Sample records for thiamin biosynthetic pathway
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Hsieh, Wei-Yu; Liao, Jo-Chien; Wang, Hsin-Tzu; Hung, Tzu-Huan; Tseng, Ching-Chih; Chung, Tsui-Yun; Hsieh, Ming-Hsiun
2017-07-01
Thiamin diphosphate (TPP, vitamin B 1 ) is an essential coenzyme present in all organisms. Animals obtain TPP from their diets, but plants synthesize TPPde novo. We isolated and characterized an Arabidopsis pale green1 (pale1) mutant that contained higher concentrations of thiamin monophosphate (TMP) and less thiamin and TPP than the wild type. Supplementation with thiamin, but not the thiazole and pyrimidine precursors, rescued the mutant phenotype, indicating that the pale1 mutant is a thiamin-deficient mutant. Map-based cloning and whole-genome sequencing revealed that the pale1 mutant has a mutation in At5g32470 encoding a TMP phosphatase of the TPP biosynthesis pathway. We further confirmed that the mutation of At5g32470 is responsible for the mutant phenotypes by complementing the pale1 mutant with constructs overexpressing full-length At5g32470. Most plant TPP biosynthetic enzymes are located in the chloroplasts and cytosol, but At5g32470-GFP localized to the mitochondrion of the root, hypocotyl, mesophyll and guard cells of the 35S:At5g32470-GFP complemented plants. The subcellular localization of a functional TMP phosphatase suggests that the complete vitamin B1 biosynthesis pathway may involve the chloroplasts, mitochondria and cytosol in plants. Analysis of PALE1 promoter-uidA activity revealed that PALE1 is mainly expressed in vascular tissues of Arabidopsis seedlings. Quantitative RT-PCR analysis of TPP biosynthesis genes and genes encoding the TPP-dependent enzymes pyruvate dehydrogenase, α-ketoglutarate dehydrogenase and transketolase revealed that the transcript levels of these genes were upregulated in the pale1 mutant. These results suggest that endogenous levels of TPP may affect the expression of genes involved in TPP biosynthesis and TPP-dependent enzymes. © 2017 The Authors The Plant Journal © 2017 John Wiley & Sons Ltd.
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Kamarudin, Amirah N.; Lai, Kok S.; Lamasudin, Dhilia U.; Idris, Abu S.; Balia Yusof, Zetty N.
2017-01-01
Thiamine, or vitamin B1 plays an indispensable role as a cofactor in crucial metabolic reactions including glycolysis, pentose phosphate pathway and the tricarboxylic acid cycle in all living organisms. Thiamine has been shown to play a role in plant adaptation toward biotic and abiotic stresses. The modulation of thiamine biosynthetic genes in oil palm seedlings was evaluated in response to root colonization by endophytic Hendersonia toruloidea. Seven-month-old oil palm seedlings were inoculated with H. toruloidea and microscopic analyses were performed to visualize the localization of endophytic H. toruloidea in oil palm roots. Transmission electron microscopy confirmed that H. toruloidea colonized cortical cells. The expression of thiamine biosynthetic genes and accumulation of total thiamine in oil palm seedlings were also evaluated. Quantitative real-time PCR was performed to measure transcript abundances of four key thiamine biosynthesis genes (THI4, THIC, TH1, and TPK) on days 1, 7, 15, and 30 in response to H. toruloidea colonization. The results showed an increase of up to 12-fold in the expression of all gene transcripts on day 1 post-inoculation. On days 7, 15, and 30 post-inoculation, the relative expression levels of these genes were shown to be downregulated. Thiamine accumulation was observed on day 7 post-colonization and subsequently decreased until day 30. This work provides the first evidence for the enhancement of thiamine biosynthesis by endophytic colonization in oil palm seedlings. PMID:29089959
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Vanillin biosynthetic pathways in plants.
Kundu, Anish
2017-06-01
The present review compiles the up-to-date knowledge on vanillin biosynthesis in plant systems to focus principally on the enzymatic reactions of in planta vanillin biosynthetic pathway and to find out its impact and prospect in future research in this field. Vanillin, a very popular flavouring compound, is widely used throughout the world. The principal natural resource of vanillin is the cured vanilla pods. Due to the high demand of vanillin as a flavouring agent, it is necessary to explore its biosynthetic enzymes and genes, so that improvement in its commercial production can be achieved through metabolic engineering. In spite of significant advancement in elucidating vanillin biosynthetic pathway in the last two decades, no conclusive demonstration had been reported yet for plant system. Several biosynthetic enzymes have been worked upon but divergences in published reports, particularly in characterizing the crucial biochemical steps of vanillin biosynthesis, such as side-chain shortening, methylation, and glucoside formation and have created a space for discussion. Recently, published reviews on vanillin biosynthesis have focused mainly on the biotechnological approaches and bioconversion in microbial systems. This review, however, aims to compile in brief the overall vanillin biosynthetic route and present a comparative as well as comprehensive description of enzymes involved in the pathway in Vanilla planifolia and other plants. Special emphasis has been given on the key enzymatic biochemical reactions that have been investigated extensively. Finally, the present standpoint and future prospects have been highlighted.
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Bioengineering natural product biosynthetic pathways for therapeutic applications.
Wu, Ming-Cheng; Law, Brian; Wilkinson, Barrie; Micklefield, Jason
2012-12-01
With the advent of next-generation DNA sequencing technologies, the number of microbial genome sequences has increased dramatically, revealing a vast array of new biosynthetic gene clusters. Genomics data provide a tremendous opportunity to discover new natural products, and also to guide the bioengineering of new and existing natural product scaffolds for therapeutic applications. Notably, it is apparent that the vast majority of biosynthetic gene clusters are either silent or produce very low quantities of the corresponding natural products. It is imperative therefore to devise methods for activating unproductive biosynthetic pathways to provide the quantities of natural products needed for further development. Moreover, on the basis of our expanding mechanistic and structural knowledge of biosynthetic assembly-line enzymes, new strategies for re-programming biosynthetic pathways have emerged, resulting in focused libraries of modified products with potentially improved biological properties. In this review we will focus on the latest bioengineering approaches that have been utilised to optimise yields and increase the structural diversity of natural product scaffolds for future clinical applications. Copyright © 2012 Elsevier Ltd. All rights reserved.
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Berrone, Elena; Beltramo, Elena; Solimine, Carmela; Ape, Alessandro Ubertalli; Porta, Massimo
2006-04-07
Hyperglycemia is a causal factor in the development of the vascular complications of diabetes. One of the biochemical mechanisms activated by excess glucose is the polyol pathway, the key enzyme of which, aldose reductase, transforms d-glucose into d-sorbitol, leading to imbalances of intracellular homeostasis. We aimed at verifying the effects of thiamine and benfotiamine on the polyol pathway, transketolase activity, and intracellular glucose in endothelial cells and pericytes under high ambient glucose. Human umbilical vein endothelial cells and bovine retinal pericytes were cultured in normal (5.6 mmol/liter) or high (28 mmol/liter) glucose, with or without thiamine or benfotiamine 50 or 100 mumol/liter. Transketolase and aldose reductase mRNA expression was determined by reverse transcription-PCR, and their activity was measured spectrophotometrically; sorbitol concentrations were quantified by gas chromatography-mass spectrometry and intracellular glucose concentrations by fluorescent enzyme-linked immunosorbent assay method. Thiamine and benfotiamine reduce aldose reductase mRNA expression, activity, sorbitol concentrations, and intracellular glucose while increasing the expression and activity of transketolase, for which it is a coenzyme, in human endothelial cells and bovine retinal pericytes cultured in high glucose. Thiamine and benfotiamine correct polyol pathway activation induced by high glucose in vascular cells. Activation of transketolase may shift excess glycolytic metabolites into the pentose phosphate cycle, accelerate the glycolytic flux, and reduce intracellular free glucose, thereby preventing its conversion to sorbitol. This effect on the polyol pathway, together with other beneficial effects reported for thiamine in high glucose, could justify testing thiamine as a potential approach to the prevention and/or treatment of diabetic complications.
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Biosynthetic Pathway and Metabolic Engineering of Plant Dihydrochalcones.
Ibdah, Mwafaq; Martens, Stefan; Gang, David R
2018-03-14
Dihydrochalcones are plant natural products containing the phenylpropanoid backbone and derived from the plant-specific phenylpropanoid pathway. Dihydrochalcone compounds are important in plant growth and response to stresses and, thus, can have large impacts on agricultural activity. In recent years, these compounds have also received increased attention from the biomedical community for their potential as anticancer treatments and other benefits for human health. However, they are typically produced at relatively low levels in plants. Therefore, an attractive alternative is to express the plant biosynthetic pathway genes in microbial hosts and to engineer the metabolic pathway/host to improve the production of these metabolites. In the present review, we discuss in detail the functions of genes and enzymes involved in the biosynthetic pathway of the dihydrochalcones and the recent strategies and achievements used in the reconstruction of multi-enzyme pathways in microorganisms in efforts to be able to attain higher amounts of desired dihydrochalcones.
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[Advance in flavonoids biosynthetic pathway and synthetic biology].
Zou, Li-Qiu; Wang, Cai-Xia; Kuang, Xue-Jun; Li, Ying; Sun, Chao
2016-11-01
Flavonoids are the valuable components in medicinal plants, which possess a variety of pharmacological activities, including anti-tumor, antioxidant and anti-inflammatory activities. There is an unambiguous understanding about flavonoids biosynthetic pathway, that is,2S-flavanones including naringenin and pinocembrin are the skeleton of other flavonoids and they can transform to other flavonoids through branched metabolic pathway. Elucidation of the flavonoids biosynthetic pathway lays a solid foundation for their synthetic biology. A few flavonoids have been produced in Escherichia coli or yeast with synthetic biological technologies, such as naringenin, pinocembrin and fisetin. Synthetic biology will provide a new way to get valuable flavonoids and promote the research and development of flavonoid drugs and health products, making flavonoids play more important roles in human diet and health. Copyright© by the Chinese Pharmaceutical Association.
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Dual biosynthetic pathways to phytosterol via cycloartenol and lanosterol in Arabidopsis.
Ohyama, Kiyoshi; Suzuki, Masashi; Kikuchi, Jun; Saito, Kazuki; Muranaka, Toshiya
2009-01-20
The differences between the biosynthesis of sterols in higher plants and yeast/mammals are believed to originate at the cyclization step of oxidosqualene, which is cyclized to cycloartenol in higher plants and lanosterol in yeast/mammals. Recently, lanosterol synthase genes were identified from dicotyledonous plant species including Arabidopsis, suggesting that higher plants possess dual biosynthetic pathways to phytosterols via lanosterol, and through cycloartenol. To identify the biosynthetic pathway to phytosterol via lanosterol, and to reveal the contributions to phytosterol biosynthesis via each cycloartenol and lanosterol, we performed feeding experiments by using [6-(13)C(2)H(3)]mevalonate with Arabidopsis seedlings. Applying (13)C-{(1)H}{(2)H} nuclear magnetic resonance (NMR) techniques, the elucidation of deuterium on C-19 behavior of phytosterol provided evidence that small amounts of phytosterol were biosynthesized via lanosterol. The levels of phytosterol increased on overexpression of LAS1, and phytosterols derived from lanosterol were not observed in a LAS1-knockout plant. This is direct evidence to indicate that the biosynthetic pathway for phytosterol via lanosterol exists in plant cells. We designate the biosynthetic pathway to phytosterols via lanosterol "the lanosterol pathway." LAS1 expression is reported to be induced by the application of jasmonate and is thought to have evolved from an ancestral cycloartenol synthase to a triterpenoid synthase, such as beta-amyrin synthase and lupeol synthase. Considering this background, the lanosterol pathway may contribute to the biosynthesis of not only phytosterols, but also steroids as secondary metabolites.
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Heterologous Expression of the Oxytetracycline Biosynthetic Pathway in Myxococcus xanthus▿
Stevens, D. Cole; Henry, Michael R.; Murphy, Kimberly A.; Boddy, Christopher N.
2010-01-01
New natural products for drug discovery may be accessed by heterologous expression of bacterial biosynthetic pathways in metagenomic DNA libraries. However, a “universal” host is needed for this experiment. Herein, we show that Myxococcus xanthus is a potential “universal” host for heterologous expression of polyketide biosynthetic gene clusters. PMID:20208031
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A nitrous acid biosynthetic pathway for diazo group formation in bacteria.
Sugai, Yoshinori; Katsuyama, Yohei; Ohnishi, Yasuo
2016-02-01
Although some diazo compounds have bioactivities of medicinal interest, little is known about diazo group formation in nature. Here we describe an unprecedented nitrous acid biosynthetic pathway responsible for the formation of a diazo group in the biosynthesis of the ortho-diazoquinone secondary metabolite cremeomycin in Streptomyces cremeus. This finding provides important insights into the biosynthetic pathways not only for diazo compounds but also for other naturally occurring compounds containing nitrogen-nitrogen bonds.
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Biosynthetic Pathways of Ergot Alkaloids
Directory of Open Access Journals (Sweden)
Nina Gerhards
2014-12-01
Full Text Available Ergot alkaloids are nitrogen-containing natural products belonging to indole alkaloids. The best known producers are fungi of the phylum Ascomycota, e.g., Claviceps, Epichloë, Penicillium and Aspergillus species. According to their structures, ergot alkaloids can be divided into three groups: clavines, lysergic acid amides and peptides (ergopeptines. All of them share the first biosynthetic steps, which lead to the formation of the tetracyclic ergoline ring system (except the simplest, tricyclic compound: chanoclavine. Different modifications on the ergoline ring by specific enzymes result in an abundance of bioactive natural products, which are used as pharmaceutical drugs or precursors thereof. From the 1950s through to recent years, most of the biosynthetic pathways have been elucidated. Gene clusters from several ergot alkaloid producers have been identified by genome mining and the functions of many of those genes have been demonstrated by knock-out experiments or biochemical investigations of the overproduced enzymes.
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Directory of Open Access Journals (Sweden)
Rapala-Kozik Maria
2012-01-01
Full Text Available Abstract Background Recent reports suggest that vitamin B1 (thiamine participates in the processes underlying plant adaptations to certain types of abiotic and biotic stress, mainly oxidative stress. Most of the genes coding for enzymes involved in thiamine biosynthesis in Arabidopsis thaliana have been identified. In our present study, we examined the expression of thiamine biosynthetic genes, of genes encoding thiamine diphosphate-dependent enzymes and the levels of thiamine compounds during the early (sensing and late (adaptation responses of Arabidopsis seedlings to oxidative, salinity and osmotic stress. The possible roles of plant hormones in the regulation of the thiamine contribution to stress responses were also explored. Results The expression of Arabidopsis genes involved in the thiamine diphosphate biosynthesis pathway, including that of THI1, THIC, TH1 and TPK, was analyzed for 48 h in seedlings subjected to NaCl or sorbitol treatment. These genes were found to be predominantly up-regulated in the early phase (2-6 h of the stress response. The changes in these gene transcript levels were further found to correlate with increases in thiamine and its diphosphate ester content in seedlings, as well as with the enhancement of gene expression for enzymes which require thiamine diphosphate as a cofactor, mainly α-ketoglutarate dehydrogenase, pyruvate dehydrogenase and transketolase. In the case of the phytohormones including the salicylic, jasmonic and abscisic acids which are known to be involved in plant stress responses, only abscisic acid was found to significantly influence the expression of thiamine biosynthetic genes, the thiamine diphosphate levels, as well as the expression of genes coding for main thiamine diphosphate-dependent enzymes. Using Arabidopsis mutant plants defective in abscisic acid production, we demonstrate that this phytohormone is important in the regulation of THI1 and THIC gene expression during salt stress
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Bettendorff, Lucien; Wins, Pierre
2009-06-01
Prokaryotes, yeasts and plants synthesize thiamin (vitamin B1) via complex pathways. Animal cells capture the vitamin through specific high-affinity transporters essential for internal thiamin homeostasis. Inside the cells, thiamin is phosphorylated to higher phosphate derivatives. Thiamin diphosphate (ThDP) is the best-known thiamin compound because of its role as an enzymatic cofactor. However, in addition to ThDP, at least three other thiamin phosphates occur naturally in most cells: thiamin monophosphate, thiamin triphosphate (ThTP) and the recently discovered adenosine thiamin triphosphate. It has been suggested that ThTP has a specific neurophysiological role, but recent data favor a much more basic metabolic function. During amino acid starvation, Escherichia coli accumulate ThTP, possibly acting as a signal involved in the adaptation of the bacteria to changing nutritional conditions. In animal cells, ThTP can phosphorylate some proteins, but the physiological significance of this mechanism remains unknown. Adenosine thiamin triphosphate, recently discovered in E. coli, accumulates during carbon starvation and might act as an alarmone. Among the proteins involved in thiamin metabolism, thiamin transporters, thiamin pyrophosphokinase and a soluble 25-kDa thiamin triphosphatase have been characterized at the molecular level, in contrast to thiamin mono- and diphosphatases whose specificities remain to be proven. A soluble enzyme catalyzing the synthesis of adenosine thiamin triphosphate from ThDP and ADP or ATP has been partially characterized in E. coli, but the mechanism of ThTP synthesis remains elusive. The data reviewed here illustrate the complexity of thiamin biochemistry, which is not restricted to the cofactor role of ThDP.
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A simple biosynthetic pathway for large product generation from small substrate amounts
Djordjevic, Marko; Djordjevic, Magdalena
2012-10-01
A recently emerging discipline of synthetic biology has the aim of constructing new biosynthetic pathways with useful biological functions. A major application of these pathways is generating a large amount of the desired product. However, toxicity due to the possible presence of toxic precursors is one of the main problems for such production. We consider here the problem of generating a large amount of product from a potentially toxic substrate. To address this, we propose a simple biosynthetic pathway, which can be induced in order to produce a large number of the product molecules, by keeping the substrate amount at low levels. Surprisingly, we show that the large product generation crucially depends on fast non-specific degradation of the substrate molecules. We derive an optimal induction strategy, which allows as much as three orders of magnitude increase in the product amount through biologically realistic parameter values. We point to a recently discovered bacterial immune system (CRISPR/Cas in E. coli) as a putative example of the pathway analysed here. We also argue that the scheme proposed here can be used not only as a stand-alone pathway, but also as a strategy to produce a large amount of the desired molecules with small perturbations of endogenous biosynthetic pathways.
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A simple biosynthetic pathway for large product generation from small substrate amounts
Energy Technology Data Exchange (ETDEWEB)
Djordjevic, Marko [Institute of Physiology and Biochemistry, Faculty of Biology, University of Belgrade (Serbia); Djordjevic, Magdalena [Institute of Physics Belgrade, University of Belgrade (Serbia)
2012-10-01
A recently emerging discipline of synthetic biology has the aim of constructing new biosynthetic pathways with useful biological functions. A major application of these pathways is generating a large amount of the desired product. However, toxicity due to the possible presence of toxic precursors is one of the main problems for such production. We consider here the problem of generating a large amount of product from a potentially toxic substrate. To address this, we propose a simple biosynthetic pathway, which can be induced in order to produce a large number of the product molecules, by keeping the substrate amount at low levels. Surprisingly, we show that the large product generation crucially depends on fast non-specific degradation of the substrate molecules. We derive an optimal induction strategy, which allows as much as three orders of magnitude increase in the product amount through biologically realistic parameter values. We point to a recently discovered bacterial immune system (CRISPR/Cas in E. coli) as a putative example of the pathway analysed here. We also argue that the scheme proposed here can be used not only as a stand-alone pathway, but also as a strategy to produce a large amount of the desired molecules with small perturbations of endogenous biosynthetic pathways. (paper)
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A simple biosynthetic pathway for large product generation from small substrate amounts
International Nuclear Information System (INIS)
Djordjevic, Marko; Djordjevic, Magdalena
2012-01-01
A recently emerging discipline of synthetic biology has the aim of constructing new biosynthetic pathways with useful biological functions. A major application of these pathways is generating a large amount of the desired product. However, toxicity due to the possible presence of toxic precursors is one of the main problems for such production. We consider here the problem of generating a large amount of product from a potentially toxic substrate. To address this, we propose a simple biosynthetic pathway, which can be induced in order to produce a large number of the product molecules, by keeping the substrate amount at low levels. Surprisingly, we show that the large product generation crucially depends on fast non-specific degradation of the substrate molecules. We derive an optimal induction strategy, which allows as much as three orders of magnitude increase in the product amount through biologically realistic parameter values. We point to a recently discovered bacterial immune system (CRISPR/Cas in E. coli) as a putative example of the pathway analysed here. We also argue that the scheme proposed here can be used not only as a stand-alone pathway, but also as a strategy to produce a large amount of the desired molecules with small perturbations of endogenous biosynthetic pathways. (paper)
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Genetic determination of the meso-diaminopimelate biosynthetic pathway of mycobacteria.
Cirillo, J D; Weisbrod, T R; Banerjee, A; Bloom, B R; Jacobs, W R
1994-07-01
The increasing incidence of multiple-drug-resistant mycobacterial infections indicates that the development of new methods for treatment of mycobacterial diseases should be a high priority. meso-Diaminopimelic acid (DAP), a key component of a highly immunogenic subunit of the mycobacterial peptidoglycan layer, has been implicated as a potential virulence factor. The mycobacterial DAP biosynthetic pathway could serve as a target for design of new antimycobacterial agents as well as the construction of in vivo selection systems. We have isolated the asd, dapA, dapB, dapD, and dapE genes involved in the DAP biosynthetic pathway of Mycobacterium bovis BCG. These genes were isolated by complementation of Escherichia coli mutations with an expression library of BCG DNA. Our analysis of these genes suggests that BCG may use more than one pathway for biosynthesis of DAP. The nucleotide sequence of the BCG dapB gene was determined. The activity of the product of this gene in Escherichia coli provided evidence that the gene may encode a novel bifunctional dihydrodipicolinate reductase and DAP dehydrogenase.
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Blockage of the pyrimidine biosynthetic pathway affects riboflavin production in Ashbya gossypii.
Silva, Rui; Aguiar, Tatiana Q; Domingues, Lucília
2015-01-10
The Ashbya gossypii riboflavin biosynthetic pathway and its connection with the purine pathway have been well studied. However, the outcome of genetic alterations in the pyrimidine pathway on riboflavin production by A. gossypii had not yet been assessed. Here, we report that the blockage of the de novo pyrimidine biosynthetic pathway in the recently generated A. gossypii Agura3 uridine/uracil auxotrophic strain led to improved riboflavin production on standard agar-solidified complex medium. When extra uridine/uracil was supplied, the production of riboflavin by this auxotroph was repressed. High concentrations of uracil hampered this (and the parent) strain growth, whereas excess uridine favored the A. gossypii Agura3 growth. Considering that the riboflavin and the pyrimidine pathways share the same precursors and that riboflavin overproduction may be triggered by nutritional stress, we suggest that overproduction of riboflavin by the A. gossypii Agura3 may occur as an outcome of a nutritional stress response and/or of an increased availability in precursors for riboflavin biosynthesis, due to their reduced consumption by the pyrimidine pathway. Copyright © 2014 Elsevier B.V. All rights reserved.
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Targeting the GPI biosynthetic pathway.
Yadav, Usha; Khan, Mohd Ashraf
2018-02-27
The GPI (Glycosylphosphatidylinositol) biosynthetic pathway is a multistep conserved pathway in eukaryotes that culminates in the generation of GPI glycolipid which in turn anchors many proteins (GPI-APs) to the cell surface. In spite of the overall conservation of the pathway, there still exist subtle differences in the GPI pathway of mammals and other eukaryotes which holds a great promise so far as the development of drugs/inhibitors against specific targets in the GPI pathway of pathogens is concerned. Many of the GPI structures and their anchored proteins in pathogenic protozoans and fungi act as pathogenicity factors. Notable examples include GPI-anchored variant surface glycoprotein (VSG) in Trypanosoma brucei, GPI-anchored merozoite surface protein 1 (MSP1) and MSP2 in Plasmodium falciparum, protein-free GPI related molecules like lipophosphoglycans (LPGs) and glycoinositolphospholipids (GIPLs) in Leishmania spp., GPI-anchored Gal/GalNAc lectin and proteophosphoglycans in Entamoeba histolytica or the GPI-anchored mannoproteins in pathogenic fungi like Candida albicans. Research in this active area has already yielded encouraging results in Trypanosoma brucei by the development of parasite-specific inhibitors of GlcNCONH 2 -β-PI, GlcNCONH 2 -(2-O-octyl)-PI and salicylic hydroxamic acid (SHAM) targeting trypanosomal GlcNAc-PI de-N-acetylase as well as the development of antifungal inhibitors like BIQ/E1210/gepinacin/G365/G884 and YW3548/M743/M720 targeting the GPI specific fungal inositol acyltransferase (Gwt1) and the phosphoethanolamine transferase-I (Mcd4), respectively. These confirm the fact that the GPI pathway continues to be the focus of researchers, given its implications for the betterment of human life.
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Elucidation and in planta reconstitution of the parthenolide biosynthetic pathway
DEFF Research Database (Denmark)
Liu, Qing; Manzano, David; Tanić, Nikola
2014-01-01
Parthenolide, the main bioactive compound of the medicinal plant feverfew (Tanacetum parthenium), is a promising anti-cancer drug. However, the biosynthetic pathway of parthenolide has not been elucidated yet. Here we report on the isolation and characterization of all the genes from feverfew tha...
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Stahlhut, Steen G; Siedler, Solvej; Malla, Sailesh; Harrison, Scott J; Maury, Jérôme; Neves, Ana Rute; Forster, Jochen
2015-09-01
Plant secondary metabolites are an underutilized pool of bioactive molecules for applications in the food, pharma and nutritional industries. One such molecule is fisetin, which is present in many fruits and vegetables and has several potential health benefits, including anti-cancer, anti-viral and anti-aging activity. Moreover, fisetin has recently been shown to prevent Alzheimer's disease in mice and to prevent complications associated with diabetes type I. Thus far the biosynthetic pathway of fisetin in plants remains elusive. Here, we present the heterologous assembly of a novel fisetin pathway in Escherichia coli. We propose a novel biosynthetic pathway from the amino acid, tyrosine, utilizing nine heterologous enzymes. The pathway proceeds via the synthesis of two flavanones never produced in microorganisms before--garbanzol and resokaempferol. We show for the first time a functional biosynthetic pathway and establish E. coli as a microbial platform strain for the production of fisetin and related flavonols. Copyright © 2015 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.
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Enhancement of Nucleoside Production in Hirsutella sinensis Based on Biosynthetic Pathway Analysis
Liu, Zhi-Qiang; Zhang, Bo; Lin, Shan; Baker, Peter James; Chen, Mao-Sheng; Xue, Ya-Ping; Wu, Hui; Xu, Feng; Yuan, Shui-Jin; Teng, Yi; Wu, Ling-Fang
2017-01-01
To enhance nucleoside production in Hirsutella sinensis, the biosynthetic pathways of purine and pyrimidine nucleosides were constructed and verified. The differential expression analysis showed that purine nucleoside phosphorylase, inosine monophosphate dehydrogenase, and guanosine monophosphate synthase genes involved in purine nucleotide biosynthesis were significantly upregulated 16.56-fold, 8-fold, and 5.43-fold, respectively. Moreover, dihydroorotate dehydrogenase, uridine nucleosidase, uridine/cytidine monophosphate kinase, and inosine triphosphate pyrophosphatase genes participating in pyrimidine nucleoside biosynthesis were upregulated 4.53-fold, 10.63-fold, 4.26-fold, and 5.98-fold, respectively. To enhance the nucleoside production, precursors for synthesis of nucleosides were added based on the analysis of biosynthetic pathways. Uridine and cytidine contents, respectively, reached 5.04 mg/g and 3.54 mg/g when adding 2 mg/mL of ribose, resulting in an increase of 28.6% and 296% compared with the control, respectively. Meanwhile, uridine and cytidine contents, respectively, reached 10.83 mg/g 2.12 mg/g when adding 0.3 mg/mL of uracil, leading to an increase of 176.3% and 137.1%, respectively. This report indicated that fermentation regulation was an effective way to enhance the nucleoside production in H. sinensis based on biosynthetic pathway analysis. PMID:29333435
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A kinetic model for the penicillin biosynthetic pathway in
DEFF Research Database (Denmark)
Nielsen, Jens; Jørgensen, Henrik
1996-01-01
A kinetic model for the first two steps in the penicillin biosynthetic pathway, i.e. the ACV synthetase (ACVS) and the isopenicillin N synthetase (IPNS) is proposed. The model is based on Michaelis-Menten type kinetics with non-competitive inhibition of the ACVS by ACV, and competitive inhibition...... of the IPNS by glutathione. The model predicted flux through the pathway corresponds well with the measured rate of penicillin biosynthesis. From the kinetic model the elasticity coefficients and the flux control coefficients are calculated throughout a fed-batch cultivation, and it is found...
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Linking metabolic QTLs with network and cis-eQTLs controlling biosynthetic pathways.
Directory of Open Access Journals (Sweden)
Adam M Wentzell
2007-09-01
Full Text Available Phenotypic variation between individuals of a species is often under quantitative genetic control. Genomic analysis of gene expression polymorphisms between individuals is rapidly gaining popularity as a way to query the underlying mechanistic causes of variation between individuals. However, there is little direct evidence of a linkage between global gene expression polymorphisms and phenotypic consequences. In this report, we have mapped quantitative trait loci (QTLs-controlling glucosinolate content in a population of 403 Arabidopsis Bay x Sha recombinant inbred lines, 211 of which were previously used to identify expression QTLs controlling the transcript levels of biosynthetic genes. In a comparative study, we have directly tested two plant biosynthetic pathways for association between polymorphisms controlling biosynthetic gene transcripts and the resulting metabolites within the Arabidopsis Bay x Sha recombinant inbred line population. In this analysis, all loci controlling expression variation also affected the accumulation of the resulting metabolites. In addition, epistasis was detected more frequently for metabolic traits compared to transcript traits, even when both traits showed similar distributions. An analysis of candidate genes for QTL-controlling networks of transcripts and metabolites suggested that the controlling factors are a mix of enzymes and regulatory factors. This analysis showed that regulatory connections can feedback from metabolism to transcripts. Surprisingly, the most likely major regulator of both transcript level for nearly the entire pathway and aliphatic glucosinolate accumulation is variation in the last enzyme in the biosynthetic pathway, AOP2. This suggests that natural variation in transcripts may significantly impact phenotypic variation, but that natural variation in metabolites or their enzymatic loci can feed back to affect the transcripts.
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Competition for vitamin B1 (thiamin) structures numerous ecological interactions.
Kraft, Clifford E; Angert, Esther R
2017-06-01
Thiamin (vitamin B1) is a cofactor required for essential biochemical reactions in all living organisms, yet free thiamin is scarce in the environment. The diversity of biochemical pathways involved in the acquisition, degradation, and synthesis of thiamin indicates that organisms have evolved numerous ecological strategies for meeting this nutritional requirement. In this review we synthesize information from multiple disciplines to show how the complex biochemistry of thiamin influences ecological outcomes of interactions between organisms in environments ranging from the open ocean and the Australian outback to the gastrointestinal tract of animals. We highlight population and ecosystem responses to the availability or absence of thiamin. These include widespread mortality of fishes, birds, and mammals, as well as the thiamin-dependent regulation of ocean productivity. Overall, we portray thiamin biochemistry as the foundation for molecularly mediated ecological interactions that influence survival and abundance of a vast array of organisms.
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Chemoreactomic analysis of thiamine disulfide, thiamine hydrochloride, and benfotiamine molecules
Directory of Open Access Journals (Sweden)
O. A. Gromova
2017-01-01
Full Text Available Objective: to analyze the interactions that could indicate the potential pharmacological properties of the molecules of thiamin, thiamine disulfide, and others.Material and methods. The investigators simulated the properties of thiamine disulfide (bistiamin versus those of the reference molecules of thiamin hydrochloride and benfotiamine. The study was performed using chemoreactomic simulation that is the newest area in post-genome pharmacology.Results and discussion. Chemoreactomic analysis has shown that thiamine disulfide can inhibit the molecular receptors involved in blood pressure regulation: adrenoceptors, vasopressin receptor, and angiotensin receptor. Thiamine disulfide can inhibit the reuptake of serotonin, increase its levels, inhibit benzodiazepine receptor and dopamine reuptake, and enhance neuronal acetylcholine release to a large extent than benfotiamine. These molecular effects are consistent with the sedative and anticonvulsant action profile of thiamine disulfide. Simulation has indicated that thiamine disulfide has neuroprotective, anti-inflammatory, normolipidemic, and antitumor activities.Conclusion. The simulation results are confirmed by the available clinical and experimental findings and indicate the virtually unstudied molecular mechanisms of action of thiamine disulfide, benfotiamine, and thiamin hydrochloride.
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Distribution of δ-aminolevulinic acid biosynthetic pathways among phototrophic and related bacteria
International Nuclear Information System (INIS)
Avissar, Y.J.; Beale, S.I.; Ormerod, J.G.
1989-01-01
Two biosynthetic pathways are known for the universal tetrapyrrole precursor, δ-aminolevulinic acid (ALA): condensation of glycine and succinyl-CoA to form ALA with the loss of C-1 of glycine as CO 2 , and conversion of the intact carbon skeleton of glutamate to ALA in a process requiring tRNA Glu , ATP, Mg 2+ , NADPH, and pyridoxal phosphate. The distribution of the two ALA biosynthetic pathways among various bacterial genera was determined, using cell-free extracts obtained from representative organisms. Evidence for the operation of the glutamate pathway was obtained by the measurement of RNase-sensitive label incorporation from glutamate into ALA using 3,4-[ 3 H]glutamate and 1-[ 14 C]glutamate as substrate. The glycine pathway was indicated by RNase-insensitive incorporation of level from 2-[ 14 C]glycine into ALA. The distribution of the two pathways among the bacteria tested was in general agreement with their previously phylogenetic relationships and clearly indicates that the glutamate pathway is the more ancient process, whereas the glycine pathway probably evolved much later. The glutamate pathway is the more widely utilized one among bacteria, while the glycine pathway is apparently limited to the α subgroup of purple bacteria (including Rhodobacter, Rhodospirillum, and Rhizobium). E. coli was found ALA via the glutamate pathway. The ALA-requiring hemA mutant of E. coli was determined to lack the dehydrogenase activity that utilizes glutamyl-tRNA as a substrate
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Pan, X H; Yang, L; Beckers, Y; Xue, F G; Tang, Z W; Jiang, L S; Xiong, B H
2017-07-01
An experiment was conducted to uncover the effects of increasing dietary grain levels on expression of thiamine transporters in ruminal epithelium, and to assess the protective effects of thiamine against high-grain-induced inflammation in dairy cows. Six rumen-fistulated, lactating Holstein dairy cows (627 ± 16.9 kg of body weight, 180 ± 6 d in milk; mean ± standard deviation) were randomly assigned to a replicated 3 × 3 Latin square design trial. Three treatments were control (20% dietary starch, dry matter basis), high-grain diet (HG, 33.2% dietary starch, DM basis), and HG diet supplemented with 180 mg of thiamine/kg of dry matter intake. On d 19 and 20 of each period, milk performance was measured. On d 21, ruminal pH, endotoxic lipopolysaccharide (LPS), and thiamine contents in rumen and blood, and plasma inflammatory cytokines were detected; a rumen papillae biopsy was taken on d 21 to determine the gene and protein expression of toll-like receptor 4 (TLR4) signaling pathways. The HG diet decreased ruminal pH (5.93 vs. 6.49), increased milk yield from 17.9 to 20.2 kg/d, and lowered milk fat and protein from 4.28 to 3.83%, and from 3.38 to 3.11%, respectively. The HG feeding reduced thiamine content in rumen (2.89 vs. 8.97 μg/L) and blood (11.66 vs. 17.63 μg/L), and the relative expression value of thiamine transporter-2 (0.37-fold) and mitochondrial thiamine pyrophosphate transporter (0.33-fold) was downregulated by HG feeding. The HG-fed cows exhibited higher endotoxin LPS in rumen fluid (134,380 vs. 11,815 endotoxin units/mL), and higher plasma concentrations of lipopolysaccharide binding protein and pro-inflammatory cytokines when compared with the control group. The gene and protein expression of tumor necrosis factor α (TNFα), IL1B, and IL6 in rumen epithelium increased when cows were fed the HG diet, indicating that local inflammation occurred. The depressions in ruminal pH, milk fat, and protein of HG-fed cows were reversed by thiamine
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Lin, Shan; Liu, Zhi-Qiang; Baker, Peter James; Yi, Ming; Wu, Hui; Xu, Feng; Teng, Yi; Zheng, Yu-Guo
2016-11-01
The addition of various sulfates for enhanced cordyceps polysaccharide (CP) production in submerged cultivation of H. sinensis was investigated, and manganese sulfate was found the most effective. 2mM of manganese sulfate on 0day (d) was investigated as the optimal adding condition, and the CP production reached optimum with 5.33%, increasing by 93.3% compared with the control. Furthermore, the consumption of three main precursors of CP was studied over cultivation under two conditions. Intracellular mannose content decreased by 43.1% throughout 6days cultivation, which corresponded to CP accumulation rate sharply increased from 0 d to 6 d, and mannose was considered as the most preferred precursor for generating CP. Subsequently, mannose biosynthetic pathway was constructed and verified for the first time in H. sinensis, which constituted the important part of CP biosynthesis, and transcriptional levels of the biosynthetic genes were studied. Transcriptional level of gene cpsA was significantly up-regulated 5.35-fold and it was a key gene involved both in mannose and CP biosynthesis. This study demonstrated that manganese sulfate addition is an efficient and simple way to improve CP production. Transcriptional analysis based on biosynthetic pathway was helpful to find key genes and better understand CP biosynthesis. Copyright © 2016 Elsevier B.V. All rights reserved.
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... deficiency. Brain disorder due to thiamine deficiency (Wernicke-Korsakoff syndrome). Thiamine helps decrease the risk and symptoms of a specific brain disorder called Wernicke-Korsakoff syndrome (WKS). This brain disorder is related to low ...
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Chemoreactomic analysis of thiamine disulfide, thiamine hydrochloride, and benfotiamine molecules
O. A. Gromova; I. Yu. Torshin; L. V. Stakhovskaya; L. E. Fedotova
2017-01-01
Objective: to analyze the interactions that could indicate the potential pharmacological properties of the molecules of thiamin, thiamine disulfide, and others.Material and methods. The investigators simulated the properties of thiamine disulfide (bistiamin) versus those of the reference molecules of thiamin hydrochloride and benfotiamine. The study was performed using chemoreactomic simulation that is the newest area in post-genome pharmacology.Results and discussion. Chemoreactomic analysis...
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Kang, Tae Sun; Korber, Darren R; Tanaka, Takuji
2014-06-01
The regulatory role of a transcriptional regulator (PocR) in the 1,3-propanediol biosynthetic pathway of Lactobacillus panis PM1 contributes to the optimization of 1,3-propanediol production by this strain, which potentially will lead to 1,3-propanediol manufacturing efficiencies. Lactobacillus panis PM1 can utilize a 1,3-propanediol (1,3-PDO) biosynthetic pathway, consisting of diol dehydratase (PduCDE) and 1,3-PDO dehydrogenase, as a NADH recycling system, to survive under various environmental conditions. In this study, we identified a key transcriptional repressor (PocR) which was annotated as a transcriptional factor of AraC family as part of the 1,3-PDO biosynthetic pathway of L. panis PM1. The over-expression of the PocR gene resulted in the significant repression (81 %) of pduC (PduCDE large subunit) transcription, and subsequently, the decreased activity of PduCDE by 22 %. As a result of the regulation of PduCDE, production of both 3-hydroxypropionaldehyde and 1,3-PDO in the PocR over-expressing strain were significantly decreased by 40 % relative to the control strain. These results clearly demonstrate the transcriptional repressor role of PocR in the 1,3-PDO biosynthetic pathway.
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Xie, Wenping; Liu, Min; Lv, Xiaomei; Lu, Wenqiang; Gu, Jiali; Yu, Hongwei
2014-01-01
Saccharomyces cerevisiae is an important platform organism for the synthesis of a great number of natural products. However, the assembly of controllable and genetically stable heterogeneous biosynthetic pathways in S. cerevisiae still remains a significant challenge. Here, we present a strategy for reconstructing controllable multi-gene pathways by employing the GAL regulatory system. A set of marker recyclable integrative plasmids (pMRI) was designed for decentralized assembly of pathways. As proof-of-principle, a controllable β-carotene biosynthesis pathway (∼16 kb) was reconstructed and optimized by repeatedly using GAL10-GAL1 bidirectional promoters with high efficiency (80-100%). By controling the switch time of the pathway, production of 11 mg/g DCW of total carotenoids (72.57 mg/L) and 7.41 mg/g DCW of β-carotene was achieved in shake-flask culture. In addition, the engineered yeast strain exhibited high genetic stability after 20 generations of subculture. The results demonstrated a controllable and genetically stable biosynthetic pathway capable of increasing the yield of target products. Furthermore, the strategy presented in this study could be extended to construct other pathways in S. cerevisisae. © 2013 Wiley Periodicals, Inc.
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Directory of Open Access Journals (Sweden)
Sebastian J. Nintemann
2017-11-01
Full Text Available Within the cell, biosynthetic pathways are embedded in protein-protein interaction networks. In Arabidopsis, the biosynthetic pathways of aliphatic and indole glucosinolate defense compounds are well-characterized. However, little is known about the spatial orchestration of these enzymes and their interplay with the cellular environment. To address these aspects, we applied two complementary, untargeted approaches—split-ubiquitin yeast 2-hybrid and co-immunoprecipitation screens—to identify proteins interacting with CYP83A1 and CYP83B1, two homologous enzymes specific for aliphatic and indole glucosinolate biosynthesis, respectively. Our analyses reveal distinct functional networks with substantial interconnection among the identified interactors for both pathway-specific markers, and add to our knowledge about how biochemical pathways are connected to cellular processes. Specifically, a group of protein interactors involved in cell death and the hypersensitive response provides a potential link between the glucosinolate defense compounds and defense against biotrophic pathogens, mediated by protein-protein interactions.
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Prevention of incipient diabetic nephropathy by high-dose thiamine and benfotiamine.
Babaei-Jadidi, Roya; Karachalias, Nikolaos; Ahmed, Naila; Battah, Sinan; Thornalley, Paul J
2003-08-01
Accumulation of triosephosphates arising from high cytosolic glucose concentrations in hyperglycemia is the trigger for biochemical dysfunction leading to the development of diabetic nephropathy-a common complication of diabetes associated with a high risk of cardiovascular disease and mortality. Here we report that stimulation of the reductive pentosephosphate pathway by high-dose therapy with thiamine and the thiamine monophosphate derivative benfotiamine countered the accumulation of triosephosphates in experimental diabetes and inhibited the development of incipient nephropathy. High-dose thiamine and benfotiamine therapy increased transketolase expression in renal glomeruli, increased the conversion of triosephosphates to ribose-5-phosphate, and strongly inhibited the development of microalbuminuria. This was associated with decreased activation of protein kinase C and decreased protein glycation and oxidative stress-three major pathways of biochemical dysfunction in hyperglycemia. Benfotiamine also inhibited diabetes-induced hyperfiltration. This was achieved without change in elevated plasma glucose concentration and glycated hemoglobin in the diabetic state. High-dose thiamine and benfotiamine therapy is a potential novel strategy for the prevention of clinical diabetic nephropathy.
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Genetic determination of the meso-diaminopimelate biosynthetic pathway of mycobacteria.
Cirillo, J. D.; Weisbrod, T. R.; Banerjee, A.; Bloom, B. R.; Jacobs, W. R.
1994-01-01
The increasing incidence of multiple-drug-resistant mycobacterial infections indicates that the development of new methods for treatment of mycobacterial diseases should be a high priority. meso-Diaminopimelic acid (DAP), a key component of a highly immunogenic subunit of the mycobacterial peptidoglycan layer, has been implicated as a potential virulence factor. The mycobacterial DAP biosynthetic pathway could serve as a target for design of new antimycobacterial agents as well as the constru...
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Molecular evolution of the lysine biosynthetic pathways.
Velasco, A M; Leguina, J I; Lazcano, A
2002-10-01
Among the different biosynthetic pathways found in extant organisms, lysine biosynthesis is peculiar because it has two different anabolic routes. One is the diaminopimelic acid pathway (DAP), and the other over the a-aminoadipic acid route (AAA). A variant of the AAA route that includes some enzymes involved in arginine and leucine biosyntheses has been recently reported in Thermus thermophilus (Nishida et al. 1999). Here we describe the results of a detailed genomic analysis of each of the sequences involved in the two lysine anabolic routes, as well as of genes from other routes related to them. No evidence was found of an evolutionary relationship between the DAP and AAA enzymes. Our results suggest that the DAP pathway is related to arginine metabolism, since the lysC, asd, dapC, dapE, and lysA genes from lysine biosynthesis are related to the argB, argC, argD, argE, and speAC genes, respectively, whose products catalyze different steps in arginine metabolism. This work supports previous reports on the relationship between AAA gene products and some enzymes involved in leucine biosynthesis and the tricarboxylic acid cycle (Irvin and Bhattacharjee 1998; Miyazaki et al. 2001). Here we discuss the significance of the recent finding that several genes involved in the arginine (Arg) and leucine (Leu) biosynthesis participate in a new alternative route of the AAA pathway (Miyazaki et al. 2001). Our results demonstrate a clear relationship between the DAP and Arg routes, and between the AAA and Leu pathways.
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Luchetti, Sabina; Bossers, Koen; Frajese, Giovanni Vanni; Swaab, Dick F.
2010-01-01
There is emerging evidence from animal studies for a neuroprotective role of sex steroids in neurodegenerative diseases, but studies in human brain are lacking. We have carried out an extensive study of the neurosteroid biosynthetic pathways in substantia nigra (SN), caudate nucleus (CN) and putamen
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Energy and glucose pathways in thiamine deficient primary rat brain microvascular endothelial cells.
Ham, D; Karska-Wysocki, B
2005-12-01
Thiamine deficiency (TD) results in lactate acidosis, which is associated with neurodegeneration. The aim of this study was to investigate this alteration in primary rat brain endothelia. Spectrophotometric analysis of culture media revealed that only a higher concentration of pyrithiamine, which accelerates the intracellular blocking of thiamine, significantly elevated the lactate level and lactate dehydrogenase activity within 7 days. The medium without pyrithiamine and with a thiamine concentration comparable to pathophysiological plasma levels mildly reduced only the activity of transketolase. This suggests that significant metabolic changes may not occur at the early phase of TD in cerebral capillary cells, while anaerobic glycolysis in capillaries may be mediated during late stage/chronic TD.
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[Thiamine and its derivatives in the regulation of cell metabolism].
Tylicki, Adam; Siemieniuk, Magdalena
2011-07-06
For over 70 years thiamine (vitamin B1) has aroused the interest of biologists, biochemists and medical doctors because of its multilateral participation in key biochemical and physiological processes. The thiamine molecule is composed of pyrimidine and thiazole rings which are linked by a methylene bridge. It is synthesized by microorganisms, fungi and plants, whereas animals and humans have to obtain it from food. There are several known forms of vitamin B1 inside cells: free thiamine, three phosphate esters (mono-, di-, and triphosphate), and the recently found adenosine thiamine triphosphate. Thiamine has a dual, coenzymatic and non-coenzymatic role. First of all, it is a precursor of thiamin diphosphate, which is a coenzyme for over 20 characterized enzymes which are involved in cell bioenergetic processes leading to the synthesis of ATP. Moreover, these enzymes take part in the biosynthesis of pentose (required for the synthesis of nucleotides), amino acids and other organic compounds of cell metabolism. On the other hand, recent discoveries show the non-coenzymatic role of thiamine derivatives in the process of regulation of gene expression (riboswitches in microorganisms and plants), the stress response, and perhaps so far unknown signal transduction pathways associated with adverse environmental conditions, or transduction of nerve signals with participation of thiamine triphosphate and adenosine thiamine triphosphate. From the clinical point of view thiamine deficiency is related to beri-beri, Parkinson disease, Alzheimer disease, Wernicke-Korsakoff syndrome and other pathologies of the nervous system, and it is successfully applied in medical practice. On the other hand, identifying new synthetic analogues of thiamine which could be used as cytostatics, herbicides or agents preventing deficiency of vitamin B1 is currently the major goal of the research. In this paper we present the current state of knowledge of thiamine and its derivatives, indicating
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DEFF Research Database (Denmark)
Rugbjerg, Peter; Naesby, Michael; Mortensen, Uffe Hasbro
2013-01-01
production in easily fermentable and genetically engineerable organisms, such as Saccharomyces cerevisiae and Escherichia coli are desirable. Rubrofusarin is an orange polyketide pigment that is a common intermediate in many different fungal biosynthetic pathways. RESULTS: In this study, we established...
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Kim, Eun Jin; Angell, Scott; Janes, Jeff; Watanabe, Coran M H
2008-06-01
Traditional approaches to natural product discovery involve cell-based screening of natural product extracts followed by compound isolation and characterization. Their importance notwithstanding, continued mining leads to depletion of natural resources and the reisolation of previously identified metabolites. Metagenomic strategies aimed at localizing the biosynthetic cluster genes and expressing them in surrogate hosts offers one possible alternative. A fundamental question that naturally arises when pursuing such a strategy is, how large must the genomic library be to effectively represent the genome of an organism(s) and the biosynthetic gene clusters they harbor? Such an issue is certainly augmented in the absence of expensive robotics to expedite colony picking and/or screening of clones. We have developed an algorism, named BPC (biosynthetic pathway coverage), supported by molecular simulations to deduce the number of BAC clones required to achieve proper coverage of the genome and their respective biosynthetic pathways. The strategy has been applied to the construction of a large-insert BAC library from a marine microorganism, Hon6 (isolated from Honokohau, Maui) thought to represent a new species. The genomic library is constructed with a BAC yeast shuttle vector pClasper lacZ paving the way for the culturing of libraries in both prokaryotic and eukaryotic hosts. Flow cytometric methods are utilized to estimate the genome size of the organism and BPC implemented to assess P-coverage or percent coverage. A genetic selection strategy is illustrated, applications of which could expedite screening efforts in the identification and localization of biosynthetic pathways from marine microbial consortia, offering a powerful complement to genome sequencing and degenerate probe strategies. Implementing this approach, we report on the biotin biosynthetic pathway from the marine microorganism Hon6.
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Brown, S.B.; Brown, L.R.; Brown, M.; Moore, K.; Villella, M.; Fitzsimons, J.D.; Williston, B.; Honeyfield, D.C.; Hinterkopf, J.P.; Tillitt, D.E.; Zajicek, J.L.; Wolgamood, M.
2005-01-01
Protocols used for therapeutic thiamine treatments in salmonine early mortality syndrome (EMS) were investigated in lake trout Salvelinus namaycush and coho salmon Oncorhynchus kisutch to assess their efficacy. At least 500 mg of thiamine HCl/L added to egg baths was required to produce a sustained elevation of thiamine content in lake trout eggs. Thiamine uptake from egg baths was not influenced by a pH ranging from 5.5 to 7.5 or by a water hardness between 2 and 200 mg CaCO3/L. There was poorer thiamine uptake when initial thiamine levels were low, suggesting that current treatment regimes may not be as effective when thiamine levels are severely depressed and that higher treatment doses are necessary. Exposure of eggs to the more lipid-soluble thiamine analog allithiamine (1,000 mg/L) during water hardening increased egg thiamine levels by 1.5-2.5 nmol/g and was completely effective at reversing EMS. Another more lipid-soluble thiamine analog, benfotiamine (100 mg/L), reduced EMS but did not produce detectable increases in egg thiamine content. Although benfotiamine may be more effective than thiamine at mitigating EMS, it is more expensive than thiamine HCl or allithiamine. In addition, there still needs to be a more thorough examination of dose-response relationships. We conclude that allithiamine is an alternative to the use of thiamine in egg baths as a therapeutic treatment for salmonid EMS. ?? Copyright by the American Fisheries Society 2005.
