrine changes induced by chronic heatexposure in broiler chickens : biological and endocrinological variables

1996 ◽  
Vol 75 (2) ◽  
pp. 205-216 ◽  
Author(s):  
P. A. Geraert ◽  
J. C. F. Padilha ◽  
S. Guillaumin
Keyword(s):  
Amino Acids ◽  
Ambient Temperature ◽  
Broiler Chickens ◽  
De Novo ◽  
Heat Exposure ◽  
Plasma Corticosterone ◽  
Plasma Metabolites ◽  
Exogenous Insulin ◽  
Plasma Insulin Concentration ◽  
Esterified Fatty Acids

Abstract:The present study was designed to investigate the effect of chronic heat exposure (32° constant) on plasma metabolites and hormone concentrations in broiler chickens. At 2 and 4 weeks of age, fifty-four male Shaver broiler chickens were allocated to one of three treatments: 22°ad lib. feeding (22AL), 32°ad lib. feeding (32AL) and 22°,pair-feeding with the 32AL group (22PF). Ambient temperature was kept constant at either 22 or 32° for 2 weeks. Plasma glucose, triacylglycerols, phospholipids, non-esterified fatty acids (NEFA), individual amino acids, uric acid, insulin, triiodothyronine (T3), thyroxine, corticosterone were determined. Sensitivity to exogenous insulin was also measured at 7 weeks of age. At 4 and 6 weeks of age, i.e. after 2 weeks at high ambient temperature, fasted 32AL chickens displayed similar concentrations of glucose and triacylglycerols to those of 22AL birds. When fed, 32AL chickens exhibited higher plasma levels of glucose and decreased concentrations of NEFAand amino acids. Feed restriction resulted in intermediate values. Concentrations of all plasmafree amino acids were decreased under heat exposure except for aspartic acid, glutamic acid andphenylalanine. At 6 weeks of age, plasma T3 was reduced irrespective of the nutritional state, while plasma corticosterone concentrations were increased in 32AL birds compared with 22AL birds. Heat exposure did not change plasma insulin concentration in either fasted or fed chickens. The 32AL chickens displayed significantly reduced sensitivity to exogenous insulin when fasted,but an enhanced response to insulin when fed, compared with both 22° groups. Such endocrinological changes could stimulate lipid accumulation through increasedde novolipogenesis, reduced lipolysis and enhanced amino acid catabolism under chronic heat exposure.

scholarly journals A role for BDNF- and NMDAR-induced lysosomal recruitment of mTORC1 in the regulation of neuronal mTORC1 activity

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Dany Khamsing ◽  
Solène Lebrun ◽  
Isabelle Fanget ◽  
Nathanaël Larochette ◽  
Christophe Tourain ◽  
...  
Keyword(s):  
Amino Acids ◽  
Nmda Receptors ◽  
Hippocampal Neurons ◽  
De Novo ◽  
Long Term Potentiation ◽  
Activation Mechanism ◽  
Functional Link ◽  
Term Potentiation ◽  
Endocytic Compartments ◽  
Mtorc1 Activation

AbstractMemory and long term potentiation require de novo protein synthesis. A key regulator of this process is mTORC1, a complex comprising the mTOR kinase. Growth factors activate mTORC1 via a pathway involving PI3-kinase, Akt, the TSC complex and the GTPase Rheb. In non-neuronal cells, translocation of mTORC1 to late endocytic compartments (LEs), where Rheb is enriched, is triggered by amino acids. However, the regulation of mTORC1 in neurons remains unclear. In mouse hippocampal neurons, we observed that BDNF and treatments activating NMDA receptors trigger a robust increase in mTORC1 activity. NMDA receptors activation induced a significant recruitment of mTOR onto lysosomes even in the absence of external amino acids, whereas mTORC1 was evenly distributed in neurons under resting conditions. NMDA receptor-induced mTOR translocation to LEs was partly dependent on the BDNF receptor TrkB, suggesting that BDNF contributes to the effect of NMDA receptors on mTORC1 translocation. In addition, the combination of Rheb overexpression and artificial mTORC1 targeting to LEs by means of a modified component of mTORC1 fused with a LE-targeting motif strongly activated mTOR. To gain spatial and temporal control over mTOR localization, we designed an optogenetic module based on light-sensitive dimerizers able to recruit mTOR on LEs. In cells expressing this optogenetic tool, mTOR was translocated to LEs upon photoactivation. In the absence of growth factor, this was not sufficient to activate mTORC1. In contrast, mTORC1 was potently activated by a combination of BDNF and photoactivation. The data demonstrate that two important triggers of synaptic plasticity, BDNF and NMDA receptors, synergistically power the two arms of the mTORC1 activation mechanism, i.e., mTORC1 translocation to LEs and Rheb activation. Moreover, they unmask a functional link between NMDA receptors and mTORC1 that could underlie the changes in the synaptic proteome associated with long-lasting changes in synaptic strength.

scholarly journals Opportunistic feeding on various organic food sources by the cold-water coral <i>Lophelia pertusa</i>

2014 ◽  
Vol 11 (1) ◽  
pp. 123-133 ◽  
Author(s):  
C. E. Mueller ◽  
A. I. Larsson ◽  
B. Veuger ◽  
J. J. Middelburg ◽  
D. van Oevelen
Keyword(s):  
Amino Acids ◽  
Fatty Acids ◽  
De Novo ◽  
Cold Water ◽  
Feeding Strategy ◽  
Food Sources ◽  
Coral Tissue ◽  
Lophelia Pertusa ◽  
Opportunistic Feeding ◽  
Water Coral

Abstract. The ability of the cold-water coral Lophelia pertusa to exploit different food sources was investigated under standardized conditions in a flume. The tested food sources, dissolved organic matter (DOM, added as dissolved free amino acids), bacteria, algae, and zooplankton (Artemia) were deliberately enriched in 13C and 15N. The incorporation of 13C and 15N was traced into bulk tissue, fatty acids, hydrolysable amino acids, and the skeleton (13C only) of L. pertusa. Incorporation rates of carbon (ranging from 0.8–2.4 μg C g−1 DW d–1) and nitrogen (0.2–0.8 μg N g−1 DW d–1) into coral tissue did not differ significantly among food sources indicating an opportunistic feeding strategy. Although total food assimilation was comparable among sources, subsequent food processing was dependent on the type of food source ingested and recovery of assimilated C in tissue compounds ranged from 17% (algae) to 35% (Artemia). De novo synthesis of individual fatty acids by L. pertusa occurred in all treatments as indicated by the 13C enrichment of individual phospholipid-derived fatty acids (PLFAs) in the coral that were absent in the added food sources. This indicates that the coral might be less dependent on its diet as a source of specific fatty acids than expected, with direct consequences for the interpretation of in situ observations on coral nutrition based on lipid profiles.

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