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Directory of Open Access Journals (Sweden)
Marjorie Gangolf
Full Text Available BACKGROUND: Thiamine (vitamin B1 is an essential molecule for all life forms because thiamine diphosphate (ThDP is an indispensable cofactor for oxidative energy metabolism. The less abundant thiamine monophosphate (ThMP, thiamine triphosphate (ThTP and adenosine thiamine triphosphate (AThTP, present in many organisms, may have still unidentified physiological functions. Diseases linked to thiamine deficiency (polyneuritis, Wernicke-Korsakoff syndrome remain frequent among alcohol abusers and other risk populations. This is the first comprehensive study on the distribution of thiamine derivatives in human biopsies, body fluids and cell lines. METHODOLOGY AND PRINCIPAL FINDINGS: Thiamine derivatives were determined by HPLC. In human tissues, the total thiamine content is lower than in other animal species. ThDP is the major thiamine compound and tissue levels decrease at high age. In semen, ThDP content correlates with the concentration of spermatozoa but not with their motility. The proportion of ThTP is higher in humans than in rodents, probably because of a lower 25-kDa ThTPase activity. The expression and activity of this enzyme seems to correlate with the degree of cell differentiation. ThTP was present in nearly all brain and muscle samples and in ∼60% of other tissue samples, in particular fetal tissue and cultured cells. A low ([ThTP]+[ThMP]/([Thiamine]+[ThMP] ratio was found in cardiovascular tissues of patients with cardiac insufficiency. AThTP was detected only sporadically in adult tissues but was found more consistently in fetal tissues and cell lines. CONCLUSIONS AND SIGNIFICANCE: The high sensitivity of humans to thiamine deficiency is probably linked to low circulating thiamine concentrations and low ThDP tissue contents. ThTP levels are relatively high in many human tissues, as a result of low expression of the 25-kDa ThTPase. Another novel finding is the presence of ThTP and AThTP in poorly differentiated fast-growing cells
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Viggiano, Annarita; Salo, Oleksandr; Ali, Hazrat; Szymanski, Wiktor; Lankhorst, Peter P; Nygård, Yvonne; Bovenberg, Roel A L; Driessen, Arnold J M
Chrysogine is a yellow pigment produced by Penicillium chrysogenum and other filamentous fungi. Although it was first isolated in 1973, the biosynthetic pathway has so far not been resolved. Here, we show that the deletion of the highly expressed non-ribosomal peptide synthetase (NRPS) gene
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Directory of Open Access Journals (Sweden)
Sandra Fresquet-Corrales
Full Text Available Proanthocyanidins (PAs, or condensed tannins, are powerful antioxidants that remove harmful free oxygen radicals from cells. To engineer the anthocyanin and proanthocyanidin biosynthetic pathways to de novo produce PAs in two Nicotiana species, we incorporated four transgenes to the plant chassis. We opted to perform a simultaneous transformation of the genes linked in a multigenic construct rather than classical breeding or retransformation approaches. We generated a GoldenBraid 2.0 multigenic construct containing two Antirrhinum majus transcription factors (AmRosea1 and AmDelila to upregulate the anthocyanin pathway in combination with two Medicago truncatula genes (MtLAR and MtANR to produce the enzymes that will derivate the biosynthetic pathway to PAs production. Transient and stable transformation of Nicotiana benthamiana and Nicotiana tabacum with the multigenic construct were respectively performed. Transient expression experiments in N. benthamiana showed the activation of the anthocyanin pathway producing a purple color in the agroinfiltrated leaves and also the effective production of 208.5 nmol (- catechin/g FW and 228.5 nmol (- epicatechin/g FW measured by the p-dimethylaminocinnamaldehyde (DMACA method. The integration capacity of the four transgenes, their respective expression levels and their heritability in the second generation were analyzed in stably transformed N. tabacum plants. DMACA and phoroglucinolysis/HPLC-MS analyses corroborated the activation of both pathways and the effective production of PAs in T0 and T1 transgenic tobacco plants up to a maximum of 3.48 mg/g DW. The possible biotechnological applications of the GB2.0 multigenic approach in forage legumes to produce "bloat-safe" plants and to improve the efficiency of conversion of plant protein into animal protein (ruminal protein bypass are discussed.
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Overexpression of the riboflavin biosynthetic pathway in Pichia pastoris
Directory of Open Access Journals (Sweden)
Mattanovich Diethard
2008-07-01
Full Text Available Abstract Background High cell density cultures of Pichia pastoris grown on methanol tend to develop yellow colored supernatants, attributed to the release of free flavins. The potential of P. pastoris for flavin overproduction is therefore given, but not pronounced when the yeast is grown on glucose. The aim of this study is to characterize the relative regulatory impact of each riboflavin synthesis gene. Deeper insight into pathway control and the potential of deregulation is established by overexpression of the single genes as well as a combined deregulation of up to all six riboflavin synthesis genes. Results Overexpression of the first gene of the riboflavin biosynthetic pathway (RIB1 is already sufficient to obtain yellow colonies and the accumulation of riboflavin in the supernatant of shake flask cultures growing on glucose. Sequential deregulation of all the genes, by exchange of their native promoter with the strong and constitutive glyceraldehyde-3-phosphate dehydrogenase promoter (PGAP increases the riboflavin accumulation significantly. Conclusion The regulation of the pathway is distributed over more than one gene. High cell density cultivations of a P. pastoris strain overexpressing all six RIB genes allow the accumulation of 175 mg/L riboflavin in the supernatant. The basis for rational engineering of riboflavin production in P. pastoris has thus been established.
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Directory of Open Access Journals (Sweden)
Wenjun Huang
2017-07-01
Full Text Available Epimedium species have been widely used both as traditional Chinese medicinal plants and ornamental perennials. Both flavonols, acting as the major bioactive components (BCs and anthocyanins, predominantly contributing to the color diversity of Epimedium flowers belong to different classes of flavonoids. It is well-acknowledged that flavonoid biosynthetic pathway is predominantly regulated by R2R3-MYB transcription factor (TF as well as bHLH TF and WD40 protein at the transcriptional level. MYB TFs specifically regulating anthocyanin or flavonol biosynthetic pathway have been already isolated and functionally characterized from Epimedium sagittatum, but a R2R3-MYB TF involved in regulating both these two pathways has not been functionally characterized to date in Epimedium plants. In this study, we report the functional characterization of EsMYB9, a R2R3-MYB TF previously isolated from E. sagittatum. The previous study indicated that EsMYB9 belongs to a small subfamily of R2R3-MYB TFs containing grape VvMYB5a and VvMYB5b TFs, which regulate flavonoid biosynthetic pathway. The present studies show that overexpression of EsMYB9 in tobacco leads to increased transcript levels of flavonoid pathway genes and increased contents of anthocyanins and flavonols. Yeast two-hybrid assay indicates that the C-terminal region of EsMYB9 contributes to the autoactivation activity, and EsMYB9 interacts with EsTT8 or AtTT8 bHLH regulator. Transient reporter assay shows that EsMYB9 slightly activates the expression of EsCHS (chalcone synthase promoter in transiently transformed leaves of Nicotiana benthamiana, but the addition of AtTT8 or EsTT8 bHLH regulator strongly enhances the transcriptional activation of EsMYB9 against five promoters of the flavonoid pathway genes except EsFLS (flavonol synthase. In addition, co-transformation of EsMYB9 and EsTT8 in transiently transfected tobacco leaves strongly induces the expressions of flavonoid biosynthetic genes. The
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Thiamine, fasting and the kidney
Klooster, Astrid
2013-01-01
Thiamine, Vasten en de Nier In dit proefschrift hebben we de rol van thiamine in ischemie-reperfusie schade van de nier onderzocht. Thiamine is cruciaal voor het behoud van optimale hoeveelheden gereduceerd glutathion (een anti-oxidant) en voor regeneratie van ATP (energiebron) in cellen. We
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Thiamine and magnesium deficiencies: keys to disease.
Lonsdale, D
2015-02-01
Thiamine deficiency (TD) is accepted as the cause of beriberi because of its action in the metabolism of simple carbohydrates, mainly as the rate limiting cofactor for the dehydrogenases of pyruvate and alpha-ketoglutarate, both being critical to the action of the citric acid cycle. Transketolase, dependent on thiamine and magnesium, occurs twice in the oxidative pentose pathway, important in production of reducing equivalents. Thiamine is also a cofactor in the dehydrogenase complex in the degradation of the branched chain amino acids, leucine, isoleucine and valine. In spite of these well accepted facts, the overall clinical effects of TD are still poorly understood. Because of the discovery of 2-hydroxyacyl-CoA lyase (HACL1) as the first peroxisomal enzyme in mammals found to be dependent on thiamine pyrophosphate (TPP) and the ability of thiamine to bind with prion protein, these factors should improve our clinical approach to TD. HACL1 has two important roles in alpha oxidation, the degradation of phytanic acid and shortening of 2-hydroxy long-chain fatty acids so that they can be degraded further by beta oxidation. The downstream effects of a lack of efficiency in this enzyme would be expected to be critical in normal brain metabolism. Although TD has been shown experimentally to produce reversible damage to mitochondria and there are many other causes of mitochondrial dysfunction, finding TD as the potential biochemical lesion would help in differential diagnosis. Stresses imposed by infection, head injury or inoculation can initiate intermittent cerebellar ataxia in thiamine deficiency/dependency. Medication or vaccine reactions appear to be more easily initiated in the more intelligent individuals when asymptomatic marginal malnutrition exists. Erythrocyte transketolase testing has shown that thiamine deficiency is widespread. It is hypothesized that the massive consumption of empty calories, particularly those derived from carbohydrate and fat, results in
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Directory of Open Access Journals (Sweden)
Floriana Gernone
2017-11-01
Full Text Available Thiamine is a water-soluble vitamin, which participates in several vital metabolic pathways involved in energy metabolism and neurotransmitter synthesis of mammals. In companion animals thiamine deficiency is classically associated with signs of diffuse encephalopathy and lesions on brainstem nuclei and mesencephalic colliculi evident on magnetic resonance imaging. This paper describes a novel clinical presentation in a thiamine-deficient dog showing multifocal, central and peripheral nervous and cardiovascular system alterations. Brain MRI showed bilateral caudate nuclei damage, with necrotic-malacic evolution, similar to the atypical MRI pattern found in Wernicke’s encephalopathy in humans. Detection of bilateral symmetrical lesions of the caudate nuclei in dogs should prompt consideration of a thiamine deficiency among the differential diagnoses.
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Thiamine content and turnover rates of some rat nervous regions, using labeled thiamine as a tracer
International Nuclear Information System (INIS)
Rindi, G.; Patrini, C.; Comincioli, V.; Reggiani, C.
1980-01-01
The content of total thiamine radioactivity in some nervous structures and liver of the rat was determined in a steady state condition, using [thiazole-2- 14 C]thiamine as a tracer. The contents were analyzed by a mamillary type compartmental model which enabled the authors to calculate the influx and efflux fractional rate constants, turnover times, turnover rates and relative accuracy. The results show in general a good agreement between turnover rate values and brain regional sensitivity to thiamine deficiency, the most vunerable areas to thiamine depletion being those with the highest turnover rates. (Auth.)
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The immediate nucleotide precursor, guanosine triphosphate, in the riboflavin biosynthetic pathway
International Nuclear Information System (INIS)
Mitsuda, Hisateru; Nakajima, Kenji; Nadamoto, Tomonori
1977-01-01
In the present paper, the nucleotide precursor of riboflavin was investigated by experiments with labeled purines using non-growing cells of Eremothecium ashbyii. The added purines, at 10 -4 M, were effectively incorporated into riboflavin at an early stage of riboflavin biosynthesis under the experimental conditions. In particular, both labeled xanthine and labeled guanine were specifically transported to guanosine nucleotides, GMP, GDP, GDP-Mannose and GTP, in the course of the riboflavin biosynthesis. A comparison of specific activities of labeled guanosine nucleotides and labeled riboflavin indicated that the nucleotide precursor of riboflavin is guanosine triphosphate. From the results obtained, a biosynthetic pathway of riboflavin is proposed. (auth.)
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Andersen-Ranberg, Johan; Kongstad, Kenneth Thermann; Nafisi, Majse; Staerk, Dan; Okkels, Finn Thyge; Mortensen, Uffe Hasbro; Lindberg Møller, Birger; Frandsen, Rasmus John Normand; Kannangara, Rubini
2017-10-05
Carminic acid is a C-glucosylated octaketide anthraquinone and the main constituent of the natural dye carmine (E120), possessing unique coloring, stability, and solubility properties. Despite being used since ancient times, longstanding efforts to elucidate its route of biosynthesis have been unsuccessful. Herein, a novel combination of enzymes derived from a plant (Aloe arborescens, Aa), a bacterium (Streptomyces sp. R1128, St), and an insect (Dactylopius coccus, Dc) that allows for the biosynthesis of the C-glucosylated anthraquinone, dcII, a precursor for carminic acid, is reported. The pathway, which consists of AaOKS, StZhuI, StZhuJ, and DcUGT2, presents an alternative biosynthetic approach for the production of polyketides by using a type III polyketide synthase (PKS) and tailoring enzymes originating from a type II PKS system. The current study showcases the power of using transient expression in Nicotiana benthamiana for efficient and rapid identification of functional biosynthetic pathways, including both soluble and membrane-bound enzymes. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.
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Pharmacokinetics of thiamine derivatives especially of benfotiamine.
Loew, D
1996-02-01
Pharmacokinetic data of orally administered lipid-soluble thiamine analogues like benfotiamine are reviewed and assessed. It is quite clear that benfotiamine is absorbed much more better than water-soluble thiamine salts: maximum plasma levels of thiamine are about 5 times higher after benfotiamine, the bioavailability is at maximum about 3.6 times as high as that of thiamine hydrochloride and better than other lipophilic thiamine derivates. The physiological activity (alphaETK) increased only after benfotiamine was given. Due to its excellent pharmacokinetic profile benfotiamine should be preferred in treatment of relevant indications.
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Widespread episodic thiamine deficiency in Northern Hemisphere wildlife
Balk, Lennart; Hägerroth, Per-Åke; Gustavsson, Hanna; Sigg, Lisa; Åkerman, Gun; Ruiz Muñoz, Yolanda; Honeyfield, Dale C.; Tjärnlund, Ulla; Oliveira, Kenneth; Ström, Karin; McCormick, Stephen D.; Karlsson, Simon; Ström, Marika; van Manen, Mathijs; Berg, Anna-Lena; Halldórsson, Halldór P.; Strömquist, Jennie; Collier, Tracy K.; Börjeson, Hans; Mörner, Torsten; Hansson, Tomas
2016-12-01
Many wildlife populations are declining at rates higher than can be explained by known threats to biodiversity. Recently, thiamine (vitamin B1) deficiency has emerged as a possible contributing cause. Here, thiamine status was systematically investigated in three animal classes: bivalves, ray-finned fishes, and birds. Thiamine diphosphate is required as a cofactor in at least five life-sustaining enzymes that are required for basic cellular metabolism. Analysis of different phosphorylated forms of thiamine, as well as of activities and amount of holoenzyme and apoenzyme forms of thiamine-dependent enzymes, revealed episodically occurring thiamine deficiency in all three animal classes. These biochemical effects were also linked to secondary effects on growth, condition, liver size, blood chemistry and composition, histopathology, swimming behaviour and endurance, parasite infestation, and reproduction. It is unlikely that the thiamine deficiency is caused by impaired phosphorylation within the cells. Rather, the results point towards insufficient amounts of thiamine in the food. By investigating a large geographic area, by extending the focus from lethal to sublethal thiamine deficiency, and by linking biochemical alterations to secondary effects, we demonstrate that the problem of thiamine deficiency is considerably more widespread and severe than previously reported.
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Widespread episodic thiamine deficiency in Northern Hemisphere wildlife
Balk, Lennart; Hägerroth, Per-Åke; Gustavsson, Hanna; Sigg, Lisa; Akerman, Gun; Ruiz Muñoz, Yolanda; Honeyfield, Dale C.; Tjarnlund, Ulla; Oliveira, Kenneth; Strom, Karin; McCormick, Stephen D.; Karlsson, Simon; Strom, Marika; van Manen, Mathijs; Berg, Anna-Lena; Halldórsson, Halldór P.; Stromquist, Jennie; Collier, Tracy K.; Borjeson, Hans; Morner, Torsten; Hansson, Tomas
2016-01-01
Many wildlife populations are declining at rates higher than can be explained by known threats to biodiversity. Recently, thiamine (vitamin B1) deficiency has emerged as a possible contributing cause. Here, thiamine status was systematically investigated in three animal classes: bivalves, ray-finned fishes, and birds. Thiamine diphosphate is required as a cofactor in at least five life-sustaining enzymes that are required for basic cellular metabolism. Analysis of different phosphorylated forms of thiamine, as well as of activities and amount of holoenzyme and apoenzyme forms of thiamine-dependent enzymes, revealed episodically occurring thiamine deficiency in all three animal classes. These biochemical effects were also linked to secondary effects on growth, condition, liver size, blood chemistry and composition, histopathology, swimming behaviour and endurance, parasite infestation, and reproduction. It is unlikely that the thiamine deficiency is caused by impaired phosphorylation within the cells. Rather, the results point towards insufficient amounts of thiamine in the food. By investigating a large geographic area, by extending the focus from lethal to sublethal thiamine deficiency, and by linking biochemical alterations to secondary effects, we demonstrate that the problem of thiamine deficiency is considerably more widespread and severe than previously reported.
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Zhu, Qinghua; Chen, Qi; Song, Yongxiang; Huang, Hongbo; Li, Jun; Ma, Junying; Li, Qinglian; Ju, Jianhua
2017-05-09
Galactose, a monosaccharide capable of assuming two possible configurational isomers (d-/l-), can exist as a six-membered ring, galactopyranose (Gal p ), or as a five-membered ring, galactofuranose (Gal f ). UDP-galactopyranose mutase (UGM) mediates the conversion of pyranose to furanose thereby providing a precursor for d-Gal f Moreover, UGM is critical to the virulence of numerous eukaryotic and prokaryotic human pathogens and thus represents an excellent antimicrobial drug target. However, the biosynthetic mechanism and relevant enzymes that drive l-Gal f production have not yet been characterized. Herein we report that efforts to decipher the sugar biosynthetic pathway and tailoring steps en route to nucleoside antibiotic A201A led to the discovery of a GDP-l-galactose mutase, MtdL. Systematic inactivation of 18 of the 33 biosynthetic genes in the A201A cluster and elucidation of 10 congeners, coupled with feeding and in vitro biochemical experiments, enabled us to: ( i ) decipher the unique enzyme, GDP-l-galactose mutase associated with production of two unique d-mannose-derived sugars, and ( ii ) assign two glycosyltransferases, four methyltransferases, and one desaturase that regiospecifically tailor the A201A scaffold and display relaxed substrate specificities. Taken together, these data provide important insight into the origin of l-Gal f -containing natural product biosynthetic pathways with likely ramifications in other organisms and possible antimicrobial drug targeting strategies.
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Thiamine absorption is not compromised in folate-deficient rats
International Nuclear Information System (INIS)
Walzem, R.L.; Clifford, A.J.
1988-01-01
Thiamine absorption and excretion were assessed in rats with severe folate deficiency (FD) by determining the fate of oral 3 H-labeled and intravenous 14 C-labeled thiamine over a 6-h test period. Thiamine status was evaluated in these same rats by measuring transketolase activity levels of blood before (TKA) and after (TPPE) addition of thiamine pyrophosphate to the incubation mixture of the assay procedure. Two additional experiments assessed active transport of thiamine and the effect of dietary succinylsulfathiazole (SST) on TKA and TPPE in rats with moderate FD. Intestinal absorption in general and thiamine absorption in particular and thiamine status were unaltered in rats with severe FD. Inanition associated with severe FD may impair thiamine status. Thiamine absorption by active transport was not compromised in FD, and dietary succinylsulfathiazole did not affect thiamine status
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Institute of Medicine, Food and Nutrition Board. Dietary Reference Intakes for Thiamin, Riboflavin, Niacin, Vitamin B6, Folate, Vitamin B12, Pantothenic Acid, Biotin, and Choline. National Academies Press. Washington, DC, 1998. PMID: 23193625 ...
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Metabolic engineering of biosynthetic pathway for production of renewable biofuels.
Singh, Vijai; Mani, Indra; Chaudhary, Dharmendra Kumar; Dhar, Pawan Kumar
2014-02-01
Metabolic engineering is an important area of research that involves editing genetic networks to overproduce a certain substance by the cells. Using a combination of genetic, metabolic, and modeling methods, useful substances have been synthesized in the past at industrial scale and in a cost-effective manner. Currently, metabolic engineering is being used to produce sufficient, economical, and eco-friendly biofuels. In the recent past, a number of efforts have been made towards engineering biosynthetic pathways for large scale and efficient production of biofuels from biomass. Given the adoption of metabolic engineering approaches by the biofuel industry, this paper reviews various approaches towards the production and enhancement of renewable biofuels such as ethanol, butanol, isopropanol, hydrogen, and biodiesel. We have also identified specific areas where more work needs to be done in the future.
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Janevska, Slavica; Tudzynski, Bettina
2018-01-01
The fungus Fusarium fujikuroi causes bakanae disease of rice due to its ability to produce the plant hormones, the gibberellins. The fungus is also known for producing harmful mycotoxins (e.g., fusaric acid and fusarins) and pigments (e.g., bikaverin and fusarubins). However, for a long time, most of these well-known products could not be linked to biosynthetic gene clusters. Recent genome sequencing has revealed altogether 47 putative gene clusters. Most of them were orphan clusters for which the encoded natural product(s) were unknown. In this review, we describe the current status of our research on identification and functional characterizations of novel secondary metabolite gene clusters. We present several examples where linking known metabolites to the respective biosynthetic genes has been achieved and describe recent strategies and methods to access new natural products, e.g., by genetic manipulation of pathway-specific or global transcritption factors. In addition, we demonstrate that deletion and over-expression of histone-modifying genes is a powerful tool to activate silent gene clusters and to discover their products.
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The flavonoid biosynthetic pathway in plants: function and evolution
International Nuclear Information System (INIS)
Koes, R.E.; Quattrocchio, F.; Mol, J.N.M.
1994-01-01
Flavonoids are a class of low molecular weight phenolic compounds that is widely distributed in the plant kingdom. They exhibit a diverse spectrum of biological functions and play an important role in the interaction between plants and their environment. Flavonoids not only protect the plant from the harmful effects of UV irradiation but also play a crucial role in the sexual reproduction process. A special class of flavonoid polymers, the tannins, plays a structural role in the plant. Yet other classes of flavonoids, flavonols and anthocyanins, have been implicated in the attraction of pollinators. Certain flavonoids participate in the interaction between plants and other organisms such as symbiotic bacteria and parasites. This raises the intriguing question as to how these different compounds arose and evolved. Based on taxonomy and molecular analysis of gene expression patterns it is possible to deduce a putative sequence of acquisition of the different branches of the biosynthetic pathway and their regulators. (author)
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The flavonoid biosynthetic pathway in plants: function and evolution
Energy Technology Data Exchange (ETDEWEB)
Koes, R. E.; Quattrocchio, F.; Mol, J. N.M. [Department of Genetics, Institute for Molecular Biological Sciences, Vrije Universiteit, BioCentrum Amsterdam, De Boelelaan 1087, 1081HV, Amsterdam (Netherlands)
1994-07-01
Flavonoids are a class of low molecular weight phenolic compounds that is widely distributed in the plant kingdom. They exhibit a diverse spectrum of biological functions and play an important role in the interaction between plants and their environment. Flavonoids not only protect the plant from the harmful effects of UV irradiation but also play a crucial role in the sexual reproduction process. A special class of flavonoid polymers, the tannins, plays a structural role in the plant. Yet other classes of flavonoids, flavonols and anthocyanins, have been implicated in the attraction of pollinators. Certain flavonoids participate in the interaction between plants and other organisms such as symbiotic bacteria and parasites. This raises the intriguing question as to how these different compounds arose and evolved. Based on taxonomy and molecular analysis of gene expression patterns it is possible to deduce a putative sequence of acquisition of the different branches of the biosynthetic pathway and their regulators. (author)
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Dietary reference values for thiamin
DEFF Research Database (Denmark)
Sjödin, Anders Mikael
2016-01-01
Following a request from the European Commission, the EFSA Panel on Dietetic Products, Nutrition and Allergies (NDA) derived dietary reference values (DRVs) for thiamin (vitamin B1). The Panel considers that data from depletion–repletion studies in adults on the amount of dietary thiamin intake...... were measured. Results from other depletion–repletion studies are in agreement with this value. The Panel agrees on the coefficient of variation of 20% used by the SCF to cover uncertainties related to distribution of thiamin requirements in the general population, and endorses the population reference...
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Namitha, Kanakapura Krishnamurthy; Archana, Surya Narayana; Negi, Pradeep Singh
2011-04-01
To study the expression pattern of carotenoid biosynthetic pathway genes, changes in their expression at different stages of maturity in tomato fruit (cv. Arka Ahuti) were investigated. The genes regulating carotenoid production were quantified by a dot blot method using a DIG (dioxigenin) labelling and detection kit. The results revealed that there was an increase in the levels of upstream genes of the carotenoid biosynthetic pathway such as 1-deoxy-d-xylulose-5-phosphate reductoisomerase (DXR), 4-hydroxy-3-methyl-but-2-enyl diphosphate reductase (Lyt B), phytoene synthase (PSY), phytoene desaturase (PDS) and ζ-carotene desaturase (ZDS) by 2-4 fold at the breaker stage as compared to leaf. The lycopene and β-carotene content was analyzed by HPLC at different stages of maturity. The lycopene (15.33 ± 0.24 mg per 100 g) and β-carotene (10.37 ± 0.46 mg per 100 g) content were found to be highest at 5 days post-breaker and 10 days post-breaker stage, respectively. The lycopene accumulation pattern also coincided with the color values at different stages of maturity. These studies may provide insight into devising gene-based strategies for enhancing carotenoid accumulation in tomato fruits.
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Volvert, Marie-Laure; Seyen, Sandrine; Piette, Marie; Evrard, Brigitte; Gangolf, Marjorie; Plumier, Jean-Christophe; Bettendorff, Lucien
2008-06-12
Lipid-soluble thiamine precursors have a much higher bioavailability than genuine thiamine and therefore are more suitable for therapeutic purposes. Benfotiamine (S-benzoylthiamine O-monophosphate), an amphiphilic S-acyl thiamine derivative, prevents the progression of diabetic complications, probably by increasing tissue levels of thiamine diphosphate and so enhancing transketolase activity. As the brain is particularly sensitive to thiamine deficiency, we wanted to test whether intracellular thiamine and thiamine phosphate levels are increased in the brain after oral benfotiamine administration. Benfotiamine that is practically insoluble in water, organic solvents or oil was solubilized in 200 mM hydroxypropyl-beta-cyclodextrin and the mice received a single oral administration of 100 mg/kg. Though thiamine levels rapidly increased in blood and liver to reach a maximum after one or two hours, no significant increase was observed in the brain. When mice received a daily oral administration of benfotiamine for 14 days, thiamine derivatives were increased significantly in the liver but not in the brain, compared to control mice. In addition, incubation of cultured neuroblastoma cells with 10 muM benfotiamine did not lead to increased intracellular thiamine levels. Moreover, in thiamine-depleted neuroblastoma cells, intracellular thiamine contents increased more rapidly after addition of thiamine to the culture medium than after addition of benfotiamine for which a lag period was observed. Our results show that, though benfotiamine strongly increases thiamine levels in blood and liver, it has no significant effect in the brain. This would explain why beneficial effects of benfotiamine have only been observed in peripheral tissues, while sulbutiamine, a lipid-soluble thiamine disulfide derivative, that increases thiamine derivatives in the brain as well as in cultured cells, acts as a central nervous system drug. We propose that benfotiamine only penetrates the cells
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Long-Distance Transport of Thiamine (Vitamin B1) Is Concomitant with That of Polyamines.
Martinis, Jacopo; Gas-Pascual, Elisabet; Szydlowski, Nicolas; Crèvecoeur, Michèle; Gisler, Alexandra; Bürkle, Lukas; Fitzpatrick, Teresa B
2016-05-01
Thiamine (vitamin B1) is ubiquitous and essential for cell energy supply in all organisms as a vital metabolic cofactor, known for over a century. In plants, it is established that biosynthesis de novo is taking place predominantly in green tissues and is furthermore limited to plastids. Therefore, transport mechanisms are required to mediate the movement of this polar metabolite from source to sink tissue to activate key enzymes in cellular energy generating pathways but are currently unknown. Similar to thiamine, polyamines are an essential set of charged molecules required for diverse aspects of growth and development, the homeostasis of which necessitates long-distance transport processes that have remained elusive. Here, a yeast-based screen allowed us to identify Arabidopsis (Arabidopsis thaliana) PUT3 as a thiamine transporter. A combination of biochemical, physiological, and genetic approaches permitted us to show that PUT3 mediates phloem transport of both thiamine and polyamines. Loss of function of PUT3 demonstrated that the tissue distribution of these metabolites is altered with growth and developmental consequences. The pivotal role of PUT3 mediated thiamine and polyamine homeostasis in plants, and its importance for plant fitness is revealed through these findings. © 2016 American Society of Plant Biologists. All Rights Reserved.
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Thamm, Antje M; Li, Gengnan; Taja-Moreno, Marlene; Gerdes, Svetlana Y; de Crécy-Lagard, Valérie; Bruner, Steven D; Hanson, Andrew D
2017-07-20
The canonical kinase (ThiD) that converts the thiamin biosynthesis intermediate hydroxymethylpyrimidine (HMP) monophosphate to the diphosphate can also very efficiently convert free HMP to the monophosphate in prokaryotes, plants, and fungi. This HMP kinase activity enables salvage of HMP, but it is not substrate-specific and so allows toxic HMP analogs and damage products to infiltrate the thiamin biosynthesis pathway. Comparative analysis of bacterial genomes uncovered a gene, thiD2 , that is often fused to the thiamin synthesis gene thiE and could potentially encode a replacement for ThiD. Standalone ThiD2 proteins and ThiD2 fusion domains are small (~130-residues) and do not belong to any previously known protein family. Genetic and biochemical analyses showed that representative standalone and fused ThiD2 proteins catalyze phosphorylation of HMP monophosphate, but not of HMP or its toxic analogs and damage products such as bacimethrin and 5-(hydroxymethyl)-2-methylpyrimidin-4-ol. As strictly monofunctional HMP monophosphate kinases, ThiD2 proteins eliminate a potentially fatal vulnerability of canonical ThiD, at the cost of the ability to reclaim HMP formed by thiamin turnover. ©2017 The Author(s).
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The role of thiamine in neurodegenerative diseases
Directory of Open Access Journals (Sweden)
Irena Bubko
2015-09-01
Full Text Available Vitamin B1 (thiamine plays an important role in metabolism. It is indispensable for normal growth and development of the organism. Thiamine has a favourable impact on a number of systems, including the digestive, cardiovascular and nervous systems. It also stimulates the brain and improves the psycho-emotional state. Hence it is often called the vitamin of “reassurance of the spirit”. Thiamine is a water-soluble vitamin. It can be present in the free form as thiamine or as its phosphate esters: mono-, di- or triphosphate. The main source of thiamine as an exogenous vitamin is certain foodstuffs, but trace amounts can be synthesised by microorganisms of the large intestine. The recommended daily intake of thiamine is about 2.0 mg. Since vitamin B1 has no ability to accumulate in the organism, manifestations of its deficiency begin to appear very quickly. The chronic state of thiamine deficiency, to a large extent, because of its function, contributes to the development of neurodegenerative diseases. It was proved that supporting vitamin B1 therapy not only constitutes neuroprotection but can also have a favourable impact on advanced neurodegenerative diseases. This article presents the current state of knowledge as regards the effects of thiamine exerted through this vitamin in a number of diseases such as Parkinson’s disease, Alzheimer’s disease, Wernicke’s encephalopathy or Wernicke-Korsakoff syndrome and Huntington’s disease.
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Haushalter, Robert W; Phelan, Ryan M; Hoh, Kristina M; Su, Cindy; Wang, George; Baidoo, Edward E K; Keasling, Jay D
2017-04-05
Dicarboxylic acids are commodity chemicals used in the production of plastics, polyesters, nylons, fragrances, and medications. Bio-based routes to dicarboxylic acids are gaining attention due to environmental concerns about petroleum-based production of these compounds. Some industrial applications require dicarboxylic acids with specific carbon chain lengths, including odd-carbon species. Biosynthetic pathways involving cytochrome P450-catalyzed oxidation of fatty acids in yeast and bacteria have been reported, but these systems produce almost exclusively even-carbon species. Here we report a novel pathway to odd-carbon dicarboxylic acids directly from glucose in Escherichia coli by employing an engineered pathway combining enzymes from biotin and fatty acid synthesis. Optimization of the pathway will lead to industrial strains for the production of valuable odd-carbon diacids.
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Genetics Home Reference: thiamine-responsive megaloblastic anemia syndrome
... Thiamine-responsive megaloblastic anemia syndrome Thiamine-responsive megaloblastic anemia syndrome Printable PDF Open All Close All Enable ... the expand/collapse boxes. Description Thiamine-responsive megaloblastic anemia syndrome is a rare condition characterized by hearing ...
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Pan, Xiaohua; Xue, Fuguang; Nan, Xuemei; Tang, Zhiwen; Wang, Kun; Beckers, Yves; Jiang, Linshu; Xiong, Benhai
2017-01-01
The requirements of thiamine in adult ruminants are mainly met by ruminal bacterial synthesis, and thiamine deficiencies will occur when dairy cows overfed with high grain diet. However, there is limited knowledge with regard to the ruminal thiamine synthesis bacteria, and whether thiamine deficiency is related to the altered bacterial community by high grain diet is still unclear. To explore thiamine synthesis bacteria and the response of ruminal microbiota to high grain feeding and thiamine supplementation, six rumen-cannulated Holstein cows were randomly assigned into a replicated 3 × 3 Latin square design trial. Three treatments were control diet (CON, 20% dietary starch, DM basis), high grain diet (HG, 33.2% dietary starch, DM basis) and high grain diet supplemented with 180 mg thiamine/kg DMI (HG+T). On day 21 of each period, rumen content samples were collected at 3 h postfeeding. Ruminal thiamine concentration was detected by high performance liquid chromatography. The microbiota composition was determined using Illumina MiSeq sequencing of 16S rRNA gene. Cows receiving thiamine supplementation had greater ruminal pH value, acetate and thiamine content in the rumen. Principal coordinate analysis and similarity analysis indicated that HG feeding and thiamine supplementation caused a strong shift in bacterial composition and structure in the rumen. At the genus level, compared with CON group, the relative abundances of 19 genera were significantly changed by HG feeding. Thiamine supplementation increased the abundance of cellulolytic bacteria including Bacteroides, Ruminococcus 1, Pyramidobacter, Succinivibrio , and Ruminobacter , and their increases enhanced the fiber degradation and ruminal acetate production in HG+T group. Christensenellaceae R7, Lachnospira, Succiniclasticum , and Ruminococcaceae NK4A214 exhibited a negative response to thiamine supplementation. Moreover, correlation analysis revealed that ruminal thiamine concentration was positively
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Directory of Open Access Journals (Sweden)
Xiaohua Pan
2017-09-01
Full Text Available The requirements of thiamine in adult ruminants are mainly met by ruminal bacterial synthesis, and thiamine deficiencies will occur when dairy cows overfed with high grain diet. However, there is limited knowledge with regard to the ruminal thiamine synthesis bacteria, and whether thiamine deficiency is related to the altered bacterial community by high grain diet is still unclear. To explore thiamine synthesis bacteria and the response of ruminal microbiota to high grain feeding and thiamine supplementation, six rumen-cannulated Holstein cows were randomly assigned into a replicated 3 × 3 Latin square design trial. Three treatments were control diet (CON, 20% dietary starch, DM basis, high grain diet (HG, 33.2% dietary starch, DM basis and high grain diet supplemented with 180 mg thiamine/kg DMI (HG+T. On day 21 of each period, rumen content samples were collected at 3 h postfeeding. Ruminal thiamine concentration was detected by high performance liquid chromatography. The microbiota composition was determined using Illumina MiSeq sequencing of 16S rRNA gene. Cows receiving thiamine supplementation had greater ruminal pH value, acetate and thiamine content in the rumen. Principal coordinate analysis and similarity analysis indicated that HG feeding and thiamine supplementation caused a strong shift in bacterial composition and structure in the rumen. At the genus level, compared with CON group, the relative abundances of 19 genera were significantly changed by HG feeding. Thiamine supplementation increased the abundance of cellulolytic bacteria including Bacteroides, Ruminococcus 1, Pyramidobacter, Succinivibrio, and Ruminobacter, and their increases enhanced the fiber degradation and ruminal acetate production in HG+T group. Christensenellaceae R7, Lachnospira, Succiniclasticum, and Ruminococcaceae NK4A214 exhibited a negative response to thiamine supplementation. Moreover, correlation analysis revealed that ruminal thiamine concentration was
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The Cremeomycin Biosynthetic Gene Cluster Encodes a Pathway for Diazo Formation.
Waldman, Abraham J; Pechersky, Yakov; Wang, Peng; Wang, Jennifer X; Balskus, Emily P
2015-10-12
Diazo groups are found in a range of natural products that possess potent biological activities. Despite longstanding interest in these metabolites, diazo group biosynthesis is not well understood, in part because of difficulties in identifying specific genes linked to diazo formation. Here we describe the discovery of the gene cluster that produces the o-diazoquinone natural product cremeomycin and its heterologous expression in Streptomyces lividans. We used stable isotope feeding experiments and in vitro characterization of biosynthetic enzymes to decipher the order of events in this pathway and establish that diazo construction involves late-stage N-N bond formation. This work represents the first successful production of a diazo-containing metabolite in a heterologous host, experimentally linking a set of genes with diazo formation. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
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DEFF Research Database (Denmark)
Ono, Hajime; Rewitz, Kim; Shinoda, Tetsu
2006-01-01
is eliminated in larvae carrying mutations in molting defective (mld), a gene encoding a nuclear zinc finger protein that is required for production of ecdysone during Drosophila larval development. Intriguingly, mld is not present in the Bombyx mori genome, and we have identified only one spook homolog in both...... Bombyx and Manduca that is expressed in both embryos and larva. These studies suggest an evolutionary split between Diptera and Lepidoptera in how the ecdysone biosynthetic pathway is regulated during development....
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Directory of Open Access Journals (Sweden)
Joon-Heum Park
2017-11-01
Full Text Available Possible crosstalk between the carotenoid and porphyrin biosynthetic pathways under photooxidative conditions was investigated by using their biosynthetic inhibitors, norflurazon (NF and oxyfluorfen (OF. High levels of protoporphyrin IX (Proto IX accumulated in rice plants treated with OF, whereas Proto IX decreased in plants treated with NF. Both NF and OF treatments resulted in greater decreases in MgProto IX, MgProto IX methyl ester, and protochlorophyllide. Activities and transcript levels of most porphyrin biosynthetic enzymes, particularly in the Mg-porphyrin branch, were greatly down-regulated in NF and OF plants. In contrast, the transcript levels of GSA, PPO1, and CHLD as well as FC2 and HO2 were up-regulated in NF-treated plants, while only moderate increases in FC2 and HO2 were observed in the early stage of OF treatment. Phytoene, antheraxanthin, and zeaxanthin showed high accumulation in NF-treated plants, whereas other carotenoid intermediates greatly decreased. Transcript levels of carotenoid biosynthetic genes, PSY1 and PDS, decreased in response to NF and OF, whereas plants in the later stage of NF treatment exhibited up-regulation of BCH and VDE as well as recovery of PDS. However, perturbed porphyrin biosynthesis by OF did not noticeably influence levels of carotenoid metabolites, regardless of the strong down-regulation of carotenoid biosynthetic genes. Both NF and OF plants appeared to provide enhanced protection against photooxidative damage, not only by scavenging of Mg-porphyrins, but also by up-regulating FC2, HO2, and Fe-chelatase, particularly with increased levels of zeaxanthin via up-regulation of BCH and VDE in NF plants. On the other hand, the up-regulation of GSA, PPO1, and CHLD under inhibition of carotenogenic flux may be derived from the necessity to recover impaired chloroplast biogenesis during photooxidative stress. Our study demonstrates that perturbations in carotenoid and porphyrin biosynthesis coordinate
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Park, Joon-Heum; Tran, Lien H; Jung, Sunyo
2017-01-01
Possible crosstalk between the carotenoid and porphyrin biosynthetic pathways under photooxidative conditions was investigated by using their biosynthetic inhibitors, norflurazon (NF) and oxyfluorfen (OF). High levels of protoporphyrin IX (Proto IX) accumulated in rice plants treated with OF, whereas Proto IX decreased in plants treated with NF. Both NF and OF treatments resulted in greater decreases in MgProto IX, MgProto IX methyl ester, and protochlorophyllide. Activities and transcript levels of most porphyrin biosynthetic enzymes, particularly in the Mg-porphyrin branch, were greatly down-regulated in NF and OF plants. In contrast, the transcript levels of GSA, PPO1 , and CHLD as well as FC2 and HO2 were up-regulated in NF-treated plants, while only moderate increases in FC2 and HO2 were observed in the early stage of OF treatment. Phytoene, antheraxanthin, and zeaxanthin showed high accumulation in NF-treated plants, whereas other carotenoid intermediates greatly decreased. Transcript levels of carotenoid biosynthetic genes, PSY1 and PDS , decreased in response to NF and OF, whereas plants in the later stage of NF treatment exhibited up-regulation of BCH and VDE as well as recovery of PDS . However, perturbed porphyrin biosynthesis by OF did not noticeably influence levels of carotenoid metabolites, regardless of the strong down-regulation of carotenoid biosynthetic genes. Both NF and OF plants appeared to provide enhanced protection against photooxidative damage, not only by scavenging of Mg - porphyrins, but also by up-regulating FC2, HO2 , and Fe-chelatase, particularly with increased levels of zeaxanthin via up-regulation of BCH and VDE in NF plants. On the other hand, the up-regulation of GSA, PPO1 , and CHLD under inhibition of carotenogenic flux may be derived from the necessity to recover impaired chloroplast biogenesis during photooxidative stress. Our study demonstrates that perturbations in carotenoid and porphyrin biosynthesis coordinate the
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Vignisse, Julie; Sambon, Margaux; Gorlova, Anna; Pavlov, Dmitrii; Caron, Nicolas; Malgrange, Brigitte; Shevtsova, Elena; Svistunov, Andrey; Anthony, Daniel C; Markova, Natalyia; Bazhenova, Natalyia; Coumans, Bernard; Lakaye, Bernard; Wins, Pierre; Strekalova, Tatyana; Bettendorff, Lucien
2017-07-01
Thiamine is essential for normal brain function and its deficiency causes metabolic impairment, specific lesions, oxidative damage and reduced adult hippocampal neurogenesis (AHN). Thiamine precursors with increased bioavailability, especially benfotiamine, exert neuroprotective effects not only for thiamine deficiency (TD), but also in mouse models of neurodegeneration. As it is known that AHN is impaired by stress in rodents, we exposed C57BL6/J mice to predator stress for 5 consecutive nights and studied the proliferation (number of Ki67-positive cells) and survival (number of BrdU-positive cells) of newborn immature neurons in the subgranular zone of the dentate gyrus. In stressed mice, the number of Ki67- and BrdU-positive cells was reduced compared to non-stressed animals. This reduction was prevented when the mice were treated (200mg/kg/day in drinking water for 20days) with thiamine or benfotiamine, that were recently found to prevent stress-induced behavioral changes and glycogen synthase kinase-3β (GSK-3β) upregulation in the CNS. Moreover, we show that thiamine and benfotiamine counteract stress-induced bodyweight loss and suppress stress-induced anxiety-like behavior. Both treatments induced a modest increase in the brain content of free thiamine while the level of thiamine diphosphate (ThDP) remained unchanged, suggesting that the beneficial effects observed are not linked to the role of this coenzyme in energy metabolism. Predator stress increased hippocampal protein carbonylation, an indicator of oxidative stress. This effect was antagonized by both thiamine and benfotiamine. Moreover, using cultured mouse neuroblastoma cells, we show that in particular benfotiamine protects against paraquat-induced oxidative stress. We therefore hypothesize that thiamine compounds may act by boosting anti-oxidant cellular defenses, by a mechanism that still remains to be unveiled. Our study demonstrates, for the first time, that thiamine and benfotiamine prevent
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Portari, Guilherme Vannucchi; Vannucchi, Helio; Jordao, Alceu Afonso
2013-03-12
Thiamine and benfotiamine are vitamin B1 and pro-vitamin B1 substances, respectively. Vitamin B1 plays an essential role in energy metabolism, and its deficiency leads to neurologic and cardiovascular pathologies, as seen in alcoholics. This study presents new data about the effects of thiamine hydrochloride or benfotiamine treatment given to rats with acute alcohol intoxication, on the distribution of thiamine and its phosphate esters in liver, plasma and erythrocytes. The treatments were effective in increasing thiamine levels in plasma, erythrocytes and liver cells. The benfotiamine-treated group had its total plasma thiamine increased by 100%. In erythrocytes, thiamine levels were 4- and 25-fold higher in the groups treated with thiamine and benfotiamine, respectively, compared with the untreated groups. Liver thiamine was increased by 60% in the treated groups compared with the untreated groups. Thus, we verified the high bioavailability especially of benfotiamine within 6h of ethanol administration. Copyright © 2013 Elsevier B.V. All rights reserved.
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Reconstruction of cytosolic fumaric acid biosynthetic pathways in Saccharomyces cerevisiae
Directory of Open Access Journals (Sweden)
Xu Guoqiang
2012-02-01
Full Text Available Abstract Background Fumaric acid is a commercially important component of foodstuffs, pharmaceuticals and industrial materials, yet the current methods of production are unsustainable and ecologically destructive. Results In this study, the fumarate biosynthetic pathway involving reductive reactions of the tricarboxylic acid cycle was exogenously introduced in S. cerevisiae by a series of simple genetic modifications. First, the Rhizopus oryzae genes for malate dehydrogenase (RoMDH and fumarase (RoFUM1 were heterologously expressed. Then, expression of the endogenous pyruvate carboxylase (PYC2 was up-regulated. The resultant yeast strain, FMME-001 ↑PYC2 + ↑RoMDH, was capable of producing significantly higher yields of fumarate in the glucose medium (3.18 ± 0.15 g liter-1 than the control strain FMME-001 empty vector. Conclusions The results presented here provide a novel strategy for fumarate biosynthesis, which represents an important advancement in producing high yields of fumarate in a sustainable and ecologically-friendly manner.
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Biosynthetic pathway of the phytohormone auxin in insects and screening of its inhibitors.
Suzuki, Hiroyoshi; Yokokura, Junpei; Ito, Tsukasa; Arai, Ryoma; Yokoyama, Chiaki; Toshima, Hiroaki; Nagata, Shinji; Asami, Tadao; Suzuki, Yoshihito
2014-10-01
Insect galls are abnormal plant tissues induced by galling insects. The galls are used for food and habitation, and the phytohormone auxin, produced by the insects, may be involved in their formation. We found that the silkworm, a non-galling insect, also produces an active form of auxin, indole-3-acetic acid (IAA), by de novo synthesis from tryptophan (Trp). A detailed metabolic analysis of IAA using IAA synthetic enzymes from silkworms indicated an IAA biosynthetic pathway composed of a three-step conversion: Trp → indole-3-acetaldoxime → indole-3-acetaldehyde (IAAld) → IAA, of which the first step is limiting IAA production. This pathway was shown to also operate in gall-inducing sawfly. Screening of a chemical library identified two compounds that showed strong inhibitory activities on the conversion step IAAld → IAA. The inhibitors can be efficiently used to demonstrate the importance of insect-synthesized auxin in gall formation in the future. Copyright © 2014 Elsevier Ltd. All rights reserved.
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Directory of Open Access Journals (Sweden)
Michael T Guarnieri
Full Text Available Biofuels derived from algal lipids represent an opportunity to dramatically impact the global energy demand for transportation fuels. Systems biology analyses of oleaginous algae could greatly accelerate the commercialization of algal-derived biofuels by elucidating the key components involved in lipid productivity and leading to the initiation of hypothesis-driven strain-improvement strategies. However, higher-level systems biology analyses, such as transcriptomics and proteomics, are highly dependent upon available genomic sequence data, and the lack of these data has hindered the pursuit of such analyses for many oleaginous microalgae. In order to examine the triacylglycerol biosynthetic pathway in the unsequenced oleaginous microalga, Chlorella vulgaris, we have established a strategy with which to bypass the necessity for genomic sequence information by using the transcriptome as a guide. Our results indicate an upregulation of both fatty acid and triacylglycerol biosynthetic machinery under oil-accumulating conditions, and demonstrate the utility of a de novo assembled transcriptome as a search model for proteomic analysis of an unsequenced microalga.
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Genetics Home Reference: biotin-thiamine-responsive basal ganglia disease
... BTBGD thiamine metabolism dysfunction syndrome 2 thiamine-responsive encephalopathy thiamine transporter-2 deficiency THMD2 Related Information How ... genetic testing? What is precision medicine? What is newborn screening? New Pages LMNA-related congenital muscular dystrophy ...
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Lee, Won-Heong; Shin, So-Yeon; Kim, Myoung-Dong; Han, Nam Soo; Seo, Jin-Ho
2012-03-01
Guanosine 5'-triphosphate (GTP) is the key substrate for biosynthesis of guanosine 5'-diphosphate (GDP)-L-fucose. In this study, improvement of GDP-L-fucose production was attempted by manipulating the biosynthetic pathway for guanosine nucleotides in recombinant Escherichia coli-producing GDP-L-fucose. The effects of overexpression of inosine 5'-monophosphate (IMP) dehydrogenase, guanosine 5'-monophosphate (GMP) synthetase (GuaB and GuaA), GMP reductase (GuaC) and guanosine-inosine kinase (Gsk) on GDP-L-fucose production were investigated in a series of fed-batch fermentations. Among the enzymes tested, overexpression of Gsk led to a significant improvement of GDP-L-fucose production. Maximum GDP-L-fucose concentration of 305.5 ± 5.3 mg l(-1) was obtained in the pH-stat fed-batch fermentation of recombinant E. coli-overexpressing Gsk, which corresponds to a 58% enhancement in the GDP-L-fucose production compared with the control strain overexpressing GDP-L-fucose biosynthetic enzymes. Such an enhancement of GDP-L-fucose production could be due to the increase in the intracellular level of GMP.
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Beriberi (thiamine deficiency and high infant mortality in northern Laos.
Directory of Open Access Journals (Sweden)
Hubert Barennes
2015-03-01
Full Text Available Infantile beriberi (thiamine deficiency occurs mainly in infants breastfed by mothers with inadequate intake of thiamine, typically among vulnerable populations. We describe possible and probable cases of infantile thiamine deficiency in northern Laos.Three surveys were conducted in Luang Namtha Province. First, we performed a retrospective survey of all infants with a diagnosis of thiamine deficiency admitted to the 5 hospitals in the province (2007-2009. Second, we prospectively recorded all infants with cardiac failure at Luang Namtha Hospital. Third, we further investigated all mothers with infants (1-6 months living in 22 villages of the thiamine deficiency patients' origin. We performed a cross-sectional survey of all mothers and infants using a pre-tested questionnaire, physical examination and squat test. Infant mortality was estimated by verbal autopsy. From March to June 2010, four suspected infants with thiamine deficiency were admitted to Luang Namtha Provincial hospital. All recovered after parenteral thiamine injection. Between 2007 and 2009, 54 infants with possible/probable thiamine deficiency were diagnosed with acute severe cardiac failure, 49 (90.2% were cured after parenteral thiamine; three died (5.6%. In the 22 villages, of 468 live born infants, 50 (10.6%, 95% CI: 8.0-13.8 died during the first year. A peak of mortality (36 deaths was reported between 1 and 3 months. Verbal autopsy suggested that 17 deaths (3.6% were due to suspected infantile thiamine deficiency. Of 127 mothers, 60 (47.2% reported edema and paresthesia as well as a positive squat test during pregnancy; 125 (98.4% respected post-partum food avoidance and all ate polished rice. Of 127 infants, 2 (1.6% had probable thiamine deficiency, and 8 (6.8% possible thiamine deficiency.Thiamine deficiency may be a major cause of infant mortality among ethnic groups in northern Laos. Mothers' and children's symptoms are compatible with thiamine deficiency. The severity
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Directory of Open Access Journals (Sweden)
Tomáš Pluskal
Full Text Available Ergothioneine is a small, sulfur-containing metabolite (229 Da synthesized by various species of bacteria and fungi, which can accumulate to millimolar levels in tissues or cells (e.g. erythrocytes of higher eukaryotes. It is commonly marketed as a dietary supplement due to its proposed protective and antioxidative functions. In this study we report the genes forming the two-step ergothioneine biosynthetic pathway in the fission yeast, Schizosaccharomyces pombe. We identified the first gene, egt1+ (SPBC1604.01, by sequence homology to previously published genes from Neurospora crassa and Mycobacterium smegmatis. We showed, using metabolomic analysis, that the Δegt1 deletion mutant completely lacked ergothioneine and its precursors (trimethyl histidine/hercynine and hercynylcysteine sulfoxide. Since the second step of ergothioneine biosynthesis has not been characterized in eukaryotes, we examined four putative homologs (Nfs1/SPBC21D10.11c, SPAC11D3.10, SPCC777.03c, and SPBC660.12c of the corresponding mycobacterial enzyme EgtE. Among deletion mutants of these genes, only one (ΔSPBC660.12c, designated Δegt2 showed a substantial decrease in ergothioneine, accompanied by accumulation of its immediate precursor, hercynylcysteine sulfoxide. Ergothioneine-deficient strains exhibited no phenotypic defects during vegetative growth or quiescence. To effectively study the role of ergothioneine, we constructed an egt1+ overexpression system by replacing its native promoter with the nmt1+ promoter, which is inducible in the absence of thiamine. We employed three versions of the nmt1 promoter with increasing strength of expression and confirmed corresponding accumulations of ergothioneine. We quantified the intracellular concentration of ergothioneine in S. pombe (0.3, 157.4, 41.6, and up to 1606.3 µM in vegetative, nitrogen-starved, glucose-starved, and egt1+-overexpressing cells, respectively and described its gradual accumulation under long
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Xie, Feifan; Cheng, Zeneng; Li, Sanwang; Liu, Xingling; Guo, Xin; Yu, Peng; Gu, Zhenkun
2014-06-01
Benfotiamine is a lipid-soluble thiamine precursor which can transform to thiamine in vivo and subsequently be metabolized to thiamine monophosphate (TMP) and thiamine diphosphate (TDP). This study investigated the pharmacokinetic profiles of thiamine and its phosphorylated metabolites after single- and multiple-dose administration of benfotiamine in healthy Chinese volunteers, and assessed the bioavailability of orally benfotiamine administration compared to thiamine hydrochloride. In addition, concentration of hippuric acid in urine which is produced in the transformation process of benfotiamine was determined. The results showed that thiamine and its phosphorylated metabolites exhibited different pharmacokinetic characteristics in plasma, blood and erythrocyte, and one-compartment model provided the best fit for pharmacokinetic profiles of thiamine. The transformation process of benfotiamine to thiamine produced large amount of hippuric acid. No accumulation of hippuric acid was observed after multiple-dose of benfotiamine. Compared to thiamine hydrochloride, the bioavailability of thiamine in plasma and TDP in erythrocyte after oral administration of benfotiamine were 1147.3 ± 490.3% and 195.8 ± 33.8%, respectively. The absorption rate and extent of benfotiamine systemic availability of thiamine were significantly increased indicating higher bioavailability of thiamine from oral dose of benfotiamine compared to oral dose of thiamine hydrochloride. © 2014, The American College of Clinical Pharmacology.
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Decoding Biosynthetic Pathways in Plants by Pulse-Chase Strategies Using 13CO2 as a Universal Tracer
Directory of Open Access Journals (Sweden)
Adelbert Bacher
2016-07-01
Full Text Available 13CO2 pulse-chase experiments monitored by high-resolution NMR spectroscopy and mass spectrometry can provide 13C-isotopologue compositions in biosynthetic products. Experiments with a variety of plant species have documented that the isotopologue profiles generated with 13CO2 pulse-chase labeling are directly comparable to those that can be generated by the application of [U-13C6]glucose to aseptically growing plants. However, the application of the 13CO2 labeling technology is not subject to the experimental limitations that one has to take into account for experiments with [U-13C6]glucose and can be applied to plants growing under physiological conditions, even in the field. In practical terms, the results of biosynthetic studies with 13CO2 consist of the detection of pairs, triples and occasionally quadruples of 13C atoms that have been jointly contributed to the target metabolite, at an abundance that is well above the stochastic occurrence of such multiples. Notably, the connectivities of jointly transferred 13C multiples can have undergone modification by skeletal rearrangements that can be diagnosed from the isotopologue data. As shown by the examples presented in this review article, the approach turns out to be powerful in decoding the carbon topology of even complex biosynthetic pathways.
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Minimum Information about a Biosynthetic Gene cluster : commentary
Medema, Marnix H; Kottmann, Renzo; Yilmaz, Pelin; Cummings, Matthew; Biggins, John B; Blin, Kai; de Bruijn, Irene; Chooi, Yit Heng; Claesen, Jan; Coates, R Cameron; Cruz-Morales, Pablo; Duddela, Srikanth; Dusterhus, Stephanie; Edwards, Daniel J; Fewer, David P; Garg, Neha; Geiger, Christoph; Gomez-Escribano, Juan Pablo; Greule, Anja; Hadjithomas, Michalis; Haines, Anthony S; Helfrich, Eric J N; Hillwig, Matthew L; Ishida, Keishi; Jones, Adam C; Jones, Carla S; Jungmann, Katrin; Kegler, Carsten; Kim, Hyun Uk; Kotter, Peter; Krug, Daniel; Masschelein, Joleen; Melnik, Alexey V; Mantovani, Simone M; Monroe, Emily A; Moore, Marcus; Moss, Nathan; Nutzmann, Hans-Wilhelm; Pan, Guohui; Pati, Amrita; Petras, Daniel; Reen, F Jerry; Rosconi, Federico; Rui, Zhe; Tian, Zhenhua; Tobias, Nicholas J; Tsunematsu, Yuta; Wiemann, Philipp; Wyckoff, Elizabeth; Yan, Xiaohui; Yim, Grace; Yu, Fengan; Xie, Yunchang; Aigle, Bertrand; Apel, Alexander K; Balibar, Carl J; Balskus, Emily P; Barona-Gomez, Francisco; Bechthold, Andreas; Bode, Helge B; Borriss, Rainer; Brady, Sean F; Brakhage, Axel A; Caffrey, Patrick; Cheng, Yi-Qiang; Clardy, Jon; Cox, Russell J; De Mot, Rene; Donadio, Stefano; Donia, Mohamed S; van der Donk, Wilfred A; Dorrestein, Pieter C; Doyle, Sean; Driessen, Arnold J M; Ehling-Schulz, Monika; Entian, Karl-Dieter; Fischbach, Michael A; Gerwick, Lena; Gerwick, William H; Gross, Harald; Gust, Bertolt; Hertweck, Christian; Hofte, Monica; Jensen, Susan E; Ju, Jianhua; Katz, Leonard; Kaysser, Leonard; Klassen, Jonathan L; Keller, Nancy P; Kormanec, Jan; Kuipers, Oscar P; Kuzuyama, Tomohisa; Kyrpides, Nikos C; Kwon, Hyung-Jin; Lautru, Sylvie; Lavigne, Rob; Lee, Chia Y; Linquan, Bai; Liu, Xinyu; Liu, Wen; Luzhetskyy, Andriy; Mahmud, Taifo; Mast, Yvonne; Mendez, Carmen; Metsa-Ketela, Mikko; Micklefield, Jason; Mitchell, Douglas A; Moore, Bradley S; Moreira, Leonilde M; Muller, Rolf; Neilan, Brett A; Nett, Markus; Nielsen, Jens; O'Gara, Fergal; Oikawa, Hideaki; Osbourn, Anne; Osburne, Marcia S; Ostash, Bohdan; Payne, Shelley M; Pernodet, Jean-Luc; Petricek, Miroslav; Piel, Jorn; Ploux, Olivier; Raaijmakers, Jos M; Salas, Jose A; Schmitt, Esther K; Scott, Barry; Seipke, Ryan F; Shen, Ben; Sherman, David H; Sivonen, Kaarina; Smanski, Michael J; Sosio, Margherita; Stegmann, Evi; Sussmuth, Roderich D; Tahlan, Kapil; Thomas, Christopher M; Tang, Yi; Truman, Andrew W; Viaud, Muriel; Walton, Jonathan D; Walsh, Christopher T; Weber, Tilmann; van Wezel, Gilles P; Wilkinson, Barrie; Willey, Joanne M; Wohlleben, Wolfgang; Wright, Gerard D; Ziemert, Nadine; Zhang, Changsheng; Zotchev, Sergey B; Breitling, Rainer; Takano, Eriko; Glockner, Frank Oliver
A wide variety of enzymatic pathways that produce specialized metabolites in bacteria, fungi and plants are known to be encoded in biosynthetic gene clusters. Information about these clusters, pathways and metabolites is currently dispersed throughout the literature, making it difficult to exploit.
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Chen, Xuelan; Tang, Li; Jiao, Haitao; Xu, Feng; Xiong, Yonghua
2013-01-04
ArgR, coded by the argR gene from Corynebacterium crenatum AS 1.542, acts as a negative regulator in arginine biosynthetic pathway. However, the effect of argR on transcriptional levels of the related biosynthetic genes has not been reported. Here, we constructed a deletion mutant of argR gene: C. crenatum AS 1.542 Delta argR using marker-less knockout technology, and compared the changes of transcriptional levels of the arginine biosynthetic genes between the mutant strain and the wild-type strain. We used marker-less knockout technology to construct C. crenatum AS 1.542 Delta argR and analyzed the changes of the relate genes at the transcriptional level using real-time fluorescence quantitative PCR. C. crenatum AS 1.542 Delta argR was successfully obtained and the transcriptional level of arginine biosynthetic genes in this mutant increased significantly with an average of about 162.1 folds. The arginine biosynthetic genes in C. crenatum are clearly controlled by the negative regulator ArgR. However, the deletion of this regulator does not result in a clear change in arginine production in the bacteria.
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Leveraging microbial biosynthetic pathways for the generation of 'drop-in' biofuels.
Zargar, Amin; Bailey, Constance B; Haushalter, Robert W; Eiben, Christopher B; Katz, Leonard; Keasling, Jay D
2017-06-01
Advances in retooling microorganisms have enabled bioproduction of 'drop-in' biofuels, fuels that are compatible with existing spark-ignition, compression-ignition, and gas-turbine engines. As the majority of petroleum consumption in the United States consists of gasoline (47%), diesel fuel and heating oil (21%), and jet fuel (8%), 'drop-in' biofuels that replace these petrochemical sources are particularly attractive. In this review, we discuss the application of aldehyde decarbonylases to produce gasoline substitutes from fatty acid products, a recently crystallized reductase that could hydrogenate jet fuel precursors from terpene synthases, and the exquisite control of polyketide synthases to produce biofuels with desired physical properties (e.g., lower freezing points). With our increased understanding of biosynthetic logic of metabolic pathways, we discuss the unique advantages of fatty acid, terpene, and polyketide synthases for the production of bio-based gasoline, diesel and jet fuel. Copyright © 2017 Elsevier Ltd. All rights reserved.
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Tabarki, Brahim; Alfadhel, Majid; AlShahwan, Saad; Hundallah, Khaled; AlShafi, Shatha; AlHashem, Amel
2015-09-01
To compare the combination of biotin plus thiamine to thiamine alone in treating patients with biotin-responsive basal ganglia disease in an open-label prospective, comparative study. twenty patients with genetically proven biotin-responsive basal ganglia disease were enrolled, and received for at least 30 months a combination of biotin plus thiamine or thiamine alone. The outcome measures included duration of the crisis, number of recurrence/admissions, the last neurological examination, the severity of dystonia using the Burke-Fahn-Marsden Dystonia Rating Scale (BFMDRS), and the brain MRI findings during the crisis and after 30 months of follow-up. Ten children with a mean age of 6 years(1/2) were recruited in the biotin plus thiamine group (group 1) and ten children (6 females and 4 males) with a mean age of 6 years and 2 months were recruited in the thiamine group (group 2). After 2 years of follow-up treatment, 6 of 20 children achieved complete remission, 10 had minimal sequelae in the form of mild dystonia and dysarthria (improvement of the BFMDRS, mean: 80%), and 4 had severe neurologic sequelae. All these 4 patients had delayed diagnosis and management. Regarding outcome measures, both groups have a similar outcome regarding the number of recurrences, the neurologic sequelae (mean BFMDS score between the groups, p = 0.84), and the brain MRI findings. The only difference was the duration of the acute crisis: group 1 had faster recovery (2 days), versus 3 days in group 2 (p = 0.005). Our study suggests that over 30 months of treatment, the combination of biotin plus thiamine is not superior to thiamine alone in the treatment of biotin-responsive basal ganglia disease. Copyright © 2015 European Paediatric Neurology Society. Published by Elsevier Ltd. All rights reserved.
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Beltramo, E; Berrone, E; Buttiglieri, S; Porta, M
2004-01-01
High glucose induces pathological alterations in small and large vessels, possibly through increased formation of AGE, activation of aldose reductase and protein kinase C, and increased flux through the hexosamine pathway. We showed previously that thiamine and benfotiamine correct delayed replication and increase lactate production in endothelial cells subjected to high glucose. We now aim at verifying the effects of thiamine and benfotiamine on cell cycle, apoptosis, and expression of adhesion molecules in endothelial cells and pericytes, under high ambient glucose. Human umbilical vein endothelial cells and bovine retinal pericytes were cultured in normal (5.6 mmol/L) or high (28 mmol/L) glucose, with or without thiamine or benfotiamine, 50 or 100 micro mol/L. Apoptosis was determined by two separate ELISA methods, measuring DNA fragmentation and caspase-3 activity, respectively. Cell cycle and integrin subunits alpha3, alpha5, and beta1 concentration were measured by flow cytometry. Apoptosis was increased in high glucose after 3 days of culture, both in endothelium and pericytes. Thiamine and benfotiamine reversed such effects. Neither cell cycle traversal nor integrin concentrations were modified in these experimental conditions. Thiamine and benfotiamine correct increased apoptosis due to high glucose in cultured vascular cells. Further elucidations of the mechanisms through which they work could help set the basis for clinical use of this vitamin in the prevention and/or treatment of diabetic microangiopathy. Copyright 2004 John Wiley & Sons, Ltd.
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Enhancement of Thiamin Content in Arabidopsis thaliana by Metabolic Engineering.
Dong, Wei; Stockwell, Virginia O; Goyer, Aymeric
2015-12-01
Thiamin is an essential nutrient in the human diet. Severe thiamin deficiency leads to beriberi, a lethal disease which is common in developing countries. Thiamin biofortification of staple food crops is a possible strategy to alleviate thiamin deficiency-related diseases. In plants, thiamin plays a role in the response to abiotic and biotic stresses, and data from the literature suggest that boosting thiamin content could increase resistance to stresses. Here, we tested an engineering strategy to increase thiamin content in Arabidopsis. Thiamin is composed of a thiazole ring linked to a pyrimidine ring by a methylene bridge. THI1 and THIC are the first committed steps in the synthesis of the thiazole and pyrimidine moieties, respectively. Arabidopsis plants were transformed with a vector containing the THI1-coding sequence under the control of a constitutive promoter. Total thiamin leaf content in THI1 plants was up approximately 2-fold compared with the wild type. THI1-overexpressing lines were then crossed with pre-existing THIC-overexpressing lines. Resulting THI1 × THIC plants accumulated up to 3.4- and 2.6-fold more total thiamin than wild-type plants in leaf and seeds, respectively. After inoculation with Pseudomonas syringae, THI1 × THIC plants had lower populations than the wild-type control. However, THI1 × THIC plants subjected to various abiotic stresses did not show any visible or biochemical changes compared with the wild type. We discuss the impact of engineering thiamin biosynthesis on the nutritional value of plants and their resistance to biotic and abiotic stresses. © The Author 2015. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.
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The relationship between lactate and thiamine levels in patients with diabetic ketoacidosis.
Moskowitz, Ari; Graver, Amanda; Giberson, Tyler; Berg, Katherine; Liu, Xiaowen; Uber, Amy; Gautam, Shiva; Donnino, Michael W
2014-02-01
Thiamine functions as an important cofactor in aerobic metabolism and thiamine deficiency can contribute to lactic acidosis. Although increased rates of thiamine deficiency have been described in diabetic outpatients, this phenomenon has not been studied in relation to diabetic ketoacidosis (DKA). In the present study, we hypothesize that thiamine deficiency is associated with elevated lactate in patients with DKA. This was a prospective observational study of patients presenting to a tertiary care center with DKA. Patient demographics, laboratory results, and outcomes were recorded. A one-time blood draw was performed and analyzed for plasma thiamine levels. Thirty-two patients were enrolled. Eight patients (25%) were thiamine deficient, with levels lower than 9 nmol/L. A negative correlation between lactic acid and plasma thiamine levels was found (r = -0.56, P = .002). This relationship remained significant after adjustment for APACHE II scores (P = .009). Thiamine levels were directly related to admission serum bicarbonate (r = 0.44, P = .019), and patients with thiamine deficiency maintained lower bicarbonate levels over the first 24 hours (slopes parallel with a difference of 4.083, P = .002). Patients with DKA had a high prevalence of thiamine deficiency. Thiamine levels were inversely related to lactate levels among patients with DKA. A study of thiamine supplementation in DKA is warranted. © 2013.
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Sato, Akiko; Shimoyama, Yoshimitsu; Ishikawa, Tomoji; Murayama, Nobuko
2011-06-01
The purpose of this study was to examine the effect of high-intensity physical activity during training on the biochemical status of thiamin and riboflavin in athletes. Thiamin and riboflavin concentrations in whole blood of a group of 19 athletes (6 men and 13 women) were measured during a low-intensity preparatory period and compared with measurements taken during a high-intensity training period. Additional variables measured included anthropometric characteristics, estimated energy expenditure during swim training, distance covered, resting energy expenditure obtained by indirect calorimetry, estimated energy requirement per day, and dietary intake of energy, thiamin, and riboflavin estimated from 3-day food records. For both male and female subjects, no major changes were observed in anthropometric characteristics or dietary intake, but energy expenditure during swim training per day significantly increased in the intensive-training period (496 ± 0 kcal in the preparation period compared with 995 ± 96 kcal in the intensive-training period for male subjects [p riboflavin was unchanged. These results suggest that intense training affects thiamin concentration, but not riboflavin concentration, in the whole blood of college swimmers.
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The relationship between lactate and thiamine levels in patients with diabetic ketoacidosis⋆
Moskowitz, Ari; Graver, Amanda; Giberson, Tyler; Berg, Katherine; Liu, Xiaowen; Uber, Amy; Gautam, Shiva; Donnino, Michael W.
2013-01-01
Purpose Thiamine functions as an important cofactor in aerobic metabolism and thiamine deficiency can contribute to lactic acidosis. Although increased rates of thiamine deficiency have been described in diabetic outpatients, this phenomenon has not been studied in relation to diabetic ketoacidosis (DKA). In the present study, we hypothesize that thiamine deficiency is associated with elevated lactate in patients with DKA. Materials and Methods This was a prospective observational study of patients presenting to a tertiary care center with DKA. Patient demographics, laboratory results, and outcomes were recorded. A one-time blood draw was performed and analyzed for plasma thiamine levels. Results Thirty-two patients were enrolled. Eight patients (25%) were thiamine deficient, with levels lower than 9 nmol/L. A negative correlation between lactic acid and plasma thiamine levels was found (r = −0.56, P = .002). This relationship remained significant after adjustment for APACHE II scores (P = .009). Thiamine levels were directly related to admission serum bicarbonate (r = 0.44, P = .019), and patients with thiamine deficiency maintained lower bicarbonate levels over the first 24 hours (slopes parallel with a difference of 4.083, P = .002). Conclusions Patients with DKA had a high prevalence of thiamine deficiency. Thiamine levels were inversely related to lactate levels among patients with DKA. A study of thiamine supplementation in DKA is warranted. PMID:23993771
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Energy Technology Data Exchange (ETDEWEB)
Haushalter, Robert W. [Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Physical Bioscience Division; Phelan, Ryan M. [Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Physical Bioscience Division; Hoh, Kristina M. [Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Physical Bioscience Division; Su, Cindy [Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Physical Bioscience Division; Wang, George [Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Physical Bioscience Division; Baidoo, Edward E. K. [Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Physical Bioscience Division; Keasling, Jay D. [Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Physical Bioscience Division
2017-03-14
Dicarboxylic acids are commodity chemicals used in the production of plastics, polyesters, nylons, fragrances, and medications. Bio-based routes to dicarboxylic acids are gaining attention due to environmental concerns about petroleum-based production of these compounds. Some industrial applications require dicarboxylic acids with specific carbon chain lengths, including odd-carbon species. Biosynthetic pathways involving cytochrome P450-catalyzed oxidation of fatty acids in yeast and bacteria have been reported, but these systems produce almost exclusively even-carbon species. Here in this paper we report a novel pathway to odd-carbon dicarboxylic acids directly from glucose in Escherichia coli by employing an engineered pathway combining enzymes from biotin and fatty acid synthesis. Optimization of the pathway will lead to industrial strains for the production of valuable odd-carbon diacids.
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Analysis of thiamine concentrations in commercial canned foods formulated for cats.
Markovich, Jessica E; Freeman, Lisa M; Heinze, Cailin R
2014-01-15
To measure thiamine concentrations in commercial canned foods formulated for cats as an initial assessment of the variation among canned foods and to determine the effects of flavor (fish vs nonfish) of the food, texture (paté vs nonpaté) of the food, country of manufacture, and size of the company on thiamine concentration. Prospective cross-sectional study. 90 canned, nontherapeutic diets formulated for cats (1 fish and 1 nonfish flavor for each of 45 brands). Each canned food was homogenized, and thiamine concentration was analyzed with a fluorometric method. Thiamine concentration was below the minimums of the Association of American Feed Control Officials in 12 of 90 (13.3%) foods and below the recommended allowance of the National Research Council in 14 of 90 (15.6%) foods. Paté foods had significantly lower thiamine concentrations than did nonpaté foods, and foods from smaller companies had significantly lower thiamine concentrations, compared with concentrations in foods from larger companies. Flavor of food and country of manufacture were not significantly associated with thiamine concentration. A wide range of thiamine concentrations was found in the foods evaluated. Thiamine concentration in a substantial percentage of commercially available canned foods was below the amount recommended for adult cats. Additional research on interlot and intralot variation in thiamine concentrations of foods formulated for cats is warranted. Companies should implement strict quality control and analysis practices regarding food products. Clinicians should consider thiamine deficiency as a differential diagnosis in a cat with acute neurologic dysfunction.
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Serum thiamine level during the initiation term of hemodialysis
Directory of Open Access Journals (Sweden)
Shuji Ono
2012-06-01
Although water-soluble vitamins such as thiamine are lost during dialysis session, serum thiamine level tended to restore spontaneously during the hospitalization (25.47±10.07 vs. 27.44±11.37, pre HD value, p=0.09. A three months-follow up measurement after initiation of HD, which is now in progress (so far n=3, revealed increase of serum thiamine level in all patients (35.33±7.37 ng/ml. It may reflect the improvement of nutritional status after the initiation of HD, and suggests the loss of thiamine by HD procedure can be covered.
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Clinically unapparent infantile thiamin deficiency in Vientiane, Laos.
Directory of Open Access Journals (Sweden)
Sengmanivong Khounnorath
2011-02-01
Full Text Available Beriberi occurs in Vientiane, Lao PDR, among breastfed infants. Clinical disease may be the tip of an iceberg with subclinical thiamin deficiency contributing to other illnesses. Thiamin treatment could improve outcome.A cohort of 778 sick infants admitted during one year without clinical evidence of beriberi were studied prospectively and erythrocyte transketolase assays (ETK performed. Biochemical thiamin deficiency was defined both in terms of the activation coefficient (α>31% and basal ETK activity 31% and 13.4 % basal ETK<0.59 micromoles/min/gHb. Infants≥2 months old had a higher frequency of biochemical markers of thiamin deficiency. Mortality was 5.5% but, among infants ≥2 months old, mortality was higher in those with basal ETK<0.59 micromoles/min/gHb (3/47, 6.4% than in those with basal ETK≥0.59 micromoles/min/gHb (1/146, 0.7% (P=0.045, relative risk=9.32 (95%CI 0.99 to 87.5. Multivariate regression analysis indicated that infant age≥2 months and fewer maternal years of schooling were independently associated with infant basal ETK<0.59 micromoles/min/gHb.Clinically unapparent thiamin deficiency is common among sick infants (≥2 months old admitted to hospital in Vientiane. This may contribute to mortality and a low clinical threshold for providing thiamin to sick infants may be needed.
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The Role of Thiamine and Effects of Deficiency in Dogs and Cats
Directory of Open Access Journals (Sweden)
Georgia Kritikos
2017-11-01
Full Text Available Recent pet food recalls for insufficient dietary thiamine have highlighted the importance of adequate thiamine intake in dogs and cats, as thiamine is an essential dietary nutrient with a critical role in energy metabolism. Prolonged thiamine deficiency leads to clinical signs that can span several organ systems, and deficiency can be fatal if not reversed. In this review, the current knowledge of thiamine metabolism will be summarized. Dietary recommendations for dogs and cats will be discussed, and the risk factors and clinical signs associated with thiamine deficiency will be examined.
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The Role of Thiamine and Effects of Deficiency in Dogs and Cats
Kritikos, Georgia; Verbrugghe, Adronie
2017-01-01
Recent pet food recalls for insufficient dietary thiamine have highlighted the importance of adequate thiamine intake in dogs and cats, as thiamine is an essential dietary nutrient with a critical role in energy metabolism. Prolonged thiamine deficiency leads to clinical signs that can span several organ systems, and deficiency can be fatal if not reversed. In this review, the current knowledge of thiamine metabolism will be summarized. Dietary recommendations for dogs and cats will be discussed, and the risk factors and clinical signs associated with thiamine deficiency will be examined.
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Park, Wan-Su; Lee, Jongtae; Hong, Taegon; Park, Gabjin; Youn, Sunil; Seo, Youngwhan; Lee, Sanghun; Han, Seunghoon
2016-10-01
Fursultiamine and benfotiamine are lipophilic thiamine derivatives used as oral sources of thiamine. Although there are many publications on the pharmacokinetic (PK) properties of thiamine-containing products, no direct comparisons between these agents . We aimed to compare the PK profiles of these lipophilic thiamine derivatives and to compare the extent of the increase in bioavailability to that of naïve thiamine. Two randomized, single-dose, 2-way crossover, full PK studies were conducted in healthy Korean male subjects (n = 24 per group). Among the test compounds, fursultiamine was compared with benfotiamine (reference A in study A) and thiamine nitrate (reference B in study B). All formulations were multivitamin preparations containing the test or reference formulation as the major thiamine source. In study A, the plasma and hemolysate concentrations of thiamine and its metabolites were measured, while only the plasma thiamine concentration was assayed in study B. The systemic thiamine exposure of the test compound was slightly greater than that of reference A, based on the geometric mean ratio (%) of the AUC last value for plasma (116.6%) and hemolysate (137.5%). The thiamine diphosphate (TDP) distribution between plasma and hemolysate showed clear differences according to the formulations, in that more TDP was present in the hemolysate when thiamine was given as the test formulation. The AUC last value of plasma thiamine showed a >300% increase when thiamine was given as the test formulation in study B. The summed total exposure to thiamine (thiamine + TDP in both plasma and hemolysate) observed as a point estimate after the administration of fursultiamine was slightly greater than that with benfotiamine; however, the 90% CI was within the conventional bioequivalence range. These findings support clear benefits of lipophilic thiamine derivatives in the absorption of thiamine in healthy volunteers. Clinical Research Information Service identifiers: KCT
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Honda, Kohsuke; Maya, Shohei; Omasa, Takeshi; Hirota, Ryuichi; Kuroda, Akio; Ohtake, Hisao
2010-08-02
Six thermophilic enzymes from Thermus thermophilus were used to construct an 'artificial bio-synthetic pathway' for the production of 2-deoxyribose 5-phosphate from fructose. By a simple operation using six recombinant Escherichia coli strains producing the thermophilic enzymes, respectively, fructose was converted to 2-deoxyribose 5-phosphate with a molar yield of 55%. Copyright 2010 Elsevier B.V. All rights reserved.
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Are brain and heart tissue prone to the development of thiamine deficiency?
Klooster, Astrid; Larkin, James R.; Wiersema-Buist, Janneke; Gans, Reinold O. B.; Thornalley, Paul J.; Navis, Gerjan; van Goor, Harry; Leuvenink, Henri G. D.; Bakker, Stephan J. L.
Thiamine deficiency is a continuing problem leading to beriberi and Wernicke's encephalopathy. The symptoms of thiamine deficiency develop in the heart, brain and neuronal tissue. Yet, it is unclear how rapid thiamine deficiency develops and which organs are prone to development of thiamine
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Thiamine in septic shock patients with alcohol use disorders: An observational pilot study.
Holmberg, Mathias Johan; Moskowitz, Ari; Patel, Parth Vijay; Grossestreuer, Anne Victoria; Uber, Amy; Stankovic, Nikola; Andersen, Lars Wiuff; Donnino, Michael William
2018-02-01
Alcohol-use disorders (AUDs) have been associated with increased sepsis-related mortality. As patients with AUDs are often thiamine deficient, we investigated practice patterns relating to thiamine administration in patients with AUDs presenting with septic shock and explored the association between receipt of thiamine and mortality. We performed a retrospective cohort study of patients presenting with septic shock between 2008 and 2014 at a single tertiary care center. We identified patients with an AUD diagnosis, orders for microbial cultures and use of antibiotics, vasopressor dependency, and lactate levels≥4mmol/L. We excluded those who received thiamine later than 48h of sepsis onset. We included 53 patients. Thirty-four (64%) patients received thiamine. Five patients (15%) received their first thiamine dose in the emergency department. The median time to thiamine administration was 9 (quartiles: 4, 18) hours. The first thiamine dose was most often given parenterally (68%) and for 100mg (88%). In those receiving thiamine, 15/34 (44%) died, compared to 15/19 (79%) of those not receiving thiamine, p=0.02. A considerable proportion of patients with AUDs admitted for septic shock do not receive thiamine. Thiamine administration in this patient population was associated with decreased mortality. Copyright © 2017 Elsevier Inc. All rights reserved.
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Qu, Ge; Wang, Wei; Chen, Ling-Ling; Qian, Shao-Song; Zhang, Hong-Yu
2009-10-01
Although cysteine (Cys) is beneficial to stabilize protein structures, it is not prevalent in thermophiles. For instance, the Cys contents in most thermophilic archaea are only around 0.7%. However, methanogenic archaea, no matter thermophilic or not, contain relatively abundant Cys, which remains elusive for a long time. Recently, Klipcan et al. correlated this intriguing property of methanogenic archaea with their unique tRNA-dependent Cys biosynthetic pathway. But, the deep reasons underlying the correlation are ambiguous. Considering the facts that free Cys is thermally labile and the tRNA-dependent Cys biosynthesis avoids the use of free Cys, we speculate that the unique Cys biosynthetic pathway represents a strategy to increase Cys contents by preventing it from thermal degradation, which may be relevant to the thermal adaptation of methanogenic archaea ancestor.
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Saifi, Monica; Nasrullah, Nazima; Ahmad, Malik Mobeen; Ali, Athar; Khan, Jawaid A; Abdin, M Z
2015-09-01
miRNAs are emerging as potential regulators of the gene expression. Their proven promising role in regulating biosynthetic pathways related gene networks may hold the key to understand the genetic regulation of these pathways which may assist in selection and manipulation to get high performing plant genotypes with better secondary metabolites yields and increased biomass. miRNAs associated with genes of steviol glycosides biosynthetic pathway, however, have not been identified so far. In this study miRNAs targeting genes of steviol glycosides biosynthetic pathway were identified for the first time whose precursors were potentially generated from ESTs and nucleotide sequences of Stevia rebaudiana. Thereafter, stem-loop coupled real time PCR based expressions of these miRNAs in different tissues of Stevia rebaudiana were investigated and their relationship pattern was analysed with the expression levels of their target mRNAs as well as steviol glycoside contents. All the miRNAs investigated showed differential expressions in all the three tissues studied, viz. leaves, flowers and stems. Out of the eleven miRNAs validated, the expression levels of nine miRNAs (miR319a, miR319b, miR319c, miR319d, miR319e, miR319f, miR319h, miRstv_7, miRstv_9) were found to be inversely related, while expression levels of the two, i.e. miR319g and miRstv_11 on the contrary, showed direct relation with the expression levels of their target mRNAs and steviol glycoside contents in the leaves, flowers and stems. This study provides a platform for better understanding of the steviol glycosides biosynthetic pathway and these miRNAs can further be employed to manipulate the biosynthesis of these metabolites to enhance their contents and yield in S. rebaudiana. Copyright © 2015 Elsevier Masson SAS. All rights reserved.
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Thiamin deficiency on fetal brain development with and without prenatal alcohol exposure.
Kloss, Olena; Eskin, N A Michael; Suh, Miyoung
2018-04-01
Adequate thiamin levels are crucial for optimal health through maintenance of homeostasis and viability of metabolic enzymes, which require thiamine as a co-factor. Thiamin deficiency occurs during pregnancy when the dietary intake is inadequate or excessive alcohol is consumed. Thiamin deficiency leads to brain dysfunction because thiamin is involved in the synthesis of myelin and neurotransmitters (e.g., acetylcholine, γ-aminobutyric acid, glutamate), and its deficiency increases oxidative stress by decreasing the production of reducing agents. Thiamin deficiency also leads to neural membrane dysfunction, because thiamin is a structural component of mitochondrial and synaptosomal membranes. Similarly, in-utero exposure to alcohol leads to fetal brain dysfunction, resulting in negative effects such as fetal alcohol spectrum disorder (FASD). Thiamin deficiency and prenatal exposure to alcohol could act synergistically to produce negative effects on fetal development; however, this area of research is currently under-studied. This minireview summarizes the evidence for the potential role of thiamin deficiency in fetal brain development, with or without prenatal exposure to alcohol. Such evidence may influence the development of new nutritional strategies for preventing or mitigating the symptoms of FASD.
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Evolutionary origins and functions of the carotenoid biosynthetic pathway in marine diatoms.
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Sacha Coesel
Full Text Available Carotenoids are produced by all photosynthetic organisms, where they play essential roles in light harvesting and photoprotection. The carotenoid biosynthetic pathway of diatoms is largely unstudied, but is of particular interest because these organisms have a very different evolutionary history with respect to the Plantae and are thought to be derived from an ancient secondary endosymbiosis between heterotrophic and autotrophic eukaryotes. Furthermore, diatoms have an additional xanthophyll-based cycle for dissipating excess light energy with respect to green algae and higher plants. To explore the origins and functions of the carotenoid pathway in diatoms we searched for genes encoding pathway components in the recently completed genome sequences of two marine diatoms. Consistent with the supplemental xanthophyll cycle in diatoms, we found more copies of the genes encoding violaxanthin de-epoxidase (VDE and zeaxanthin epoxidase (ZEP enzymes compared with other photosynthetic eukaryotes. However, the similarity of these enzymes with those of higher plants indicates that they had very probably diversified before the secondary endosymbiosis had occurred, implying that VDE and ZEP represent early eukaryotic innovations in the Plantae. Consequently, the diatom chromist lineage likely obtained all paralogues of ZEP and VDE genes during the process of secondary endosymbiosis by gene transfer from the nucleus of the algal endosymbiont to the host nucleus. Furthermore, the presence of a ZEP gene in Tetrahymena thermophila provides the first evidence for a secondary plastid gene encoded in a heterotrophic ciliate, providing support for the chromalveolate hypothesis. Protein domain structures and expression analyses in the pennate diatom Phaeodactylum tricornutum indicate diverse roles for the different ZEP and VDE isoforms and demonstrate that they are differentially regulated by light. These studies therefore reveal the ancient origins of several
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Evolutionary origins and functions of the carotenoid biosynthetic pathway in marine diatoms.
Coesel, Sacha; Oborník, Miroslav; Varela, Joao; Falciatore, Angela; Bowler, Chris
2008-08-06
Carotenoids are produced by all photosynthetic organisms, where they play essential roles in light harvesting and photoprotection. The carotenoid biosynthetic pathway of diatoms is largely unstudied, but is of particular interest because these organisms have a very different evolutionary history with respect to the Plantae and are thought to be derived from an ancient secondary endosymbiosis between heterotrophic and autotrophic eukaryotes. Furthermore, diatoms have an additional xanthophyll-based cycle for dissipating excess light energy with respect to green algae and higher plants. To explore the origins and functions of the carotenoid pathway in diatoms we searched for genes encoding pathway components in the recently completed genome sequences of two marine diatoms. Consistent with the supplemental xanthophyll cycle in diatoms, we found more copies of the genes encoding violaxanthin de-epoxidase (VDE) and zeaxanthin epoxidase (ZEP) enzymes compared with other photosynthetic eukaryotes. However, the similarity of these enzymes with those of higher plants indicates that they had very probably diversified before the secondary endosymbiosis had occurred, implying that VDE and ZEP represent early eukaryotic innovations in the Plantae. Consequently, the diatom chromist lineage likely obtained all paralogues of ZEP and VDE genes during the process of secondary endosymbiosis by gene transfer from the nucleus of the algal endosymbiont to the host nucleus. Furthermore, the presence of a ZEP gene in Tetrahymena thermophila provides the first evidence for a secondary plastid gene encoded in a heterotrophic ciliate, providing support for the chromalveolate hypothesis. Protein domain structures and expression analyses in the pennate diatom Phaeodactylum tricornutum indicate diverse roles for the different ZEP and VDE isoforms and demonstrate that they are differentially regulated by light. These studies therefore reveal the ancient origins of several components of the
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Thiamine deficiency and its correlation with dyslipidaemia in diabetics with microalbuminuria
International Nuclear Information System (INIS)
Waheed, P.; Naveed, A. K.; Ahmed, T.
2013-01-01
Objective: To measure and correlate the levels of thiamine and dyslipidaemia in microalbuminuric diabetics. Methods: Cross-sectional comparative study was conducted at the Department of Biochemistry and Molecular Biology, Army Medical College, Rawalpindi, from January 2009 to December 2010, and comprised 60 known diabetic patients, who were inducted from diabetic clinics of Rawalpindi. These patients were divided into three equal groups, with group I (n=20) being normal healthy individuals, group II comprised of microalbuminurics type 2 diabetics (n=20) and group III (n=20) were macroalbuminuric type 2 diabetics, based on their albumin excretion rate. The healthy volunteers (n=20) had blood glucose less than 6 mmol/L and were inducted as the comparison group. Fasting blood samples of diabetic and control groups were analysed for glucose, glycosylated haemoglobin, lipid profile, thiamine chloride and thiamine monophosphate. Besides, 24-hour urine samples were analysed for microalbuminuria, thiamine chloride and thiamine monophosphate. Results: Plasma thiamine chloride and thiamine monophosphate levels were found to be significantly (p<0.001) reduced in the diabetics (n=60) compared to the controls (n=20). Furthermore, there was a progressive decline in these levels with increasing albuminuria; the lowest being in the macroalbuminuric group (group IV). Urinary thiamine levels were significantly (p<0.001) higher in the diabetics compared to the controls. These changes were more pronounced as albuminuria level increased; the highest being in group IV. The parameters of lipid profile, including triglycerides, total cholesterol and low-density lipoprotein cholesterol, were significantly (p<0.001) higher in diabetics and showed progressive increase with worsening albuminuria. Whereas, the high-density lipoprotein cholesterol levels were significantly (p<0.001) reduced in diabetics and showed progressive decline as the microalbuminuria status worsened. Furthermore, a
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Thiamine deficiency impairs common eider (Somateria mollissima) reproduction in the field.
Mörner, Torsten; Hansson, Tomas; Carlsson, Le; Berg, Anna-Lena; Ruiz Muñoz, Yolanda; Gustavsson, Hanna; Mattsson, Roland; Balk, Lennart
2017-10-31
The Baltic Sea population of the common eider (Somateria mollissima) has declined dramatically during the last two decades. Recently, widespread episodic thiamine (vitamin B 1 ) deficiency has been demonstrated in feral birds and suggested to contribute significantly to declining populations. Here we show that the decline of the common eider population in the Baltic Sea is paralleled by high mortality of the pulli a few days after hatch, owing to thiamine deficiency and probably also thereby associated abnormal behaviour resulting in high gull predation. An experiment with artificially incubated common eider eggs collected in the field revealed that thiamine treatment of pulli had a therapeutic effect on the thiamine status of the brain and prevented death. The mortality was 53% in untreated specimens, whereas it was only 7% in thiamine treated specimens. Inability to dive was also linked to brain damage typical for thiamine deficiency. Our results demonstrate how thiamine deficiency causes a range of symptoms in the common eider pulli, as well as massive die-offs a few days after hatch, which probably are the major explanation of the recent dramatic population declines.
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Prostatic acid phosphatase is required for the antinociceptive effects of thiamine and benfotiamine.
Hurt, Julie K; Coleman, Jennifer L; Fitzpatrick, Brendan J; Taylor-Blake, Bonnie; Bridges, Arlene S; Vihko, Pirkko; Zylka, Mark J
2012-01-01
Thiamine (Vitamin B1) is an essential vitamin that must be obtained from the diet for proper neurological function. At higher doses, thiamine and benfotiamine (S-benzoylthiamine O-monophosphate, BT)-a phosphorylated derivative of thiamine-have antinociceptive effects in animals and humans, although how these compounds inhibit pain is unknown. Here, we found that Prostatic acid phosphatase (PAP, ACPP) can dephosphorylate BT in vitro, in dorsal root ganglia (DRG) neurons and in primary-afferent axon terminals in the dorsal spinal cord. The dephosphorylated product S-benzoylthiamine (S-BT) then decomposes to O-benzoylthiamine (O-BT) and to thiamine in a pH-dependent manner, independent of additional enzymes. This unique reaction mechanism reveals that BT only requires a phosphatase for conversion to thiamine. However, we found that the antinociceptive effects of BT, thiamine monophosphate (TMP) and thiamine-a compound that is not phosphorylated-were entirely dependent on PAP at the spinal level. Moreover, pharmacokinetic studies with wild-type and Pap(-/-) mice revealed that PAP is not required for the conversion of BT to thiamine in vivo. Taken together, our study highlights an obligatory role for PAP in the antinociceptive effects of thiamine and phosphorylated thiamine analogs, and suggests a novel phosphatase-independent function for PAP.
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Prostatic acid phosphatase is required for the antinociceptive effects of thiamine and benfotiamine.
Directory of Open Access Journals (Sweden)
Julie K Hurt
Full Text Available Thiamine (Vitamin B1 is an essential vitamin that must be obtained from the diet for proper neurological function. At higher doses, thiamine and benfotiamine (S-benzoylthiamine O-monophosphate, BT-a phosphorylated derivative of thiamine-have antinociceptive effects in animals and humans, although how these compounds inhibit pain is unknown. Here, we found that Prostatic acid phosphatase (PAP, ACPP can dephosphorylate BT in vitro, in dorsal root ganglia (DRG neurons and in primary-afferent axon terminals in the dorsal spinal cord. The dephosphorylated product S-benzoylthiamine (S-BT then decomposes to O-benzoylthiamine (O-BT and to thiamine in a pH-dependent manner, independent of additional enzymes. This unique reaction mechanism reveals that BT only requires a phosphatase for conversion to thiamine. However, we found that the antinociceptive effects of BT, thiamine monophosphate (TMP and thiamine-a compound that is not phosphorylated-were entirely dependent on PAP at the spinal level. Moreover, pharmacokinetic studies with wild-type and Pap(-/- mice revealed that PAP is not required for the conversion of BT to thiamine in vivo. Taken together, our study highlights an obligatory role for PAP in the antinociceptive effects of thiamine and phosphorylated thiamine analogs, and suggests a novel phosphatase-independent function for PAP.
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Pyeon, Hye-Rim; Nah, Hee-Ju; Kang, Seung-Hoon; Choi, Si-Sun; Kim, Eung-Soo
2017-05-31
Heterologous expression of biosynthetic gene clusters of natural microbial products has become an essential strategy for titer improvement and pathway engineering of various potentially-valuable natural products. A Streptomyces artificial chromosomal conjugation vector, pSBAC, was previously successfully applied for precise cloning and tandem integration of a large polyketide tautomycetin (TMC) biosynthetic gene cluster (Nah et al. in Microb Cell Fact 14(1):1, 2015), implying that this strategy could be employed to develop a custom overexpression scheme of natural product pathway clusters present in actinomycetes. To validate the pSBAC system as a generally-applicable heterologous overexpression system for a large-sized polyketide biosynthetic gene cluster in Streptomyces, another model polyketide compound, the pikromycin biosynthetic gene cluster, was preciously cloned and heterologously expressed using the pSBAC system. A unique HindIII restriction site was precisely inserted at one of the border regions of the pikromycin biosynthetic gene cluster within the chromosome of Streptomyces venezuelae, followed by site-specific recombination of pSBAC into the flanking region of the pikromycin gene cluster. Unlike the previous cloning process, one HindIII site integration step was skipped through pSBAC modification. pPik001, a pSBAC containing the pikromycin biosynthetic gene cluster, was directly introduced into two heterologous hosts, Streptomyces lividans and Streptomyces coelicolor, resulting in the production of 10-deoxymethynolide, a major pikromycin derivative. When two entire pikromycin biosynthetic gene clusters were tandemly introduced into the S. lividans chromosome, overproduction of 10-deoxymethynolide and the presence of pikromycin, which was previously not detected, were both confirmed. Moreover, comparative qRT-PCR results confirmed that the transcription of pikromycin biosynthetic genes was significantly upregulated in S. lividans containing tandem
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International Nuclear Information System (INIS)
Park, Joon-Heum; Jung, Sunyo
2017-01-01
In this study, we used the biosynthetic inhibitors of carotenoid and tetrapyrrole biosynthetic pathways, norflurazon (NF) and oxyfluorfen (OF), as tools to gain insight into mechanisms of photooxidation in rice plants. NF resulted in bleaching symptom on leaves of the treated plants, whereas OF treatment developed a fast symptom of an apparent necrotic phenotype. Both plants exhibited decreases in photosynthetic efficiency, as indicated by F v /F m . NF caused severe disruption in thylakoid membranes, whereas OF-treated plants exhibited disruption of chloroplast envelope and plasma membrane. Levels of Lhca and Lhcb proteins in photosystem I (PSI) and PSII were reduced by photooxidative stress in NF- and OF-treated plants, with a greater decrease in NF plants. The down-regulation of nuclear-encoded photosynthesis genes Lhcb and rbcS was also found in both NF- and OF-treated plants, whereas plastid-encoded photosynthetic genes including RbcL, PsaC, and PsbD accumulated normally in NF plants but decreased drastically in OF plants. This proposes that the plastids in NF plants retain their potential to develop thylakoid membranes and that photobleaching is mainly controlled by nuclear genes. Distinct photooxidation patterns between NF- and OF-treated plants developed differential signaling, which might enable the plant to coordinate the expression of photosynthetic genes from the nuclear and plastidic genomes. - Highlights: • Two modes of photooxidation by carotenoid and tetrapyrrole biosynthetic inhibitors. • We examine differential alterations in chloroplast function and plastid signaling. • NF and OF cause differential alterations in chloroplast ultrastructure and function. • Photooxidation coordinates photosynthetic gene expression from nucleus and plastid.
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Directory of Open Access Journals (Sweden)
Wenjun Huang
2016-07-01
Full Text Available Flavonols as plant secondary metabolites with vital roles in plant development and defense against UV light, have been demonstrated to be the main bioactive components in the genus Epimedium plants, several species of which are used as materials for Herba Epimedii, an important traditional Chinese medicine. The flavonol biosynthetic pathway genes had been already isolated from E. sagittatum, but a R2R3-MYB transcription factor regulating the flavonol synthesis has not been functionally characterized so far in Epimedium plants. In this study, we isolated and characterized the R2R3-MYB transcription factor EsMYBF1 involved in regulation of the flavonol biosynthetic pathway from E. sagittatum. Sequence analysis indicated that EsMYBF1 belongs to the subgroup 7 of R2R3-MYB family which contains the flavonol-specific MYB regulators identified to date. Transient reporter assay showed that EsMYBF1 strongly activated the promoters of EsF3H (flavanone 3-hydroxylase and EsFLS (flavonol synthase, but not the promoters of EsDFRs (dihydroflavonol 4-reductase and EsANS (anthocyanidin synthase in transiently transformed Nicotiana benthamiana leaves. Both yeast two-hybrid assay and transient reporter assay validated EsMYBF1 to be independent of EsTT8, or AtTT8 bHLH regulators of the flavonoid pathway as cofactors. Ectopic expression of EsMYBF1 in transgenic tobacco resulted in the increased flavonol content and the decreased anthocyanin content in flowers. Correspondingly, the structural genes involved in flavonol synthesis were upregulated in the EsMYBF1 overexpression lines, including NtCHS (chalcone synthase, NtCHI (chalcone isomerase, NtF3H and NtFLS, whereas the late biosynthetic genes of the anthocyanin pathway (NtDFR and NtANS were remarkably downregulated, compared to the controls. These results suggest that EsMYBF1 is a flavonol-specific R2R3-MYB regulator, and involved in regulation of the biosynthesis of the flavonol-derived bioactive components in E
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Acute thiamine deficiency and refeeding syndrome: Similar findings but different pathogenesis.
Maiorana, Arianna; Vergine, Gianluca; Coletti, Valentina; Luciani, Matteo; Rizzo, Cristiano; Emma, Francesco; Dionisi-Vici, Carlo
2014-01-01
Refeeding syndrome can occur in several contexts of relative malnutrition in which an overaggressive nutritional support is started. The consequences are life threatening with multiorgan impairment, and severe electrolyte imbalances. During refeeding, glucose-involved insulin secretion causes abrupt reverse of lipolysis and a switch from catabolism to anabolism. This creates a sudden cellular demand for electrolytes (phosphate, potassium, and magnesium) necessary for synthesis of adenosine triphosphate, glucose transport, and other synthesis reactions, resulting in decreased serum levels. Laboratory findings and multiorgan impairment similar to refeeding syndrome also are observed in acute thiamine deficiency. The aim of this study was to determine whether thiamine deficiency was responsible for the electrolyte imbalance caused by tubular electrolyte losses. We describe two patients with leukemia who developed acute thiamine deficiency with an electrolyte pattern suggestive of refeeding syndrome, severe lactic acidosis, and evidence of proximal renal tubular dysfunction. A single thiamine administration led to rapid resolution of the tubular dysfunction and normalization of acidosis and electrolyte imbalance. This demonstrated that thiamine deficiency was responsible for the electrolyte imbalance, caused by tubular electrolyte losses. Our study indicates that, despite sharing many laboratory similarities, refeeding syndrome and acute thiamine deficiency should be viewed as separate entities in which the electrolyte abnormalities reported in cases of refeeding syndrome with thiamine deficiency and refractory lactic acidosis may be due to renal tubular losses instead of a shifting from extracellular to intracellular compartments. In oncologic and malnourished patients, individuals at particular risk for developing refeeding syndrome, in the presence of these biochemical abnormalities, acute thiamine deficiency should be suspected and treated because it promptly
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Functional conservation of coenzyme Q biosynthetic genes among yeasts, plants, and humans.
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Kazuhiro Hayashi
Full Text Available Coenzyme Q (CoQ is an essential factor for aerobic growth and oxidative phosphorylation in the electron transport system. The biosynthetic pathway for CoQ has been proposed mainly from biochemical and genetic analyses of Escherichia coli and Saccharomyces cerevisiae; however, the biosynthetic pathway in higher eukaryotes has been explored in only a limited number of studies. We previously reported the roles of several genes involved in CoQ synthesis in the fission yeast Schizosaccharomyces pombe. Here, we expand these findings by identifying ten genes (dps1, dlp1, ppt1, and coq3-9 that are required for CoQ synthesis. CoQ10-deficient S. pombe coq deletion strains were generated and characterized. All mutant fission yeast strains were sensitive to oxidative stress, produced a large amount of sulfide, required an antioxidant to grow on minimal medium, and did not survive at the stationary phase. To compare the biosynthetic pathway of CoQ in fission yeast with that in higher eukaryotes, the ability of CoQ biosynthetic genes from humans and plants (Arabidopsis thaliana to functionally complement the S. pombe coq deletion strains was determined. With the exception of COQ9, expression of all other human and plant COQ genes recovered CoQ10 production by the fission yeast coq deletion strains, although the addition of a mitochondrial targeting sequence was required for human COQ3 and COQ7, as well as A. thaliana COQ6. In summary, this study describes the functional conservation of CoQ biosynthetic genes between yeasts, humans, and plants.
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Thiamine Deficiency Induces Anorexia by Inhibiting Hypothalamic AMPK
Liu, Mei; Alimov, Alexander; Wang, Haiping; Frank, Jacqueline A.; Katz, Wendy; Xu, Mei; Ke, Zun-Ji; Luo, Jia
2014-01-01
Obesity and eating disorders are prevailing health concerns worldwide. It is important to understand the regulation of food intake and energy metabolism. Thiamine (vitamin B1) is an essential nutrient. Thiamine deficiency (TD) can cause a number of disorders in humans, such as Beriberi and Wernicke-Korsakoff syndrome. We demonstrated here that TD caused anorexia in C57BL/6 mice. After feeding a TD diet for 16 days, the mice displayed a significant decrease in food intake and an increase in resting energy expenditure (REE), which resulted in a severe weight loss. At the 22nd day, the food intake was reduced by 69% and 74% for male and female mice, respectively in TD group. The REE increased by 9 folds in TD group. The loss of body weight (17–24%) was similar between male and female animals and mainly resulted from the reduction of fat mass (49% decrease). Re-supplementation of thiamine (benfotiamine) restored animal's appetite, leading to a total recovery of body weight. The hypothalamic AMPK is a critical regulator of food intake. TD inhibited the phosphorylation of AMPK in the arcuate nucleus (ARN) and paraventricular nucleus (PVN) of the hypothalamus without affecting its expression. TD-induced inhibition of AMPK phosphorylation was reversed once thiamine was re-supplemented. In contrast, TD increased AMPK phosphorylation in the skeletal muscle and upregulated the uncoupling protein (UCP)-1 in brown adipose tissues which was consistent with increased basal energy expenditure. Re-administration of thiamine stabilized AMPK phosphorylation in the skeletal muscle as well as energy expenditure. Taken together, TD may induce anorexia by inhibiting hypothalamic AMPK activity. With a simultaneous increase in energy expenditure, TD caused an overall body weight loss. The results suggest that the status of thiamine levels in the body may affect food intake and body weight. PMID:24607345
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Infantile beriberi, a potentially fatal disease caused by thiamine deficiency, is a public health concern in Cambodia and regions where thiamine-poor white rice is a staple food. Low maternal thiamine intake reduces breast milk thiamine placing breastfed infants at risk of beriberi. The objective wa...
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In vitro immune functions in thiamine-replete and -depleted lake trout (Salvelinus namaycush)
Ottinger, Christopher A.; Honeyfield, Dale C.; Densmore, Christine L.; Iwanowicz, Luke R.
2014-01-01
In this study we examined the impacts of in vivo thiamine deficiency on lake trout leukocyte function measured in vitro. When compared outside the context of individual-specific thiamine concentrations no significant differences were observed in leukocyte bactericidal activity or in concanavalin A (Con A), and phytohemagglutinin-P (PHA-P) stimulated leukocyte proliferation. Placing immune functions into context with the ratio of in vivo liver thiamine monophosphate (TMP – biologically inactive form) to thiamine pyrophosphate (TPP – biologically active form) proved to be the best indicator of thiamine depletion impacts as determined using regression modeling. These observed relationships indicated differential effects on the immune measures with bactericidal activity exhibiting an inverse relationship with TMP to TPP ratios, Con A stimulated mitogenesis exhibiting a positive relationship with TMP to TPP ratios and PHA-P stimulated mitogenesis exhibiting no significant relationships. In addition, these relationships showed considerable complexity which included the consistent observation of a thiamine-replete subgroup with characteristics similar to those seen in the leukocytes from thiamine-depleted fish. When considered together, our observations indicate that lake trout leukocytes experience cell-type specific impacts as well as an altered physiologic environment when confronted with a thiamine-limited state.
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Directory of Open Access Journals (Sweden)
Bingji Ma
2017-12-01
Full Text Available The unusual amino acid l-cyclopropylalanine was isolated from the mushroom Amanita virgineoides after detection in an anti-fungal screening test. l-Cyclopropylalanine was found to exhibit broad-spectrum inhibition against fungi and bacteria. The anti-fungal activity was found to be abolished in the presence of the amino acid l-leucine, but not any other amino acids, indicating that l-cyclopropylalanine may block the biosynthesis of the essential amino acid l-leucine, thereby inhibiting fungal and bacteria growth. Further biochemical studies found l-cyclopropylalanine indeed inhibits α-isopropylmalate synthase (α-IMPS, the enzyme that catalyzes the rate-limiting step in the biosynthetic pathway of l-leucine. Inhibition of essential l-leucine synthesis in fungal and bacteria organisms, a pathway absent in host organisms such as humans, may represent a novel antibiotic mechanism to counter the ever-increasing problem of drug resistance to existing antibiotics.
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Examining strategies to facilitate vitamin B1 biofortification of plants by genetic engineering
Directory of Open Access Journals (Sweden)
Lucille ePourcel
2013-05-01
Full Text Available Thiamin (vitamin B1 is made by plants and microorganisms but is an essential micronutrient in the human diet. All organisms require it as a cofactor in its form as thiamin pyrophosphate (TPP for the activity of key enzymes of central metabolism. In humans, deficiency is widespread particularly in populations where polished rice is a major component of the diet. Considerable progress has been made on the elucidation of the biosynthesis pathway within the last few years enabling concrete strategies for biofortification purposes to be devised, with a particular focus here on genetic engineering. Furthermore, the vitamin has been shown to play a role in both abiotic and biotic stress responses. The precursors for de novo biosynthesis of thiamin differ between microorganisms and plants. Bacteria use intermediates derived from purine and isoprenoid biosynthesis, whereas the pathway in yeast involves the use of compounds from the vitamin B3 and B6 groups. Plants on the other hand use a combination of the bacterial and yeast pathways and there is subcellular partitioning of the biosynthesis steps. Specifically, thiamin biosynthesis occurs in the chloroplast of plants through the separate formation of the pyrimidine and thiazole moieties, which are then coupled to form thiamin monophosphate (TMP. Phosphorylation of thiamin to form TPP occurs in the cytosol. Therefore, thiamin (or TMP must be exported from the chloroplast to the cytosol for the latter step to be executed. The regulation of biosynthesis is mediated through riboswitches, where binding of the product TPP to the pre-mRNA of a biosynthetic gene modulates expression. Here we examine and hypothesize on genetic engineering approaches attempting to increase the thiamin content employing knowledge gained with the model plant Arabidopsis thaliana. We will discuss the regulatory steps that need to be taken into consideration and can be used a prerequisite for devising such strategies in crop plants.
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Sex pheromone biosynthetic pathways are conserved between moths and the butterfly Bicyclus anynana
Liénard, Marjorie A; Wang, Hong-Lei; Lassance, Jean-Marc; Löfstedt, Christer
2014-01-01
Although phylogenetically nested within the moths, butterflies have diverged extensively in a number of life history traits. Whereas moths rely greatly on chemical signals, visual advertisement is the hallmark of mate finding in butterflies. In the context of courtship, however, male chemical signals are widespread in both groups although they likely have multiple evolutionary origins. Here, we report that in males of the butterfly Bicyclus anynana, courtship scents are produced de novo via biosynthetic pathways shared with females of many moth species. We show that two of the pheromone components that play a major role in mate choice, namely the (Z)-9-tetradecenol and hexadecanal, are produced through the activity of a fatty acyl Δ11-desaturase and two specialized alcohol-forming fatty acyl reductases. Our study provides the first evidence of conservation and sharing of ancestral genetic modules for the production of FA-derived pheromones over a long evolutionary timeframe thereby reconciling mate communication in moths and butterflies. PMID:24862548
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Expression profile of genes coding for carotenoid biosynthetic ...
Indian Academy of Sciences (India)
Expression profile of genes coding for carotenoid biosynthetic pathway during ripening and their association with accumulation of lycopene in tomato fruits. Shuchi Smita, Ravi Rajwanshi, Sangram Keshari Lenka, Amit Katiyar, Viswanathan Chinnusamy and. Kailash Chander Bansal. J. Genet. 92, 363–368. Table 1.
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A rapid solid-phase extraction fluorometric method for thiamine and riboflavin in salmonid eggs
Zajicek, James L.; Tillitt, Donald E.; Brown, Scott B.; Brown, Lisa R.; Honeyfield, Dale C.; Fitzsimons, John D.
2005-01-01
A new method has been developed and successfully applied to the selective measurement of thiamine (nonphosphorylated), total thiamine (sum of thiamine, thiamine monophosphate [TMP], thiamine diphosphate [TDP], and thiamine triphosphate [TTP]), and potentially interfering riboflavin in acidic (2% trichloroacetic acid) extracts of selected salmonid and walleye egg samples. Acidic extracts of eggs were applied directly to end-capped C18, reversed-phase solid-phase extraction (SPE) columns and separated into three fractions by elution with mixtures of PO4 buffer (pH 2), methanol (10%), and acetonitrile (20%). All thiamine compounds recovered in the first two fractions were oxidized to their corresponding thiochromes with alkaline potassium hexacyanoferrate, and we measured the thiochrome fluorescence (excitation at 360 nm, emission at 460 nm) in a 96-well microplate reader. Riboflavin, recovered in third fraction (eluted with pH 2, 20% acetonitrile), was analyzed directly by measuring the fluorescence of this fraction (excitation at 450 nm, emission at 530 nm). Significant portions of the phosphate esters of thiamine (TMP, TDP, and presumably TTP), when present at low concentrations (extract thiamine compounds into 2% trichlororacetic acid solution; an inexpensive, commercially available SPE column; small amounts of sample (0.5-1 g); microliter volumes of solvents per sample; a traditional, relatively nonhazardous, oxidation of thiamine compounds to fluorescent thiochromes; and an ultraviolet-visible-wavelength-filter fluorometer for the measurements. ?? Copyright by the American Fisheries Society 2005.
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Directory of Open Access Journals (Sweden)
Nikseresht S
2010-08-01
Full Text Available "nBackground: Postpartum depression is a mood disorder that has harmful effects on mothers, infants, family and relationships. Acute decrease of progesterone after delivery has been proposed as a cause for postpartum depression. This hormone can affect neurotransmitters' function. Zinc (Zn and magnesium (Mg as trace elements exert their antidepressant effects through neurotransmitter pathways. On the other hand, thiamin (Vit B1 deficiency leads to depression in animal models. The aim of this study was to evaluate effects of combination of zinc, magnesium and thiamine on postpartum depression and role of nitrergic system. "n"nMethods: One hundred ten female mice in five groups were used. Postpartum depression was conducted using progesterone injections. Combinations of Zinc chloride, magnesium chloride and thiamine HCL were administered 30 minutes before open field and forced swimming test (FST. In order to investigate role of nitrergic system, L-arginine and LNAME were administered. "n"nResults: All treatment groups spent less immobility time than the control group (p< 0.05. Combined administration of Zn+ Mg+ Vit B1 caused the most reduction in immobility time. Administration of L-NAME in Zn+ Mg+ Vit B1 group caused reduction in immobility time while administration of L-arginine caused increase in immobility time in the same group. "nConclusion: Zinc, magnesium and thiamine can improve depressive symptoms by nitrergic pathway. These elements as supplement compounds could be alternatives for antidepressants in postpartum period.
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The effects of thiamin on lead metabolism: organ distribution of lead 203.
Kim, J S; Hamilton, D L; Blakley, B R; Rousseaux, C G
1992-01-01
The effect of thiamin on the organ distribution of lead was evaluated in CD-1 mice exposed intragastrically or intraperitoneally to a single dose of lead acetate (100 micrograms) containing 100 microCi lead 203. They were treated with either thiamin (25 or 50 mg/kg body weight), calcium ethylenediaminetetraacetic acid (CaEDTA) (50 mg/kg body weight), or combinations of thiamin and CaEDTA. The whole body retention and the organ distribution of lead 203 varied depending upon the route of lead administration, dose of thiamin and the specific treatment combination. Thiamin (25 or 50 mg/kg) treatment increased the whole body retention of both intragastric and intraperitoneal lead by approximately 10% in each instance. Calcium ethylenediaminetetraacetic acid, either alone or in combination with thiamin (50 mg/kg) reduced the whole body retention of lead by as much as 14% regardless of route of lead exposure. The relative retention of lead by the liver, kidney and spleen was greater in mice exposed to lead by the intragastric route. Regardless of route, CaEDTA in the combined treatment reduced the relative retention of lead in both the liver and kidney, whereas thiamin alone only reduced the retention of lead in the kidney. The results of this study indicate that thiamin in combination with CaEDTA alters the distribution and retention of lead in a manner which may have therapeutic application as it relates to chelation therapy. PMID:1423063
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Osmotic stress upregulates the transcription of thiamine (vitamin B1 ...
African Journals Online (AJOL)
Osmotic stress upregulates the transcription of thiamine (vitamin B1) ... Oil palm's responses in terms of the expression profiles of these two thiamine biosynthesis genes to an osmotic stress inducer, polyethylene glycol ... from 32 Countries:.
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Energy Technology Data Exchange (ETDEWEB)
Kyrpides, Nikos; Anderson, Iain; Rodriguez, Jason; Susanti, Dwi; Porat, Iris; Reich, Claudia; Ulrich, Luke E.; Elkins, James G.; Mavromatis, Kostas; Lykidis, Athanasios; Kim, Edwin; Thompson, Linda S.; Nolan, Matt; Land, Miriam; Copeland, Alex; Lapidus, Alla; Lucas, Susan; Detter, Chris; Zhulin, Igor B.; Olsen, Gary J.; Whitman, William; Mukhopadhyay, Biswarup; Bristow, James; Kyrpides, Nikos
2008-01-01
We report the complete genome of Thermofilum pendens, a deep-branching, hyperthermophilic member of the order Thermoproteales within the archaeal kingdom Crenarchaeota. T. pendens is a sulfur-dependent, anaerobic heterotroph isolated from a solfatara in Iceland. It is an extracellular commensal, requiring an extract of Thermoproteus tenax for growth, and the genome sequence reveals that biosynthetic pathways for purines, most amino acids, and most cofactors are absent. In fact T. pendens has fewer biosynthetic enzymes than obligate intracellular parasites, although it does not display other features common among obligate parasites and thus does not appear to be in the process of becoming a parasite. It appears that T. pendens has adapted to life in an environment rich in nutrients. T. pendens was known to utilize peptides as an energy source, but the genome reveals substantial ability to grow on carbohydrates. T. pendens is the first crenarchaeote and only the second archaeon found to have a transporter of the phosphotransferase system. In addition to fermentation, T. pendens may gain energy from sulfur reduction with hydrogen and formate as electron donors. It may also be capable of sulfur-independent growth on formate with formate hydrogenlyase. Additional novel features are the presence of a monomethylamine:corrinoid methyltransferase, the first time this enzyme has been found outside of Methanosarcinales, and a presenilin-related protein. Predicted highly expressed proteins do not include housekeeping genes, and instead include ABC transporters for carbohydrates and peptides, and CRISPR-associated proteins.
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In vitro immune functions in thiamine-replete and -depleted lake trout (Salvelinus namaycush).
Ottinger, Christopher A; Honeyfield, Dale C; Densmore, Christine L; Iwanowicz, Luke R
2014-05-01
In this study we examined the impacts of in vivo thiamine deficiency on lake trout leukocyte function measured in vitro. When compared outside the context of individual-specific thiamine concentrations no significant differences were observed in leukocyte bactericidal activity or in concanavalin A (Con A), and phytohemagglutinin-P (PHA-P) stimulated leukocyte proliferation. Placing immune functions into context with the ratio of in vivo liver thiamine monophosphate (TMP--biologically inactive form) to thiamine pyrophosphate (TPP--biologically active form) proved to be the best indicator of thiamine depletion impacts as determined using regression modeling. These observed relationships indicated differential effects on the immune measures with bactericidal activity exhibiting an inverse relationship with TMP to TPP ratios, Con A stimulated mitogenesis exhibiting a positive relationship with TMP to TPP ratios and PHA-P stimulated mitogenesis exhibiting no significant relationships. In addition, these relationships showed considerable complexity which included the consistent observation of a thiamine-replete subgroup with characteristics similar to those seen in the leukocytes from thiamine-depleted fish. When considered together, our observations indicate that lake trout leukocytes experience cell-type specific impacts as well as an altered physiologic environment when confronted with a thiamine-limited state. Published by Elsevier Ltd.
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Presumed primary thiamine deficiency in a young African lion (Panthera leo).
DiGesualdo, Cynthia L; Hoover, John P; Lorenz, Michael D
2005-09-01
A 1-yr-old intact male African lion (Panthera leo) fed only beef muscle meat was evaluated for episodes of hypermetric ataxia, generalized weakness and tonic-clonic front limb movements. A hemogram, biochemical profile, blood lead, electrocardiogram, survey radiographs, and brain computed tomography were normal. Cerebral spinal fluid analyses suggested mild inflammation. Acetylcholine receptor antibody and serologic tests for all infectious agents tested were negative. Clinical signs resolved completely 9 days after instituting oral thiamine (3 mg/kg/day) and a completely nutritional diet. This lion's pretreatment thiamine blood value (11 nmol/L) was markedly lower than that of a healthy lion (191 nmol/L) and a proposed reference range for adult African lions (160-350 nmol/L). The lion remained clinically normal 2 yr later when his blood thiamine value was 340 nmol/L. African lions can develop clinical primary thiamine deficiency and may respond favorably when thiamine treatment and adequate diet are instituted prior to irreversible neuronal necrosis.
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Directory of Open Access Journals (Sweden)
Girish Deshpande
Full Text Available The Drosophila embryonic gonad is assembled from two distinct cell types, the Primordial Germ Cells (PGCs and the Somatic Gonadal Precursor cells (SGPs. The PGCs form at the posterior of blastoderm stage embryos and are subsequently carried inside the embryo during gastrulation. To reach the SGPs, the PGCs must traverse the midgut wall and then migrate through the mesoderm. A combination of local repulsive cues and attractive signals emanating from the SGPs guide migration. We have investigated the role of the hedgehog (hh pathway gene shifted (shf in directing PGC migration. shf encodes a secreted protein that facilitates the long distance transmission of Hh through the proteoglycan matrix after it is released from basolateral membranes of Hh expressing cells in the wing imaginal disc. shf is expressed in the gonadal mesoderm, and loss- and gain-of-function experiments demonstrate that it is required for PGC migration. Previous studies have established that the hmgcr-dependent isoprenoid biosynthetic pathway plays a pivotal role in generating the PGC attractant both by the SGPs and by other tissues when hmgcr is ectopically expressed. We show that production of this PGC attractant depends upon shf as well as a second hh pathway gene gγ1. Further linking the PGC attractant to Hh, we present evidence indicating that ectopic expression of hmgcr in the nervous system promotes the release/transmission of the Hh ligand from these cells into and through the underlying mesodermal cell layer, where Hh can contact migrating PGCs. Finally, potentiation of Hh by hmgcr appears to depend upon cholesterol modification.
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Isolation and Purification of Thiamine Binding Protein from Mung Bean
Directory of Open Access Journals (Sweden)
DWIRINI RETNO GUNARTI
2013-03-01
Full Text Available Thiamine has fundamental role in energy metabolism. The organs mostly sensitive to the lack of thiamine levels in the body are the nervous system and the heart. Thiamine deficiency causes symptoms of polyneuritis and cardiovascular diseases. Because of its importance in the metabolism of carbohydrates, we need to measure the levels of thiamine in the body fluids by using an easy and inexpensive way without compromising the sensitivity and selectivity. An option to it is thiamine measurement based on the principle of which is analogous to ELISA, in which a thiamine binding protein (TBP act by replacing antibodies. The presence of TBP in several seeds have been reported by previous researchers, but the presence of TBP in mung beans has not been studied. This study was aimed to isolate and purify TBP from mung bean. The protein was isolated from mung bean through salting out by ammonium sulphate of 40, 70, and 90% (w/v. TBP has a negative charge as shown by cellulose acetate electrophoresis. The result obtained after salting out by ammonium sulphate was further purified bymeans of DEAE-cellulose chromatography and affinity chromatography. In precipitation of 90% of salting out method, one peak protein was obtained by using affinity chromatography. The protein was analyzed by SDS PAGE electrophoresis. The result of SDS PAGE electrophoresis showed that TBP has a molecular weight of 72.63 kDa.
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Blood thiamin status and determinants in the population of Seychelles (Indian Ocean).
Bovet, P; Larue, D; Fayol, V; Paccaud, F
1998-04-01
Micronutrient deficiencies have become rare in industrialised countries as availability of fresh food, supplementation, and fortification have improved but a less favourable situation may still prevail in many developing countries. Blood thiamin status and determinants were therefore investigated in the Seychelles in view of the high incidence of dilated cardiomyopathy and as the staple diet is polished rice that is deficient in thiamin. This was a cross sectional population study using an age and sex stratified random sample. Seychelles Islands (Indian Ocean). A subsample of 206 subjects aged 25-64 years from the population of Seychelles. Measurement of total thiamin concentration in whole blood using high performance liquid chromatography. Dietary variables measured using a face to face semi-quantitative food questionnaire. Mean (SD) whole blood thiamin concentration was 77.9 (22.4) nmol/l and low concentration (< 70 nmol/l) was found in 37% of the subjects (95% CI: 31%, 44%). Blood thiamin was significantly related to education and diet but not to age, sex, smoking, and body mass index. Blood thiamin was associated positively with meat, vegetable, salad, and tea intake and negatively with alcohol and fish intake. However, no combination of the examined variables could explain more than 15% of the observed variance in blood thiamin values. These data suggest that the distribution of blood thiamin in the sampled population is shifted to lower values compared with that generally accepted as normal in European populations. Further research should establish the significance of such lower values in this specific population to facilitate clinical and public health action as necessary.
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Ottinger, Christopher A.; Honeyfield, Dale C.; Densmore, Christine L.; Iwanowicz, Luke R.
2012-01-01
Lake trout Salvelinus namaycush on thiamine-replete and thiamine-depleted diets were evaluated for the effects of thiamine status on in vivo responses to the T-dependent antigen trinitophenol (TNP)-keyhole limpet hemocyanin (TNP-KLH), the T-independent antigen trinitrophenol-lipolysaccaharide (TNP-LPS), or Dulbecco's phosphate-buffered saline (DPBS; negative control fish). Plasma antibody concentrations were evaluated for possible differences in total anti-TNP activity as well as differences in response kinetics. Associations between anti-TNP activity and muscle and liver thiamine concentrations as well as ratios of muscle-to-liver thiamine to anti-TNP activity were also examined. Thiamine-depleted lake trout that were injected with TNP-LPS exhibited significantly more anti-TNP activity than thiamine-replete fish. The depleted fish injected with TNP-LPS also exhibited significantly different response kinetics relative to thiamine-replete lake trout. No differences in activity or kinetics were observed between the thiamine-replete and -depleted fish injected with TNP-KLH or in the DPBS negative controls. Anti-TNP activity in thiamine-depleted lake trout injected with TNP-KLH was positively associated with muscle thiamine pyrophosphate (thiamine diphosphate; TPP) concentration. A negative association was observed between the ratio of muscle-to-liver TPP and T-independent responses. No significant associations between anti-TNP activity and tissue thiamine concentration were observed in the thiamine-replete fish. We demonstrated that thiamine deficiency leads to alterations in both T-dependent and T-independent immune responses in lake trout.
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Ottinger, Christopher A; Honeyfield, Dale C; Densmore, Christine L; Iwanowicz, Luke R
2012-12-01
Lake trout Salvelinus namaycush on thiamine-replete and thiamine-depleted diets were evaluated for the effects of thiamine status on in vivo responses to the T-dependent antigen trinitophenol (TNP)-keyhole limpet hemocyanin (TNP-KLH), the T-independent antigen trinitrophenol-lipolysaccaharide (TNP-LPS), or Dulbecco's phosphate-buffered saline (DPBS; negative control fish). Plasma antibody concentrations were evaluated for possible differences in total anti-TNP activity as well as differences in response kinetics. Associations between anti-TNP activity and muscle and liver thiamine concentrations as well as ratios of muscle-to-liver thiamine to anti-TNP activity were also examined. Thiamine-depleted lake trout that were injected with TNP-LPS exhibited significantly more anti-TNP activity than thiamine-replete fish. The depleted fish injected with TNP-LPS also exhibited significantly different response kinetics relative to thiamine-replete lake trout. No differences in activity or kinetics were observed between the thiamine-replete and -depleted fish injected with TNP-KLH or in the DPBS negative controls. Anti-TNP activity in thiamine-depleted lake trout injected with TNP-KLH was positively associated with muscle thiamine pyrophosphate (thiamine diphosphate; TPP) concentration. A negative association was observed between the ratio of muscle-to-liver TPP and T-independent responses. No significant associations between anti-TNP activity and tissue thiamine concentration were observed in the thiamine-replete fish. We demonstrated that thiamine deficiency leads to alterations in both T-dependent and T-independent immune responses in lake trout.
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Infantile beriberi, a disease caused by thiamin (vitamin B1) deficiency, remains a public health concern in Cambodia and other parts of Southeast Asia. Infantile beriberi presents during the exclusive breastfeeding period and without treatment commonly results in death within *24 hours of clinical p...
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Ketola, H.G.; Isaacs, G.R.; Robins, J.S.; Lloyd, R.C.
2008-01-01
We investigated the feasibility of enhancing the reproduction of steelhead Oncorhynchus mykiss and landlocked Atlantic salmon Salmo salar in lakes where the consumption of alewives Alosa pseudoharengus and other forage fishes containing thiaminase can cause them to become thiamine deficient and thereby reduce the survival of their fry. We evaluated feeding fingerling steelhead excess thiamine hydrochloride (THCl) for 1 or 2 weeks or equimolar amounts of thiamine mononitrate, thiamine-tetrahydrofurfuryl-disulfide, benfotiamine, or dibenzoyl thiamine (DBT). We found minimal internal reserves of thiamine after 6 months. We also compared the ability of injections of thiamine and its analogs to prevent mortality in thiamine-deficient steelhead and Atlantic salmon sac fry and found all forms to be effective, although benfotiamine was the least effective on an equimolar basis. Further, we injected yearling steelhead and found that DBT was tolerated at approximately 11,200 nmol/g of body weight, about 10 times more than thiamine in any other form. When yearling steelhead were injected with near-maximal doses of thiamine hydrochloride and several analogs and then fed a thiamine-deficient diet, DBT was retained for approximately 2 years - in contrast to other forms, which were retained for less than about 6 months. Therefore, these results suggest that neither feeding nor injecting young hatchery salmonids with DBT is likely to enhance their reproduction for more than 2 years after stocking. However, injecting DBT in nearly mature fish (either cultured fish from hatcheries or wild fish captured in lakes) may provide them with enough thiamine to successfully spawn within 2 years even though they consume mainly thiaminase-containing forage fishes. ?? Copyright by the American Fisheries Society 2008.
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Whitfield, Kyly C; Karakochuk, Crystal D; Kroeun, Hou; Sokhoing, Ly; Chan, Benny B; Borath, Mam; Sophonneary, Prak; Moore, Kirsten; Tong, Jeffery K T; McLean, Judy; Talukder, Aminuzzaman; Lynd, Larry D; Li-Chan, Eunice C Y; Kitts, David D; Green, Tim J
2017-02-01
To assess whether ad libitum consumption of thiamin-fortified fish sauce over 6 months yields higher erythrocyte thiamin diphosphate concentrations (eTDP) among women of childbearing age and their children aged 12-59 months compared with control sauce containing no thiamin. In this double-blind, randomized controlled efficacy trial, 276 nonpregnant, nonlactating women (18-45 years of age) and their families in Prey Veng, Cambodia, were randomized to receive 1 of 3 fish sauce formulations: low thiamin concentration (low, 2 g/L), high thiamin concentration (high, 8 g/L), or a control (no thiamin) fish sauce. Baseline (t = 0) and endline (t = 6 months) eTDP were measured with the use of high-performance liquid chromatography with a fluorescence detector. Fish sauce consumption did not differ between treatment groups (P = .19). In intent-to-treat analysis, women's baseline-adjusted endline eTDP (mean; 95% CI) was higher among women in the low (259; 245-274 nmol/L) and high (257; 237-276 nmol/L) groups compared with control (184; 169-198 nmol/L; P sauce appears to be an efficacious means of improving biochemical thiamin status in nonpregnant, nonlactating women and their children (1-5 years of age) living in rural Cambodia. ClinicalTrials.gov: NCT02221063. Copyright © 2016 Elsevier Inc. All rights reserved.
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Adenylate kinase 1 knockout mice have normal thiamine triphosphate levels.
Makarchikov, A.F.; Wins, P.; Janssen, E.E.W.; Wieringa, B.; Grisar, T.; Bettendorff, L.
2002-01-01
Thiamine triphosphate (ThTP) is found at low concentrations in most animal tissues and it may act as a phosphate donor for the phosphorylation of proteins, suggesting a potential role in cell signaling. Two mechanisms have been proposed for the enzymatic synthesis of ThTP. A thiamine diphosphate
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Regulation of Flavonoid Biosynthetic Genes in Germinating Arabidopsis Seedlings.
Kubasek, WL; Shirley, BW; McKillop, A; Goodman, HM; Briggs, W; Ausubel, FM
1992-01-01
Many higher plants, including Arabidopsis, transiently display purple anthocyanin pigments just after seed germination. We observed that steady state levels of mRNAs encoded by four flavonoid biosynthetic genes, PAL1 (encoding phenylalanine ammonia-lyase 1), CHS (encoding chalcone synthase), CHI (encoding chalcone isomerase), and DFR (encoding dihydroflavonol reductase), were temporally regulated, peaking in 3-day-old seedlings grown in continuous white light. Except for the case of PAL1 mRNA, mRNA levels for these flavonoid genes were very low in seedlings grown in darkness. Light induction studies using seedlings grown in darkness showed that PAL1 mRNA began to accumulate before CHS and CHI mRNAs, which, in turn, began to accumulate before DFR mRNA. This order of induction is the same as the order of the biosynthetic steps in flavonoid biosynthesis. Our results suggest that the flavonoid biosynthetic pathway is coordinately regulated by a developmental timing mechanism during germination. Blue light and UVB light induction experiments using red light- and dark-grown seedlings showed that the flavonoid biosynthetic genes are induced most effectively by UVB light and that blue light induction is mediated by a specific blue light receptor. PMID:12297632
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Whitfield, Kyly C; Karakochuk, Crystal D; Kroeun, Hou; Hampel, Daniela; Sokhoing, Ly; Chan, Benny B; Borath, Mam; Sophonneary, Prak; McLean, Judy; Talukder, Aminuzzaman; Lynd, Larry D; Li-Chan, Eunice C Y; Kitts, David D; Allen, Lindsay H; Green, Timothy J
2016-10-03
Infantile beriberi, a potentially fatal disease caused by thiamine deficiency, remains a public health concern in Cambodia and regions where thiamine-poor white rice is a staple food. Low maternal thiamine intake reduces breast milk thiamine concentrations, placing breastfed infants at risk of beriberi. To determine if consumption of thiamine-fortified fish sauce yields higher erythrocyte thiamine diphosphate concentrations (eTDP) among lactating women and newborn infants and higher breast milk thiamine concentrations compared with a control sauce. In this double-blind randomized clinical trial, 90 pregnant women were recruited in the Prey Veng province, Cambodia. The study took place between October 2014 and April 2015. Women were randomized to 1 of 3 groups (n = 30) for ad libitum fish sauce consumption for 6 months: control (no thiamine), low-concentration (2 g/L), or high-concentration (8 g/L) fish sauce. Maternal eTDP was assessed at baseline (October 2014) and endline (April 2015). Secondary outcomes, breast milk thiamine concentration and infant eTDP, were measured at endline. Women's mean (SD) age and gestational stage were 26 (5) years and 23 (7) weeks, respectively. April 2015 eTDP was measured among 28 women (93%), 29 women (97%), and 23 women (77%) in the control, low-concentration, and high-concentration groups, respectively. In modified intent-to-treat analysis, mean baseline-adjusted endline eTDP was higher among women in the low-concentration (282nM; 95% CI, 235nM to 310nM) and high-concentration (254nM; 95% CI, 225nM to 284nM) groups compared with the control group (193nM; 95% CI, 164nM to 222M; P sauce through pregnancy and early lactation had higher eTDP and breast milk thiamine concentrations and their infants had higher eTDP, which was more pronounced in the high group. Thiamine-fortified fish sauce has the potential to prevent infantile beriberi in this population. Clinicaltrials.gov Identifier: NCT02221063.
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Disruption of thiamine uptake and growth of cells by feline leukemia virus subgroup A.
Mendoza, Ramon; Miller, A Dusty; Overbaugh, Julie
2013-03-01
Feline leukemia virus (FeLV) is still a major cause of morbidity and mortality in domestic cats and some wild cats despite the availability of relatively effective vaccines against the virus. FeLV subgroup A (FeLV-A) is transmitted in natural infections, and FeLV subgroups B, C, and T can evolve directly from FeLV-A by mutation and/or recombination with endogenous retroviruses in domestic cats, resulting in a variety of pathogenic outcomes. The cell surface entry receptor for FeLV-A is a putative thiamine transporter (THTR1). Here, we have addressed whether FeLV-A infection might disrupt thiamine uptake into cells and, because thiamine is an essential nutrient, whether this disruption might have pathological consequences. First, we cloned the cat ortholog of the other of the two known thiamine transporters in mammals, THTR2, and we show that feline THTR1 (feTHTR1) and feTHTR2 both mediate thiamine uptake, but feTHTR2 does not function as a receptor for FeLV-A. We found that feTHTR1 is widely expressed in cat tissues and in cell lines, while expression of feTHTR2 is restricted. Thiamine uptake mediated by feTHTR1 was indeed blocked by FeLV-A infection, and in feline fibroblasts that naturally express feTHTR1 and not feTHTR2, this blockade resulted in a growth arrest at physiological concentrations of extracellular thiamine. The growth arrest was reversed at high extracellular concentrations of thiamine. Our results show that FeLV-A infection can indeed disrupt thiamine uptake with pathological consequences. A prediction of these experiments is that raising the plasma levels of thiamine in FeLV-infected cats may ameliorate the pathogenic effects of infection.
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Zhao, Shicheng; Park, Chang Ha; Li, Xiaohua; Kim, Yeon Bok; Yang, Jingli; Sung, Gyoo Byung; Park, Nam Il; Kim, Soonok; Park, Sang Un
2015-09-30
Mulberry (Morus alba L.) is used in traditional Chinese medicine and is the sole food source of the silkworm. Here, 21 cDNAs encoding phenylpropanoid biosynthetic genes and 21 cDNAs encoding triterpene biosynthetic genes were isolated from mulberry. The expression levels of genes involved in these biosynthetic pathways and the accumulation of rutin, betulin, and betulinic acid, important secondary metabolites, were investigated in different plant organs. Most phenylpropanoid and triterpene biosynthetic genes were highly expressed in leaves and/or fruit, and most genes were downregulated during fruit ripening. The accumulation of rutin was more than fivefold higher in leaves than in other organs, and higher levels of betulin and betulinic acid were found in roots and leaves than in fruit. By comparing the contents of these compounds with gene expression levels, we speculate that MaUGT78D1 and MaLUS play important regulatory roles in the rutin and betulin biosynthetic pathways.
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Park, Joon-Heum; Jung, Sunyo
2017-01-22
In this study, we used the biosynthetic inhibitors of carotenoid and tetrapyrrole biosynthetic pathways, norflurazon (NF) and oxyfluorfen (OF), as tools to gain insight into mechanisms of photooxidation in rice plants. NF resulted in bleaching symptom on leaves of the treated plants, whereas OF treatment developed a fast symptom of an apparent necrotic phenotype. Both plants exhibited decreases in photosynthetic efficiency, as indicated by F v /F m . NF caused severe disruption in thylakoid membranes, whereas OF-treated plants exhibited disruption of chloroplast envelope and plasma membrane. Levels of Lhca and Lhcb proteins in photosystem I (PSI) and PSII were reduced by photooxidative stress in NF- and OF-treated plants, with a greater decrease in NF plants. The down-regulation of nuclear-encoded photosynthesis genes Lhcb and rbcS was also found in both NF- and OF-treated plants, whereas plastid-encoded photosynthetic genes including RbcL, PsaC, and PsbD accumulated normally in NF plants but decreased drastically in OF plants. This proposes that the plastids in NF plants retain their potential to develop thylakoid membranes and that photobleaching is mainly controlled by nuclear genes. Distinct photooxidation patterns between NF- and OF-treated plants developed differential signaling, which might enable the plant to coordinate the expression of photosynthetic genes from the nuclear and plastidic genomes. Copyright © 2016 Elsevier Inc. All rights reserved.
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A novel thiamine-derived pigment, pyrizepine, formed by the Maillard reaction.
Igoshi, Asuka; Noda, Kyoko; Murata, Masatsune
2018-04-26
To find a Maillard pigment derived from thiamine, a solution containing glucose and thiamine was heated and analyzed with high-performance liquid chromatography equipped with diode-array detection. As a result, a unique peak showing an absorption maximum at 380 nm was detected. This peak was then isolated from a reaction solution containing glucose, lysine and thiamine, and was identified as 1-(2-methyl-6,9-dihydro-5H-pyrimido[4,5-e][1,4]diazepin-7-yl)ethan-1-one using instrumental analyses. This compound, named pyrizepine, was a novel yellow pigment having a fused ring consisting of pyrimidine and diazepine. Pyrizepine was a major low-molecular-weight pigment in the reaction solution. The structure suggests that pyrizepine is formed by condensation reaction between a degradation product of thiamine and a tetrosone derivative formed from glucose by the Maillard reaction.
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Abnormal thiamine-dependent processes in Alzheimer's Disease. Lessons from diabetes.
Gibson, Gary E; Hirsch, Joseph A; Cirio, Rosanna T; Jordan, Barry D; Fonzetti, Pasquale; Elder, Jessica
2013-07-01
Reduced glucose metabolism is an invariant feature of Alzheimer's Disease (AD) and an outstanding biomarker of disease progression. Glucose metabolism may be an attractive therapeutic target, whether the decline initiates AD pathophysiology or is a critical component of a cascade. The cause of cerebral regional glucose hypometabolism remains unclear. Thiamine-dependent processes are critical in glucose metabolism and are diminished in brains of AD patients at autopsy. Further, the reductions in thiamine-dependent processes are highly correlated to the decline in clinical dementia rating scales. In animal models, thiamine deficiency exacerbates plaque formation, promotes phosphorylation of tau and impairs memory. In contrast, treatment of mouse models of AD with the thiamine derivative benfotiamine diminishes plaques, decreases phosphorylation of tau and reverses memory deficits. Diabetes predisposes to AD, which suggests they may share some common mechanisms. Benfotiamine diminishes peripheral neuropathy in diabetic humans and animals. In diabetes, benfotiamine induces key thiamine-dependent enzymes of the pentose shunt to reduce accumulation of toxic metabolites including advanced glycation end products (AGE). Related mechanisms may lead to reversal of plaque formation by benfotiamine in animals. If so, the use of benfotiamine could provide a safe intervention to reverse biological and clinical processes of AD progression. This article is part of a Special Issue entitled 'Mitochondrial function and dysfunction in neurodegeneration'. Copyright © 2012 Elsevier Inc. All rights reserved.
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Hiffler, Laurent; Rakotoambinina, Benjamin; Lafferty, Nadia; Martinez Garcia, Daniel
2016-01-01
In humans, thiamine is a micronutrient prone to depletion that may result in severe clinical abnormalities. This narrative review summarizes current knowledge on thiamine deficiency (TD) and bridges the gap between pathophysiology and clinical presentation by integrating thiamine metabolism at subcellular level with its function to vital organs. The broad clinical spectrum of TD is outlined, with emphasis on conditions encountered in tropical pediatric practice. In particular, TD is associated with type B lactic acidosis and classic forms of beriberi in children, but it is often unrecognized. Other severe acute conditions are associated with hypermetabolism, inducing a functional TD. The crucial role of thiamine in infant cognitive development is also highlighted in this review, along with analysis of the potential impact of TD in refeeding syndrome during severe acute malnutrition (SAM). This review aims to increase clinical awareness of TD in tropical settings where access to diagnostic tests is poor, and advocates for an early therapeutic thiamine challenge in resource-limited settings. Moreover, it provides evidence for thiamine as treatment in critical conditions requiring metabolic resuscitation, and gives rationale to the consideration of increased thiamine supplementation in therapeutic foods for malnourished children. PMID:27379239
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Prevalence of thiamine deficiency in heart failure patients on long-term diuretic therapy
Directory of Open Access Journals (Sweden)
Shrenik Doshi
2015-01-01
Full Text Available Background: Loop diuretics are an integral part of heart failure management. It has been shown that loop diuretics cause thiamine deficiency (TD by increasing its urinary loss. The aim of this study was to determine the prevalence of TD in heart failure patients on long-term oral loop diuretics. Methods: Heart failure patients (cases on oral loop diuretics (furosemide ≥40 mg/day or torsemide ≥20 mg/day, irrespective of the cause of heart failure, were compared to non-heart failure patients (controls not on loop diuretics in a 1:1 ratio. Whole blood free thiamine level was determined by liquid chromatography-tandem mass spectrometry method. Results: A total of 100 subjects were enrolled in a 1:1 ratio (50 cases and 50 controls. 67% of the total study population had TD, (defined as whole blood free thiamine level <0.7 ng/ml. There was no difference in mean thiamine level between cases and controls. On comparing patients with TD in both groups, patients on diuretics had significantly lower thiamine level compared to the patients, not on diuretics (P < 0.0001. Conclusions: There was no difference in the thiamine level when patients on loop diuretics were compared to controls. However, very low thiamine levels (<0.1 ng/ml was significantly more common in patients on loop diuretics.
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Bitsch, R; Wolf, M; Möller, J; Heuzeroth, L; Grüneklee, D
1991-01-01
The bioequivalence of thiamin in 2 therapeutically used preparations was tested in 10 healthy young men. Thiamin was orally administered either as lipophilic benfotiamine or as water-soluble thiamin mononitrate. Biokinetic data, measured as area under the curve and maximal concentration in plasma and hemolysate after ingestion, demonstrated a significantly improved bioavailability from the lipophilic derivative despite an ingested dose of only 40% as compared with the water-soluble salt. A superior cellular efficacy of benfotiamine was also concluded from the short-term stimulation of the thiamin-dependent transketolase activity in erythrocytes.
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Energy Technology Data Exchange (ETDEWEB)
Tran, Timothy H. [Cornell University, Ithaca, New York 14853-1301 (United States); Krishnamoorthy, Kalyanaraman; Begley, Tadhg P., E-mail: begley@tamu.edu [Texas A& M University, College Station, TX 77842 (United States); Ealick, Steven E., E-mail: begley@tamu.edu [Cornell University, Ithaca, New York 14853-1301 (United States)
2011-10-01
MetY is the first reported structure of an O-acetylhomoserine sulfhydrylase that utilizes a protein thiocarboxylate intermediate as the sulfur source in a novel methionine-biosynthetic pathway instead of catalyzing a direct sulfhydrylation reaction. O-Acetylhomoserine sulfhydrylase (OAHS) is a pyridoxal 5′-phosphate (PLP) dependent sulfide-utilizing enzyme in the l-cysteine and l-methionine biosynthetic pathways of various enteric bacteria and fungi. OAHS catalyzes the conversion of O-acetylhomoserine to homocysteine using sulfide in a process known as direct sulfhydrylation. However, the source of the sulfur has not been identified and no structures of OAHS have been reported in the literature. Here, the crystal structure of Wolinella succinogenes OAHS (MetY) determined at 2.2 Å resolution is reported. MetY crystallized in space group C2 with two monomers in the asymmetric unit. Size-exclusion chromatography, dynamic light scattering and crystal packing indicate that the biological unit is a tetramer in solution. This is further supported by the crystal structure, in which a tetramer is formed using a combination of noncrystallographic and crystallographic twofold axes. A search for structurally homologous proteins revealed that MetY has the same fold as cystathionine γ-lyase and methionine γ-lyase. The active sites of these enzymes, which are also PLP-dependent, share a high degree of structural similarity, suggesting that MetY belongs to the γ-elimination subclass of the Cys/Met metabolism PLP-dependent family of enzymes. The structure of MetY, together with biochemical data, provides insight into the mechanism of sulfur transfer to a small molecule via a protein thiocarboxylate intermediate.
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Abnormal Thiamine-Dependent Processes in Alzheimer’s Disease. Lessons from Diabetes
Gibson, Gary E.; Hirsch, Joseph A.; Cirio, Rosanna T.; Jordan, Barry D.; Fonzetti, Pasquale; Elder, Jessica
2013-01-01
Reduced glucose metabolism is an invariant feature of Alzheimer’s Disease (AD) and an outstanding biomarker of disease progression. Glucose metabolism may be an attractive therapeutic target, whether the decline initiates AD pathophysiology or is a critical component of a cascade. The cause of cerebral regional glucose hypometabolism remains unclear. Thiamine-dependent processes are critical in glucose metabolism and are diminished in brains of AD patients at autopsy. Further, the reductions in thiamine-dependent processes are highly correlated to the decline in clinical dementia rating scales. In animal models, thiamine deficiency exacerbates plaque formation, promotes phosphorylation of tau and impairs memory. In contrast, treatment of mouse models of AD with the thiamine derivative benfotiamine diminishes plaques, decreases phosphorylation of tau and reverses memory deficits. Diabetes predisposes to AD, which suggests they may share some common mechanisms. Benfotiamine diminishes peripheral neuropathy in diabetic humans and animals. In diabetes, benfotiamine induces key thiamine-dependent enzymes of the pentose shunt to reduce accumulation of toxic metabolites including advanced glycation end products (AGE). Related mechanisms may lead to reversal of plaque formation by benfotiamine in animals. If so, the use of benfotiamine could provide a safe intervention to reverse biological and clinical processes of AD progression. PMID:22982063
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A kinetic model of thiamine biosynthesis in Escherichia coli
DEFF Research Database (Denmark)
Matos, Marta; Herrgard, Markus; Andersen, Mikael Rørdam
2014-01-01
Thiamine can only be synthesized by prokaryotes and some eukaryotes, humans for example get it through their diet. Yet, it is key for the correct functioning of the carbohydrate and amino acid metabolism, and thiamine deficiency in humans can cause beriberi, which can result in muscle weakness...
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Importance: Infantile beriberi, a potentially fatal disease caused by thiamin deficiency, remains a public health concern in Cambodia and regions where B-vitamin poor, polished white rice is a staple food. Low maternal thiamin intake reduces breast milk thiamin concentrations, placing breastfed infa...
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Psychological Assessment of Patients With Biotin-Thiamine-Responsive Basal Ganglia Disease.
Alfadhel, Majid; Al-Bluwi, Amal
2017-01-01
Biotin-thiamine-responsive basal ganglia disease is a devastating autosomal recessive inherited neurological disorder. We conducted a retrospective chart review of all patients with biotin-thiamine-responsive basal ganglia disease who underwent a formal psychological assessment. Six females and 3 males were included. Five patients (56%) had an average IQ, two patients (22%) had mild delay, and two (22%) had severe delay. A normal outcome was directly related to the time of diagnosis and initiation of treatment. Early diagnosis and immediate commencement of treatment were associated with a favorable outcome and vice versa. The most affected domain was visual motor integration, while understanding and mathematical problem-solving were the least affected. In summary, this is the first study discussing the psychological assessment of patients with biotin-thiamine-responsive basal ganglia disease. The results of this study alert clinicians to consider prompt initiation of biotin and thiamine in any patient presenting with neuroregression and a basal ganglia lesion on a brain magnetic resonance imaging.
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Ruiz-López, Noemi; Sayanova, Olga; Napier, Johnathan A; Haslam, Richard P
2012-04-01
Omega-3 (ω-3) very long chain polyunsaturated fatty acids (VLC-PUFAs) such as eicosapentaenoic acid (EPA; 20:5 Δ5,8,11,14,17) and docosahexaenoic acid (DHA; 22:6 Δ4,7,10,13,16,19) have been shown to have significant roles in human health. Currently the primary dietary source of these fatty acids are marine fish; however, the increasing demand for fish and fish oil (in particular the expansion of the aquaculture industry) is placing enormous pressure on diminishing marine stocks. Such overfishing and concerns related to pollution in the marine environment have directed research towards the development of a viable alternative sustainable source of VLC-PUFAs. As a result, the last decade has seen many genes encoding the primary VLC-PUFA biosynthetic activities identified and characterized. This has allowed the reconstitution of the VLC-PUFA biosynthetic pathway in oilseed crops, producing transgenic plants engineered to accumulate ω-3 VLC-PUFAs at levels approaching those found in native marine organisms. Moreover, as a result of these engineering activities, knowledge of the fundamental processes surrounding acyl exchange and lipid remodelling has progressed. The application of new technologies, for example lipidomics and next-generation sequencing, is providing a better understanding of seed oil biosynthesis and opportunities for increasing the production of unusual fatty acids. Certainly, it is now possible to modify the composition of plant oils successfully, and, in this review, the most recent developments in this field and the challenges of producing VLC-PUFAs in the seed oil of higher plants will be described.
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The effects of thiamin on lead metabolism: organ distribution of lead 203.
Kim, J S; Hamilton, D L; Blakley, B R; Rousseaux, C G
1992-01-01
The effect of thiamin on the organ distribution of lead was evaluated in CD-1 mice exposed intragastrically or intraperitoneally to a single dose of lead acetate (100 micrograms) containing 100 microCi lead 203. They were treated with either thiamin (25 or 50 mg/kg body weight), calcium ethylenediaminetetraacetic acid (CaEDTA) (50 mg/kg body weight), or combinations of thiamin and CaEDTA. The whole body retention and the organ distribution of lead 203 varied depending upon the route of lead a...
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Masini, T; Lacy, B; Monjas, L; Hawksley, D; de Voogd, A R; Illarionov, B; Iqbal, A; Leeper, F J; Fischer, M; Kontoyianni, M; Hirsch, A K H
2015-12-14
The enzyme DXS catalyzes the first, rate-limiting step of the 2-C-methyl-d-erythritol-4-phosphate (MEP, 1) pathway using thiamine diphosphate (ThDP) as cofactor; the DXS-catalyzed reaction constitutes also the first step in vitamin B1 and B6 metabolism in bacteria. DXS is the least studied among the enzymes of this pathway in terms of crystallographic information, with only one complete crystal structure deposited in the Protein Data Bank (Deinococcus radiodurans DXS, PDB: ). We synthesized a series of thiamine and ThDP derivatives and tested them for their biochemical activity against two DXS orthologues, namely D. radiodurans DXS and Mycobacterium tuberculosis DXS. These experimental results, combined with advanced docking studies, led to the development and validation of a homology model of M. tuberculosis DXS, which, in turn, will guide medicinal chemists in rationally designing potential inhibitors for M. tuberculosis DXS.
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Netzer, Roman; Stafsnes, Marit H; Andreassen, Trygve; Goksøyr, Audun; Bruheim, Per; Brautaset, Trygve
2010-11-01
We report the cloning and characterization of the biosynthetic gene cluster (crtE, crtB, crtI, crtE2, crtYg, crtYh, and crtX) of the γ-cyclic C(50) carotenoid sarcinaxanthin in Micrococcus luteus NCTC2665. Expression of the complete and partial gene cluster in Escherichia coli hosts revealed that sarcinaxanthin biosynthesis from the precursor molecule farnesyl pyrophosphate (FPP) proceeds via C(40) lycopene, C(45) nonaflavuxanthin, C(50) flavuxanthin, and C(50) sarcinaxanthin. Glucosylation of sarcinaxanthin was accomplished by the crtX gene product. This is the first report describing the biosynthetic pathway of a γ-cyclic C(50) carotenoid. Expression of the corresponding genes from the marine M. luteus isolate Otnes7 in a lycopene-producing E. coli host resulted in the production of up to 2.5 mg/g cell dry weight sarcinaxanthin in shake flasks. In an attempt to experimentally understand the specific difference between the biosynthetic pathways of sarcinaxanthin and the structurally related ε-cyclic decaprenoxanthin, we constructed a hybrid gene cluster with the γ-cyclic C(50) carotenoid cyclase genes crtYg and crtYh from M. luteus replaced with the analogous ε-cyclic C(50) carotenoid cyclase genes crtYe and crtYf from the natural decaprenoxanthin producer Corynebacterium glutamicum. Surprisingly, expression of this hybrid gene cluster in an E. coli host resulted in accumulation of not only decaprenoxanthin, but also sarcinaxanthin and the asymmetric ε- and γ-cyclic C(50) carotenoid sarprenoxanthin, described for the first time in this work. Together, these data contributed to new insight into the diverse and multiple functions of bacterial C(50) carotenoid cyclases as key catalysts for the synthesis of structurally different carotenoids.
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Directory of Open Access Journals (Sweden)
Juwairiah Remali
2017-11-01
Full Text Available Background Streptomyces are well known for their capability to produce many bioactive secondary metabolites with medical and industrial importance. Here we report a novel bioactive phenazine compound, 6-((2-hydroxy-4-methoxyphenoxy carbonyl phenazine-1-carboxylic acid (HCPCA extracted from Streptomyces kebangsaanensis, an endophyte isolated from the ethnomedicinal Portulaca oleracea. Methods The HCPCA chemical structure was determined using nuclear magnetic resonance spectroscopy. We conducted whole genome sequencing for the identification of the gene cluster(s believed to be responsible for phenazine biosynthesis in order to map its corresponding pathway, in addition to bioinformatics analysis to assess the potential of S. kebangsaanensis in producing other useful secondary metabolites. Results The S. kebangsaanensis genome comprises an 8,328,719 bp linear chromosome with high GC content (71.35% consisting of 12 rRNA operons, 81 tRNA, and 7,558 protein coding genes. We identified 24 gene clusters involved in polyketide, nonribosomal peptide, terpene, bacteriocin, and siderophore biosynthesis, as well as a gene cluster predicted to be responsible for phenazine biosynthesis. Discussion The HCPCA phenazine structure was hypothesized to derive from the combination of two biosynthetic pathways, phenazine-1,6-dicarboxylic acid and 4-methoxybenzene-1,2-diol, originated from the shikimic acid pathway. The identification of a biosynthesis pathway gene cluster for phenazine antibiotics might facilitate future genetic engineering design of new synthetic phenazine antibiotics. Additionally, these findings confirm the potential of S. kebangsaanensis for producing various antibiotics and secondary metabolites.
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Thiamine deficiency in Cambodian infants with and without beriberi.
Coats, Debra; Shelton-Dodge, Kelsey; Ou, Kevanna; Khun, Vannara; Seab, Sommon; Sok, Kimsan; Prou, Chiva; Tortorelli, Silvia; Moyer, Thomas P; Cooper, Lisa E; Begley, Tadhg P; Enders, Felicity; Fischer, Philip R; Topazian, Mark
2012-11-01
To test the hypothesis that heavy metal toxicity and consumption of thiaminase-containing foods predispose to symptomatic thiamine deficiency. In a case-control study, thiamine diphosphate (TDP) blood concentrations were measured in 27 infants diagnosed with beriberi at a rural clinic, as well as their mothers and healthy Cambodian and American controls. Blood and urine levels of lead, arsenic, cadmium, mercury, and thallium were measured. Local food samples were analyzed for thiaminase activity. Mean TDP level among cases and Cambodian controls was 48 and 56 nmol/L, respectively (P = .08) and was 132 nmol/L in American controls (P < .0001 compared with both Cambodian groups). Mean TDP level of mothers of cases and Cambodian controls was 57 and 57 nmol/L (P = .92), and was 126 nmol/L in American mothers (P < .0001 compared with both Cambodian groups). Cases (but not controls) had lower blood TDP levels than their mothers (P = .02). Infant TDP level decreased with infant age and was positively associated with maternal TDP level. Specific diagnostic criteria for beriberi did not correlate with TDP level. There was no correlation between heavy metal levels and either TDP level or case/control status. No thiaminase activity was observed in food samples. Thiamine deficiency is endemic among infants and nursing mothers in rural southeastern Cambodia and is often clinically inapparent. Neither heavy metal toxicity nor consumption of thiaminase-containing foods account for thiamine deficiency in this region. Copyright © 2012 Mosby, Inc. All rights reserved.
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A genomics based discovery of secondary metabolite biosynthetic gene clusters in Aspergillus ustus.
Directory of Open Access Journals (Sweden)
Borui Pi
Full Text Available Secondary metabolites (SMs produced by Aspergillus have been extensively studied for their crucial roles in human health, medicine and industrial production. However, the resulting information is almost exclusively derived from a few model organisms, including A. nidulans and A. fumigatus, but little is known about rare pathogens. In this study, we performed a genomics based discovery of SM biosynthetic gene clusters in Aspergillus ustus, a rare human pathogen. A total of 52 gene clusters were identified in the draft genome of A. ustus 3.3904, such as the sterigmatocystin biosynthesis pathway that was commonly found in Aspergillus species. In addition, several SM biosynthetic gene clusters were firstly identified in Aspergillus that were possibly acquired by horizontal gene transfer, including the vrt cluster that is responsible for viridicatumtoxin production. Comparative genomics revealed that A. ustus shared the largest number of SM biosynthetic gene clusters with A. nidulans, but much fewer with other Aspergilli like A. niger and A. oryzae. These findings would help to understand the diversity and evolution of SM biosynthesis pathways in genus Aspergillus, and we hope they will also promote the development of fungal identification methodology in clinic.
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A Genomics Based Discovery of Secondary Metabolite Biosynthetic Gene Clusters in Aspergillus ustus
Pi, Borui; Yu, Dongliang; Dai, Fangwei; Song, Xiaoming; Zhu, Congyi; Li, Hongye; Yu, Yunsong
2015-01-01
Secondary metabolites (SMs) produced by Aspergillus have been extensively studied for their crucial roles in human health, medicine and industrial production. However, the resulting information is almost exclusively derived from a few model organisms, including A. nidulans and A. fumigatus, but little is known about rare pathogens. In this study, we performed a genomics based discovery of SM biosynthetic gene clusters in Aspergillus ustus, a rare human pathogen. A total of 52 gene clusters were identified in the draft genome of A. ustus 3.3904, such as the sterigmatocystin biosynthesis pathway that was commonly found in Aspergillus species. In addition, several SM biosynthetic gene clusters were firstly identified in Aspergillus that were possibly acquired by horizontal gene transfer, including the vrt cluster that is responsible for viridicatumtoxin production. Comparative genomics revealed that A. ustus shared the largest number of SM biosynthetic gene clusters with A. nidulans, but much fewer with other Aspergilli like A. niger and A. oryzae. These findings would help to understand the diversity and evolution of SM biosynthesis pathways in genus Aspergillus, and we hope they will also promote the development of fungal identification methodology in clinic. PMID:25706180
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Thiamin and Salicylic Acid as Biological Alternatives for Controlling Broad Bean Rot Disease
International Nuclear Information System (INIS)
AlHakimi, A.M.A; Alghalibi, Saeed M.S
2007-01-01
The interactive effects of fungi (Fusarium solani and Rhizoctonia solani) infection and thiamin or salicylic acid on growth rate, membrane stability, K+ efflux, UV-absorbing metabolites, photosynthetic pigments, cell wall components and lipid fractions of broad bean plants (30-day-old) were studied. Fungal infection induced a reduction in growth rate, membrane stability and content of photosynthetic pigments. Application of thiamin or salicylic acid increased growth rate, membrane stability and content of photosynthetic pigments. The K+ efflux and the leakage of UV-absorbing metabolites were stimulated with fungal infection. However, thiamin and salicylic acid treatment partially retarded the stimulatory effect on leakage of K+ and UV-absorbing metabolites of fungal infected plants. Fungal infection produced a reduction in the content of pectin and cellulose, total lipid, glycolipids and sterols fraction of shoots and roots and phospholipids of roots. On the other hand, the contents of hemicellulose and lignin of shoots and roots and phospholipids of shoots were stimulated by fungal infection. Soaking seeds in thiamin or salicylic acid counteracts partially or completely the adverse effect of fungal infection on pectin and cellulose composition, total lipid, glycolipids and sterols of either shoots or roots. On the other side, thiamin or salicylic acid treatments retarded the phospholipids accumulation in shoots of infected plants, and in roots the phospholipids accumulation was partially or completely alleviated. The content of hemicellulose and lignin of shoots and roots were antagonistically lowered by the application of thiamin or salicylic acid. (author)
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Gamma irradiation effects on thiamin and riboflavin in beef, lamb, pork, and turkey
International Nuclear Information System (INIS)
Fox, J.B. Jr.; Lakritz, L.; Hampson, J.; Richardson, R.; Ward, K.; Thayer, D.W.
1995-01-01
A study was made of the loss of thiamin and riboflavin due to gamma irradiation of beef, lamb and pork longissimus dorsi, turkey breast and leg muscles. Thiamin losses averaged 11%/kiloGray (kGy) and riboflavin losses 2.5%/kGy above three kGy. The rate of loss of thiamin in beef was higher than that in lamb, pork and turkey leg, but not turkey breast, with losses of 16%/kGy in beef and 8%/kGy in lamb. The rate of thiamin loss was not related to sulfhydryl, protein, moisture, fat or water content, pH or reducing capacity by redox titration. Loss of riboflavin was not different among species. Any detriment from such slight losses would seem to be more than compensated by the advantage of controlling bacteriological contamination by irradiation processing
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Connelly, L; Price, J
1996-04-01
Alcoholic Wernicke's encephalopathy has been commonplace in Australia for many years and, as this syndrome is attributed to a deficiency in the diet, it should be preventable. This study employs conventional cost-effectiveness methodology to compare the economic efficiency of several thiamin-supplementation alternatives that have been proposed for the prevention of Wernicke's encephalopathy. A series of rankings of these measures is derived from an estimated cost per case averted for each of the alternatives studied. These rankings identify the least cost-effective thiamin-supplementation alternative as that of enriching bread-making flour with thiamin.
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Rai, Ashutosh Kumar; Dubey, Ashutosh Prakash; Kumar, Santosh; Dutta, Debashis; Mishra, Mukti Nath; Singh, Bhupendra Narain; Tripathi, Anil Kumar
2016-11-01
Carotenoids constitute an important component of the defense system against photooxidative stress in bacteria. In Azospirillum brasilense Sp7, a nonphotosynthetic rhizobacterium, carotenoid synthesis is controlled by a pair of extracytoplasmic function sigma factors (RpoEs) and their cognate zinc-binding anti-sigma factors (ChrRs). Its genome harbors two copies of the gene encoding geranylgeranyl pyrophosphate synthase (CrtE), the first critical step in the carotenoid biosynthetic pathway in bacteria. Inactivation of each of two crtE paralogs found in A. brasilense caused reduction in carotenoid content, suggesting their involvement in carotenoid synthesis. However, the effect of crtE1 deletion was more pronounced than that of crtE2 deletion. Out of the five paralogs of rpoH in A. brasilense, overexpression of rpoH1 and rpoH2 enhanced carotenoid synthesis. Promoters of crtE2 and rpoH2 were found to be dependent on RpoH2 and RpoE1, respectively. Using a two-plasmid system in Escherichia coli, we have shown that the crtE2 gene of A. brasilense Sp7 is regulated by two cascades of sigma factors: one consisting of RpoE1and RpoH2 and the other consisting of RpoE2 and RpoH1. In addition, expression of crtE1 was upregulated indirectly by RpoE1 and RpoE2. This study shows, for the first time in any carotenoid-producing bacterium, that the regulation of carotenoid biosynthetic pathway involves a network of multiple cascades of alternative sigma factors. Carotenoids play a very important role in coping with photooxidative stress in prokaryotes and eukaryotes. Although extracytoplasmic function (ECF) sigma factors are known to directly regulate the expression of carotenoid biosynthetic genes in bacteria, regulation of carotenoid biosynthesis by one or multiple cascades of sigma factors had not been reported. This study provides the first evidence of the involvement of multiple cascades of sigma factors in the regulation of carotenoid synthesis in any bacterium by showing the
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Thiamine plays a critical role in the acid tolerance of Listeria monocytogenes.
Madeo, Moira; O'Riordan, Niamh; Fuchs, Thilo M; Utratna, Marta; Karatzas, Kimon Andreas G; O'Byrne, Conor P
2012-01-01
Understanding the molecular basis of acid tolerance in the food-borne pathogen Listeria monocytogenes is important as this property contributes to survival in the food-chain and enhances survival within infected hosts. The aim of this study was to identify genes contributing to acid tolerance in L. monocytogenes using transposon mutagenesis and subsequently to elucidate the physiological role of these genes in acid tolerance. One mutant harboring a Tn917 insertion in the thiT gene (formerly lmo1429), which encodes a thiamine (vitamin B1) uptake system, was found to be highly sensitive to acid. The acid-sensitive phenotype associated with loss of this gene was confirmed with an independently isolated mutant, from which the thiT gene was deleted (∆thiT). Cells of both wild-type and ∆thiT mutant that were thiamine depleted were found to be significantly more acid sensitive than control cultures. Thiamine-depleted cultures failed to produce significant concentrations of acetoin, consistent with the known thiamine dependence of acetolactate synthase, an enzyme required for acetoin synthesis from pyruvate. As acetoin synthesis is a proton-consuming process, we suggest that the acid sensitivity observed in thiamine-depleted cultures may be owing to an inability to produce acetoin. © 2011 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.
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Pácal, Lukáš; Kuricová, Katarína; Kaňková, Kateřina
2014-01-01
Growing prevalence of diabetes (type 2 as well as type 1) and its related morbidity due to vascular complications creates a large burden on medical care worldwide. Understanding the molecular pathogenesis of chronic micro-, macro- and avascular complications mediated by hyperglycemia is of crucial importance since novel therapeutic targets can be identified and tested. Thiamine (vitamin B1) is an essential cofactor of several enzymes involved in carbohydrate metabolism and published data suggest that thiamine metabolism in diabetes is deficient. This review aims to point out the physiological role of thiamine in metabolism of glucose and amino acids, to present overview of thiamine metabolism and to describe the consequences of thiamine deficiency (either clinically manifest or latent). Furthermore, we want to explain why thiamine demands are increased in diabetes and to summarise data indicating thiamine mishandling in diabetics (by review of the studies mapping the prevalence and the degree of thiamine deficiency in diabetics). Finally, we would like to summarise the evidence for the beneficial effect of thiamine supplementation in progression of hyperglycemia-related pathology and, therefore, to justify its importance in determining the harmful impact of hyperglycemia in diabetes. Based on the data presented it could be concluded that although experimental studies mostly resulted in beneficial effects, clinical studies of appropriate size and duration focusing on the effect of thiamine supplementation/therapy on hard endpoints are missing at present. Moreover, it is not currently clear which mechanisms contribute to the deficient action of thiamine in diabetes most. Experimental studies on the molecular mechanisms of thiamine deficiency in diabetes are critically needed before clear answer to diabetes community could be given. PMID:24936250
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Pácal, Lukáš; Kuricová, Katarína; Kaňková, Kateřina
2014-06-15
Growing prevalence of diabetes (type 2 as well as type 1) and its related morbidity due to vascular complications creates a large burden on medical care worldwide. Understanding the molecular pathogenesis of chronic micro-, macro- and avascular complications mediated by hyperglycemia is of crucial importance since novel therapeutic targets can be identified and tested. Thiamine (vitamin B1) is an essential cofactor of several enzymes involved in carbohydrate metabolism and published data suggest that thiamine metabolism in diabetes is deficient. This review aims to point out the physiological role of thiamine in metabolism of glucose and amino acids, to present overview of thiamine metabolism and to describe the consequences of thiamine deficiency (either clinically manifest or latent). Furthermore, we want to explain why thiamine demands are increased in diabetes and to summarise data indicating thiamine mishandling in diabetics (by review of the studies mapping the prevalence and the degree of thiamine deficiency in diabetics). Finally, we would like to summarise the evidence for the beneficial effect of thiamine supplementation in progression of hyperglycemia-related pathology and, therefore, to justify its importance in determining the harmful impact of hyperglycemia in diabetes. Based on the data presented it could be concluded that although experimental studies mostly resulted in beneficial effects, clinical studies of appropriate size and duration focusing on the effect of thiamine supplementation/therapy on hard endpoints are missing at present. Moreover, it is not currently clear which mechanisms contribute to the deficient action of thiamine in diabetes most. Experimental studies on the molecular mechanisms of thiamine deficiency in diabetes are critically needed before clear answer to diabetes community could be given.
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DEFF Research Database (Denmark)
Blin, Kai; Kim, Hyun Uk; Medema, Marnix H.
2017-01-01
Many drugs are derived from small molecules produced by microorganisms and plants, so-called natural products. Natural products have diverse chemical structures, but the biosynthetic pathways producing those compounds are often organized as biosynthetic gene clusters (BGCs) and follow a highly...... conserved biosynthetic logic. This allows for the identification of core biosynthetic enzymes using genome mining strategies that are based on the sequence similarity of the involved enzymes/genes. However, mining for a variety of BGCs quickly approaches a complexity level where manual analyses...... are no longer possible and require the use of automated genome mining pipelines, such as the antiSMASH software. In this review, we discuss the principles underlying the predictions of antiSMASH and other tools and provide practical advice for their application. Furthermore, we discuss important caveats...
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Differential hexosamine biosynthetic pathway gene expression with type 2 diabetes
Directory of Open Access Journals (Sweden)
Megan Coomer
2014-01-01
Full Text Available The hexosamine biosynthetic pathway (HBP culminates in the attachment of O-linked β-N-acetylglucosamine (O-GlcNAc onto serine/threonine residues of target proteins. The HBP is regulated by several modulators, i.e. O-linked β-N-acetylglucosaminyl transferase (OGT and β-N-acetylglucosaminidase (OGA catalyze the addition and removal of O-GlcNAc moieties, respectively; while flux is controlled by the rate-limiting enzyme glutamine:fructose-6-phosphate amidotransferase (GFPT, transcribed by two genes, GFPT1 and GFPT2. Since increased HBP flux is glucose-responsive and linked to insulin resistance/type 2 diabetes onset, we hypothesized that diabetic individuals exhibit differential expression of HBP regulatory genes. Volunteers (n = 60; n = 20 Mixed Ancestry, n = 40 Caucasian were recruited from Stellenbosch and Paarl (Western Cape, South Africa and classified as control, pre- or diabetic according to fasting plasma glucose and HbA1c levels, respectively. RNA was purified from leukocytes isolated from collected blood samples and OGT, OGA, GFPT1 and GFPT2 expressions determined by quantitative real-time PCR. The data reveal lower OGA expression in diabetic individuals (P < 0.01, while pre- and diabetic subjects displayed attenuated OGT expression vs. controls (P < 0.01 and P < 0.001, respectively. Moreover, GFPT2 expression decreased in pre- and diabetic Caucasians vs. controls (P < 0.05 and P < 0.01, respectively. We also found ethnic differences, i.e. Mixed Ancestry individuals exhibited a 2.4-fold increase in GFPT2 expression vs. Caucasians, despite diagnosis (P < 0.01. Gene expression of HBP regulators differs between diabetic and non-diabetic individuals, together with distinct ethnic-specific gene profiles. Thus differential HBP gene regulation may offer diagnostic utility and provide candidate susceptibility genes for different ethnic groupings.
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Wang, Zhao V; Deng, Yingfeng; Gao, Ningguo; Pedrozo, Zully; Li, Dan L; Morales, Cyndi R; Criollo, Alfredo; Luo, Xiang; Tan, Wei; Jiang, Nan; Lehrman, Mark A; Rothermel, Beverly A; Lee, Ann-Hwee; Lavandero, Sergio; Mammen, Pradeep P A; Ferdous, Anwarul; Gillette, Thomas G; Scherer, Philipp E; Hill, Joseph A
2014-03-13
The hexosamine biosynthetic pathway (HBP) generates uridine diphosphate N-acetylglucosamine (UDP-GlcNAc) for glycan synthesis and O-linked GlcNAc (O-GlcNAc) protein modifications. Despite the established role of the HBP in metabolism and multiple diseases, regulation of the HBP remains largely undefined. Here, we show that spliced X-box binding protein 1 (Xbp1s), the most conserved signal transducer of the unfolded protein response (UPR), is a direct transcriptional activator of the HBP. We demonstrate that the UPR triggers HBP activation via Xbp1s-dependent transcription of genes coding for key, rate-limiting enzymes. We further establish that this previously unrecognized UPR-HBP axis is triggered in a variety of stress conditions. Finally, we demonstrate a physiologic role for the UPR-HBP axis by showing that acute stimulation of Xbp1s in heart by ischemia/reperfusion confers robust cardioprotection in part through induction of the HBP. Collectively, these studies reveal that Xbp1s couples the UPR to the HBP to protect cells under stress. Copyright © 2014 Elsevier Inc. All rights reserved.
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Factors affecting a cyanogen bromide-based assay of thiamin.
Wyatt, D T; Lee, M; Hillman, R E
1989-11-01
We analyzed extensively a modified thiochrome method for thiamin analysis. Acid phosphatase (EC 3.1.3.2) from potato was superior to either alpha-amylase or acid phosphatase from wheat germ as a dephosphorylating agent. Timing of cyanogen bromide exposure was important, but the assay had good precision and accuracy. The standard curve was linear from 10 to 3000 nmol/L. The within-run and between-run coefficients of variation for total thiamin in whole blood were 3.6% and 7.4%, respectively. Analytical recoveries for low, intermediate, and high additions of thiamin to whole blood were 93-109%. Sample yield was increased by 41% (+/- 29% SD) with pre-assay freezing. Samples were stable for two days at room temperature, for seven days when refrigerated, and for two years when frozen. Previously unreported interference was seen with penicillin derivatives, and with several commonly used diuretic and antiepileptic medications. This assay may be suitable for population screening; 200 samples could be analyzed weekly at a cost of +0.20 per sample.
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Benfotiamine is similar to thiamine in correcting endothelial cell defects induced by high glucose.
Pomero, F; Molinar Min, A; La Selva, M; Allione, A; Molinatti, G M; Porta, M
2001-01-01
We investigated the hypothesis that benfotiamine, a lipophilic derivative of thiamine, affects replication delay and generation of advanced glycosylation end-products (AGE) in human umbilical vein endothelial cells cultured in the presence of high glucose. Cells were grown in physiological (5.6 mM) and high (28.0 mM) concentrations of D-glucose, with and without 150 microM thiamine or benfotiamine. Cell proliferation was measured by mitochondrial dehydrogenase activity. AGE generation after 20 days was assessed fluorimetrically. Cell replication was impaired by high glucose (72.3%+/-5.1% of that in physiological glucose, p=0.001). This was corrected by the addition of either thiamine (80.6%+/-2.4%, p=0.005) or benfotiamine (87.5%+/-8.9%, p=0.006), although it not was completely normalized (p=0.001 and p=0.008, respectively) to that in physiological glucose. Increased AGE production in high glucose (159.7%+/-38.9% of fluorescence in physiological glucose, p=0.003) was reduced by thiamine (113.2%+/-16.3%, p=0.008 vs. high glucose alone) or benfotiamine (135.6%+/-49.8%, p=0.03 vs. high glucose alone) to levels similar to those observed in physiological glucose. Benfotiamine, a derivative of thiamine with better bioavailability, corrects defective replication and increased AGE generation in endothelial cells cultured in high glucose, to a similar extent as thiamine. These effects may result from normalization of accelerated glycolysis and the consequent decrease in metabolites that are extremely active in generating nonenzymatic protein glycation. The potential role of thiamine administration in the prevention or treatment of vascular complications of diabetes deserves further investigation.
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Energy Technology Data Exchange (ETDEWEB)
Zhai, Ying; Bai, Silei; Liu, Jingjing; Yang, Liyuan [National Key Laboratory of Agricultural Microbiology, College of Life Science and Technology, Huazhong Agricultural University, Wuhan 430070 (China); Han, Li; Huang, Xueshi [Institute of Microbial Pharmaceuticals, College of Life and Health Sciences, Northeastern University, Shenyang 110819 (China); He, Jing, E-mail: hejingjj@mail.hzau.edu.cn [National Key Laboratory of Agricultural Microbiology, College of Life Science and Technology, Huazhong Agricultural University, Wuhan 430070 (China)
2016-04-22
Dithiolopyrrolone group antibiotics characterized by an electronically unique dithiolopyrrolone heterobicyclic core are known for their antibacterial, antifungal, insecticidal and antitumor activities. Recently the biosynthetic gene clusters for two dithiolopyrrolone compounds, holomycin and thiomarinol, have been identified respectively in different bacterial species. Here, we report a novel dithiolopyrrolone biosynthetic gene cluster (aut) isolated from Streptomyces thioluteus DSM 40027 which produces two pyrrothine derivatives, aureothricin and thiolutin. By comparison with other characterized dithiolopyrrolone clusters, eight genes in the aut cluster were verified to be responsible for the assembly of dithiolopyrrolone core. The aut cluster was further confirmed by heterologous expression and in-frame gene deletion experiments. Intriguingly, we found that the heterogenetic thioesterase HlmK derived from the holomycin (hlm) gene cluster in Streptomyces clavuligerus significantly improved heterologous biosynthesis of dithiolopyrrolones in Streptomyces albus through coexpression with the aut cluster. In the previous studies, HlmK was considered invalid because it has a Ser to Gly point mutation within the canonical Ser-His-Asp catalytic triad of thioesterases. However, gene inactivation and complementation experiments in our study unequivocally demonstrated that HlmK is an active distinctive type II thioesterase that plays a beneficial role in dithiolopyrrolone biosynthesis. - Highlights: • Cloning of the aureothricin biosynthetic gene cluster from Streptomyces thioluteus DSM 40027. • Identification of the aureothricin gene cluster by heterologous expression and in-frame gene deletion. • The heterogenetic thioesterase HlmK significantly improved dithiolopyrrolones production of the aureothricin gene cluster. • Identification of HlmK as an unusual type II thioesterase.
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International Nuclear Information System (INIS)
Zhai, Ying; Bai, Silei; Liu, Jingjing; Yang, Liyuan; Han, Li; Huang, Xueshi; He, Jing
2016-01-01
Dithiolopyrrolone group antibiotics characterized by an electronically unique dithiolopyrrolone heterobicyclic core are known for their antibacterial, antifungal, insecticidal and antitumor activities. Recently the biosynthetic gene clusters for two dithiolopyrrolone compounds, holomycin and thiomarinol, have been identified respectively in different bacterial species. Here, we report a novel dithiolopyrrolone biosynthetic gene cluster (aut) isolated from Streptomyces thioluteus DSM 40027 which produces two pyrrothine derivatives, aureothricin and thiolutin. By comparison with other characterized dithiolopyrrolone clusters, eight genes in the aut cluster were verified to be responsible for the assembly of dithiolopyrrolone core. The aut cluster was further confirmed by heterologous expression and in-frame gene deletion experiments. Intriguingly, we found that the heterogenetic thioesterase HlmK derived from the holomycin (hlm) gene cluster in Streptomyces clavuligerus significantly improved heterologous biosynthesis of dithiolopyrrolones in Streptomyces albus through coexpression with the aut cluster. In the previous studies, HlmK was considered invalid because it has a Ser to Gly point mutation within the canonical Ser-His-Asp catalytic triad of thioesterases. However, gene inactivation and complementation experiments in our study unequivocally demonstrated that HlmK is an active distinctive type II thioesterase that plays a beneficial role in dithiolopyrrolone biosynthesis. - Highlights: • Cloning of the aureothricin biosynthetic gene cluster from Streptomyces thioluteus DSM 40027. • Identification of the aureothricin gene cluster by heterologous expression and in-frame gene deletion. • The heterogenetic thioesterase HlmK significantly improved dithiolopyrrolones production of the aureothricin gene cluster. • Identification of HlmK as an unusual type II thioesterase.
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Identification of Genes Encoding the Folate- and Thiamine-Binding Membrane Proteins in Firmicutes
Eudes, Aymerick; Erkens, Guus B.; Slotboom, Dirk J.; Rodionov, Dmitry A.; Naponelli, Valeria; Hanson, Andrew D.
Genes encoding high-affinity folate- and thiamine-binding proteins (FolT, ThiT) were identified in the Lactobacillus casei genome, expressed in Lactococcus lactis, and functionally characterized. Similar genes occur in many Firmicutes, sometimes next to folate or thiamine salvage genes. Most thiT
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Directory of Open Access Journals (Sweden)
Margaret B. E. Livingstone
2011-01-01
Full Text Available In the early part of the rainy season in 1988, an outbreak of beriberi occurred in free-living adults in a relatively small area in the North Bank region of The Gambia. In 1995 we selected two compounds in a village called Chilla situated within the affected district to retrospectively examine dietary factors potentially contributing to the outbreak. There had previously been cases of beriberi in one compound (BBC but not in the other (NBC. We measured energy and thiamin intakes for four days on six occasions during the year. We calculated energy and thiamin intakes of people living in the two compounds and foods were collected for thiamin analysis through the year. Thiamin:Energy ratios only met international recommendations in the immediate post‑harvest season when energy and thiamin intakes were highest and then fell through the year. In the rainy season when food was short and labour was heaviest, energy intakes were lower in the NBC but thiamin:energy ratios were lower in BBC. Records of rainfall in 1988 collected near the village indicated that the amount in August was twice the average. We suggest the heavy rainfall may have increased farm workload and reduced income from outside-village work activity. The lower energy intakes in the NBC may have forced adults to rest thus sparing thiamin demands and delaying onset of beriberi. In contrast, the higher energy intake of adults in the BBC may have enabled them to continue working, thus increasing demands for thiamin and inducing the earlier onset of beriberi.
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Karachalias, Nikolaos; Babaei-Jadidi, Roya; Kupich, Christian; Ahmed, Naila; Thornalley, Paul J
2005-06-01
The streptozotocin-induced (STZ) diabetic rat experimental model of diabetes on insulin maintenance therapy exhibits dyslipidemia, mild thiamine deficiency, and increased plasma protein advanced glycation end products (AGEs). The reversal of thiamine deficiency by high-dose thiamine and S-benzoylthiamine monophosphate (benfotiamine) prevented the development of incipient nephropathy. Recently, we reported that high-dose thiamine (but not benfotiamine) countered diabetic dyslipidemia. To understand further the differences between the effects of thiamine and benfotiamine therapy, we quantified the levels of the AGEs in plasma protein. We found hydroimidazolone AGE residues derived from glyoxal and methylglyoxal, G-H1 and MG-H1, were increased 115% and 68% in STZ diabetic rats, with respect to normal controls, and were normalized by both thiamine and benfotiamine; whereas N-carboxymethyl-lysine (CML) and N-carboxyethyl-lysine (CEL) residues were increased 74% and 118% in STZ diabetic rats and were normalized by thiamine only. The lack of effect of benfotiamine on plasma CML and CEL residue concentrations suggests there may be important precursors of plasma protein CML and CEL residues other than glyoxal and methylglyoxal. These are probably lipid-derived aldehydes.
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Al-Daghri, Nasser M; Alharbi, Mohammed; Wani, Kaiser; Abd-Alrahman, Sherif H; Sheshah, Eman; Alokail, Majed S
2015-01-01
Thiamine (vitamin B1) is an essential enzyme cofactor in most organisms required at several stages of anabolic and catabolic intermediary metabolism. However, little is known on the positive effects of thiamine in diabetic type 1 (DMT1) patients. The objectives of this study were to evaluate the biochemical changes related to thiamine deficiency in patients with DMT1 outcomes among Saudi adults. We hypothesized that blood thiamine deficiency in patients with DMT1 manifestations might lead to an increase in metabolic syndrome. A total of 77 patients with DMT1 (age 35.8 ± 5.5) and 81 controls (age 45.0 ± 18.1) (total N = 158) were randomly selected from the Riyadh Cohort Study for inclusion. Saudi adults with diabetes type 1, a significant decrease in systolic (P < 0.001), and diastolic blood pressure (P = 0.008) and microalbuminuria (P = 0.02). Moreover, cholesterol, glucose and triglycerides were significantly increased (P 0.001, 0.001 and 0.008, respectively) in patients with diabetes type 1 compared to controls. On the other hand, HDL, TMP, TDP and thiamine, were significantly decreased in patients with diabetes type 1 (P 0.005, 0.002, 0.005, and 0.002), respectively. A strong association between blood thiamine level and diabetes type 1 was detected in our study population. The results confirmed the role of thiamine and thiamine phosphate esters, in preventing metabolic changes and complications of diabetes type 1. The levels of these thiamine and thiamine phosphate esters were correlated with diabetes related biomarkers including HDL, glucose, triglycerides and cholesterol, as well as microalbuminuria, LDL and urine thiamine. The results support a pivotal role of blood thiamine and its phosphate esters in preventing the biochemical changes and complications in patients with DMT1.
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Wolterink-van Loo, S.
2009-01-01
Aldolases are enzymes that catalyze reactions in both degradation and biosynthetic pathways in vivo and have been discovered in all domains of life. they. An interesting property of aldolases is that they can synthesize carbon-carbon bonds, generating a new stereogenic centre. As enzymes are
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Whitfield, Kyly C; Karakochuk, Crystal D; Liu, Yazheng; McCann, Adrian; Talukder, Aminuzzaman; Kroeun, Hou; Ward, Mary; McNulty, Helene; Lynd, Larry D; Kitts, David D; Li-Chan, Eunice C Y; McLean, Judy; Green, Timothy J
2015-03-01
Thiamin deficiency in infancy is the underlying cause of beriberi, which can be fatal without rapid treatment. Reports of thiamin deficiency are common in Cambodia; however, population representative data are unavailable. Because B-complex vitamin deficiencies commonly occur in combination, riboflavin was also investigated. We determined the biomarker status of thiamin and riboflavin in women of childbearing age in rural and urban Cambodia. We measured thiamin (erythrocyte thiamin diphosphate; TDP) and riboflavin (erythrocyte glutathione reductase activity coefficient; EGRac) status in a representative sample of Cambodian women (aged 20-45 y) in urban Phnom Penh (n = 146) and rural Prey Veng (n = 156), Cambodia, and, for comparison purposes, in a convenience sample of women in urban Vancouver, British Columbia, Canada (n = 49). Thiamin insufficiency (TDP ≤ 90 nmol/L) was common among both urban (39%) and rural (59%) Cambodian women (P Cambodia. The unexpected finding of high riboflavin inadequacy status in Vancouver women warrants further investigation. © 2015 American Society for Nutrition.
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Directory of Open Access Journals (Sweden)
Gratton SM
2014-05-01
Full Text Available Sean M Gratton, Byron L LamBascom Palmer Eye Institute, University of Miami, Miller School of Medicine, Miami, FL, USAAbstract: Visual loss due to optic neuropathy is a rare manifestation of thiamine deficiency. We report a case of a 39-year-old woman with a body mass index (BMI of 29 kg/m2 who developed visual loss and bilateral optic nerve head swelling after a short, self-limited gastrointestinal illness. She was disoriented and inattentive and had absent ankle jerk reflexes, diminished sensation in both legs below the knees, and marked truncal ataxia. Magnetic resonance imaging (MRI showed increased T2-signal in the medial thalami and mammillary bodies. The serum thiamine level was 8 nmol/L (normal 8–30. The diagnosis of thiamine deficiency was made, and the patient’s vision and neurologic symptoms improved significantly with intramuscular thiamine treatment. Thiamine deficiency can occur in the absence of an obvious predisposing factor such as alcoholism or low body weight. The clinician must be aware of the factors that govern vitamin availability and maintain a high index of suspicion to make the diagnosis in such cases.Keywords: optic neuropathy, nutritional deficiency
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International Nuclear Information System (INIS)
Azrolan, N.
1987-01-01
Cholesterol biosynthesis was characterized in cell-free post-mitochondrial supernatant (PMS) systems prepared from both normal rat liver and Morris hepatoma 3924A. Per cell, the rate of cholesterol synthesis from either 14 C-citrate of 14 -acetate in the hepatoma system was 9-fold greater than that observed in the liver system. Furthermore, the ratio of sterol-to-fatty acid synthesis rates from 14 C-citrate was more than 3-fold greater in the tumor than in the normal liver system. Incubations using radiolabeled acetate and mevalonate have demonstrated the loss of a normally rate-limiting control site within the early portion of the cholesterol biosynthetic pathway in the tumor system. Upon analysis of the steady-state levels of early lipogenic intermediates, the specific site of decontrol in the tumor was identified as the 3-hydroxy-3-methylglutaryl-CoA → mevalonate site of this pathway. In contrast, this reaction appeared to retain its rate-limiting properties in the cell-free system from normal liver
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Frank, T; Bitsch, R; Maiwald, J; Stein, G
2000-06-01
The influence of either orally administered S-benzoylthiamine-O-monophosphate (benfotiamine) or thiamine nitrate on the thiamine status was tested in a randomised, two-group comparison study in 20 end-stage renal disease (ESRD) patients. Main outcome measures were the pharmacokinetics of thiamine diphosphate (TDP) in blood, the in vitro erythrocyte transketolase activity, its activation coefficient (alpha-ETK) and the TDP concentration in erythrocytes. After ingestion of a single dose of either 100 mg thiamine nitrate (corresponding to 305 micromol thiamine) or 100 mg benfotiamine (corresponding to 214 micromol thiamine), the blood levels of thiamine phosphate esters were analysed by means of high-performance liquid chromatography for a 24-h period. The TDP concentration in erythrocytes was calculated using the haematocrit and TDP concentration in blood. Erythrocyte transketolase activity and alpha-ETK were measured before and 10 h after administration. The pharmacokinetics of TDP in blood were compared with healthy subjects of other studies retrieved from database query. Regarding the blood concentrations of TDP, the patients with ESRD had a 4.3 times higher area under the concentration time curve after benfotiamine administration than after thiamine nitrate. After benfotiamine administration, the peak plasma concentration of TDP exceeded that in healthy subjects by 51%. In the ESRD patients, after 24 h, the mean TDP concentration in erythrocytes increased from 158.7+/-30.9 ng/ml initially to 325.8+/-50.9 ng/ml after administration of benfotiamine and from 166.2+/-51.9 ng/ml to 200.5+/-50.0 ng/ml after thiamine nitrate administration. The ratio between the maximum erythrocyte TDP concentration and basal concentration was 2.66+/-0.6 in the benfotiamine group and 1.44+/-0.2 in the group receiving thiamine nitrate (P benfotiamine intake (P = 0.02) and from 3.71+/-0.8 microkat/l to 4.02+/-0.7 microkat/l after thiamine nitrate intake (P = 0.08). Likewise, alpha
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Paparrigopoulos, Thomas; Tzavellas, Elias; Karaiskos, Dimitris; Kouzoupis, Anastasios; Liappas, Ioannis
2010-01-01
Wernicke-Korsakoff syndrome (WKS) is a neuropsychiatric condition which results from thiamine deficiency, most commonly due to alcohol abuse. The prognosis of WKS is poor and its outcome depends mainly on prompt treatment. A 52-year-old male with a ten-year history of heavy alcohol abuse was admitted in hospital and treated for WKS. Ataxic and oculomotor symptoms promptly reversed following standard treatment but no change was observed in higher mental functioning. Although the protracted WK symptoms made the patient's improvement unlikely, aggressive treatment with thiamine (600 mg/day orally and 300 mg/day intramuscularly) fully reversed the condition within two months. Even though prolongation of undertreatment of WKS typically precludes significant improvement of symptoms due to irreversible damage of the brain, at least in some cases, higher thiamine doses (over 500 mg/day) for a longer period (at least three months) than usually recommended should be tried.
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Markova, Nataliia; Bazhenova, Nataliia; Anthony, Daniel C; Vignisse, Julie; Svistunov, Andrey; Lesch, Klaus-Peter; Bettendorff, Lucien; Strekalova, Tatyana
2017-04-03
Thiamine (vitamin B1) deficiency in the brain has been implicated in the development of dementia and symptoms of depression. Indirect evidence suggests that thiamine may contribute to these pathologies by controlling the activities of glycogen synthase kinase (GSK)-3β. While decreased GSK-3β activity appears to impair memory, increased GSK-3β activity is associated with the distressed/depressed state. However, hitherto direct evidence for the effects of thiamine on GSK-3β function has not been reported. Here, we administered thiamine or, the more bioavailable precursor, benfotiamine at 200mg/kg/day for 2weeks to C57BL/6J mice, to determine whether treatment might affect behaviours that are known to be sensitive to GSK-3β activity and whether such administration impacts on GSK-3β expression within the brain. The mice were tested in models of contextual conditioning and extinction, a 5-day rat exposure stress test, and a modified swim test with repeated testing. The tricyclic antidepressant imipramine (7.5mg/kg/day), was administered as a positive control for the effects of thiamine or benfotiamine. As for imipramine, both compounds inhibited the upregulation of GSK-3β induced by predator stress or repeated swimming, and reduced floating scores and the predator stress-induced behavioural changes in anxiety and exploration. Coincident, thiamine and benfotiamine improved learning and extinction of contextual fear, and the acquisition of the step-down avoidance task. Our data indicate that thiamine and benfotiamine have antidepressant/anti-stress effects in naïve animals that are associated with reduced GSK-3β expression and conditioning of adverse memories. Thus thiamine and benfotiamine may modulate GSK-3β functions in a manner that is dependent on whether the contextual conditioning is adaptive or maladaptive. Copyright © 2016 Elsevier Inc. All rights reserved.
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The Role of Thiamine Pyrophosphate in Prevention of Cisplatin Ototoxicity in an Animal Model
Directory of Open Access Journals (Sweden)
Ozan Kuduban
2013-01-01
Full Text Available Objective. The aim of this study was to evaluate the effectiveness of thiamine pyrophosphate against cisplatin-induced ototoxicity in guinea pigs. Materials and Methods. Healthy guinea pigs (n=18 were randomly divided into three groups. Group 1 (n=6 received an intraperitoneal injection of saline solution and cisplatin for 7 days, group 2 (n=6 received an intraperitoneal injection of thiamine pyrophosphate and cisplatin for 7 days, and group 3 (n=6 received only intraperitoneal injection of saline for 7 days. The animals in all groups were sacrificed under anesthesia, and their cochleas were harvested for morphological and biochemical observations. Results. In group 1, receiving only cisplatin, cochlear glutathione concentrations, superoxide dismutase, and glutathione peroxidase activities significantly decreased (P<0.05 and malondialdehyde concentrations significantly increased (P<0.05 compared to the control group. In group 2, receiving thiamine pyrophosphate and cisplatin, the concentrations of enzymes were near those of the control group. Microscopic examination showed that outer hair cells, spiral ganglion cells, and stria vascularis were preserved in group 2. Conclusion. Systemic administration of thiamine pyrophosphate yielded statistically significant protection to the cochlea of guinea pigs from cisplatin toxicity. Further experimental animal studies are essential to determine the appropriate indications of thiamine pyrophosphate before clinical use.
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Ortega, Rosa M; López-Sobaler, Ana M; Andrés, Pedro; Rodríguez-Rodríguez, Elena; Aparicio, Aránzazu; Bermejo, Laura M
2009-01-01
The aim of this study was to analyse the thiamine status in a group of young, overweight/obese women and to analyse the changes in this status produced by two weight-control programmes based on approximating the diet to the theoretical ideal, increasing the relative consumption of vegetables (V) or cereals (C) (especially breakfast cereals). The study subjects were 57 Spanish women with a body mass index (BMI) of 24-35 kg/m(2), all of whom were randomly assigned to one of two slightly hypocaloric diets. Dietetic, anthropometric and biochemical data were collected at the beginning of the study and at 2 and 6 weeks. C and V subjects showed a reduction in their energy intake, body weight and BMI, both at 2 and 6 weeks. Thiamine intake and blood thiamine levels only increased with diet C (both at 2 and 6 weeks). 21.8% cf the women, 21.8% had blood thiamine levels diet C, as compared with 30.8% of the women in diet V, showed blood thiamine levels diet C and diet V induced weight loss and a reduction in BMI, diet C would appear to be more useful in the maintenance of an adequate thiamine status.
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Directory of Open Access Journals (Sweden)
Padmanabhan Srinivasan
Full Text Available Thiamin (vitamin B1, a member of the water-soluble family of vitamins, is essential for normal cellular functions; its deficiency results in oxidative stress and mitochondrial dysfunction. Pancreatic acinar cells (PAC obtain thiamin from the circulation using a specific carrier-mediated process mediated by both thiamin transporters -1 and -2 (THTR-1 and THTR-2; encoded by the SLC19A2 and SLC19A3 genes, respectively. The aim of the current study was to examine the effect of chronic exposure of mouse PAC in vivo and human PAC in vitro to nicotine (a major component of cigarette smoke that has been implicated in pancreatic diseases on thiamin uptake and to delineate the mechanism involved. The results showed that chronic exposure of mice to nicotine significantly inhibits thiamin uptake in murine PAC, and that this inhibition is associated with a marked decrease in expression of THTR-1 and THTR-2 at the protein, mRNA and hnRNAs level. Furthermore, expression of the important thiamin-metabolizing enzyme, thiamin pyrophosphokinase (TPKase, was significantly reduced in PAC of mice exposed to nicotine. Similarly, chronic exposure of cultured human PAC to nicotine (0.5 μM, 48 h significantly inhibited thiamin uptake, which was also associated with a decrease in expression of THTR-1 and THTR-2 proteins and mRNAs. This study demonstrates that chronic exposure of PAC to nicotine impairs the physiology and the molecular biology of the thiamin uptake process. Furthermore, the study suggests that the effect is, in part, mediated through transcriptional mechanism(s affecting the SLC19A2 and SLC19A3 genes.
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Directory of Open Access Journals (Sweden)
Wenjun Huang
Full Text Available Herba epimedii (Epimedium, a traditional Chinese medicine, has been widely used as a kidney tonic and antirheumatic medicine for thousands of years. The bioactive components in herba epimedii are mainly prenylated flavonol glycosides, end-products of the flavonoid pathway. Epimedium species are also used as garden plants due to the colorful flowers and leaves. Many R2R3-MYB transcription factors (TFs have been identified to regulate the flavonoid and anthocyanin biosynthetic pathways. However, little is known about the R2R3-MYB TFs involved in regulation of the flavonoid pathway in Epimedium. Here, we reported the isolation and functional characterization of the first R2R3-MYB TF (EsMYBA1 from Epimedium sagittatum (Sieb. Et Zucc. Maxim. Conserved domains and phylogenetic analysis showed that EsMYBA1 belonged to the subgroup 6 clade (anthocyanin-related MYB clade of R2R3-MYB family, which includes Arabidopsis AtPAP1, apple MdMYB10 and legume MtLAP1. EsMYBA1 was preferentially expressed in leaves, especially in red leaves that contain higher content of anthocyanin. Alternative splicing of EsMYBA1 resulted in three transcripts and two of them encoded a MYB-related protein. Yeast two-hybrid and transient luciferase expression assay showed that EsMYBA1 can interact with several bHLH regulators of the flavonoid pathway and activate the promoters of dihydroflavonol 4-reductase (DFR and anthocyanidin synthase (ANS. In both transgenic tobacco and Arabidopsis, overexpression of EsMYBA1 induced strong anthocyanin accumulation in reproductive and/or vegetative tissues via up-regulation of the main flavonoid-related genes. Furthermore, transient expression of EsMYBA1 in E. sagittatum leaves by Agrobacterium infiltration also induced anthocyanin accumulation in the wounded area. This first functional characterization of R2R3-MYB TFs in Epimedium species will promote further studies of the flavonoid biosynthesis and regulation in medicinal plants.
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Huang, Wenjun; Sun, Wei; Lv, Haiyan; Luo, Ming; Zeng, Shaohua; Pattanaik, Sitakanta; Yuan, Ling; Wang, Ying
2013-01-01
Herba epimedii (Epimedium), a traditional Chinese medicine, has been widely used as a kidney tonic and antirheumatic medicine for thousands of years. The bioactive components in herba epimedii are mainly prenylated flavonol glycosides, end-products of the flavonoid pathway. Epimedium species are also used as garden plants due to the colorful flowers and leaves. Many R2R3-MYB transcription factors (TFs) have been identified to regulate the flavonoid and anthocyanin biosynthetic pathways. However, little is known about the R2R3-MYB TFs involved in regulation of the flavonoid pathway in Epimedium. Here, we reported the isolation and functional characterization of the first R2R3-MYB TF (EsMYBA1) from Epimedium sagittatum (Sieb. Et Zucc.) Maxim. Conserved domains and phylogenetic analysis showed that EsMYBA1 belonged to the subgroup 6 clade (anthocyanin-related MYB clade) of R2R3-MYB family, which includes Arabidopsis AtPAP1, apple MdMYB10 and legume MtLAP1. EsMYBA1 was preferentially expressed in leaves, especially in red leaves that contain higher content of anthocyanin. Alternative splicing of EsMYBA1 resulted in three transcripts and two of them encoded a MYB-related protein. Yeast two-hybrid and transient luciferase expression assay showed that EsMYBA1 can interact with several bHLH regulators of the flavonoid pathway and activate the promoters of dihydroflavonol 4-reductase (DFR) and anthocyanidin synthase (ANS). In both transgenic tobacco and Arabidopsis, overexpression of EsMYBA1 induced strong anthocyanin accumulation in reproductive and/or vegetative tissues via up-regulation of the main flavonoid-related genes. Furthermore, transient expression of EsMYBA1 in E. sagittatum leaves by Agrobacterium infiltration also induced anthocyanin accumulation in the wounded area. This first functional characterization of R2R3-MYB TFs in Epimedium species will promote further studies of the flavonoid biosynthesis and regulation in medicinal plants.
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DEFF Research Database (Denmark)
Liu, Chengwei; Tagami, Koichi; Minami, Atsushi
2015-01-01
KULNJ). Importantly, without conventional gene disruption, reconstitution of the biosynthetic machinery provided sufficient data to determine the pathway. It was thus demonstrated that the Aspergillus oryzae reconstitution system is a powerful method for studying the biosynthesis of complex natural products....
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Directory of Open Access Journals (Sweden)
Dian Anggraini Suroto
Full Text Available Phthoxazolin A, an oxazole-containing polyketide, has a broad spectrum of anti-oomycete activity and herbicidal activity. We recently identified phthoxazolin A as a cryptic metabolite of Streptomyces avermitilis that produces the important anthelmintic agent avermectin. Even though genome data of S. avermitilis is publicly available, no plausible biosynthetic gene cluster for phthoxazolin A is apparent in the sequence data. Here, we identified and characterized the phthoxazolin A (ptx biosynthetic gene cluster through genome sequencing, comparative genomic analysis, and gene disruption. Sequence analysis uncovered that the putative ptx biosynthetic genes are laid on an extra genomic region that is not found in the public database, and 8 open reading frames in the extra genomic region could be assigned roles in the biosynthesis of the oxazole ring, triene polyketide and carbamoyl moieties. Disruption of the ptxA gene encoding a discrete acyltransferase resulted in a complete loss of phthoxazolin A production, confirming that the trans-AT type I PKS system is responsible for the phthoxazolin A biosynthesis. Based on the predicted functional domains in the ptx assembly line, we propose the biosynthetic pathway of phthoxazolin A.
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Directory of Open Access Journals (Sweden)
Samiha Sioud
2007-01-01
Full Text Available We have previously isolated a new actinomycete strain from Tunisian soil called Streptomyces sp. US24, and have shown that it produces two bioactive molecules including a Cyclo (L-Phe, L-Pro diketopiperazine (DKP. To identify the structural genes responsible for the synthesis of this DKP derivative, a PCR amplification (696 bp was carried out using the Streptomyces sp. US24 genomic DNA as template and two degenerate oligonucleotides designed by analogy with genes encoding peptide synthetases (NRPS. The detection of DKP derivative biosynthetic pathway of the Streptomyces sp. US24 strain was then achieved by gene disruption via homologous recombination using a suicide vector derived from the conjugative plasmid pSET152 and containing the PCR product. Chromatography analysis, biological tests and spectroscopic studies of supernatant cultures of the wild-type Streptomyces sp. US24 strain and three mutants obtained by this gene targeting disruption approach showed that the amplified DNA fragment is required for Cyclo (L-Phe, L-Pro biosynthesis in Streptomyces sp. US24 strain. This DKP derivative seems to be produced either directly via a nonribosomal pathway or as a side product in the course of nonribosomal synthesis of a longer peptide.
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Directory of Open Access Journals (Sweden)
Çetin KADAKAL
2017-10-01
Full Text Available Abstract In this paper, the loss of L-ascorbic acid, thiamine and riboflavin in rosehip nectar with the heating periods (0, 5, 10, 15, 20 and 30 min at temperatures ranging from 70 to 95 °C is analyzed and experimental results are presented. Firstly, dried rosehip fruits were processed to rosehip nectar and then thermal treatment is performed. Liquid chromatographic (HPLC method was used for the analysis of the contents of L-ascorbic acid, thiamine and riboflavin and examined compounds are thoroughly separated within 25 min. During thermal processing, degradation of L-ascorbic acid, thiamine and riboflavin in rosehip nectar were fitted to a first-order reaction kinetic model. Arrhenius relationship was used for the description of temperature dependence of reaction. Activation energies for L-ascorbic acid, thiamine and riboflavin between 70 to 95 ºC were found to be 55.30, 36.38 and 37.15 kJ/mol, respectively. To the best of the author’s knowledge, due to lack of study on the thermal degradation of L-ascorbic acid, thiamine and riboflavin in rosehip nectar, this manuscript will be the first reported study to enable future analysis.
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Can long-term thiamine treatment improve the clinical outcomes of myotonic dystrophy type 1?
Directory of Open Access Journals (Sweden)
Antonio Costantini
2016-01-01
Full Text Available Myotonic dystrophy type 1, also known as Steinert′s disease, is an autosomal dominant disorder with multisystemic clinical features affecting the skeletal and cardiac muscles, the eyes, and the endocrine system. Thiamine (vitamin B1 is a cofactor of fundamental enzymes involved in the energetic cell metabolism; recent studies described its role in oxidative stress, protein processing, peroxisomal function, and gene expression. Thiamine deficiency is critical mainly in the central and peripheral nervous system, as well as in the muscular cells. Our aim was to investigate the potential therapeutical effects of long-term treatment with thiamine in myotonic dystrophy type 1 in an observational open-label pilot study. We described two patients with myotonic dystrophy type 1 treated with intramuscular thiamine 100 mg twice a week for 12 or 11 months. We evaluated the patients using the grading of muscle strength according to Medical Research Council (MRC, the Muscular Impairment Rating Scale (MIRS, and the Modified Barthel index. High-dose thiamine treatment was well tolerated and effective in improving the motor symptomatology, particularly the muscle strength evaluated with the MRC scale, and the patients′ activities of daily living using the Modified Barthel Index. At the end of treatment, the MRC score was 5 in the proximal muscles and 2-4 in the distal muscles (the MRC score before the treatment was 3-4 and 1-3, respectively. The MIRS grade improved by 25% compared to baseline for both patients. In patient #1, the Modified Barthel Index improved by 44%, and in patient #2 by 29%. These findings suggest that clinical outcomes are improved by long-term thiamine treatment.
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Can long-term thiamine treatment improve the clinical outcomes of myotonic dystrophy type 1?
Costantini, Antonio; Trevi, Erika; Pala, Maria Immacolata; Fancellu, Roberto
2016-09-01
Myotonic dystrophy type 1, also known as Steinert's disease, is an autosomal dominant disorder with multisystemic clinical features affecting the skeletal and cardiac muscles, the eyes, and the endocrine system. Thiamine (vitamin B1) is a cofactor of fundamental enzymes involved in the energetic cell metabolism; recent studies described its role in oxidative stress, protein processing, peroxisomal function, and gene expression. Thiamine deficiency is critical mainly in the central and peripheral nervous system, as well as in the muscular cells. Our aim was to investigate the potential therapeutical effects of long-term treatment with thiamine in myotonic dystrophy type 1 in an observational open-label pilot study. We described two patients with myotonic dystrophy type 1 treated with intramuscular thiamine 100 mg twice a week for 12 or 11 months. We evaluated the patients using the grading of muscle strength according to Medical Research Council (MRC), the Muscular Impairment Rating Scale (MIRS), and the Modified Barthel index. High-dose thiamine treatment was well tolerated and effective in improving the motor symptomatology, particularly the muscle strength evaluated with the MRC scale, and the patients' activities of daily living using the Modified Barthel Index. At the end of treatment, the MRC score was 5 in the proximal muscles and 2-4 in the distal muscles (the MRC score before the treatment was 3-4 and 1-3, respectively). The MIRS grade improved by 25% compared to baseline for both patients. In patient #1, the Modified Barthel Index improved by 44%, and in patient #2 by 29%. These findings suggest that clinical outcomes are improved by long-term thiamine treatment.
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Effects of thiamine on growth, aflatoxin production, and aflr gene expression in A. parasiticus
Directory of Open Access Journals (Sweden)
Ladan Nazemi
2015-01-01
Results: The minimum inhibitory concentration was yielded as > 500 mg/ml. However, HPLC analysis results showed that aflatoxin production reduced in samples treated with 500 mg/ml of thiamine. In addition, the level of afIR gene expression was significantly reduced after treating with 500 and 250 mg/ml of vitamin B1. Conclusion: Based on the obtained results, thiamine could not inhibit the fungal growth completely. However, the rate of afIR gene expression and aflatoxin production was significantly reduced after fungal treating with thiamine. Consequently, using natural compounds such as vitamins may be regarded as potential antitoxic agent in food industry and the industries related to agriculture.
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Thiamine for prevention and treatment of Wernicke-Korsakoff Syndrome in people who abuse alcohol.
Day, Ed; Bentham, Peter W; Callaghan, Rhiannon; Kuruvilla, Tarun; George, Sanju
2013-07-01
Autopsy studies suggest that Wernicke-Korsakoff syndrome (WKS) is not a rare disorder, particularly in individuals who abuse alcohol. Thiamine has been established as the treatment of choice for over 50 years, but uncertainty remains about appropriate dosage and duration. Current practice guidelines are based on case reports and clinical experience. This is an update of a review first published in 2004 and last updated in 2008. • To assess the efficacy of thiamine in preventing and treating the manifestations of WKS due to excess alcohol consumption. • To determine the optimum form, dose and duration of thiamine treatment for this indication. ALOIS, the Specialized Register of the Cochrane Dementia and Cognitive Improvement Group (CDCIG), The Cochrane Library, MEDLINE, EMBASE, PsycINFO, CINAHL and LILACS were searched on 6 September 2012 using the term thiamine OR aneurine. ALOIS contains records from all major health care databases (The Cochrane Library, MEDLINE, EMBASE, PsycINFO, CINAHL, LILACS) as well as from many trial databases and grey literature sources. Any randomised trials comparing thiamine with alternative interventions or comparing different thiamine regimens (varying in formulation, dose or duration of administration). All abstracts were independently inspected by two reviewers (ED and PWB), and relevant articles were retrieved and assessed for methodological quality using criteria provided in the Cochrane Handbook for Systematic Reviews of Interventions. Two studies were identified that met the inclusion criteria, but only one contained sufficient data for quantitative analysis. Ambrose (2001) randomly assigned participants (n = 107) to one of five doses of intramuscular thiamine and measured outcomes after 2 days of treatment. We compared the lowest dose (5 mg/day) with each of the other four doses. A significant difference favoured 200 mg/day compared with the 5-mg/day dose in determining the number of trials needed to meet inclusion criteria
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Beltramo, Elena; Berrone, Elena; Tarallo, Sonia; Porta, Massimo
2008-09-01
Thiamine (vitamin B1) is an essential cofactor in most organisms and is required at several stages of anabolic and catabolic intermediary metabolism, such as intracellular glucose metabolism, and is also a modulator of neuronal and neuro-muscular transmission. Lack of thiamine or defects in its intracellular transport can cause a number of severe disorders. Thiamine acts as a coenzyme for transketolase (TK) and for the pyruvate dehydrogenase and alpha-ketoglutarate dehydrogenase complexes, enzymes which play a fundamental role for intracellular glucose metabolism. In particular, TK is able to shift excess fructose-6-phosphate and glycerhaldeyde-3-phosphate from glycolysis into the pentose-phosphate shunt, thus eliminating these potentially damaging metabolites from the cytosol. Diabetes might be considered a thiamine-deficient state, if not in absolute terms at least relative to the increased requirements deriving from accelerated and amplified glucose metabolism in non-insulin dependent tissues that, like the vessel wall, are prone to complications. A thiamine/TK activity deficiency has been described in diabetic patients, the correction of which by thiamine and/or its lipophilic derivative, benfotiamine, has been demonstrated in vitro to counteract the damaging effects of hyperglycaemia on vascular cells. Little is known, however, on the positive effects of thiamine/benfotiamine administration in diabetic patients, apart from the possible amelioration of neuropathic symptoms. Clinical trials on diabetic patients would be necessary to test this vitamin as a potential and inexpensive approach to the prevention and/or treatment of diabetic vascular complications.
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Tarallo, Sonia; Beltramo, Elena; Berrone, Elena; Dentelli, Patrizia; Porta, Massimo
2010-06-01
Pericyte survival in diabetic retinopathy depends also on interactions with extracellular matrix (ECM) proteins, which are degraded by matrix metalloproteinases (MMP). Elevated glucose influences ECM turnover, through expression of MMP and their tissue inhibitors, TIMP. We reported on reduced pericyte adhesion to high glucose-conditioned ECM and correction by thiamine. We aimed at verifying the effects of thiamine and benfotiamine on MMP-2, MMP-9 and TIMP expression and activity in human vascular cells with high glucose. In HRP, MMP-2 activity, though not expression, increased with high glucose and decreased with thiamine and benfotiamine; TIMP-1 expression increased with high glucose plus thiamine and benfotiamine; MMP-9 was not expressed. In EC, MMP-9 and MMP-2 expression and activity increased with high glucose, but thiamine and benfotiamine had no effects; TIMP-1 expression was unchanged. Neither glucose nor thiamine modified TIMP-2 and TIMP-3 expression. TIMP-1 concentrations did not change in either HRP or EC. High glucose imbalances MMP/TIMP regulation, leading to increased ECM turnover. Thiamine and benfotiamine correct the increase in MMP-2 activity due to high glucose in HRP, while increasing TIMP-1.
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Xue, Fuguang; Pan, Xiaohua; Jiang, Linshu; Guo, Yuming; Xiong, Benhai
2018-01-01
Thiamine is known to attenuate high-concentrate diet induced subacute ruminal acidosis (SARA) in dairy cows, however, the underlying mechanisms remain unclear. The major objective of this study was to investigate the metabolic mechanisms of thiamine supplementation on high-concentrate diet induced SARA. Six multiparous, rumen-fistulated Holstein cows were used in a replicated 3 × 3 Latin square design. The treatments included a control diet (CON; 20% starch, dry matter basis), a SARA-inducing diet (SAID; 33.2% starch, dry matter basis) and SARA-inducing diet supplemented with 180 mg of thiamine/kg of dry matter intake (SAID + T). On d21 of each period, ruminal fluid samples were collected at 3 h post feeding, and GC/MS was used to analyze rumen fluid samples. PCA and OPLS-DA analysis demonstrated that the ruminal metabolite profile were different in three treatments. Compared with CON treatment, SAID feeding significantly decreased rumen pH, acetate, succinic acid, increased propionate, pyruvate, lactate, glycine and biogenic amines including spermidine and putrescine. Thiamine supplementation significantly decreased rumen content of propionate, pyruvate, lactate, glycine and spermidine; increase rumen pH, acetate and some medium-chain fatty acids. The enrichment analysis of different metabolites indicated that thiamine supplementation mainly affected carbohydrates, amino acids, pyruvate and thiamine metabolism compared with SAID treatment. These findings revealed that thiamine supplementation could attenuate high-concentrate diet induced SARA by increasing pyruvate formate-lyase activity to promote pyruvate to generate acetyl-CoA and inhibit lactate generation. Besides, thiamine reduced biogenic amines to alleviate ruminal epithelial inflammatory response.
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Liu, Dexiang; Ke, Zunji; Luo, Jia
2017-09-01
Thiamine (vitamin B1) is an essential nutrient and indispensable for normal growth and development of the organism due to its multilateral participation in key biochemical and physiological processes. Humans must obtain thiamine from their diet since it is synthesized only in bacteria, fungi, and plants. Thiamine deficiency (TD) can result from inadequate intake, increased requirement, excessive deletion, and chronic alcohol consumption. TD affects multiple organ systems, including the cardiovascular, muscular, gastrointestinal, and central and peripheral nervous systems. In the brain, TD causes a cascade of events including mild impairment of oxidative metabolism, neuroinflammation, and neurodegeneration, which are commonly observed in neurodegenerative diseases, such as Alzheimer's disease (AD), Parkinson's disease (PD), and Huntington's disease (HD). Thiamine metabolites may serve as promising biomarkers for neurodegenerative diseases, and thiamine supplementations exhibit therapeutic potential for patients of some neurodegenerative diseases. Experimental TD has been used to model aging-related neurodegenerative diseases. However, to date, the cellular and molecular mechanisms underlying TD-induced neurodegeneration are not clear. Recent research evidence indicates that TD causes oxidative stress, endoplasmic reticulum (ER) stress, and autophagy in the brain, which are known to contribute to the pathogenesis of various neurodegenerative diseases. In this review, we discuss the role of oxidative stress, ER stress, and autophagy in TD-mediated neurodegeneration. We propose that it is the interplay of oxidative stress, ER stress, and autophagy that contributes to TD-mediated neurodegeneration.
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International Nuclear Information System (INIS)
Tanhindarto, Rindy P.; Fox, J.B.; Lakritz, L.; Thayer, D.W.
2002-01-01
An experiment have been conducted on the effect of gamma irradiation on thiamine and riboflavin in fresh tuna and salmon. Samples were irradiated at (20 ± 0,5) oC by gamma rays with doses of 0; 2,5 and 5 kGy. The purpose of the present experiment was to study on changes of thiamine and riboflavin contents on extraction of fresh tuna and salmon after and before irradiation. The results showed that irradiation up to 2,5 kGy changed the thiamine and riboflavin contents on fresh tuna as well as their destruction which was significantly increasing caused by irradiated treatment with the dose of 5 kGy. The fresh salmon did not significantly change of the thiamine and riboflavin contents treated by irradiation up to 5 kGy
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Gonzalez, Antonio; Zhao, Mingzhe; Leavitt, John M; Lloyd, Alan M
2008-03-01
In all higher plants studied to date, the anthocyanin pigment pathway is regulated by a suite of transcription factors that include Myb, bHLH and WD-repeat proteins. However, in Arabidopsis thaliana, the Myb regulators remain to be conclusively identified, and little is known about anthocyanin pathway regulation by TTG1-dependent transcriptional complexes. Previous overexpression of the PAP1 Myb suggested that genes from the entire phenylpropanoid pathway are targets of regulation by Myb/bHLH/WD-repeat complexes in Arabidopsis, in contrast to other plants. Here we demonstrate that overexpression of Myb113 or Myb114 results in substantial increases in pigment production similar to those previously seen as a result of over-expression of PAP1, and pigment production in these overexpressors remains TTG1- and bHLH-dependent. Also, plants harboring an RNAi construct targeting PAP1 and three Myb candidates (PAP2, Myb113 and Myb114) showed downregulated Myb gene expression and obvious anthocyanin deficiencies. Correlated with these anthocyanin deficiencies is downregulation of the same late anthocyanin structural genes that are downregulated in ttg1 and bHLH anthocyanin mutants. Expression studies using GL3:GR and TTG1:GR fusions revealed direct regulation of the late biosynthetic genes only. Functional diversification between GL3 and EGL3 with regard to activation of gene targets was revealed by GL3:GR studies in single and double bHLH mutant seedlings. Expression profiles for Myb and bHLH regulators are also presented in the context of pigment production in young seedlings.
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Costa, Nara Aline; Gut, Ana Lúcia; de Souza Dorna, Mariana; Pimentel, José Alexandre Coelho; Cozzolino, Silvia Maria Franciscato; Azevedo, Paula Schmidt; Fernandes, Ana Angélica Henrique; Zornoff, Leonardo Antonio Mamede; de Paiva, Sergio Alberto Rupp; Minicucci, Marcos Ferreira
2014-04-01
The purpose of the study is to determine the influence of serum thiamine, glutathione peroxidase (GPx) activity, and serum protein carbonyl concentrations in hospital mortality in patients with septic shock. This prospective study included all patients with septic shock on admission or during intensive care unit (ICU) stay, older than 18 years, admitted to 1 of the 3 ICUs of the Botucatu Medical School, from January to August 2012. Demographic information, clinical evaluation, and blood sample were taken within the first 72 hours of the patient's admission or within 72 hours after septic shock diagnosis for serum thiamine, GPx activity, and protein carbonyl determination. One hundred eight consecutive patients were evaluated. The mean age was 57.5 ± 16.0 years, 63% were male, 54.6% died in the ICU, and 71.3% had thiamine deficiency. Thiamine was not associated with oxidative stress. Neither vitamin B1 levels nor the GPx activity was associated with outcomes in these patients. However, protein carbonyl concentration was associated with increased mortality. In patients with septic shock, oxidative stress was associated with mortality. On the other hand, thiamine was not associated with oxidative stress or mortality in these patients. Copyright © 2014 Elsevier Inc. All rights reserved.
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Dias, Fernando M V; Silva, Danielle Marra de Freitas; Doyle, Flavia Costa de Proença; Ribeiro, Angela Maria
2013-01-01
The acquisition of cognitive, sensory-motor and social emotional functions depend on a proper development of the Central Nervous System (CNS). This set of functions, known as intelligence, allows a better adaptation to the environment. In the last decades, an increase in the average of intelligence has been reported. However, such an increase cannot be observed in an equivalent way in economically and social underprivileged regions. Children from those regions are in great risk of being affected by mental retardation or impaired cognitive development. In later life they will, probably, be unable to transform and improve themselves and their communities, perpetuating the poverty of the region. Therefore, knowledge of factors involved in CNS development is a matter of health closely related to social improvement. Malnutrition throughout pregnancy and breastfeeding is clearly identifiable as a cause of damage in CNS development. Vitamin B1 (Thiamine) is a micronutrient important to the growth and maturity of the CNS. Thiamine shortcoming may affect 50% of pregnant women. Thiamine function in cerebral development is still not well known. There is a gap in the literature regarding systematical research about the blood thiamine concentration throughout the periods of gestation and breastfeeding. These studies are relevant in populations with a high level of nutritional vulnerability, because in a follow up offspring cognitive exam they could reveal if the maternal thiamine deficiency is related to child CNS impairment. This paper introduce the hypothesis that thiamine shortcoming during pregnancy and breastfeeding is directly related to cognitive impairment of child. Data about the neurophysiological role of thiamine, consequences of its shortcoming in experimental models, populations under the risk of thiamine shortcoming are presented. The hypothesis that maternal thiamine shortcoming causes damage related to child cognitive development needs to be considered. Thus
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DEFF Research Database (Denmark)
Jakobsen, Jette
2008-01-01
The aim of this study was first to optimise and validate a method using an enzyme-mixture to liberate protein- and phosphate-bound thiamin and riboflavin in food by the use of ultrasonication and HPLC, and second to include the quantitation of the vitamin B-1 active compound 2-(1-hydroxyethyl...
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Kudo, Fumitaka; Matsuura, Yasunori; Hayashi, Takaaki; Fukushima, Masayuki; Eguchi, Tadashi
2016-07-01
Sordarin is a glycoside antibiotic with a unique tetracyclic diterpene aglycone structure called sordaricin. To understand its intriguing biosynthetic pathway that may include a Diels-Alder-type [4+2]cycloaddition, genome mining of the gene cluster from the draft genome sequence of the producer strain, Sordaria araneosa Cain ATCC 36386, was carried out. A contiguous 67 kb gene cluster consisting of 20 open reading frames encoding a putative diterpene cyclase, a glycosyltransferase, a type I polyketide synthase, and six cytochrome P450 monooxygenases were identified. In vitro enzymatic analysis of the putative diterpene cyclase SdnA showed that it catalyzes the transformation of geranylgeranyl diphosphate to cycloaraneosene, a known biosynthetic intermediate of sordarin. Furthermore, a putative glycosyltransferase SdnJ was found to catalyze the glycosylation of sordaricin in the presence of GDP-6-deoxy-d-altrose to give 4'-O-demethylsordarin. These results suggest that the identified sdn gene cluster is responsible for the biosynthesis of sordarin. Based on the isolated potential biosynthetic intermediates and bioinformatics analysis, a plausible biosynthetic pathway for sordarin is proposed.
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Portari, Guilherme Vannucchi; Ovidio, Paula Payão; Deminice, Rafael; Jordão, Alceu Afonso
2016-10-01
The aim of this study was to evaluate possible beneficial effects of treatment with thiamine or benfotiamine in an animal model of acute ethanol intoxication. Thirty male Wistar rats were separated at random into three groups of 10 animals each: Ethanol (E), Ethanol treated with thiamine (T) and Ethanol treated with benfotiamine (BE). Rats were gavaged with single dose of ethanol (5g/kg, 40% v:v). After 30min of ethanol gavage the animals were treated with thiamine or benfotiamine. Six hours after first gavage, the animals were euthanized and blood and liver samples were collected for ethanol and oxidative stress biomarkers quantification. Serum ethanol levels were higher in animals treated with thiamine or benfotiamine while hepatic alcohol levels were higher in animals of the group treated with benfotiamine comparing to controls or thiamine treated groups. The lipid peroxidation biomarkers were diminished for the groups treated with thiamine or benfotiamine comparing to E animals. Concerning protein oxidative damage parameters, they were enhanced for animals treated with benfotiamine in relation to other groups. In conclusion, the treatment with thiamine or benfotiamine even 30min after the massive dose of ethanol has proven to be beneficial against liver damage. Improved results were obtained with benfotiamine in relation to oxidative damage from aqueous compartments. Copyright © 2016 Elsevier Inc. All rights reserved.
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Directory of Open Access Journals (Sweden)
Gutha Linga R
2010-08-01
Full Text Available Abstract Background Symptoms of grapevine leafroll disease (GLRD in red-fruited wine grape (Vitis vinifera L. cultivars consist of green veins and red and reddish-purple discoloration of inter-veinal areas of leaves. The reddish-purple color of symptomatic leaves may be due to the accumulation of anthocyanins and could reflect an up-regulation of genes involved in their biosynthesis. Results We examined six putative constitutively expressed genes, Ubiquitin, Actin, GAPDH, EF1-a, SAND and NAD5, for their potential as references for normalization of gene expression in reverse transcription-quantitative real-time polymerase chain reaction (RT-qPCR. Using the geNorm program, a combination of two genes (Actin and NAD5 was identified as the stable set of reference genes for normalization of gene expression data obtained from grapevine leaves. By using gene-specific RT-qPCR in combination with a reliable normalization factor, we compared relative expression of the flavonoid biosynthetic pathway genes between leaves infected with Grapevine leafroll-associated virus 3 (GLRaV-3 and exhibiting GLRD symptoms and virus-free green leaves obtained from a red-fruited wine grape cultivar (cv. Merlot. The expression levels of these different genes ranged from two- to fifty-fold increase in virus-infected leaves. Among them, CHS3, F3'5'H, F3H1, LDOX, LAR1 and MybA1 showed greater than 10-fold increase suggesting that they were expressed at significantly higher levels in virus-infected symptomatic leaves. HPLC profiling of anthocyanins extracted from leaves indicated the presence of cyanidin-3-glucoside and malvidin-3-glucoside only in virus-infected symptomatic leaves. The results also showed 24% higher levels of flavonols in virus-infected symptomatic leaves than in virus-free green leaves, with quercetin followed by myricetin being the predominant compounds. Proanthocyanidins, estimated as total tannins by protein precipitation method, were 36% higher in virus
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International Nuclear Information System (INIS)
Reinold, S.; Hahlbrock, K.
1997-01-01
Using in situ RNA/RNA hybridization, enzyme immunolocalization, and histochemical techniques, several phenylpropanoid biosynthetic activities and products were localized in tissue sections from various aerial parts of parsley (Petroselinum crispum) plants at different developmental stages. The enzymes and corresponding mRNAs analyzed included two representatives of general phenylpropanoid metabolism: phenylalanine ammonia-lyase (PAL) and 4-coumarate: CoA ligase (4CL), and one representative each from two distinct branch pathways: chalcone synthase (CHS; flavonoids) and S-adenosyl-L-methionine: bergaptol O-methyltransferase (BMT; furanocoumarins). In almost all cases, the relative timing of accumulation differed greatly for mRNA and protein and indicated short expression periods and short half-lives for all mRNAs as compared to the proteins. PAL and 4CL occurred almost ubiquitously in cell type-specific patterns, and their mRNAs and proteins were always coordinately expressed, whereas the cell type-specific localization of flavonoid and furanocoumarin biosynthetic activities was to a large extent mutually exclusive. However, the distribution patterns of CHS and BMT, when superimposed, closely matched those of PAL and 4CL in nearly all tissues analysed, suggesting that the flavonoid and furanocoumarin pathways together constituted a large majority of the total phenylpropanoid biosynthetic activity. Differential sites of synthesis and accumulation indicating intercellular translocation were observed both for flavonoids and for furanocoumarins in oil ducts and the surrounding tissue. The widespread occurrence of both classes of compounds, as well as selected, pathway-specific mRNAs and enzymes, in many cell types of all parsley organs including various flower parts suggests additional functions beyond the previously established roles of flavonoids in UV protection and furanocoumarins in pathogen defence. (author)
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Sepulveda, M.S.; Wiebe, J.J.; Honeyfield, D.C.; Rauschenberger, H.R.; Hinterkopf, J.P.; Johnson, W.E.; Gross, T.S.
2004-01-01
Thiamine deficiency has been linked to early mortality syndrome in salmonids in the Great Lakes. This study was conducted to compare thiamine concentrations in American alligators (Alligator mississippiensis) and Florida largemouth bass (Micropterus salmoides floridanus) eggs from sites with high embryo mortality and high exposure to organochlorine pesticides (OCPs) (Lakes Apopka and Griffin, and Emeralda Marsh, Florida, USA) to those from sites that have historically exhibited low embryo mortality and low OCPs (Lakes Woodruff and Orange, Florida). During June-July 2000, 20 alligator clutches were collected from these sites, artificially incubated, and monitored for embryo mortality. Thiamine and OCPs were measured in one egg/clutch. During February 2002, 10 adult female bass were collected from Emeralda Marsh and Lake Woodruff and mature ovaries analyzed for thiamine and OCP concentrations. Although ovaries from the Emeralda Marsh bass contained almost 1,000-fold more OCPs compared with the reference site, Lake Woodruff, there were no differences in thiamine concentrations between sites (11,710 vs. 11,857 pmol/g). In contrast, alligator eggs from the reference site had five times the amount of thiamine compared with the contaminated sites (3,123 vs. 617 pmol/g). Similarly, clutches with > 55% hatch rates had significantly higher concentrations of thiamine compared with clutches with eggs. ?? Wildlife Disease Association 2004.
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Directory of Open Access Journals (Sweden)
Сніжана Сергіївна Чернадчук
2015-05-01
Full Text Available It is studied an aminotransferase activity during injection of thiamin bromide in rat tissues in normal and hypoxic enclosed space. After injection of thiamine bromide we have set reduction of AST and ALT activity, relative to control, except by the brain tissue, where there was an increase of investigated indicators. The decrease of activity of the investigated elements is occurred in animals which before hypoxia were injection of thiamine bromide
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Chen, Zhenya; Shen, Xiaolin; Wang, Jian; Wang, Jia; Yuan, Qipeng; Yan, Yajun
2017-11-01
Gallic acid (GA) is a naturally occurring phytochemical that has strong antioxidant and antibacterial activities. It is also used as a potential platform chemical for the synthesis of diverse high-value compounds. Hydrolytic degradation of tannins by acids, bases or microorganisms serves as a major way for GA production, which however, might cause environmental pollution and low yield and efficiency. Here, we report a novel approach for efficient microbial production of GA. First, structure-based rational engineering of PobA, a p-hydroxybenzoate hydroxylase from Pseudomonas aeruginosa, generated a new mutant, Y385F/T294A PobA, which displayed much higher activity toward 3,4-dihydroxybenzoic acid (3,4-DHBA) than the wild-type and any other reported mutants. Remarkably, expression of this mutant in Escherichia coli enabled generation of 1149.59 mg/L GA from 1000 mg/L 4-hydroxybenzoic acid (4-HBA), representing a 93% molar conversion ratio. Based on that, we designed and reconstituted a novel artificial biosynthetic pathway of GA and achieved 440.53 mg/L GA production from simple carbon sources in E. coli. Further enhancement of precursor supply through reinforcing shikimate pathway was able to improve GA de novo production to 1266.39 mg/L in shake flasks. Overall, this study not only led to the development of a highly active PobA variant for hydroxylating 3,4-DHBA into GA via structure-based protein engineering approach, but also demonstrated a promising pathway for bio-based manufacturing of GA and its derived compounds. Biotechnol. Bioeng. 2017;114: 2571-2580. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.
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International Nuclear Information System (INIS)
Chuaqui-Offermanns, N.; Shoemaker, L.; McDougall, T.
1989-01-01
The effects of heat and radiation on thiamine stability are being studied both singly and in combination. Heat, γ-radiation and a combination of them were applied to a model system consisting of 2 x 10 -5 M thiamine hydrochloride in 0.01N HCl (pH=2.5), and their effects are reported. The effects of these two agents on thiamine in two food matrices, concentrated orange juice and green peas, are also reported. Heat was not found to have a significant effect on thiamine in the model system at temperatures up to 120 0 C for up to 60 min of treatment. A small, but significant heat effect was found in the two foods. The retention of thiamine in the model system and in the two foods decreased exponentially as the radiation dose increased. The degradation of thiamine by γ-radiation in both foods was a factor of 10 less than that observed in the model system. A small, but significant synergistic effect was found when samples of the model system were heated at 120 0 C for one hour 24 h after irradiation. (author)
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Nahas, John V; Iosue, Christine L; Shaik, Noor F; Selhorst, Kathleen; He, Bin Z; Wykoff, Dennis D
2018-05-10
Convergent evolution is often due to selective pressures generating a similar phenotype. We observe relatively recent duplications in a spectrum of Saccharomycetaceae yeast species resulting in multiple phosphatases that are regulated by different nutrient conditions - thiamine and phosphate starvation. This specialization is both transcriptional and at the level of phosphatase substrate specificity. In Candida glabrata , loss of the ancestral phosphatase family was compensated by the co-option of a different histidine phosphatase family with three paralogs. Using RNA-seq and functional assays, we identify one of these paralogs, CgPMU3 , as a thiamine phosphatase. We further determine that the 81% identical paralog CgPMU2 does not encode thiamine phosphatase activity; however, both are capable of cleaving the phosphatase substrate, 1-napthyl-phosphate. We functionally demonstrate that members of this family evolved novel enzymatic functions for phosphate and thiamine starvation, and are regulated transcriptionally by either nutrient condition, and observe similar trends in other yeast species. This independent, parallel evolution involving two different families of histidine phosphatases suggests that there were likely similar selective pressures on multiple yeast species to recycle thiamine and phosphate. In this work, we focused on duplication and specialization, but there is also repeated loss of phosphatases, indicating that the expansion and contraction of the phosphatase family is dynamic in many Ascomycetes. The dynamic evolution of the phosphatase gene families is perhaps just one example of how gene duplication, co-option, and transcriptional and functional specialization together allow species to adapt to their environment with existing genetic resources. Copyright © 2018, G3: Genes, Genomes, Genetics.
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Anttila, Matthew; Strangman, Wendy; York, Robert; Tomas, Carmelo; Wright, Jeffrey L C
2016-03-25
Understanding the biosynthesis of dinoflagellate polyketides presents many unique challenges. Because of the remaining hurdles to dinoflagellate genome sequencing, precursor labeling studies remain the only viable way to investigate dinoflagellate biosynthesis. However, prior studies have shown that polyketide chain assembly does not follow any of the established processes. Additionally, acetate, the common precursor for polyketides, is frequently scrambled, thus compromising interpretation. These factors are further compounded by low production yields of the compounds of interest. A recent report on the biosynthesis of spirolides, a group belonging to the growing class of toxic spiroimines, provided some insight into the polyketide assembly process based on acetate labeling studies, but many details were left uncertain. By feeding (13)C methyl-labeled methionine to cultures of Alexandrium ostenfeldii, the producing organism of 13-desmethylspirolide C, and application of the odd-even methylation rule, the complete biosynthetic pathway has been established.
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Directory of Open Access Journals (Sweden)
Ahmad-Faris Seman-Kamarulzaman
Full Text Available Juvenile Hormone III is of great concern due to negative effects on major developmental and reproductive maturation in insect pests. Thus, the elucidation of enzymes involved JH III biosynthetic pathway has become increasing important in recent years. One of the enzymes in the JH III biosynthetic pathway that remains to be isolated and characterized is farnesal dehydrogenase, an enzyme responsible to catalyze the oxidation of farnesal into farnesoic acid. A novel NAD+-farnesal dehydrogenase of Polygonum minus was purified (315-fold to apparent homogeneity in five chromatographic steps. The purification procedures included Gigacap S-Toyopearl 650M, Gigacap Q-Toyopearl 650M, and AF-Blue Toyopearl 650ML, followed by TSK Gel G3000SW chromatographies. The enzyme, with isoelectric point of 6.6 is a monomeric enzyme with a molecular mass of 70 kDa. The enzyme was relatively active at 40°C, but was rapidly inactivated above 45°C. The optimal temperature and pH of the enzyme were found to be 35°C and 9.5, respectively. The enzyme activity was inhibited by sulfhydryl agent, chelating agent, and metal ion. The enzyme was highly specific for farnesal and NAD+. Other terpene aldehydes such as trans- cinnamaldehyde, citral and α- methyl cinnamaldehyde were also oxidized but in lower activity. The Km values for farnesal, citral, trans- cinnamaldehyde, α- methyl cinnamaldehyde and NAD+ were 0.13, 0.69, 0.86, 1.28 and 0.31 mM, respectively. The putative P. minus farnesal dehydrogenase that's highly specific towards farnesal but not to aliphatic aldehydes substrates suggested that the enzyme is significantly different from other aldehyde dehydrogenases that have been reported. The MALDI-TOF/TOF-MS/MS spectrometry further identified two peptides that share similarity to those of previously reported aldehyde dehydrogenases. In conclusion, the P. minus farnesal dehydrogenase may represent a novel plant farnesal dehydrogenase that exhibits distinctive substrate
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Directory of Open Access Journals (Sweden)
Wenjun eHuang
2015-09-01
Full Text Available Herba epimedii (Epimedium, a traditional Chinese medicine, has been widely used as a kidney tonic and antirheumatic medicine for thousands of years. In Epimedium, flavonoids have been demonstrated to be the main bioactive components (BCs. However, the molecular biosynthetic and regulatory mechanisms of flavonoid-derived BCs remain obscure. In this study, we isolated twelve structural genes and two putative transcription factors (TFs in the flavonoid pathway. Phytochemical analysis showed that the total content of four representative BCs (epimedin A, B, C and icariin decreased slightly or dramatically in two lines of E. sagittatum during leaf development. Transcriptional analysis revealed that two R2R3-MYB TFs (EsMYBA1 and EsMYBF1, together with a bHLH TF (EsGL3 and WD40 protein (EsTTG1, were supposed to coordinately regulate the anthocyanin and flavonol-derived BCs biosynthesis in leaves. Overexpression of EsFLS (flavonol synthase in tobacco resulted in increased flavonols content and decreased anthocyanins content in flowers. Moreover, EsMYB12 negatively correlated with the accumulation of the four BCs, and might act as a transcriptional repressor in the flavonoid pathway. Therefore, the anthocyanin pathway may coordinate with the flavonol-derived BCs pathway in Epimedium leaves. A better understanding of the flavonoid biosynthetic and regulatory mechanisms in E. sagittatum will facilitate functional characterization, metabolic engineering and molecular breeding studies of Epimedium species.
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Thiamin, riboflavin and alpha-tocopherol retention in processed and stored irradiated pork
International Nuclear Information System (INIS)
Fox, J.B. Jr.; Lakritz, L.; Thayer, D.W.
1997-01-01
Combination treatments for preservation of irradiated pork were investigated with respect to vitamin loss. Ground pork was prepared under nitrogen and packaged in anaerobic foil. The samples were enzyme denatured by heating before and after irradiation, then cooked and stored. Irradiation resulted in thiamin loss, but neither riboflavin nor alpha-tocopherol was affected. Neither thiamin nor riboflavin was affected by heat denaturation, cooking or storage, but heating and cooking increased the measured alpha-tocopherol. The lack of loss of the vitamins was attributed to the exclusion of oxygen
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Bahuguna, Rajeev Nayan; Joshi, Rohit; Shukla, Alok; Pandey, Mayank; Kumar, J
2012-08-01
A novel pathogen defense strategy by thiamine priming was evaluated for its efficacy against sheath blight pathogen, Rhizoctonia solani AG-1A, of rice and compared with that of systemic fungicide, carbendazim (BCM). Seeds of semidwarf, high yielding, basmati rice variety Vasumati were treated with thiamine (50 mM) and BCM (4 mM). The pot cultured plants were challenge inoculated with R. solani after 40 days of sowing and effect of thiamine and BCM on rice growth and yield traits was examined. Higher hydrogen peroxide content, total phenolics accumulation, phenylalanine ammonia lyase (PAL) activity and superoxide dismutase (SOD) activity under thiamine treatment displayed elevated level of systemic resistance, which was further augmented under challenging pathogen infection. High transcript level of phenylalanine ammonia lyase (PAL) and manganese superoxide dismutase (MnSOD) validated mode of thiamine primed defense. Though minimum disease severity was observed under BCM treatment, thiamine produced comparable results, with 18.12 per cent lower efficacy. Along with fortifying defense components and minor influence on photosynthetic pigments and nitrate reductase (NR) activity, thiamine treatment significantly reduced pathogen-induced loss in photosynthesis, stomatal conductance, chlorophyll fluorescence, NR activity and NR transcript level. Physiological traits affected under pathogen infection were found signatory for characterizing plant's response under disease and were detectable at early stage of infection. These findings provide a novel paradigm for developing alternative, environmentally safe strategies to control plant diseases. Copyright © 2012 Elsevier Masson SAS. All rights reserved.
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Sepúlveda, Maria S; Wiebe, Jon J; Honeyfield, Dale C; Rauschenberger, Heath R; Hinterkopf, Joy P; Johnson, William E; Gross, Timothy S
2004-10-01
Thiamine deficiency has been linked to early mortality syndrome in salmonids in the Great Lakes. This study was conducted to compare thiamine concentrations in American alligators (Alligator mississippiensis) and Florida largemouth bass (Micropterus salmoides floridanus) eggs from sites with high embryo mortality and high exposure to organochlorine pesticides (OCPs) (Lakes Apopka and Griffin, and Emeralda Marsh, Florida, USA) to those from sites that have historically exhibited low embryo mortality and low OCPs (Lakes Woodruff and Orange, Florida). During June-July 2000, 20 alligator clutches were collected from these sites, artificially incubated, and monitored for embryo mortality. Thiamine and OCPs were measured in one egg/clutch. During February 2002, 10 adult female bass were collected from Emeralda Marsh and Lake Woodruff and mature ovaries analyzed for thiamine and OCP concentrations. Although ovaries from the Emeralda Marsh bass contained almost 1,000-fold more OCPs compared with the reference site, Lake Woodruff, there were no differences in thiamine concentrations between sites (11,710 vs. 11,857 pmol/g). In contrast, alligator eggs from the reference site had five times the amount of thiamine compared with the contaminated sites (3,123 vs. 617 pmol/g). Similarly, clutches with >55% hatch rates had significantly higher concentrations of thiamine compared with clutches with reproductive failure and recruitment of largemouth bass. The cause(s) of this thiamine deficiency are unknown but might be related to differences in the nutritional value of prey items across the sites studied and/or to the presence of high concentration of contaminants in eggs.
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International Nuclear Information System (INIS)
Meghal, S.K.; O'Neal, R.M.; Koeppe, R.E.
1977-01-01
Rats were fed either a thiamine-deficient diet or diets containing pyrithiamine or oxythiamine. When symptoms of thiamine deficiency appeared, the animals were injected intraperitoneally with [2- 14 C] pyruvate six to twelve minutes prior to sacrifice. Free glutamic and aspartic acids were isolated from liver and brain and degraded. The results indicate that, in thiamine-deficient or oxythiamine-treated rats, pyruvate metabolism in liver and brain is similar to that in normal animals. In contrast, pyrithinamine drastically decreases the oxidative decarboxylation of pyruvate by rat liver. (auth.)
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Directory of Open Access Journals (Sweden)
Wei Sun
Full Text Available Chalcone synthase (CHS catalyzes the first committed step in the flavonoid biosynthetic pathway. In this study, the cDNA (FhCHS1 encoding CHS from Freesia hybrida was successfully isolated and analyzed. Multiple sequence alignments showed that both the conserved CHS active site residues and CHS signature sequence were found in the deduced amino acid sequence of FhCHS1. Meanwhile, crystallographic analysis revealed that protein structure of FhCHS1 is highly similar to that of alfalfa CHS2, and the biochemical analysis results indicated that it has an enzymatic role in naringenin biosynthesis. Moreover, quantitative real-time PCR was performed to detect the transcript levels of FhCHS1 in flowers and different tissues, and patterns of FhCHS1 expression in flowers showed significant correlation to the accumulation patterns of anthocyanin during flower development. To further characterize the functionality of FhCHS1, its ectopic expression in Arabidopsis thaliana tt4 mutants and Petunia hybrida was performed. The results showed that overexpression of FhCHS1 in tt4 mutants fully restored the pigmentation phenotype of the seed coats, cotyledons and hypocotyls, while transgenic petunia expressing FhCHS1 showed flower color alteration from white to pink. In summary, these results suggest that FhCHS1 plays an essential role in the biosynthesis of flavonoid in Freesia hybrida and may be used to modify the components of flavonoids in other plants.
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Anaphylactic shock and cardiac arrest caused by thiamine infusion
DEFF Research Database (Denmark)
Juel, Jacob; Pareek, Manan; Langfrits, Christian Sigvald
2013-01-01
intoxication and developed cardiac arrest due to anaphylactic shock following intravenous thiamine infusion. The patient was successfully resuscitated after 15 min and repeated epinephrine administrations. He was discharged in good health after 14 days. This case report emphasises both the importance...
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Bunik, Victoria I; Tylicki, Adam; Lukashev, Nikolay V
2013-12-01
Bringing a knowledge of enzymology into research in vivo and in situ is of great importance in understanding systems biology and metabolic regulation. The central metabolic significance of thiamin (vitamin B1 ) and its diphosphorylated derivative (thiamin diphosphate; ThDP), and the fundamental differences in the ThDP-dependent enzymes of metabolic networks in mammals versus plants, fungi and bacteria, or in health versus disease, suggest that these enzymes are promising targets for biotechnological and medical applications. Here, the in vivo action of known regulators of ThDP-dependent enzymes, such as synthetic structural analogs of the enzyme substrates and thiamin, is analyzed in light of the enzymological data accumulated during half a century of research. Mimicking the enzyme-specific catalytic intermediates, the phosphonate analogs of 2-oxo acids selectively inhibit particular ThDP-dependent enzymes. Because of their selectivity, use of these compounds in cellular and animal models of ThDP-dependent enzyme malfunctions improves the validity of the model and its predictive power when compared with the nonselective and enzymatically less characterized oxythiamin and pyrithiamin. In vitro studies of the interaction of thiamin analogs and their biological derivatives with potential in vivo targets are necessary to identify and attenuate the analog selectivity. For both the substrate and thiamin synthetic analogs, in vitro reactivities with potential targets are highly relevant in vivo. However, effective concentrations in vivo are often higher than in vitro studies would suggest. The significance of specific inihibition of the ThDP-dependent enzymes for the development of herbicides, antibiotics, anticancer and neuroprotective strategies is discussed. © 2013 FEBS.
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Stracke, H; Hammes, H P; Werkmann, D; Mavrakis, K; Bitsch, I; Netzel, M; Geyer, J; Köpcke, W; Sauerland, C; Bretzel, R G; Federlin, K F
2001-01-01
In rats with streptozotocin (STZ) induced diabetes the effect of (watersoluble) thiamine nitrate and of (lipidsoluble) benfotiamine on peripheral nerve function (motor nerve conduction velocity) as well as on the formation of advanced glycation end-products in peripheral nerve tissue was studied. In one group of animals drug administration was started immediately after diabetes induction (prevention study) and in another group two months after diabetes induction (treatment study). Motor nerve conduction velocity (NCV) dropped by 10.5% in diabetic animals, carboxymethyl-lysine (CML) rose to a 3.5fold concentration, deoxyglucosone (3DG)-type AGE formation was increased 5.1fold compared with controls. After three months preventive administration of both vitamin B(1) preparations NCV had increased substantially compared with results in diabetic controls. It was nearly normal after six months with benfotiamine, while the administration of thiamine nitrate resulted in no further amelioration. NCV was nearly normalized after six months of benfotiamine application but not with thiamine. Furthermore, benfotiamine induced a major inhibition of neural imidazole-type AGE formation and completely prevented diabetes induced glycoxidation products (CML). Treatment with thiamine did not significantly affect AGE or cmL levels. Unlike treatment with water-soluble thiamine nitrate timely administration of liposoluble prodrug benfotiamine was effective in the prevention of functional damage and of AGE and cmL formation in nerves of diabetic rats.
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Energy Technology Data Exchange (ETDEWEB)
Chuaqui-Offermanns, N.; Shoemaker, L.; McDougall, T. (Atomic Energy of Canada Ltd., Pinawa, MB (Canada). Whiteshell Nuclear Research Establishment)
1989-01-01
The effects of heat and radiation on thiamine stability are being studied both singly and in combination. Heat, {gamma}-radiation and a combination of them were applied to a model system consisting of 2 x 10{sup -5}M thiamine hydrochloride in 0.01N HCl (pH=2.5), and their effects are reported. The effects of these two agents on thiamine in two food matrices, concentrated orange juice and green peas, are also reported. Heat was not found to have a significant effect on thiamine in the model system at temperatures up to 120{sup 0}C for up to 60 min of treatment. A small, but significant heat effect was found in the two foods. The retention of thiamine in the model system and in the two foods decreased exponentially as the radiation dose increased. The degradation of thiamine by {gamma}-radiation in both foods was a factor of 10 less than that observed in the model system. A small, but significant synergistic effect was found when samples of the model system were heated at 120{sup 0}C for one hour 24 h after irradiation. (author).
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Can long-term thiamine treatment improve the clinical outcomes of myotonic dystrophy type 1?
Costantini, Antonio; Trevi, Erika; Pala, Maria Immacolata; Fancellu, Roberto
2016-01-01
Myotonic dystrophy type 1, also known as Steinert′s disease, is an autosomal dominant disorder with multisystemic clinical features affecting the skeletal and cardiac muscles, the eyes, and the endocrine system. Thiamine (vitamin B1) is a cofactor of fundamental enzymes involved in the energetic cell metabolism; recent studies described its role in oxidative stress, protein processing, peroxisomal function, and gene expression. Thiamine deficiency is critical mainly in the central and periphe...
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Thiamine hydrochloride: An efficient catalyst for one-pot synthesis of ...
Indian Academy of Sciences (India)
thiamine hydrochloride (VB1) as an inexpensive, non-toxic and metal ion free catalyst at ambient temperature. Keywords. ... from practical applications due to environmental and economic ... filtered and purified by column chromatography on.
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International Nuclear Information System (INIS)
Katsumi Iida
2013-01-01
The biosynthetic pathways leading to δ-aminolevulinic acid (ALA) from the Shemin precursor glycine via the C5 pathway in Arthrobacter hyalinus were quantitatively evaluated by means of feeding experiments with [2- 13 C]glycine, sodium [1- 13 C]acetate, and sodium [2- 13 C]acetate, followed by analysis of the labeling patterns of coproporphyrinogen III (Copro'gen III) (biosynthesized from ALA) using 13 C NMR spectroscopy. Two biosynthetic pathways leading to ALA from glycine via the C5 pathway were identified: i.e., transformation of glycine to l-serine catalyzed by glycine hydroxymethyltransferase, and glycine synthase-catalyzed catabolism of glycine to N 5 , N 10 -methylene-tetrahydrofolic acid (THF), which reacts with another molecule of glycine to afford l-serine. l-Serine is transformed to acetyl-CoA via pyruvic acid. Acetyl-CoA enters the tricarboxylic acid cycle, affording 2-oxoglutaric acid, which in turn is transformed to l-glutamic acid. The l-glutamic acid enters the C5 pathway, affording ALA in A. hyalinus. A 13 C NMR spectroscopic comparison of the labeling patterns of Copro'gen III obtained after feeding of [2- 13 C]glycine, sodium [1- 13 C]acetate, and sodium [2- 13 C]acetate showed that [2- 13 C]glycine transformation and [2- 13 C]glycine catabolism in A. hyalinus proceed in the ratio of 52 and 48 %. The reaction of [2- 13 C]glycine and N 5 , N 10 -methylene-THF, that of glycine and N 5 , N 10 -[methylene- 13 C]methylene-THF generated from the [2- 13 C]glycine catabolism, and that of [2- 13 C]glycine and N 5 , N 10 -[methylene- 13 C]methylene-THF transformed the fed [2- 13 C]glycine to [1- 13 C]acetyl-CoA, [2- 13 C]acetyl-CoA, and [1,2- 13 C 2 ]acetyl-CoA in the ratios of 42, 37, and 21 %, respectively. These labeled acetyl-CoAs were then incorporated into ALA. Our results provide a quantitative picture of the pathways of biosynthetic transformation to ALA from glycine in A. hyalinus. (author)
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Bou, Marta; Østbye, Tone-Kari; Berge, Gerd M; Ruyter, Bente
2017-03-01
The aim of the present study was to investigate how EPA, DHA, and lipoic acid (LA) influence the different metabolic steps in the n-3 fatty acid (FA) biosynthetic pathway in hepatocytes from Atlantic salmon fed four dietary levels (0, 0.5, 1.0 and 2.0%) of EPA, DHA or a 1:1 mixture of these FA. The hepatocytes were incubated with [1- 14 C] 18:3n-3 in the presence or absence of LA (0.2 mM). Increased endogenous levels of EPA and/or DHA and LA exposure both led to similar responses in cells with reduced desaturation and elongation of [1- 14 C] 18:3n-3 to 18:4n-3, 20:4n-3, and EPA, in agreement with reduced expression of the Δ6 desaturase gene involved in the first step of conversion. DHA production, on the other hand, was maintained even in groups with high endogenous levels of DHA, possibly due to a more complex regulation of this last step in the n-3 metabolic pathway. Inhibition of the Δ6 desaturase pathway led to increased direct elongation to 20:3n-3 by both DHA and LA. Possibly the route by 20:3n-3 and then Δ8 desaturation to 20:4n-3, bypassing the first Δ6 desaturase step, can partly explain the maintained or even increased levels of DHA production. LA increased DHA production in the phospholipid fraction of hepatocytes isolated from fish fed 0 and 0.5% EPA and/or DHA, indicating that LA has the potential to further increase the production of this health-beneficial FA in fish fed diets with low levels of EPA and/or DHA.
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Directory of Open Access Journals (Sweden)
Bo Wu
2009-07-01
Full Text Available Filarial parasites (e.g., Brugia malayi, Onchocerca volvulus, and Wuchereria bancrofti are causative agents of lymphatic filariasis and onchocerciasis, which are among the most disabling of neglected tropical diseases. There is an urgent need to develop macro-filaricidal drugs, as current anti-filarial chemotherapy (e.g., diethylcarbamazine [DEC], ivermectin and albendazole can interrupt transmission predominantly by killing microfilariae (mf larvae, but is less effective on adult worms, which can live for decades in the human host. All medically relevant human filarial parasites appear to contain an obligate endosymbiotic bacterium, Wolbachia. This alpha-proteobacterial mutualist has been recognized as a potential target for filarial nematode life cycle intervention, as antibiotic treatments of filarial worms harboring Wolbachia result in the loss of worm fertility and viability upon antibiotic treatments both in vitro and in vivo. Human trials have confirmed this approach, although the length of treatments, high doses required and medical counter-indications for young children and pregnant women warrant the identification of additional anti-Wolbachia drugs.Genome sequence analysis indicated that enzymes involved in heme biosynthesis might constitute a potential anti-Wolbachia target set. We tested different heme biosynthetic pathway inhibitors in ex vivo B. malayi viability assays and report a specific effect of N-methyl mesoporphyrin (NMMP, which targets ferrochelatase (FC, the last step. Our phylogenetic analysis indicates evolutionarily significant divergence between Wolbachia heme genes and their human homologues. We therefore undertook the cloning, overexpression and analysis of several enzymes of this pathway alongside their human homologues, and prepared proteins for drug targeting. In vitro enzyme assays revealed a approximately 600-fold difference in drug sensitivities to succinyl acetone (SA between Wolbachia and human 5
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While thiamin and riboflavin in breast milk have been analyzed for over 50 years, less attention has been given to the different forms of each vitamin. Thiamin-monophosphate (TMP) and free thiamin contribute to total thiamin content; flavin adenine-dinucleotide (FAD) and free riboflavin are the main...
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Guo, X; Geng, P; Bai, F; Bai, G; Sun, T; Li, X; Shi, L; Zhong, Q
2012-08-01
The aims of this study are to obtain the draft genome sequence of Streptomyces coelicoflavus ZG0656, which produces novel acarviostatin family α-amylase inhibitors, and then to reveal the putative acarviostatin-related gene cluster and the biosynthetic pathway. The draft genome sequence of S. coelicoflavus ZG0656 was generated using a shotgun approach employing a combination of 454 and Solexa sequencing technologies. Genome analysis revealed a putative gene cluster for acarviostatin biosynthesis, termed sct-cluster. The cluster contains 13 acarviostatin synthetic genes, six transporter genes, four starch degrading or transglycosylation enzyme genes and two regulator genes. On the basis of bioinformatic analysis, we proposed a putative biosynthetic pathway of acarviostatins. The intracellular steps produce a structural core, acarviostatin I00-7-P, and the extracellular assemblies lead to diverse acarviostatin end products. The draft genome sequence of S. coelicoflavus ZG0656 revealed the putative biosynthetic gene cluster of acarviostatins and a putative pathway of acarviostatin production. To our knowledge, S. coelicoflavus ZG0656 is the first strain in this species for which a genome sequence has been reported. The analysis of sct-cluster provided important insights into the biosynthesis of acarviostatins. This work will be a platform for producing novel variants and yield improvement. © 2012 The Authors. Letters in Applied Microbiology © 2012 The Society for Applied Microbiology.
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Nozaki, T; Asai, T; Sanchez, L B; Kobayashi, S; Nakazawa, M; Takeuchi, T
1999-11-05
The enteric protist parasites Entamoeba histolytica and Entamoeba dispar possess a cysteine biosynthetic pathway, unlike their mammalian host, and are capable of de novo production of L-cysteine. We cloned and characterized cDNAs that encode the regulated enzyme serine acetyltransferase (SAT) in this pathway from these amoebae by genetic complementation of a cysteine-auxotrophic Escherichia coli strain with the amoebic cDNA libraries. The deduced amino acid sequences of the amoebic SATs exhibited, within the most conserved region, 36-52% identities with the bacterial and plant SATs. The amoebic SATs contain a unique insertion of eight amino acids, also found in the corresponding region of a plasmid-encoded SAT from Synechococcus sp., which showed the highest overall identities to the amoebic SATs. Phylogenetic reconstruction also revealed a close kinship of the amoebic SATs with cyanobacterial SATs. Biochemical characterization of the recombinant E. histolytica SAT revealed several enzymatic features that distinguished the amoebic enzyme from the bacterial and plant enzymes: 1) inhibition by L-cysteine in a competitive manner with L-serine; 2) inhibition by L-cystine; and 3) no association with cysteine synthase. Genetically engineered amoeba strains that overproduced cysteine synthase and SAT were created. The cysteine synthase-overproducing amoebae had a higher level of cysteine synthase activity and total thiol content and revealed increased resistance to hydrogen peroxide. These results indicate that the cysteine biosynthetic pathway plays an important role in antioxidative defense of these enteric parasites.
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Study of pyruvate decarboxylase and thiamine kinase from brewer's yeast by SERS
Maskevich, Sergei A.; Chernikevich, Ivan P.; Gachko, Gennedy A.; Kivach, Leonid N.; Strekal, Nataliya D.
1993-06-01
The Surface Enhanced Raman Scattering (SERS) spectra of holopyruvate decarboxylase (PDC) and thiamine kinase (ThK) adsorbed on silver electrode were obtained. In contrast to the Raman, the SERS spectrum of PDC contained no modes of tryptophan residues, it indicates a removal of this moiety from the surface. In the SERS spectrum of ThK the bands belonging to ligands bound to the protein were observed. A correlation between the SERS signal intensity and the enzymatic activity of the ThK separate fraction and found. The influence of amino acids on SERS spectra of thiamine (Th) was studied to determine the possible composition on microsurrounding of coenzyme.
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Nosaka, Kazuto; Esaki, Hiroyoshi; Onozuka, Mari; Konno, Hiroyuki; Hattori, Yasunao; Akaji, Kenichi
2012-05-01
In Saccharomyces cerevisiae, genes involved in thiamin pyrophosphate (TPP) synthesis (THI genes) and the pyruvate decarboxylase structural gene PDC5 are transcriptionally induced in response to thiamin starvation. Three positive regulatory factors (Thi2p, Thi3p, and Pdc2p) are involved in the expression of THI genes, whereas only Pdc2p is required for the expression of PDC5. Thi2p and Pdc2p serve as transcriptional activators and each factor can interact with Thi3p. The target consensus DNA sequence of Thi2p has been deduced. When TPP is not bound to Thi3p, the interactions between the regulatory factors are increased and THI gene expression is upregulated. In this study, we demonstrated that Pdc2p interacts with the upstream region of THI genes and PDC5. The association of Pdc2p or Thi2p with THI gene promoters was enhanced by thiamin starvation, suggesting that Pdc2p and Thi2p assist each other in their recruitment to the THI promoters via interaction with Thi3p. It is highly likely that, under thiamin-deprived conditions, a ternary Thi2p/Thi3p/Pdc2p complex is formed and transactivates THI genes in yeast cells. On the other hand, the association of Pdc2p with PDC5 was unaffected by thiamin. We also identified a DNA element in the upstream region of PDC5, which can bind to Pdc2p and is required for the expression of PDC5. © 2012 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.
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Szymandera-Buszka, Krystyna; Hęś, Marzanna; Waszkowiak, Katarzyna; Jędrusek-Golińska, Anna
2014-01-01
The aim of the study was to determine the effect of pasteurisation and sterilization of model systems of minced chicken meat in the presence of low or high-oxidised pork lard, soy and sunflower oil, as well as casein hydrolysate and rosemary extract, on losses of thiamine in model systems. In the samples, the thiamine content was analysed periodically by thiochromium method, as well as rate of lipid oxidation based on measurement of peroxide value (PV) by iodometric method and p-anisidine value (AV) by spectrophotometric method. It was observed that the thiamine losses in model systems of minced chicken after pasteurisation (61-71%) were higher than after sterilization (57-67%). Introduction of high-oxidised fat increased the total thiamine losses both during thermal processing and storage of meat samples (to 23%). A strong relationship was established between thiamine losses and rate of fat oxidation. The lowest total thiamine losses were observed in the samples with low-oxidised pork lard. Antioxidant addition (rosemary extract or casein hydrolysate) into meat samples limited the thiamine losses. However, the effect depended on oxidation of fat that was mixed with meat. In the samples with low-oxidised fat, higher protective effect was found for rosemary extract (7-11%). In the samples with high-oxidised fat, casein hydrolysate was superior to rosemary extract (14%). In order to increase the stability of thiamine in pasteurized or sterilized meat products with fats, the influence of fat type and its oxidative stability should be taken under consideration. Moreover, the addition of rosemary extract or casein hydrolysate has impact on the thiamine losses since it slows down lipid oxidation to a significant extent.
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Directory of Open Access Journals (Sweden)
F. Baniasadi
2016-07-01
Full Text Available Introduction: Catharanthus roseus (L. belongs to Apocynaceae family is an important medicinal plant and also cultivated as an ornamental plant almost throughout the tropical and subtropical areas all over the world. Recently, the uses of natural substances are considered very helpful to improve plant growth and development. Application of plant growth regulators (PGRs and vitamins are reported in many horticultural crops. Ascorbic acid is the most abundant antioxidant in plant which protects plant cells. This substance affects cell differentiation and growth. Now a day it is considered as a plant growth regulator. Thiamine or vitamin B1 is water soluble and one of the B complex vitamins. This study was conducted to investigate the effect of the ascorbic acid, thiamine, BA and GA on growth and biochemical parameters of periwinkle. Materials and Methods: The experiment was perform as a completely randomized design with 5 replications at research greenhouse of Shahid Bahonar University of Kerman in 2014.The treatments used include spraying water (control, ascorbic acid 100 mg.l-1, gibberellic acid 100 mg.l-1, thiamine 100 mg.l-1, BA 200 mg. l-1 and combinations of two, three and four of them to determine the response of plants to these substances alone or in combination of them. When the seedlings become on 6-leavs stage, the first foliar spray was carried out. In other two steps was repeated with interval of 10 days. In this study, longevity of the flower, flower diameter, the number of lateral branches, branches length, fresh and dry weight, chlorophyll content and reduced sugars were measured. Chlorophyll content and reduced sugars were measured according to method of Somogy (1952 and Lichtenthder (1987, respectively. Results and Discussion: The results showed that the effect of thiamine on the number of flowers was more pronounced than other substances. The lowest number of flowers per plant was found in plants treated with gibberellic acid
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Regulation of the Omega-3 Fatty Acid Biosynthetic Pathway in Atlantic Salmon Hepatocytes.
Directory of Open Access Journals (Sweden)
Marte Avranden Kjær
Full Text Available Limited availability of the n-3 fatty acids EPA and DHA have led to an interest in better understanding of the n-3 biosynthetic pathway and its regulation. The biosynthesis of alpha-linolenic acid to EPA and DHA involves several complex reaction steps including desaturation-, elongation- and peroxisomal beta-oxidation enzymes. The aims of the present experiments were to gain more knowledge on how this biosynthesis is regulated over time by different doses and fatty acid combinations. Hepatocytes isolated from salmon were incubated with various levels and combinations of oleic acid, EPA and DHA. Oleic acid led to a higher expression of the Δ6 fatty acid desaturase (fad genes Δ6fad_a, Δ6fad_b, Δ6fad_c and the elongase genes elovl2 compared with cells cultured in medium enriched with DHA. Further, the study showed rhythmic variations in expression over time. Levels were reached where a further increase in specific fatty acids given to the cells not stimulated the conversion further. The gene expression of Δ6fad_a_and Δ6fad_b responded similar to fatty acid treatment, suggesting a co-regulation of these genes, whereas Δ5fad and Δ6fad_c showed a different regulation pattern. EPA and DHA induced different gene expression patterns, especially of Δ6fad_a. Addition of radiolabelled alpha-linolenic acid to the hepatocytes confirmed a higher degree of elongation and desaturation in cells treated with oleic acid compared to cells treated with DHA. This study suggests a complex regulation of the conversion process of n-3 fatty acids. Several factors, such as that the various gene copies are differently regulated, the gene expression show rhythmic variations and gene expression only affected to a certain level, determines when you get the maximum conversion of the beneficial n-3 fatty acids.
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Effect of overall feedback inhibition in unbranched biosynthetic pathways.
Alves, R; Savageau, M A
2000-11-01
We have determined the effects of control by overall feedback inhibition on the systemic behavior of unbranched metabolic pathways with an arbitrary pattern of other feedback inhibitions by using a recently developed numerical generalization of Mathematically Controlled Comparisons, a method for comparing the function of alternative molecular designs. This method allows the rigorous determination of the changes in systemic properties that can be exclusively attributed to overall feedback inhibition. Analytical results show that the unbranched pathway can achieve the same steady-state flux, concentrations, and logarithmic gains with respect to changes in substrate, with or without overall feedback inhibition. The analytical approach also shows that control by overall feedback inhibition amplifies the regulation of flux by the demand for end product while attenuating the sensitivity of the concentrations to the same demand. This approach does not provide a clear answer regarding the effect of overall feedback inhibition on the robustness, stability, and transient time of the pathway. However, the generalized numerical method we have used does clarify the answers to these questions. On average, an unbranched pathway with control by overall feedback inhibition is less sensitive to perturbations in the values of the parameters that define the system. The difference in robustness can range from a few percent to fifty percent or more, depending on the length of the pathway and on the metabolite one considers. On average, overall feedback inhibition decreases the stability margins by a minimal amount (typically less than 5%). Finally, and again on average, stable systems with overall feedback inhibition respond faster to fluctuations in the metabolite concentrations. Taken together, these results show that control by overall feedback inhibition confers several functional advantages upon unbranched pathways. These advantages provide a rationale for the prevalence of this
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Directory of Open Access Journals (Sweden)
R Cameron Coates
Full Text Available Cyanobacteria possess the unique capacity to naturally produce hydrocarbons from fatty acids. Hydrocarbon compositions of thirty-two strains of cyanobacteria were characterized to reveal novel structural features and insights into hydrocarbon biosynthesis in cyanobacteria. This investigation revealed new double bond (2- and 3-heptadecene and methyl group positions (3-, 4- and 5-methylheptadecane for a variety of strains. Additionally, results from this study and literature reports indicate that hydrocarbon production is a universal phenomenon in cyanobacteria. All cyanobacteria possess the capacity to produce hydrocarbons from fatty acids yet not all accomplish this through the same metabolic pathway. One pathway comprises a two-step conversion of fatty acids first to fatty aldehydes and then alkanes that involves a fatty acyl ACP reductase (FAAR and aldehyde deformylating oxygenase (ADO. The second involves a polyketide synthase (PKS pathway that first elongates the acyl chain followed by decarboxylation to produce a terminal alkene (olefin synthase, OLS. Sixty-one strains possessing the FAAR/ADO pathway and twelve strains possessing the OLS pathway were newly identified through bioinformatic analyses. Strains possessing the OLS pathway formed a cohesive phylogenetic clade with the exception of three Moorea strains and Leptolyngbya sp. PCC 6406 which may have acquired the OLS pathway via horizontal gene transfer. Hydrocarbon pathways were identified in one-hundred-forty-two strains of cyanobacteria over a broad phylogenetic range and there were no instances where both the FAAR/ADO and the OLS pathways were found together in the same genome, suggesting an unknown selective pressure maintains one or the other pathway, but not both.
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In order to determine the genetic basis for loss of fumonisin B¬2 (FB2) biosynthesis in FB2 non-producing A. niger strains, we developed multiplex PCR primer sets to amplify fragments of eight fumonisin biosynthetic pathway (fum) genes. Fragments of all eight fum genes were amplified in FB2-produci...
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Hampel, Daniela; Shahab-Ferdows, Setareh; Adair, Linda S; Bentley, Margaret E; Flax, Valerie L; Jamieson, Denise J; Ellington, Sascha R; Tegha, Gerald; Chasela, Charles S; Kamwendo, Debbie; Allen, Lindsay H
2016-01-01
While thiamin and riboflavin in breast milk have been analyzed for over 50 years, less attention has been given to the different forms of each vitamin. Thiamin-monophosphate (TMP) and free thiamin contribute to total thiamin content; flavin adenine-dinucleotide (FAD) and free riboflavin are the main contributors to total riboflavin. We analyzed milk collected at 2 (n = 258) or 6 (n = 104), and 24 weeks (n = 362) from HIV-infected Malawian mothers within the Breastfeeding, Antiretrovirals and Nutrition (BAN) study, randomly assigned at delivery to lipid-based nutrient supplements (LNS) or a control group, to investigate each vitamer's contribution to total milk vitamin content and the effects of supplementation on the different thiamin and riboflavin vitamers at early and later stages of lactation, and obtain insight into the transport and distribution of these vitamers in human milk. Thiamin vitamers were derivatized into thiochrome-esters and analyzed by high-performance liquid-chromatography-fluorescence-detection (HPLC-FLD). Riboflavin and FAD were analyzed by ultra-performance liquid-chromatography-tandem-mass-spectrometry (ULPC-MS/MS). Thiamin-pyrophosphate (TPP), identified here for the first time in breast milk, contributed 1.9-4.5% to total thiamin. Free thiamin increased significantly from 2/6 to 24 weeks regardless of treatment indicating an active transport of this vitamer in milk. LNS significantly increased TMP and free thiamin only at 2 weeks compared to the control: median 170 versus 151 μg/L (TMP), 13.3 versus 10.5 μg/L (free thiamin, priboflavin was consistently and significantly increased with LNS (range: 14.8-19.6 μg/L (LNS) versus 5.0-7.4 μg/L (control), priboflavin relative amounts from 92-94:6-8% to 85:15%, indicating a preferred secretion of the free form into breast milk. The continuous presence of FAD in breast milk suggests an active transport and secretion system for this vitamer or possibly formation of this co-enymatic form in the
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Bhat, Javeed Iqbal; Rather, Hilal Ahmad; Ahangar, Ambreen Ali; Qureshi, Umar Amin; Dar, Parvez; Ahmed, Qazi Iqbal; Charoo, Bashir Ahmed; Ali, Syed Wajid
To study the effect of thiamine administration on the resolution of pulmonary hypertension in exclusively breastfed infants. Prospective cohort study. Hospital based study of a tertiary care hospital. A total of 29 infants with 17 males (58.6%) and 12 females (41.4%) were included in the study. In addition to the management of shock, right heart failure and renal failure, patients received intravenous thiamine 100mg/kg IV followed by 10mg/day till introduction of supplementary feeds. Resolution of shock, metabolic complications and pulmonary hypertension. Mean age at presentation was 78.45±30.7 days. All infants were exclusively breastfed. 86.2% of mothers were on customary dietary restrictions. Biventricular failure and tachycardia was commonly present. There were four deaths in our series. Acute metabolic acidosis was a universal feature with a mean pH of 7.21±0.15. Pulmonary hypertension was present in all patients on admission. Intravenous thiamine 100mg/kg IV stat was given immediately after documenting pulmonary hypertension. Repeat echocardiography showed complete resolution of pulmonary hypertension. Many infants present to us with Shoshin beriberi with unusually high pulmonary pressures. These patients respond to thiamine challenge with prompt resolution of metabolic complications and reversal of pulmonary hypertension. We believe this is first of its kind from the region, which is reported. Copyright © 2016. Published by Elsevier B.V.
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Role of riboflavin and thiamine in induced resistance against ...
African Journals Online (AJOL)
The effect of two inducer chemicals, that is, riboflavin (B2) and thiamine (B1) on the induction of systemic resistance in soybean against charcoal rot disease as well as biochemical changes associated with these treatments in soybean plants was investigated. Under greenhouse condition, the dose effect of 0.1 to 15 mM ...
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Ketola, H.G.; Rinchard, J.; O'Gorman, R.; Begnoche, L.J.; Bishop, D.L.; Greulich, A.W.
2009-01-01
Early mortality syndrome in fry of Great Lakes salmonines is linked to reduced levels of thiamine in eggs, which reflects maternal consumption of forage fishes such as alewife (Alosa pseudoharengus) that contain thiaminase, an enzyme that destroys thiamine. We assessed annual variations in abundance and condition of alewives and thiamine status of coho salmon (Oncorhynchus kisutch) in Lake Ontario. We analyzed total thiamine in eggs of 20 coho salmon collected annually between 2003 and 2006 at the Salmon River Hatchery on the Salmon River, New York. Alewife abundance was assessed annually in southern and eastern Lake Ontario with bottom trawls during late April and early May. Mean thiamine concentration in eggs varied annually, with those collected in 2003 (2.5 nmol/g) being significantly higher than those collected in 2004 to 2006 (1.5 to 1.7 nmol/g). Although we did not test survival of fry, if reported threshold levels of thiamine for preventing mortality of Lake Michigan coho salmon fry apply, then many or most Lake Ontario coho salmon produced fry were likely to incur thiamine-deficiency mortality, especially during years 2004 to 2006. Comparison to indices of annual abundance of alewife in Lake Ontario with thiamine concentration in coho salmon eggs failed to show any significant correlations (P > 0.05). However, total length of female spawning coho salmon was positively correlated (P < 0.05) with increasing condition and estimated energy content of adult alewives in the previous spring. These results suggest that growth of coho salmon in Lake Ontario was first limited by energy intake, whereas the amount of thiamine provided by alewives was sufficient for growth (in length) but not for producing thiamine-adequate eggs.
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Biotechnological Potential of Yarrowia lipolytica Grown under Thiamine Limitation
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Maria N. Chiglintseva
2012-01-01
Full Text Available During the cultivation of a thiamine-auxotrophic yeast strain Yarrowia lipolytica VKM Y-2412 on ethanol, the growth limitation by thiamine leads to the production of α-ketoglutaric acid. The α-ketoglutaric acid synthesis has been studied in dependence on pH, oxygen supply and ethanol, zinc and iron concentrations. Under optimal conditions, Y. lipolytica produced 88.7 g/L of α-ketoglutaric acid. The culture broth containing α-ketoglutaric acid was subjected to chemical treatment with hydrogen peroxide, which led to the formation of succinic acid in significant quantities (71.7 g/L. Further direct esterification of succinic acid with excess absolute ethanol yielded diethyl succinate. Biomass of Y. lipolytica, a superproducer of α-ketoglutaric acid, was characterized by a high content of protein and essential amino acids, free amino acids, and unusually large amount of γ-aminobutyric acid. The unique amino acid composition of the producer makes it possible to use this biomass as a component of parenteral nutrition mixtures and as a basis for neuroleptics.
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Gober, Claire Marie
Roquefortine C is one of the most ubiquitous indoline alkaloids of fungal origin. It has been isolated from over 30 different species of Penicillium fungi and has garnered attention in recent years for its role as a biosynthetic precursor to the triazaspirocyclic natural products glandicoline B, meleagrin, and oxaline. The triazaspirocyclic motif, which encompasses three nitrogen atoms attached to one quaternary carbon forming a spirocyclic scaffold, is a unique chemical moiety that has been shown to impart a wide array of biological activity, from anti-bacterial activity and antiproliferative activity against cancer cell lines to anti-biofouling against marine organisms. Despite the promise of these compounds in the pharmaceutical and materials industries, few syntheses of triazaspirocycles exist in the literature. The biosynthesis of roquefortine C-derived triazaspirocycles, however, provides inspiration for the synthesis of these compounds, namely through a nitrone-promoted transannular rearrangement. This type of internal rearrangement has never been carried out synthetically and would provide an efficient stereoselective synthesis of triazaspirocycles. This work encompasses efforts towards elucidating the biosynthetic pathway of roquefortine C-derived triazaspirocycles as well as synthetic efforts towards the construction of triazaspirocycles. Chapter 1 will discuss a large-scale fermentation procedure for the production of roquefortine C from Penicillium crustosum. Chapters 2 and 3 explore (through enzymatic and synthetic means, respectively) the formation of the key indoline nitrone moiety required for the proposed transannular rearrangement. Finally, chapter 4 will discuss synthetic efforts towards the synthesis of triazaspirocycles. This work has considerably enhanced our understanding of the roquefortine C biosynthetic pathway and the unique chemistry of this natural product, and our efforts towards the synthesis of triazaspirocycles will facilitate the
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Directory of Open Access Journals (Sweden)
Freda Koh
2015-04-01
Full Text Available The effectiveness of nutritional interventions to prevent and maintain cognitive functioning in older adults has been gaining interest due to global population ageing. A systematic literature review was conducted to obtain and appraise relevant studies on the effects of dietary protein or thiamine on cognitive function in healthy older adults. Studies that reported on the use of nutritional supplementations and/or populations with significant cognitive impairment were excluded. Seventeen eligible studies were included. Evidence supporting an association between higher protein and/or thiamine intakes and better cognitive function is weak. There was no evidence to support the role of specific protein food sources, such as types of meat, on cognitive function. Some cross-sectional and case-control studies reported better cognition in those with higher dietary thiamine intakes, but the data remains inconclusive. Adequate protein and thiamine intake is more likely associated with achieving a good overall nutritional status which affects cognitive function rather than single nutrients. A lack of experimental studies in this area prevents the translation of these dietary messages for optimal cognitive functioning and delaying the decline in cognition with advancing age.
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Norman, Shirley M.; Maier, Vincent P.; Echols, Linda C.
1981-01-01
Abscisic acid production by Cercospora rosicola Passerini in liquid shake culture was measured with different amino acids in combination and singly as nitrogen sources and with different amounts of thiamine in the media. Production of abscisic acid was highest with aspartic acid-glutamic acid and aspartic acid-glutamic acid-serine mixtures as nitrogen sources. Single amino acids that supported the highest production of abscisic acid were asparagine and monosodium glutamate. Thiamine was impor...
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Directory of Open Access Journals (Sweden)
Cynthia A Dick
2011-04-01
Full Text Available Intra- and interspecific variation in flower color is a hallmark of angiosperm diversity. The evolutionary forces underlying the variety of flower colors can be nearly as diverse as the colors themselves. In addition to pollinator preferences, non-pollinator agents of selection can have a major influence on the evolution of flower color polymorphisms, especially when the pigments in question are also expressed in vegetative tissues. In such cases, identifying the target(s of selection starts with determining the biochemical and molecular basis for the flower color variation and examining any pleiotropic effects manifested in vegetative tissues. Herein, we describe a widespread purple-white flower color polymorphism in the mustard Parrya nudicaulis spanning Alaska. The frequency of white-flowered individuals increases with increasing growing-season temperature, consistent with the role of anthocyanin pigments in stress tolerance. White petals fail to produce the stress responsive flavonoid intermediates in the anthocyanin biosynthetic pathway (ABP, suggesting an early pathway blockage. Petal cDNA sequences did not reveal blockages in any of the eight enzyme-coding genes in white-flowered individuals, nor any color differentiating SNPs. A qRT-PCR analysis of white petals identified a 24-fold reduction in chalcone synthase (CHS at the threshold of the ABP, but no change in CHS expression in leaves and sepals. This arctic species has avoided the deleterious effects associated with the loss of flavonoid intermediates in vegetative tissues by decoupling CHS expression in petals and leaves, yet the correlation of flower color and climate suggests that the loss of flavonoids in the petals alone may affect the tolerance of white-flowered individuals to colder environments.
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Directory of Open Access Journals (Sweden)
Raffaele Nardone
2013-01-01
Full Text Available Nutritional deficiency can cause, mainly in chronic alcoholic subjects, the Wernicke encephalopathy and its chronic neurological sequela, the Wernicke-Korsakoff syndrome (WKS. Long-term chronic ethanol abuse results in hippocampal and cortical cell loss. Thiamine deficiency also alters principally hippocampal- and frontal cortical-dependent neurochemistry; moreover in WKS patients, important pathological damage to the diencephalon can occur. In fact, the amnesic syndrome typical for WKS is mainly due to the damage in the diencephalic-hippocampal circuitry, including thalamic nuclei and mammillary bodies. The loss of cholinergic cells in the basal forebrain region results in decreased cholinergic input to the hippocampus and the cortex and reduced choline acetyltransferase and acetylcholinesterase activities and function, as well as in acetylcholine receptor downregulation within these brain regions. In this narrative review, we will focus on the neurochemical, neuroanatomical, and neuropsychological studies shedding light on the effects of thiamine deficiency in experimental models and in humans.
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Antiglycation activity of thiamin-HCl and benfotiamine in diabetic condition.
Kousar, Samina; Sheikh, Munir Ahmad; Asghar, Muhammad
2012-10-01
To observe the antiglycation effect of water-soluble vitamin, thiamin-hydrochloride (B1) and its fat-soluble derivative benfotiamine and their comparison with two different glycation assays in diabetics. Plasma of both the normal's and diabetic persons was taken for this experimental study which was conducted in the Department of Chemistry and Biochemistry, University of Agriculture, Faisalabad in 2008. Varying concentrations of both the glycation inhibitors and glucose were incubated for 5 weeks at 37 degrees C. Thiobarbituric acid and periodate borohydride assays were used to measure the antiglycation activity. The increase in glycation was observed from 1st to 3rd week of incubation, while it decreased after the 5th week due to the formation of advanced glycation end products. It was observed that 10mM concentration of benfotiamine and 5mM and 1mM concentrations of thiamine-hydrochloride produced fairly good response to decreased glycation. Comparison between two assays proved periodate borohydride to be more reliable and sensitive than thiobarbituric acid.
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Conditional sterility in plants
Meagher, Richard B.; McKinney, Elizabeth; Kim, Tehryung
2010-02-23
The present disclosure provides methods, recombinant DNA molecules, recombinant host cells containing the DNA molecules, and transgenic plant cells, plant tissue and plants which contain and express at least one antisense or interference RNA specific for a thiamine biosynthetic coding sequence or a thiamine binding protein or a thiamine-degrading protein, wherein the RNA or thiamine binding protein is expressed under the regulatory control of a transcription regulatory sequence which directs expression in male and/or female reproductive tissue. These transgenic plants are conditionally sterile; i.e., they are fertile only in the presence of exogenous thiamine. Such plants are especially appropriate for use in the seed industry or in the environment, for example, for use in revegetation of contaminated soils or phytoremediation, especially when those transgenic plants also contain and express one or more chimeric genes which confer resistance to contaminants.
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Afzal, Muhammad; Kazmi, Imran; Khan, Ruqaiyah; Rana, Poonam; Kumar, Vikas; Al-Abbasi, Fahad A; Zamzami, Mazin A; Anwar, Firoz
2017-06-01
Present study, was an effort to scrutinize the molecular and biochemical role of ibuprofen and thiamine combination in diethylnitrosamine (DEN)-induced HCC in Wistar rats. Single intraperitoneal injection of DEN (200 mg/kg) was used for induction of HCC in rats. The rats were divided into eight various groups. DEN induced rats were treated with pure ibuprofen (40 mg/kg) and thiamine in combination for the period of 12th weeks. The protocol was terminated after the 16th week. Exposure of DEN up-regulated the levels of different serum biochemical parameters, antioxidant enzyme level, Alfa-fetoprotein (AFP) and reduced the level of High density lipoprotein (HDL) in Wistar rats along with the alteration in pro-inflammatory cytokines viz., interlukin-6 (IL-6), Tumor necrosis factor (TNF-α) and Interleukin-1β (IL-1β) with decrease in body weight. Macroscopic evaluation, revealed DEN group rats confirmed the expansion of hepatic nodules, which were reduced by the individual treatment of ibuprofen and thiamine, but the synergistic treatment of ibuprofen and thiamine confirm the significant reduction of hepatic nodules. Further, this combination possesses the significant chemoprotective effect in DEN-induced HCC by restoring the hepatic enzymes and other biomarkers along with an alteration in pro-inflammatory cytokines. The above result concludes that ibuprofen and thiamine combination possess potent anti-cancerous activity. Copyright © 2017 Elsevier Inc. All rights reserved.
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Parkhomenko, Yulia M; Kudryavtsev, Pavel A; Pylypchuk, Svetlana Yu; Chekhivska, Lilia I; Stepanenko, Svetlana P; Sergiichuk, Andrej A; Bunik, Victoria I
2011-06-01
Thiamine-dependent changes in alcoholic brain were studied using a rat model. Brain thiamine and its mono- and diphosphates were not reduced after 20 weeks of alcohol exposure. However, alcoholism increased both synaptosomal thiamine uptake and thiamine diphosphate synthesis in brain, pointing to mechanisms preserving thiamine diphosphate in the alcoholic brain. In spite of the unchanged level of the coenzyme thiamine diphosphate, activities of the mitochondrial 2-oxoglutarate and pyruvate dehydrogenase complexes decreased in alcoholic brain. The inactivation of pyruvate dehydrogenase complex was caused by its increased phosphorylation. The inactivation of 2-oxoglutarate dehydrogenase complex (OGDHC) correlated with a decrease in free thiols resulting from an elevation of reactive oxygen species. Abstinence from alcohol following exposure to alcohol reactivated OGDHC along with restoration of the free thiol content. However, restoration of enzyme activity occurred before normalization of reactive oxygen species levels. Hence, the redox status of cellular thiols mediates the action of oxidative stress on OGDHC in alcoholic brain. As a result, upon chronic alcohol consumption, physiological mechanisms to counteract the thiamine deficiency and silence pyruvate dehydrogenase are activated in rat brain, whereas OGDHC is inactivated due to impaired antioxidant ability. © 2011 The Authors. Journal of Neurochemistry © 2011 International Society for Neurochemistry.
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Yuge, Shinya; Richter, Catherine A.; Wright-Osment, Maureen K.; Nicks, Diane; Saloka, Stephanie K.; Tillitt, Donald E.; Li, Weiming
2012-01-01
Thiamin pyrophosphokinase (TPK) converts thiamin to its active form, thiamin diphosphate. In humans, TPK expression is down-regulated in some thiamin deficiency related syndrome, and enhanced during pregnancy. Rainbow trout are also vulnerable to thiamin deficiency in wild life and are useful models for thiamin metabolism research. We identified the tpk gene transcript including seven splice variants in the rainbow trout. Almost all cell lines and tissues examined showed co-expression of several tpk splice variants including a potentially major one at both mRNA and protein levels. However, relative to other tissues, the longest variant mRNA expression was predominant in the ovary and abundant in embryos. During embryogenesis, total tpk transcripts increased abruptly in early development, and decreased to about half of the peak shortly after hatching. In rainbow trout, the tpk transcript complex is ubiquitously expressed for all tissues and cells examined, and its increase in expression could be important in the early-middle embryonic stages. Moreover, decimated tpk expression in a hepatoma cell line relative to hepatic and gonadal cell lines appears to be consistent with previously reported down-regulation of thiamin metabolism in cancer.
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Directory of Open Access Journals (Sweden)
Ginsburg Hagai
2005-03-01
Full Text Available Abstract The general paradigm that emerges from the analysis of the transcriptome of the malaria parasite Plasmodium falciparum is that the expression clusters of genes that code for enzymes engaged in the same cellular function is coordinated. Here the consistency of this perception is examined by analysing specific pathways that metabolically-linked. The pentose phosphate pathway (PPP is a fundamental element of cell biochemistry since it is the major pathway for the recycling of NADP+ to NADPH and for the production of ribose-5-phosphate that is needed for the synthesis of nucleotides. The function of PPP depends on the synthesis of NADP+ and thiamine pyrophosphate, a co-enzyme of the PPP enzyme transketolase. In this essay, the transcription of gene coding for enzymes involved in the PPP, thiamine and NAD(P+ syntheses are analysed. The genes coding for two essential enzymes in these pathways, transaldolase and NAD+ kinase could not be found in the genome of P. falciparum. It is found that the transcription of the genes of each pathway is not always coordinated and there is usually a gene whose transcription sets the latest time for the full deployment of the pathway's activity. The activity of PPP seems to involve only the oxidative arm of PPP that is geared for maximal NADP+ reduction and ribose-5-phosphate production during the early stages of parasite development. The synthesis of thiamine diphosphate is predicted to occur much later than the expression of transketolase. Later in the parasite cycle, the non-oxidative arm of PPP that can use fructose-6-phosphate and glyceraldehyde-3-phosphate supplied by glycolysis, becomes fully deployed allowing to maximize the production of ribose-5-phosphate. These discrepancies require direct biochemical investigations to test the activities of the various enzymes in the developing parasite. Notably, several transcripts of PPP enzyme-coding genes display biphasic pattern of transcription unlike most
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Effect of high dose thiamine therapy on activity and molecular ...
African Journals Online (AJOL)
Commonest form of diabetes mellitus is Type 2, treated with oral hypoglycemic agents, which often carry potential adverse effects and do not address the intracellular metabolism of glucose. Thiamine is an essential co-factor for vital subcellular enzymes and has potential to benefit Type 2 diabetics. This study was therefore ...
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Directory of Open Access Journals (Sweden)
Guadalupe Albarrán
2014-10-01
Full Text Available Vitamin loss during irradiation has been claimed as a critical area in food irradiation technology, especially that of thiamine (B1, which has been considered as the most sensitive to radiation. Although it has been suggested that no vitamin deficiency could result from consuming irradiated food, a long debate on the loss of vitamins and other nutrients during food irradiation has been maintained by the lack of experimental studies monitoring decomposition rates at different concentrations and doses. Since thiamine, riboflavin, and pyridoxine are labile vitamins, this study has focused on their radiolytic decomposition in dilute aqueous solutions in the presence of air. The decomposition process was followed by HPLC and UV-spectroscopy. The results obtained in aqueous solutions showed a dependence of the decomposition as a nonlinear function of the dose. Of these three compounds, the decomposition was higher for thiamine than for riboflavin and even less in pyridoxine.
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antiSMASH 3.0—a comprehensive resource for the genome mining of biosynthetic gene clusters
DEFF Research Database (Denmark)
Weber, Tilmann; Blin, Kai; Duddela, Srikanth
2015-01-01
Microbial secondary metabolism constitutes a rich source of antibiotics, chemotherapeutics, insecticides and other high-value chemicals. Genome mining of gene clusters that encode the biosynthetic pathways for these metabolites has become a key methodology for novel compound discovery. In 2011, we...... introduced antiSMASH, a web server and stand-alone tool for the automatic genomic identification and analysis of biosynthetic gene clusters, available at http://antismash.secondarymetabolites.org. Here, we present version 3.0 of antiSMASH, which has undergone major improvements. A full integration...... of the recently published ClusterFinder algorithm now allows using this probabilistic algorithm to detect putative gene clusters of unknown types. Also, a new dereplication variant of the ClusterBlast module now identifies similarities of identified clusters to any of 1172 clusters with known end products...
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Moses, Tessa; Papadopoulou, Kalliope K.
2014-01-01
Saponins are widely distributed plant natural products with vast structural and functional diversity. They are typically composed of a hydrophobic aglycone, which is extensively decorated with functional groups prior to the addition of hydrophilic sugar moieties, to result in surface-active amphipathic compounds. The saponins are broadly classified as triterpenoids, steroids or steroidal glycoalkaloids, based on the aglycone structure from which they are derived. The saponins and their biosynthetic intermediates display a variety of biological activities of interest to the pharmaceutical, cosmetic and food sectors. Although their relevance in industrial applications has long been recognized, their role in plants is underexplored. Recent research on modulating native pathway flux in saponin biosynthesis has demonstrated the roles of saponins and their biosynthetic intermediates in plant growth and development. Here, we review the literature on the effects of these molecules on plant physiology, which collectively implicate them in plant primary processes. The industrial uses and potential of saponins are discussed with respect to structure and activity, highlighting the undoubted value of these molecules as therapeutics. PMID:25286183
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Engineering the spatial organization of metabolic pathways
DEFF Research Database (Denmark)
Albertsen, Line; Maury, Jerome; Bach, Lars Stougaard
One of the goals of metabolic engineering is to optimize the production of valuable metabolites in cell factories. In this context, modulating the gene expression and activity of enzymes are tools that have been extensively used. Another approach that is gaining interest is the engineering...... of the spatial organization of biosynthetic pathways. Several natural systems for ensuring optimal spatial arrangement of biosynthetic enzymes exist. Sequentially acting enzymes can for example be positioned in close proximity by attachment to cellular structures, up-concentration in membrane enclosed organelles...... or assembly into large complexes. The vision is that by positioning sequentially acting enzymes in close proximity, the cell can accelerate reaction rates and thereby prevent loss of intermediates through diffusion, degradation or competing pathways. The production of valuable metabolites in cell factories...
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International Nuclear Information System (INIS)
Graham, W.D.; Stevenson, M.H.; Stewart, E.M.
1998-01-01
The usefulness of ionising radiation for the elimination of pathogenic bacteria in poultry meat has been well documented as have the effects of this processing treatment on the nutritional status of the food, in particular, the vitamins. Unfortunately, much of the earlier research carried out on the effect of irradiation on vitamins was carried out in solution or in model systems at doses much greater than those used commercially thereby resulting in considerable destruction of these compounds. Thus, those opposed to the process of food irradiation labelled the treated food as nutritionally poor. However, in reality, due to the complexity of food systems the effects of irradiation on vitamins are generally not as marked and many processes, for example cooking, cause the same degree of change to the vitamins. Thiamin (vitamin B1) is the most radiation sensitive of the water-soluble vitamins and is therefore a good indicator of the effect of irradiation treatment. In this study the effects of irradiation at either 4°C or −20°C followed by cooking on the thiamin content of chicken breast meat was determined. Results showed that whilst both irradiation and cooking resulted in a decrease in thiamin concentration, the losses incurred were unlikely to be of nutritional significance and could be further minimised by irradiating the chicken meat at a low temperature. Thiamin analyses were carried out using high-performance liquid chromatography since this technique is faster and more selective than the chemical or microbiological methods more commonly employed. Total thiamin, both free and combined form, was determined following acid and enzyme hydrolysis. © 1998 Society of Chemical Industry
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Flg22-Triggered Immunity Negatively Regulates Key BR Biosynthetic Genes.
Jiménez-Góngora, Tamara; Kim, Seong-Ki; Lozano-Durán, Rosa; Zipfel, Cyril
2015-01-01
In plants, activation of growth and activation of immunity are opposing processes that define a trade-off. In the past few years, the growth-promoting hormones brassinosteroids (BR) have emerged as negative regulators of pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI), promoting growth at the expense of defense. The crosstalk between BR and PTI signaling was described as negative and unidirectional, since activation of PTI does not affect several analyzed steps in the BR signaling pathway. In this work, we describe that activation of PTI by the bacterial PAMP flg22 results in the reduced expression of BR biosynthetic genes. This effect does not require BR perception or signaling, and occurs within 15 min of flg22 treatment. Since the described PTI-induced repression of gene expression may result in a reduction in BR biosynthesis, the crosstalk between PTI and BR could actually be negative and bidirectional, a possibility that should be taken into account when considering the interaction between these two pathways.
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Hucker, Barry; Wakeling, Lara; Vriesekoop, Frank
2011-12-14
This research aimed to develop a simple and effective method for analyzing thiamin (B(1)), riboflavin (B(2)) and their respective vitamers by high performance liquid chromatography (HPLC) in fermented alcoholic beverages. The method developed here employs a phosphate buffer/methanol gradient elution on a single reverse phase column, coupled with independent fluorescent detection regimes. It also employs a precolumn derivatization to convert thiamin to thiochrome via an alkaline potassium ferricyanide solution. The method described here allowed a spike recovery of better than 97%, with a typical linear detection range (R(2) ≥ 0.9997) between ≤ 5 and ≥ 500 μg/L for all vitamers studied. Lager style beers were found to contain significantly (p porters, 104.4 μg/L; wheat beers, 130.7 μg/L), which may be due to the raw material and extensive processing that occurs for this style. There was no statistical difference (p = 0.608) between the riboflavin content of each beer style. Furthermore, wines and ciders contain less thiamin and riboflavin than beer, which is also likely to be due to the base materials used and the differences in processing steps to produce these beverages.
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Biosynthetic origin of acetic acid using SNIF-NMR
International Nuclear Information System (INIS)
Boffo, Elisangela Fabiana; Ferreira, Antonio Gilberto
2006-01-01
The main purpose of this work is to describe the use of the technique Site-Specific Natural Isotopic Fractionation of hydrogen (SNIF-NMR), using 2 H and 1 H NMR spectroscopy, to investigate the biosynthetic origin of acetic acid in commercial samples of Brazilian vinegar. This method is based on the deuterium to hydrogen ratio at a specific position (methyl group) of acetic acid obtained by fermentation, through different biosynthetic mechanisms, which result in different isotopic ratios. We measured the isotopic ratio of vinegars obtained through C 3 , C 4 , and CAM biosynthetic mechanisms, blends of C 3 and C 4 (agrins) and synthetic acetic acid. (author)
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Karachalias, N; Babaei-Jadidi, R; Rabbani, N; Thornalley, P J
2010-07-01
The aim of this study was to quantify protein damage by glycation, oxidation and nitration in a rat model of diabetes at the sites of development of microvascular complications, including the effects of thiamine and benfotiamine therapy. Diabetes was induced in male Sprague-Dawley rats by 55 mg/kg streptozotocin and moderated by insulin (2 U twice daily). Diabetic and control rats were given thiamine or benfotiamine (7 or 70 mg kg(-1) day(-1)) over 24 weeks. Plasma, urine and tissues were collected and analysed for protein damage by stable isotopic dilution analysis MS. There were two- to fourfold increases in fructosyl-lysine and AGE content of glomerular, retinal, sciatic nerve and plasma protein in diabetes. Increases in AGEs were reversed by thiamine and benfotiamine therapy but increases in fructosyl-lysine were not. Methionine sulfoxide content of plasma protein and 3-nitrotyrosine content of sciatic nerve protein were increased in diabetes. Plasma glycation free adducts were increased up to twofold in diabetes; the increases were reversed by thiamine. Urinary excretion of glycation, oxidation and nitration free adducts was increased by seven- to 27-fold in diabetes. These increases were reversed by thiamine and benfotiamine therapy. AGEs, particularly arginine-derived hydroimidazolones, accumulate at sites of microvascular complication development and have markedly increased urinary excretion rates in experimental diabetes. Thiamine and benfotiamine supplementation prevented tissue accumulation and increased urinary excretion of protein glycation, oxidation and nitration adducts. Similar effects may contribute to the reversal of early-stage clinical diabetic nephropathy by thiamine.
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Bâ, Abdoulaye
2012-09-01
The present study compared the effects of thiamine (vitamin B1) deficiency (TD) on the patterns of food intake and body weight in adult female and neonatal Wistar rats. The adults weighed 250-270 g at the start and were fed for 60 days either with a synthetic TD diet (211 B1) or with the same synthetic diet+thiamine (210 B1). TD led to a marked reduction in food intake and the body weight set point, both recovering rapidly to their initial level in only 3 days after dietetic reversion. The effects of TD in developing rats were evaluated by subjecting pregnant rats to thiamine restriction during different time windows: prenatal (3 days before mating to parturition); perinatal (7 days after mating to the 10th postnatal day); and postnatal (from parturition to weaning). The effect of TD on the occurrence of low birth weight and ponderal growth retardation was examined from postnatal days 1 to 45. Only perinatal TD significantly decreased birth weight relative to untreated or pair-fed controls. Moreover, compared with the control treatments, ponderal growth retardation was not induced by prenatal TD, whereas induction of TD from perinatal into postnatal periods did cause ponderal growth retardation, with long-lasting effects persisting in adulthood. The results suggest a major physiological role of thiamine in the homeostasis of body weight programming, increment, and set point regulation in both offspring and adult female rats.
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Ostrovskiĭ KuM; Voskoboev, A I; Gorenshtenĭn, B I; Dosta, G A
1979-09-01
The molecula-kinetic parameters (Km, Ki) of three thiamine enzymes, e. g. thiamine pyrophosphokinase (EC 2.7.6.2), pyruvate dehydrogenase (EC 1.2.4.1) and transketolase (EC 2.2.1.1) with respect to the effects of the thiamine antimetabolite hydroxythiamine in the whole animal organism have been compared. It has been shown that only the first two enzymes, which interact competitively with the vitamin, antivitamin or their pyrophosphate ethers, obey the kinetic parameters obtained for the purified enzymes in vitro. The anticoenzymic effect of hydroxythiamine pyrophosphate with respect to transketolase is not observed in vivo at maximal concentration of the anticoenzyme in tissues due to the absence of competitive interactions with thiamine pyrophosphate. The incorporation of the true and false coenzymes into transketolase occurs only during de novo transketolase synthesis (the apoform is absent in tissues, with the exception of erythrocytes) and proceeds slowly with a half-life time equal to 24--30 hrs. After a single injection of hydroxythiamine at a large dose (70--400 mg/kg) the maximal inhibition of the transketolase activity in tissues (liver, heart, kidney, muscle, spleen, lungs adrenal grands) manifests itself by the 48th--72nd hour, when the concentration of free hydroxythiamine and its pyrophosphate is minimal and the whole anticoenzyme is tightly bound to the protein, forming the false holoenzyme. The use of hydroxythiamine for inhibition of pyruvate dehydrogenase or transketolase in animal organism is discussed.
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Palacios, Oskar A; Gomez-Anduro, Gracia; Bashan, Yoav; de-Bashan, Luz E
2016-06-01
During synthetic mutualistic interactions between the microalga Chlorella sorokiniana and the plant growth-promoting bacterium (PGPB) Azospirillum brasilense, mutual exchange of resources involved in producing and releasing the phytohormone indole-3-acetic acid (IAA) by the bacterium, using tryptophan and thiamine released by the microalga, were measured. Although increased activities of tryptophan synthase in C. sorokiniana and indole pyruvate decarboxylase (IPDC) in A. brasilense were observed, we could not detect tryptophan or IAA in the culture medium when both organisms were co-immobilized. This indicates that no extra tryptophan or IAA is produced, apart from the quantities required to sustain the interaction. Over-expression of the ipdC gene occurs at different incubation times: after 48 h, when A. brasilense was immobilized alone and grown in exudates of C. sorokiniana and at 96 h, when A. brasilense was co-immobilized with the microalga. When A. brasilense was cultured in exudates of C. sorokiniana, increased expression of the ipdC gene, corresponding increase in activity of IPDC encoded by the ipdC gene, and increase in IAA production were measured during the first 48 h of incubation. IAA production and release by A. brasilense was found only when tryptophan and thiamine were present in a synthetic growth medium (SGM). The absence of thiamine in SGM yielded no detectable IAA. In summary, this study demonstrates that C. sorokiniana can exude sufficient tryptophan and thiamine to allow IAA production by a PGPB during their interaction. Thiamine is essential for IAA production by A. brasilense and these three metabolites are part of a communication between the two microorganisms. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.
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On the influence of thiamine and ammonium ions on alcoholic fermentation
Maesen, Th.J.M.
1953-01-01
1. 1. In a glucose-sodium acetate medium of pH 5.6 the fermentation rate of bakers' yeast remains constant at a low level during several hours, while in the presence of ammonium sulphate it gradually increases. The rise is steeper in the presence of thiamine. 2. 2. After the ammonium ions have
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The oxylipin pathway in Arabidopsis.
Creelman, Robert A; Mulpuri, Rao
2002-01-01
Oxylipins are acyclic or cyclic oxidation products derived from the catabolism of fatty acids which regulate many defense and developmental pathways in plants. The dramatic increase in the volume of publications and reviews on these compounds since 1997 documents the increasing interest in this compound and its role in plants. Research on this topic has solidified our understanding of the chemistry and biosynthetic pathways for oxylipin production. However, more information is still needed on how free fatty acids are produced and the role of beta-oxidation in the biosynthetic pathway for oxylipins. It is also becoming apparent that oxylipin content and composition changes during growth and development and during pathogen or insect attack. Oxylipins such as jasmonic acid (JA) or 12-oxo-phytodienoic acid modulate the expression of numerous genes and influence specific aspects of plant growth, development and responses to abiotic and biotic stresses. Although oxylipins are believed to act alone, several examples were presented to illustrate that JA-induced responses are modulated by the type and the nature of crosstalk with other signaling molecules such as ethylene and salicylic acid. How oxylipins cause changes in gene expression and instigate a physiological response is becoming understood with the isolation of mutations in both positive and negative regulators in the jasmonate signaling pathway and the use of cDNA microarrays.
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Lukacik, Petra; Lobley, Carina M C; Bumann, Mario; Arena de Souza, Victoria; Owens, Raymond J; O'Toole, Paul W; Walsh, Martin A
2015-10-01
Probiotic bacterial strains have been shown to enhance the health of the host through a range of mechanisms including colonization, resistance against pathogens, secretion of antimicrobial compounds and modulation of the activity of the innate immune system. Lactobacillus salivarius UCC118 is a well characterized probiotic strain which survives intestinal transit and has many desirable host-interaction properties. Probiotic bacteria display a wide range of catabolic activities, which determine their competitiveness in vivo. Some lactobacilli are heterofermentative and can metabolize pentoses, using a pathway in which transketolase and transaldolase are key enzymes. L. salivarius UCC118 is capable of pentose utilization because it encodes the key enzymes on a megaplasmid. The crystal structures of the megaplasmid-encoded transketolase with and without the enzyme cofactor thiamine pyrophosphate have been determined. Comparisons with other known transketolase structures reveal a high degree of structural conservation in both the catalytic site and the overall conformation. This work extends structural knowledge of the transketolases to the industrially and commercially important Lactobacillus genus.
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Evolution and Diversity of Biosynthetic Gene Clusters in Fusarium
Directory of Open Access Journals (Sweden)
Koen Hoogendoorn
2018-06-01
Full Text Available Plant pathogenic fungi in the Fusarium genus cause severe damage to crops, resulting in great financial losses and health hazards. Specialized metabolites synthesized by these fungi are known to play key roles in the infection process, and to provide survival advantages inside and outside the host. However, systematic studies of the evolution of specialized metabolite-coding potential across Fusarium have been scarce. Here, we apply a combination of bioinformatic approaches to identify biosynthetic gene clusters (BGCs across publicly available genomes from Fusarium, to group them into annotated families and to study gain/loss events of BGC families throughout the history of the genus. Comparison with MIBiG reference BGCs allowed assignment of 29 gene cluster families (GCFs to pathways responsible for the production of known compounds, while for 57 GCFs, the molecular products remain unknown. Comparative analysis of BGC repertoires using ancestral state reconstruction raised several new hypotheses on how BGCs contribute to Fusarium pathogenicity or host specificity, sometimes surprisingly so: for example, a gene cluster for the biosynthesis of hexadehydro-astechrome was identified in the genome of the biocontrol strain Fusarium oxysporum Fo47, while being absent in that of the tomato pathogen F. oxysporum f.sp. lycopersici. Several BGCs were also identified on supernumerary chromosomes; heterologous expression of genes for three terpene synthases encoded on the Fusarium poae supernumerary chromosome and subsequent GC/MS analysis showed that these genes are functional and encode enzymes that each are able to synthesize koraiol; this observed functional redundancy supports the hypothesis that localization of copies of BGCs on supernumerary chromosomes provides freedom for evolutionary innovations to occur, while the original function remains conserved. Altogether, this systematic overview of biosynthetic diversity in Fusarium paves the way for
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antiSMASH 3.0-a comprehensive resource for the genome mining of biosynthetic gene clusters.
Weber, Tilmann; Blin, Kai; Duddela, Srikanth; Krug, Daniel; Kim, Hyun Uk; Bruccoleri, Robert; Lee, Sang Yup; Fischbach, Michael A; Müller, Rolf; Wohlleben, Wolfgang; Breitling, Rainer; Takano, Eriko; Medema, Marnix H
2015-07-01
Microbial secondary metabolism constitutes a rich source of antibiotics, chemotherapeutics, insecticides and other high-value chemicals. Genome mining of gene clusters that encode the biosynthetic pathways for these metabolites has become a key methodology for novel compound discovery. In 2011, we introduced antiSMASH, a web server and stand-alone tool for the automatic genomic identification and analysis of biosynthetic gene clusters, available at http://antismash.secondarymetabolites.org. Here, we present version 3.0 of antiSMASH, which has undergone major improvements. A full integration of the recently published ClusterFinder algorithm now allows using this probabilistic algorithm to detect putative gene clusters of unknown types. Also, a new dereplication variant of the ClusterBlast module now identifies similarities of identified clusters to any of 1172 clusters with known end products. At the enzyme level, active sites of key biosynthetic enzymes are now pinpointed through a curated pattern-matching procedure and Enzyme Commission numbers are assigned to functionally classify all enzyme-coding genes. Additionally, chemical structure prediction has been improved by incorporating polyketide reduction states. Finally, in order for users to be able to organize and analyze multiple antiSMASH outputs in a private setting, a new XML output module allows offline editing of antiSMASH annotations within the Geneious software. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.
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Isenberg-Grzeda, Elie; Alici, Yesne; Hatzoglou, Vaios; Nelson, Christian; Breitbart, William
2016-01-01
Wernicke-Korsakoff Syndrome (WKS) is a neuropsychiatric syndrome caused by thiamine deficiency. Cancer predisposes to thiamine deficiency through various mechanisms. Although many case reports exist on nonalcoholic WKS in cancer, larger qualitative studies are lacking. Retrospective study of patients admitted to a cancer hospital and diagnosed with WKS during routine care on a psychiatric consultation service. Only patients with at least 1 additional supporting feature (magnetic resonance imaging findings, low serum thiamine concentrations, or response to treatment) were included. Data pertaining to demographics, risk factors, phenomenology, and outcomes were abstracted from medical records by chart review. In all, 18 patients were included. All patients developed WKS during cancer treatment. Hematologic malignancy, gastrointestinal tract tumors, low oral intake, and weight loss were common risk factors. All patients presented with cognitive dysfunction, most commonly impaired alertness, attention, and short-term memory. All were diagnosed by operational criteria proposed by Caine et al., 1997 (where 2 of the following are required: nutritional deficiency, ocular signs, cerebellar signs, and either altered mental status or mild memory impairment). Few exhibited Wernicke's classic triad. Diagnostic and treatment delay were common. Only 3 patients recovered fully. Nonalcoholic WKS can occur during cancer treatment and manifests clinically as delirium. Diagnosis should be made using operational criteria, not Wernicke's triad. Most patients were not underweight and had normal serum concentration of vitamin B12 and folate. A variety of mechanisms might predispose to thiamine deficiency and WKS in cancer. Given the high frequency of residual morbidity, studies should focus on decreasing diagnostic and treatment delay. Copyright © 2016 The Academy of Psychosomatic Medicine. Published by Elsevier Inc. All rights reserved.
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Molecular basis of the evolution of alternative tyrosine biosynthetic routes in plants
Energy Technology Data Exchange (ETDEWEB)
Schenck, Craig A.; Holland, Cynthia K.; Schneider, Matthew R.; Men, Yusen; Lee, Soon Goo; Jez, Joseph M.; Maeda , Hiroshi A. (UW); (WU)
2017-06-26
L-Tyrosine (Tyr) is essential for protein synthesis and is a precursor of numerous specialized metabolites crucial for plant and human health. Tyr can be synthesized via two alternative routes by different key regulatory TyrA family enzymes, prephenate dehydrogenase (PDH, also known as TyrAp) or arogenate dehydrogenase (ADH, also known as TyrAa), representing a unique divergence of primary metabolic pathways. The molecular foundation underlying the evolution of these alternative Tyr pathways is currently unknown. Here we characterized recently diverged plant PDH and ADH enzymes, obtained the X-ray crystal structure of soybean PDH, and identified a single amino acid residue that defines TyrA substrate specificity and regulation. Structures of mutated PDHs co-crystallized with Tyr indicate that substitutions of Asn222 confer ADH activity and Tyr sensitivity. Reciprocal mutagenesis of the corresponding residue in divergent plant ADHs further introduced PDH activity and relaxed Tyr sensitivity, highlighting the critical role of this residue in TyrA substrate specificity that underlies the evolution of alternative Tyr biosynthetic pathways in plants.
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Pfab, Alexander; Breindl, Matthias; Grasser, Klaus D
2018-03-01
The histone chaperone FACT is involved in the expression of genes encoding anthocyanin biosynthetic enzymes also upon induction by moderate high-light and therefore contributes to the stress-induced plant pigmentation. The histone chaperone FACT consists of the SSRP1 and SPT16 proteins and associates with transcribing RNAPII (RNAPII) along the transcribed region of genes. FACT can promote transcriptional elongation by destabilising nucleosomes in the path of RNA polymerase II, thereby facilitating efficient transcription of chromatin templates. Transcript profiling of Arabidopsis plants depleted in SSRP1 or SPT16 demonstrates that only a small subset of genes is differentially expressed relative to wild type. The majority of these genes is either up- or down-regulated in both the ssrp1 and spt16 plants. Among the down-regulated genes, those encoding enzymes of the biosynthetic pathway of the plant secondary metabolites termed anthocyanins (but not regulators of the pathway) are overrepresented. Upon exposure to moderate high-light stress several of these genes are up-regulated to a lesser extent in ssrp1/spt16 compared to wild type plants, and accordingly the mutant plants accumulate lower amounts of anthocyanin pigments. Moreover, the expression of SSRP1 and SPT16 is induced under these conditions. Therefore, our findings indicate that FACT is a novel factor required for the accumulation of anthocyanins in response to light-induction.
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Directory of Open Access Journals (Sweden)
Kristy Zera
Full Text Available Thiamine is an essential enzyme cofactor required for proper metabolic function and maintenance of metabolism and energy production in the brain. In developed countries, thiamine deficiency (TD is most often manifested following chronic alcohol consumption leading to impaired mitochondrial function, oxidative stress, inflammation and excitotoxicity. These biochemical lesions result in apoptotic cell death in both neurons and astrocytes. Comparable histological injuries in patients with hypoxia/ischemia and TD have been described in the thalamus and mammillary bodies, suggesting a congruency between the cellular responses to these stresses. Consistent with hypoxia/ischemia, TD stabilizes and activates Hypoxia Inducible Factor-1α (HIF-1α under physiological oxygen levels. However, the role of TD-induced HIF-1α in neurological injury is currently unknown. Using Western blot analysis and RT-PCR, we have demonstrated that TD induces HIF-1α expression and activity in primary mouse astrocytes. We observed a time-dependent increase in mRNA and protein expression of the pro-apoptotic and pro-inflammatory HIF-1α target genes MCP1, BNIP3, Nix and Noxa during TD. We also observed apoptotic cell death in TD as demonstrated by PI/Annexin V staining, TUNEL assay, and Cell Death ELISA. Pharmacological inhibition of HIF-1α activity using YC1 and thiamine repletion both reduced expression of pro-apoptotic HIF-1α target genes and apoptotic cell death in TD. These results demonstrate that induction of HIF-1α mediated transcriptional up-regulation of pro-apoptotic/inflammatory signaling contributes to astrocyte cell death during thiamine deficiency.
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Ishibashi, S; Yokota, T; Shiojiri, T; Matunaga, T; Tanaka, H; Nishina, K; Hirota, H; Inaba, A; Yamada, M; Kanda, T; Mizusawa, H
2003-05-01
Acute axonal polyneuropathy and Wernicke-Korsakoff encephalopathy developed simultaneously in three patients. Nerve conduction studies (NCS) detected markedly decreased compound muscle action potentials (CMAPs) and sensory nerve action potentials (SNAPs) with minimal conduction slowing; sympathetic skin responses (SSRs) were also notably decreased. Sural nerve biopsies showed only mild axonal degeneration with scattered myelin ovoid formation. The symptoms of neuropathy lessened within two weeks after an intravenous thiamine infusion. CMAPs, SNAPs, and SSRs also increased considerably. We suggest that this is a new type of peripheral nerve impairment: physiological conduction failure with minimal conduction delay due to thiamine deficiency.
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Drywień, Małgorzata; Frąckiewicz, Joanna; Górnicka, Magdalena; Gadek, Joanna; Jałosińska, Małgorzata
2015-01-01
Fermented milk drinks are unique products due to content of Lactobacillus and Bifidobacterium that are recognized as probiotics. They are a natural component of the colon microbiota as well as commonly used probiotics in functional food. The effects of the storage time and prebiotic type (inuline or oligofructose) were studied in banana-milk drink after fermentation by Lactobacillus casei KNE-1 on the thiamine and riboflavin concentrations. The material for the study was fermented fruit milk drinks: banana-milk prepared in laboratory conditions and fruit milk drinks purchased in a local shop, as a comparative material. The thiamine was determined by thiochrome method and the riboflavin was determined by fluorometric method. The storage time after the end of the fermentation process did not increase the content of thiamine and riboflavin in fermented banana-milk drink more than the output level. The addition of oligofructose significantly affected the synthesis of thiamine by Lactobacillus casei KNE-1 irrespectively of the storage time. The storage time but not the type of prebiotic affected the riboflavin concentration. Taking into account the highest content of both vitamins, the banana-milk drink fermented by Lactobacillus casei KNE-1 should be consumed immediately or 24 days after fermentation. This information could be used by manufacturers for the planning of technological process. The content of thiamine and riboflavin in the fermented milk drinks is the result of the type of prebiotic, the individual bacterial strain properties as well as the storage time. These factors should be investigated to optimize the content of B vitamins in fermented milk drinks in the future.
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Directory of Open Access Journals (Sweden)
nazi nadernejad
2017-08-01
Full Text Available Soybean (Glycin max L. is one of the most important oily seeds in the world. This plant is rich in protein and unsaturated fats, and plays a significant role in human health with phenolic compounds and flavonoids. Indole Butyric Acid (IBA is a plant growth regulator that plays a key role in producing phenolic compounds and increasing the antioxidant capacity of the plant. Thiamine is one of the important vitamins in strengthening the immune system and increasing the resistance to environmental stresses in the plant's growth stages. Regarding the effect of hormone auxin and thiamine on the production of phenolic compounds as one of the antioxidant compounds in growth stages, the aim of this study was to investigate the effect of the two compounds in two stages of soybean growth and compare their effect on phenolic compounds changes in order to Detecting higher antioxidant capacity in environmental stress tolerance. For this purpose, the DPX cultivar of soybean seeds were prepared from Dezful Agriculture Research Center and planted in perlite containing flowers. The plants were planted under factorial design under IBA treatments with three concentrations of 0, 10 and 50 and thiamine with three concentrations of 0, 50 and 200. Extraction and evaluation of phenolic compounds, anthocyanins and pigments in leaves were performed. Data were analyzed using Duncan's test at a significant level of 5%. The results showed that the combined use of auxin and thiamine increased the carotenoid content in both phases and caused a significant increase in phenolic content. Application of auxin alone reduced auxin and thiamine the anthocyanin content significantly in both phases, but did not have a significant effect on phenolic content. The results showed that the PAL activity of the phenolic and anthocyanin content increased significantly in the 9-leaf stage compared to 3-leaf. Generally, the results showed that interaction effect between auxin and thiamine on
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Directory of Open Access Journals (Sweden)
moslem salehi
2017-02-01
Full Text Available Introduction: Bedding plants, especially petunia is important element for urban landscaping and attracted the attention of landscapers. This is due to some properties such as growth habit and color. The petunia (Petunia hybrida L. belongs to Solanaceae family that has annual and perennial varieties. This plant is originally from Argentina, Brazil and Uruguay. Some plant growth regulators such as gibberellic acid (GA3 and vitamins including thiamine and ascorbic acid affect plant growth and development and may extend flowering period. Vitamin C affects cell division and cell growth in plants and is effective on the feeding cycle activity in higher plants and it has an important role in electron transport system. The concentrations of 50 and 100 ppm of vitamin C and thiamine can increase the plant height, leaf number, leaf area, fresh and dry weight, and chemical compounds of the Syngonium plant. The application of 100 mg/l of GA3 significantly increased plant height and the number of leaves of gladiolus. Material and methods: The experiment was arranged in a factorial based on a completely randomized design with five replications. In this research, growth characteristics (lateral branch number, flower number, flower diameter, stem diameter, root length, and lateral branch length and biochemical characteristics (chlorophyll a, chlorophyll b, total chlorophyll, carotenoids, and reducing sugar were measured. After seeding and transplanting the seedling at 6 leaf stage, plants sprayed at 4 various growth stages with following treatments: 1-\tControl 2-\tVitamin C(100 mg/l 3-\tThiamin (100 mg/l 4-\tGibberellic acid (100 mg/l 5-\tVitamin C and Thiamin (The concentration of both 100 mg/l 6-\tVitamin C and Gibberellic acid (The concentration of both 100 mg/l 7-\tThiamin and Gibberellic acid (The concentration of both 100 mg/l 8-\tVitamin C, Thiamin and Gibberellic acid (The concentration of three 100 mg/l Data obtained from the measured parameters
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Sobhani Najafabadi, Ahmad; Naghavi, Mohammad Reza; Farahmand, Hamid; Abbasi, Alireza
2017-11-01
Ferula gummosa Boiss. is an industrial and pharmaceutical plant that has been highly recognized for its valuable oleo-gum-resin, namely galbanum. Despite the fabulous value of galbanum, very little information on the genetic and biochemical mechanisms of its production existed. In the present study, the oleo-gum-resin and four organs (root, flower, stem, and leaf) of F. gummosa were assessed in terms of metabolic compositions and the expression of genes involved in their biosynthetic pathways. Results showed that the most accumulation of resin and essential oils were occurred in the roots (13.99 mg/g) and flowers (6.01 mg/g), respectively. While the most dominant compound of the resin was β-amyrin from triterpenes, the most abundant compounds of the essential oils were α-pinene and β-pinene from monoterpenes and α-eudesmol and germacrene-D from sesquiterpenes. Transcriptome analysis was performed by RNA sequencing (RNA-seq) for the plant roots and flowers. Differential gene expression analysis showed that 1172 unigenes were differential between two organs that 934 (79.6%) of them were up-regulated in the flowers and 238 (20.4%) unigenes were up-regulated in the roots (FDR ≤0.001). The most important up-regulated unigenes in the roots were involved in the biosynthesis of the major components of galbanum, including myrcene, germacrene-D, α-terpineol, and β-amyrin. The results obtained by RNA-Seq were confirmed by qPCR. These analyses showed that different organs of F. gummosa are involved in the production of oleo-gum-resin, but the roots are more active than other organs in terms of the biosynthesis of triterpenes and some mono- and sesquiterpenes. This study provides rich molecular and biochemical resources for further studies on molecular genetics and functional genomics of oleo-gum-resin production in F. gummosa.
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Beltramo, Elena; Berrone, Elena; Tarallo, Sonia; Porta, Massimo
2009-09-01
Vascular cells in diabetes are subjected to daily fluctuations from high to low glucose. We aimed at investigating whether pulsed exposure to different glucose concentrations influences apoptosis in human retinal pericytes (HRP) versus bovine retinal pericytes (BRP), with consequences on the onset of diabetic retinopathy, and the possible protective role of thiamine. BRP and HRP (wild-type and immortalized) were grown in physiological/high glucose for 7 days, and then returned to physiological glucose for another 24, 48 or 72 h. Cells were also kept intermittently at 48-h intervals in high/normal glucose for 8 days, with/without thiamine/benfotiamine. Apoptosis was determined through ELISA, TUNEL, Bcl-2, Bax and p53 expression/concentration. Continuous exposure to high glucose increased apoptosis in BRP, but not HRP. BRP apoptosis normalized within 24 h of physiological glucose re-entry, while HRP apoptosis increased within 24-48 h of re-entry. Intermittent exposure to high glucose increased apoptosis in HRP and BRP. Bcl-2/Bax results were consistent with DNA fragmentation, while p53 was unchanged. Thiamine and benfotiamine countered intermittent high glucose-induced apoptosis. Human pericytes are less prone to apoptosis induced by persistently high glucose than bovine cells. However, while BRP recover after returning to physiological levels, HRP are more vulnerable to both downwardly fluctuating glucose levels and intermittent exposure. These findings reinforce the hypotheses that (1) glycaemic fluctuations play a role in the development of diabetic retinopathy and (2) species-specific models are needed. Thiamine and benfotiamine prevent human pericyte apoptosis, indicating this vitamin as an inexpensive approach to the prevention and/or treatment of diabetic complications.
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Tarigh, Ghazale Daneshvar; Shemirani, Farzaneh
2014-06-01
A simple and rapid method for the simultaneous in situ derivatizaion, preconcentration and extraction of thiamine (vitamin B1) as a model analyte was developed by a novel quantitative method, namely ultrasound-assisted dispersive magnetic solid phase extraction spectrofluorimetry (USA-DMSPE-FL) from different real samples. This method consists of sample preparation, in situ derivatization, exhaustive extraction and clean up by a single process. High extraction efficiency and in situ derivatization in a short period of time is the main advantages of this procedure. For this purpose, the reusable magnetic multi-wall carbon nanotube (MMWCNT) nanocomposite was used as an adsorbent for preconcentration and determination of thiamine. Thiamine was, simultaneously, in situ derivatized as thiochrome by potassium hexacyanoferrate (III) and adsorbed on MMWCNT in an ultrasonic water bath. The MMWCNTs were then collected using an external magnetic field. Subsequently, the extracted thiochrome was washed from the surface of the adsorbent and determined by spectrofluorimetry. The developed method, which has been analytically characterized under its optimal operating conditions, allows the detection of the analyte in the samples with method detection limits of 0.37 µg L(-1). The repeatability of the method, expressed as the relative standard deviation (RSD, n=6), varies between 2.0% and 4.8% in different real samples, while the enhancement factor is 197. The proposed procedure has been applied for the determination of thiamine in biological (serum and urine), pharmaceutical (multivitamin tablet and B complex syrup) and foodstuff samples (cereal, wheat flour, banana and honey) with the good recoveries in the range from 90% to 105%. Copyright © 2014 Elsevier B.V. All rights reserved.
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Mozzillo, Enza; Melis, Daniela; Falco, Mariateresa; Fattorusso, Valentina; Taurisano, Roberta; Flanagan, Sarah E; Ellard, Sian; Franzese, Adriana
2013-08-01
Thiamine responsive megaloblastic anemia (TRMA) is an autosomal recessive disease caused by loss of function mutations in the SLC19A2 gene. TRMA is characterized by anemia, deafness, and diabetes. In some cases, optic atrophy or more rarely retinitis pigmentosa is noted. We now report two sisters, the eldest of which presented to a different hospital during childhood with sensorineural deafness, which was treated with a hearing prosthesis, insulin requiring diabetes, retinitis pigmentosa, optic atrophy, and macrocytic anemia. These features initially suggested a clinical diagnosis of Wolfram syndrome (WS). Therapy with thiamine was initiated which resulted in the resolution of the anemia. The younger sister, who was affected with sensorineural deafness, was referred to our hospital for non-autoimmune diabetes. She was found to have macrocytosis and ocular abnormalities. Because a diagnosis of TRMA was suspected, therapy with insulin and thiamine was started. Sequencing analysis of the SLC19A2 gene identified a compound heterozygous mutation p.Y81X/p.L457X (c.242insA/c.1370delT) in both sisters. Non-autoimmune diabetes associated with deafness and macrocytosis, without anemia, suggests a diagnosis of TRMA. Patients clinically diagnosed with WS with anemia and/or macrocytosis should be reevaluated for TRMA. © 2012 John Wiley & Sons A/S.
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Fox, J. B.; Thayer, D. W.; Phillips, J. G.
The effect of low dose γ-irradiation on the thiamin content of ground pork was studied in the range of 0-14 kGy at 2°C and at radiation doses from 0.5 to 7 kGy at temperatures -20, 10, 0, 10 and 20°C. The detailed study at 2°C showed that loss of thiamin was exponential down to 0kGy. An exponential expression was derived for the effect of radiation dose and temperature of irradiation on thiamin loss, and compared with a previously derived general linear expression. Both models were accurate depictions of the data, but the exponential expression showed a significant decrease in the rate of loss between 0 and -10°C. This is the range over which water in meat freezes, the decrease being due to the immobolization of reactive radiolytic products of water in ice crystals.
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International Nuclear Information System (INIS)
Wajda, L.; Korzonek, M.
1979-01-01
Influence of thiamine within the investigated range of concentrations 1.5 to 12.0 mg/l on the postirradiation effect caused by fast neutrons or gamma irradiation was found in cultures of Haplopappus gracilis callus. The lowest sensitivity to fast neutrons was noticed in callus grown on nutrient combination 1.5 to 3.0 and 12.0 to 3.0 mg/l of thiamine. On the contrary the highest sensitivity to gamma radiation was shown by cultures grown on nutrient media: 1.5 to 3.0 mg/l; all the other investigated thiamine concentrations caused a marked decrease in the sensitivity of the callus. (author)
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Menendez-Bravo, Simón; Comba, Santiago; Sabatini, Martín; Arabolaza, Ana; Gramajo, Hugo
2014-07-01
Microbial fatty acid (FA)-derived molecules have emerged as promising alternatives to petroleum-based chemicals for reducing dependence on fossil hydrocarbons. However, native FA biosynthetic pathways often yield limited structural diversity, and therefore restricted physicochemical properties, of the end products by providing only a limited variety of usually linear hydrocarbons. Here we have engineered into Escherichia coli a mycocerosic polyketide synthase-based biosynthetic pathway from Mycobacterium tuberculosis and redefined its biological role towards the production of multi-methyl-branched-esters (MBEs) with novel chemical structures. Expression of FadD28, Mas and PapA5 enzymes enabled the biosynthesis of multi-methyl-branched-FA and their further esterification to an alcohol. The high substrate tolerance of these enzymes towards different FA and alcohol moieties resulted in the biosynthesis of a broad range of MBE. Further metabolic engineering of the MBE producer strain coupled this system to long-chain-alcohol biosynthetic pathways resulting in de novo production of branched wax esters following addition of only propionate. Copyright © 2014 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.
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Hammes, Hans-Peter; Du, Xueliang; Edelstein, Diane; Taguchi, Tetsuya; Matsumura, Takeshi; Ju, Qida; Lin, Jihong; Bierhaus, Angelika; Nawroth, Peter; Hannak, Dieter; Neumaier, Michael; Bergfeld, Regine; Giardino, Ida; Brownlee, Michael
2003-03-01
Three of the major biochemical pathways implicated in the pathogenesis of hyperglycemia induced vascular damage (the hexosamine pathway, the advanced glycation end product (AGE) formation pathway and the diacylglycerol (DAG)-protein kinase C (PKC) pathway) are activated by increased availability of the glycolytic metabolites glyceraldehyde-3-phosphate and fructose-6-phosphate. We have discovered that the lipid-soluble thiamine derivative benfotiamine can inhibit these three pathways, as well as hyperglycemia-associated NF-kappaB activation, by activating the pentose phosphate pathway enzyme transketolase, which converts glyceraldehyde-3-phosphate and fructose-6-phosphate into pentose-5-phosphates and other sugars. In retinas of diabetic animals, benfotiamine treatment inhibited these three pathways and NF-kappaB activation by activating transketolase, and also prevented experimental diabetic retinopathy. The ability of benfotiamine to inhibit three major pathways simultaneously might be clinically useful in preventing the development and progression of diabetic complications.
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Novel metabolic pathways in Archaea.
Sato, Takaaki; Atomi, Haruyuki
2011-06-01
The Archaea harbor many metabolic pathways that differ to previously recognized classical pathways. Glycolysis is carried out by modified versions of the Embden-Meyerhof and Entner-Doudoroff pathways. Thermophilic archaea have recently been found to harbor a bi-functional fructose-1,6-bisphosphate aldolase/phosphatase for gluconeogenesis. A number of novel pentose-degrading pathways have also been recently identified. In terms of anabolic metabolism, a pathway for acetate assimilation, the methylaspartate cycle, and two CO2-fixing pathways, the 3-hydroxypropionate/4-hydroxybutyrate cycle and the dicarboxylate/4-hydroxybutyrate cycle, have been elucidated. As for biosynthetic pathways, recent studies have clarified the enzymes responsible for several steps involved in the biosynthesis of inositol phospholipids, polyamine, coenzyme A, flavin adeninedinucleotide and heme. By examining the presence/absence of homologs of these enzymes on genome sequences, we have found that the majority of these enzymes and pathways are specific to the Archaea. Copyright © 2011 Elsevier Ltd. All rights reserved.
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Lin-Wang, Kui; Bolitho, Karen; Grafton, Karryn; Kortstee, Anne; Karunairetnam, Sakuntala; McGhie, Tony K; Espley, Richard V; Hellens, Roger P; Allan, Andrew C
2010-03-21
The control of plant anthocyanin accumulation is via transcriptional regulation of the genes encoding the biosynthetic enzymes. A key activator appears to be an R2R3 MYB transcription factor. In apple fruit, skin anthocyanin levels are controlled by a gene called MYBA or MYB1, while the gene determining fruit flesh and foliage anthocyanin has been termed MYB10. In order to further understand tissue-specific anthocyanin regulation we have isolated orthologous MYB genes from all the commercially important rosaceous species. We use gene specific primers to show that the three MYB activators of apple anthocyanin (MYB10/MYB1/MYBA) are likely alleles of each other. MYB transcription factors, with high sequence identity to the apple gene were isolated from across the rosaceous family (e.g. apples, pears, plums, cherries, peaches, raspberries, rose, strawberry). Key identifying amino acid residues were found in both the DNA-binding and C-terminal domains of these MYBs. The expression of these MYB10 genes correlates with fruit and flower anthocyanin levels. Their function was tested in tobacco and strawberry. In tobacco, these MYBs were shown to induce the anthocyanin pathway when co-expressed with bHLHs, while over-expression of strawberry and apple genes in the crop of origin elevates anthocyanins. This family-wide study of rosaceous R2R3 MYBs provides insight into the evolution of this plant trait. It has implications for the development of new coloured fruit and flowers, as well as aiding the understanding of temporal-spatial colour change.
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Directory of Open Access Journals (Sweden)
McGhie Tony K
2010-03-01
Full Text Available Abstract Background The control of plant anthocyanin accumulation is via transcriptional regulation of the genes encoding the biosynthetic enzymes. A key activator appears to be an R2R3 MYB transcription factor. In apple fruit, skin anthocyanin levels are controlled by a gene called MYBA or MYB1, while the gene determining fruit flesh and foliage anthocyanin has been termed MYB10. In order to further understand tissue-specific anthocyanin regulation we have isolated orthologous MYB genes from all the commercially important rosaceous species. Results We use gene specific primers to show that the three MYB activators of apple anthocyanin (MYB10/MYB1/MYBA are likely alleles of each other. MYB transcription factors, with high sequence identity to the apple gene were isolated from across the rosaceous family (e.g. apples, pears, plums, cherries, peaches, raspberries, rose, strawberry. Key identifying amino acid residues were found in both the DNA-binding and C-terminal domains of these MYBs. The expression of these MYB10 genes correlates with fruit and flower anthocyanin levels. Their function was tested in tobacco and strawberry. In tobacco, these MYBs were shown to induce the anthocyanin pathway when co-expressed with bHLHs, while over-expression of strawberry and apple genes in the crop of origin elevates anthocyanins. Conclusions This family-wide study of rosaceous R2R3 MYBs provides insight into the evolution of this plant trait. It has implications for the development of new coloured fruit and flowers, as well as aiding the understanding of temporal-spatial colour change.
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Directory of Open Access Journals (Sweden)
Belshoff Alex C
2011-05-01
Full Text Available Abstract Background Stable isotope tracing is a powerful technique for following the fate of individual atoms through metabolic pathways. Measuring isotopic enrichment in metabolites provides quantitative insights into the biosynthetic network and enables flux analysis as a function of external perturbations. NMR and mass spectrometry are the techniques of choice for global profiling of stable isotope labeling patterns in cellular metabolites. However, meaningful biochemical interpretation of the labeling data requires both quantitative analysis and complex modeling. Here, we demonstrate a novel approach that involved acquiring and modeling the timecourses of 13C isotopologue data for UDP-N-acetyl-D-glucosamine (UDP-GlcNAc synthesized from [U-13C]-glucose in human prostate cancer LnCaP-LN3 cells. UDP-GlcNAc is an activated building block for protein glycosylation, which is an important regulatory mechanism in the development of many prominent human diseases including cancer and diabetes. Results We utilized a stable isotope resolved metabolomics (SIRM approach to determine the timecourse of 13C incorporation from [U-13C]-glucose into UDP-GlcNAc in LnCaP-LN3 cells. 13C Positional isotopomers and isotopologues of UDP-GlcNAc were determined by high resolution NMR and Fourier transform-ion cyclotron resonance-mass spectrometry. A novel simulated annealing/genetic algorithm, called 'Genetic Algorithm for Isotopologues in Metabolic Systems' (GAIMS was developed to find the optimal solutions to a set of simultaneous equations that represent the isotopologue compositions, which is a mixture of isotopomer species. The best model was selected based on information theory. The output comprises the timecourse of the individual labeled species, which was deconvoluted into labeled metabolic units, namely glucose, ribose, acetyl and uracil. The performance of the algorithm was demonstrated by validating the computed fractional 13C enrichment in these subunits
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Wang, Kui-Lin; Bolitho, Karen; Grafton, Karryn; Kortstee, A.J.; Karunairetnam, Sakuntala; McGhie, T.K.; Espley, R.V.; Hellens, R.P.; Allan, A.C.
2010-01-01
Background - The control of plant anthocyanin accumulation is via transcriptional regulation of the genes encoding the biosynthetic enzymes. A key activator appears to be an R2R3 MYB transcription factor. In apple fruit, skin anthocyanin levels are controlled by a gene called MYBA or MYB1, while the
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Honeyfield, D.C.; Ross, J.P.; Carbonneau, D.A.; Terrell, S.P.; Woodward, A.R.; Schoeb, T.R.; Perceval, H.F.; Hinterkopf, J.P.
2008-01-01
An investigation of adult alligator (Alligator mississippiensis) mortalities in Lake Griffin, central Florida, was conducted from 1998-2004. Alligator mortality was highest in the months of April and May and annual death count peaked in 2000. Bacterial pathogens, heavy metals, and pesticides were not linked with the mortalities. Blood chemistry did not point to any clinical diagnosis, although differences between impaired and normal animals were noted. Captured alligators with signs of neurologic impairment displayed unresponsive and uncoordinated behavior. Three of 21 impaired Lake Griffin alligators were found to have neural lesions characteristic of thiamine deficiency in the telencephalon, particularly the dorsal ventricular ridge. In some cases, lesions were found in the thalamus, and parts of the midbrain. Liver and muscle tissue concentrations of thiamine (vitamin B"1) were lowest in impaired Lake Griffin alligators when compared to unimpaired alligators or to alligators from Lake Woodruff. The consumption of thiaminase-positive gizzard shad (Dorosoma cepedianum) is thought to have been the cause of the low tissue thiamine and resulting mortalities. ?? Wildlife Disease Association 2008.
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Novel key metabolites reveal further branching of the roquefortine/meleagrin biosynthetic pathway.
Ries, Marco I; Ali, Hazrat; Lankhorst, Peter P; Hankemeier, Thomas; Bovenberg, Roel A L; Driessen, Arnold J M; Vreeken, Rob J
2013-12-27
Metabolic profiling and structural elucidation of novel secondary metabolites obtained from derived deletion strains of the filamentous fungus Penicillium chrysogenum were used to reassign various previously ascribed synthetase genes of the roquefortine/meleagrin pathway to their corresponding products. Next to the structural characterization of roquefortine F and neoxaline, which are for the first time reported for P. chrysogenum, we identified the novel metabolite roquefortine L, including its degradation products, harboring remarkable chemical structures. Their biosynthesis is discussed, questioning the exclusive role of glandicoline A as key intermediate in the pathway. The results reveal that further enzymes of this pathway are rather unspecific and catalyze more than one reaction, leading to excessive branching in the pathway with meleagrin and neoxaline as end products of two branches.
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Thiamin is decomposed due to Anaphe spp. entomophagy in seasonal ataxia patients in Nigeria.
Nishimune, T; Watanabe, Y; Okazaki, H; Akai, H
2000-06-01
A fairly high activity of a relatively heat-resistant thiaminase was detected and characterized from the pupae of an African silkworm Anaphe spp. which had been the putative cause of a seasonal ataxia and impaired consciousness in Nigerians. The thiaminase in the buffer extract of Anaphe pupae was type I (thiamin: base 2-methyl-4-aminopyrimidine methyl transferase EC 2.5.1.2), and the optimal temperature and pH were 70 degrees C and 8.0-8.5, respectively. Based on gel filtration chromatography, the molecules were estimated to be 200 kDa. Second substrates which could be utilized by the thiaminase were pyridoxine, amino acids, glutathione, taurine and 4-aminopyridine. Thiamin phosphate esters were inactive as substrates. This is the first report describing an insect thiaminase. Our results indicate the necessity of thorough heat treatment for the detoxification of the African silkworm, making the worm a safe source of high-quality protein.
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Harper, Clive G; Sheedy, Donna L; Lara, Ana I; Garrick, Therese M; Hilton, John M; Raisanen, Jack
2012-01-01
Objective To determine the prevalence of the Wernicke-Korsakoff syndrome (WKS) in Australia and compare this with previous studies. Design and setting Prospective autopsy study at the New South Wales Institute of Forensic Medicine, 1996-1997. Methods Brains of deceased people (aged over 15 years) derived from 2212 sequential autopsies performed between 1 January 1996 and 31 December 1997 were studied macroscopically and microscopically to identify cases of WKS. Main outcome measures Standard histological criteria for WKS and any available clinical data. Results Twenty-five cases of WKS were identified (prevalence, 1.1%), mostly among the 5.9% of the 2212 people who had a history suggestive of alcohol abuse. Only four cases (16%) had been diagnosed during life. Conclusions There has been a significant reduction in the prevalence of WKS in Australia since the introduction of thiamine enrichment of bread flour. While the prevalence is still higher than in most other Western countries, further research is needed before adding thiamine to alcoholic beverages can be recommended. PMID:9640303
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Wang, Yu; Dou, Ying; Wang, Rui; Guan, Xuelian; Hu, Zenghui; Zheng, Jian
2017-11-30
The flower color of Syringa oblata Lindl., which is often modulated by the flavonoid content, varies and is an important ornamental feature. Chalcone synthase (CHS) catalyzes the first key step in the flavonoid biosynthetic pathway. However, little is known about the role of S. oblata CHS (SoCHS) in flavonoid biosynthesis in this species. Here, we isolate and analyze the cDNA (SoCHS1) that encodes CHS in S. oblata. We also sought to analyzed the molecular characteristics and function of flavonoid metabolism by SoCHS1. We successfully isolated the CHS-encoding genomic DNA (gDNA) in S. oblata (SoCHS1), and the gene structural analysis indicated it had no intron. The opening reading frame (ORF) sequence of SoCHS1 was 1170bp long and encoded a 389-amino acid polypeptide. Multiple sequence alignment revealed that both the conserved CHS active site residues and CHS signature sequence were in the deduced amino acid sequence of SoCHS1. Crystallographic analysis revealed that the protein structure of SoCHS1 is highly similar to that of FnCHS1 in Freesia hybrida. The quantitative real-time polymerase chain reaction (PCR) performed to detect the SoCHS1 transcript expression levels in flowers, and other tissues revealed the expression was significantly correlated with anthocyanin accumulation during flower development. The ectopic expression results of Nicotiana tabacum showed that SoCHS1 overexpression in transgenic tobacco changed the flower color from pale pink to pink. In conclusion, these results suggest that SoCHS1 plays an essential role in flavonoid biosynthesis in S. oblata, and could be used to modify flavonoid components in other plant species. Copyright © 2017. Published by Elsevier B.V.
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International Nuclear Information System (INIS)
Nara, Takeshi; Hashimoto, Muneaki; Hirawake, Hiroko; Liao, Chien-Wei; Fukai, Yoshihisa; Suzuki, Shigeo; Tsubouchi, Akiko; Morales, Jorge; Takamiya, Shinzaburo; Fujimura, Tsutomu; Taka, Hikari; Mineki, Reiko; Fan, Chia-Kwung; Inaoka, Daniel Ken; Inoue, Masayuki; Tanaka, Akiko; Harada, Shigeharu; Kita, Kiyoshi
2012-01-01
Highlights: ► An Escherichia coli strain co-expressing CPSII, ATC, and DHO of Trypanosoma cruzi was constructed. ► Molecular interactions between CPSII, ATC, and DHO of T. cruzi were demonstrated. ► CPSII bound with both ATC and DHO. ► ATC bound with both CPSII and DHO. ► A functional tri-enzyme complex might precede the establishment of the fused enzyme. -- Abstract: The first 3 reaction steps of the de novo pyrimidine biosynthetic pathway are catalyzed by carbamoyl-phosphate synthetase II (CPSII), aspartate transcarbamoylase (ATC), and dihydroorotase (DHO), respectively. In eukaryotes, these enzymes are structurally classified into 2 types: (1) a CPSII-DHO-ATC fusion enzyme (CAD) found in animals, fungi, and amoebozoa, and (2) stand-alone enzymes found in plants and the protist groups. In the present study, we demonstrate direct intermolecular interactions between CPSII, ATC, and DHO of the parasitic protist Trypanosoma cruzi, which is the causative agent of Chagas disease. The 3 enzymes were expressed in a bacterial expression system and their interactions were examined. Immunoprecipitation using an antibody specific for each enzyme coupled with Western blotting-based detection using antibodies for the counterpart enzymes showed co-precipitation of all 3 enzymes. From an evolutionary viewpoint, the formation of a functional tri-enzyme complex may have preceded—and led to—gene fusion to produce the CAD protein. This is the first report to demonstrate the structural basis of these 3 enzymes as a model of CAD. Moreover, in conjunction with the essentiality of de novo pyrimidine biosynthesis in the parasite, our findings provide a rationale for new strategies for developing drugs for Chagas disease, which target the intermolecular interactions of these 3 enzymes.
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Liu, Fenghong; Wang, Lei; Gu, Liang; Zhao, Wei; Su, Hongyan; Cheng, Xianhao
2015-12-01
In our preliminary study, the ripe fruits of two highbush blueberry (Vaccinium corymbosum L.) cultivars, cv 'Berkeley' and cv 'Bluecrop', were found to contain different levels of ascorbic acid. However, factors responsible for these differences are still unknown. In the present study, ascorbic acid content in fruits was compared with expression profiles of ascorbic acid biosynthetic and recycling genes between 'Bluecrop' and 'Berkeley' cultivars. The results indicated that the l-galactose pathway was the predominant route of ascorbic acid biosynthesis in blueberry fruits. Moreover, higher expression levels of the ascorbic acid biosynthetic genes GME, GGP, and GLDH, as well as the recycling genes MDHAR and DHAR, were associated with higher ascorbic acid content in 'Bluecrop' compared with 'Berkeley', which indicated that a higher efficiency ascorbic acid biosynthesis and regeneration was likely to be responsible for the higher ascorbic acid accumulation in 'Bluecrop'. Copyright © 2015 Elsevier Ltd. All rights reserved.
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The preliminary research for biosynthetic engineering by radiation fusion technology
Energy Technology Data Exchange (ETDEWEB)
Roh, Chang Hyun; Jung, U Hee; Park, Hae Ran [KAERI, Daejeon (Korea, Republic of)
2012-01-15
The purpose of this project is to elucidate the solution to the production of bioactive substance using biotransformation process from core technology of biosynthetic engineering by radiation fusion technology. And, this strategy will provide core technology for development of drugs as new concept and category. Research scopes and contents of project include 1) The development of mutant for biosynthetic engineering by radiation fusion technology 2) The development of host for biosynthetic engineering by radiation fusion technology 3) The preliminary study for biosynthetic engineering of isoflavone by radiation fusion technology. The results are as follows. Isoflavone compounds(daidzein, hydroxylated isoflavone) were analyzed by GC-MS. The study of radiation doses and p-NCA high-throughput screening for mutant development were elucidated. And, it was carried out the study of radiation doses for host development. Furthermore, the study of redox partner and construction of recombinant strain for region-specific hydroxylation(P450, redox partner). In addition, the biological effect of 6,7,4'-trihydroxyisoflavone as an anti-obesity agent was elucidated in this study.
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Metabolic profiling of alternative NAD biosynthetic routes in mouse tissues.
Directory of Open Access Journals (Sweden)
Valerio Mori
Full Text Available NAD plays essential redox and non-redox roles in cell biology. In mammals, its de novo and recycling biosynthetic pathways encompass two independent branches, the "amidated" and "deamidated" routes. Here we focused on the indispensable enzymes gating these two routes, i.e. nicotinamide mononucleotide adenylyltransferase (NMNAT, which in mammals comprises three distinct isozymes, and NAD synthetase (NADS. First, we measured the in vitro activity of the enzymes, and the levels of all their substrates and products in a number of tissues from the C57BL/6 mouse. Second, from these data, we derived in vivo estimates of enzymes'rates and quantitative contributions to NAD homeostasis. The NMNAT activity, mainly represented by nuclear NMNAT1, appears to be high and nonrate-limiting in all examined tissues, except in blood. The NADS activity, however, appears rate-limiting in lung and skeletal muscle, where its undetectable levels parallel a relative accumulation of the enzyme's substrate NaAD (nicotinic acid adenine dinucleotide. In all tissues, the amidated NAD route was predominant, displaying highest rates in liver and kidney, and lowest in blood. In contrast, the minor deamidated route showed higher relative proportions in blood and small intestine, and higher absolute values in liver and small intestine. Such results provide the first comprehensive picture of the balance of the two alternative NAD biosynthetic routes in different mammalian tissues under physiological conditions. This fills a gap in the current knowledge of NAD biosynthesis, and provides a crucial information for the study of NAD metabolism and its role in disease.
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DEFF Research Database (Denmark)
Nielsen, Jens Christian; Grijseels, Sietske; Prigent, Sylvain
2017-01-01
Filamentous fungi produce a wide range of bioactive compounds with important pharmaceutical applications, such as antibiotic penicillins and cholesterol-lowering statins. However, less attention has been paid to fungal secondary metabolites compared to those from bacteria. In this study, we...... sequenced the genomes of 9 Penicillium species and, together with 15 published genomes, we investigated the secondary metabolism of Penicillium and identified an immense, unexploited potential for producing secondary metabolites by this genus. A total of 1,317 putative biosynthetic gene clusters (BGCs) were......-referenced the predicted pathways with published data on the production of secondary metabolites and experimentally validated the production of antibiotic yanuthones in Penicillia and identified a previously undescribed compound from the yanuthone pathway. This study is the first genus-wide analysis of the genomic...
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The role of trace elements, thiamin (e) and transketolase in autism and autistic spectrum disorder.
Obrenovich, Mark E; Shola, Dorjee; Schroedel, Kathleen; Agrahari, Aditya; Lonsdale, Derrick
2015-01-01
Although there has been much research into autism or autistic spectrum disorder (ASD), there is room for considerable conjecture regarding the etiology of these developmental brain disorders. ASD is marked by a complex interaction between environmental factors and genetic predisposition, including epistasis. This manuscript argues that changes in oxidative metabolism, thiamine homeostasis, heavy metal deposition and cellular immunity have a role in the etiopathogenesis of autism and ASD. Recent findings from our group and others provide evidence for abnormal thiol metabolism, marked by significant alteration in the deposition of several trace heavy metal species. Together with these, we find differences in thiamine homeostasis in ASD patients, which can be corrected by supplementation. We hypothesize that altered thiol metabolism from heavy metal toxicity, one of the key mechanisms for oxidative stress production, may be responsible for the biochemical alterations in transketolase, dysautonomia and abnormal thiamine homeostasis. Although it is unknown why these particular metals accumulate, we suspect that children with ASD and forms of autism may have particular trouble excreting thiol-toxic heavy metal species, many of which exist as divalent cations. We maintain mercury accumulation is evidence of altered clearance. Together with concomitant oxidative stress, these findings may offer an intriguing component or possible mechanism for oxidative stress-mediated neurodegeneration in ASD patients. Regardless of the exact cause, these factors may be more important to the etiology of this symptomatically diverse disease spectrum. Here, we offer insight into new avenues of exploration as well as the development of novel treatment approaches for these growing and devastating diseases.
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Directory of Open Access Journals (Sweden)
Yang Haihua
2009-11-01
Full Text Available Abstract Background Nikkomycins are a group of peptidyl nucleoside antibiotics and act as potent inhibitors of chitin synthases in fungi and insects. Nikkomycin X and Z are the main components produced by Streptomyces ansochromogenes. Of them, nikkomycin Z is a promising antifungal agent with clinical significance. Since highly structural similarities between nikkomycin Z and X, separation of nikkomycin Z from the culture medium of S. ansochromogenes is difficult. Thus, generating a nikkomycin Z selectively producing strain is vital to scale up the nikkomycin Z yields for clinical trials. Results A nikkomycin Z producing strain (sanPDM was constructed by blocking the imidazolone biosynthetic pathway of nikkomycin X via genetic manipulation and yielded 300 mg/L nikkomycin Z and abolished the nikkomycin X production. To further increase the yield of nikkomycin Z, the effects of different precursors on its production were investigated. Precursors of nucleoside moiety (uracil or uridine had a stimulatory effect on nikkomycin Z production while precursors of peptidyl moiety (L-lysine and L-glutamate had no effect. sanPDM produced the maximum yields of nikkomycin Z (800 mg/L in the presence of uracil at the concentration of 2 g/L and it was approximately 2.6-fold higher than that of the parent strain. Conclusion A high nikkomycin Z selectively producing was obtained by genetic manipulation combined with precursors feeding. The strategy presented here might be applicable in other bacteria to selectively produce targeted antibiotics.
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Energy Technology Data Exchange (ETDEWEB)
Nemeria, Natalia S; Arjunan, Palaniappa; Chandrasekhar, Krishnamoorthy; Mossad, Madouna; Tittmann, Kai; Furey, William; Jordan, Frank [Pitt; (Goettingen); (VA); (Rutgers)
2010-11-03
Kinetic, spectroscopic, and structural analysis tested the hypothesis that a chain of residues connecting the 4{prime}-aminopyrimidine N1{prime} atoms of thiamin diphosphates (ThDPs) in the two active centers of the Escherichia coli pyruvate dehydrogenase complex E1 component provides a signal transduction pathway. Substitution of the three acidic residues (Glu{sup 571}, Glu{sup 235}, and Glu{sup 237}) and Arg{sup 606} resulted in impaired binding of the second ThDP, once the first active center was filled, suggesting a pathway for communication between the two ThDPs. (1) Steady-state kinetic and fluorescence quenching studies revealed that upon E571A, E235A, E237A, and R606A substitutions, ThDP binding in the second active center was affected. (2) Analysis of the kinetics of thiazolium C2 hydrogen/deuterium exchange of enzyme-bound ThDP suggests half-of-the-sites reactivity for the E1 component, with fast (activated site) and slow exchanging sites (dormant site). The E235A and E571A variants gave no evidence for the slow exchanging site, indicating that only one of two active sites is filled with ThDP. (3) Titration of the E235A and E237A variants with methyl acetylphosphonate monitored by circular dichroism suggested that only half of the active sites were filled with a covalent predecarboxylation intermediate analog. (4) Crystal structures of E235A and E571A in complex with ThDP revealed the structural basis for the spectroscopic and kinetic observations and showed that either substitution affects cofactor binding, despite the fact that Glu{sup 235} makes no direct contact with the cofactor. The role of the conserved Glu{sup 571} residue in both catalysis and cofactor orientation is revealed by the combined results for the first time.
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SnapShot: O-Glycosylation Pathways across Kingdoms
DEFF Research Database (Denmark)
Joshi, Hiren J.; Narimatsu, Yoshiki; Schjoldager, Katrine T.
2018-01-01
O-glycosylation is one of the most abundant and diverse types of post-translational modifications of proteins. O-glycans modulate the structure, stability, and function of proteins and serve generalized as well as highly specific roles in most biological processes. This ShapShot presents types of......-glycans found in different organisms and their principle biosynthetic pathways...
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Biosynthetic origin of the isoprene units in chromenes of Piper aduncum (Piperaceae)
International Nuclear Information System (INIS)
Leite, Ana C.; Lopes, Adriana A.; Bolzani, Vanderlan da S.; Furlan, Maysa; Kato, Massuo J.
2007-01-01
Metabolic studies involving the incorporation of [1- 13 C]-D-glucose into intact leaves of Piper aduncum (Piperaceae) have indicated that both the mevalonate (MVA) and the pyruvate-triose (MEP) non-mevalonate pathways are implicated in the biosynthesis of isoprene moieties present in methyl 2,2-dimethyl-2H-1-chromene-6-carboxylate (1) and methyl 2,2-dimethyl-8-(3'-methyl- 2'-butenyl)-2H-1-chromene-6-carboxylate (2). The pattern of incorporation of label from [1- 13 C]-D-glucose into these chromenes was determined by quantitative 13 C NMR spectroscopy. The results confirmed that biosynthetic compartment of 1 and 2 could either be the plastid and/ or the cytosol or, possibly, an additional compartment such as the plastid inter-membrane space. (author)
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Directory of Open Access Journals (Sweden)
Irene Flønes
Full Text Available Biotin-thiamine responsive basal ganglia disease is a severe, but potentially treatable disorder caused by mutations in the SLC19A3 gene. Although the disease is inherited in an autosomal recessive manner, patients with typical phenotypes carrying single heterozygous mutations have been reported. This makes the diagnosis uncertain and may delay treatment.In two siblings with early-onset encephalopathy dystonia and epilepsy, whole-exome sequencing revealed a novel single heterozygous SLC19A3 mutation (c.337T>C. Although Sanger-sequencing and copy-number analysis revealed no other aberrations, RNA-sequencing in brain tissue suggested the second allele was silenced. Whole-genome sequencing resolved the genetic defect by revealing a novel 45,049 bp deletion in the 5'-UTR region of the gene abolishing the promoter. High dose thiamine and biotin therapy was started in the surviving sibling who remains stable. In another patient two novel compound heterozygous SLC19A3 mutations were found. He improved substantially on thiamine and biotin therapy.We show that large genomic deletions occur in the regulatory region of SLC19A3 and should be considered in genetic testing. Moreover, our study highlights the power of whole-genome sequencing as a diagnostic tool for rare genetic disorders across a wide spectrum of mutations including non-coding large genomic rearrangements.
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Paraptosis Cell Death Induction by the Thiamine Analog Benfotiamine in Leukemia Cells
Sugimori, Naomi; Espinoza, J. Luis; Trung, Ly Quoc; Takami, Akiyoshi; Kondo, Yukio; An, Dao Thi; Sasaki, Motoko; Wakayama, Tomohiko; Nakao, Shinji
2015-01-01
Benfotiamine is a synthetic thiamine analogue that stimulates transketolase, a cellular enzyme essential for glucose metabolism. Currently, benfotiamine is used to treat diabetic neuropathy. We recently reported that oral benfotiamine induced a temporary but remarkable recovery from acute myeloid leukemia in an elderly patient who was ineligible for standard chemotherapy due to dementia and renal failure. In the present study we present evidences that benfotiamine possess antitumor activity a...
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Miras-Moreno, Begoña; Almagro, Lorena; Pedreño, María Angeles; Sabater-Jara, Ana Belén
2018-04-21
Terbinafine induced a significant increase of squalene production. Terbinafine increased the expression levels of squalene synthase. Cyclodextrins did not work as elicitors due to the gene expression levels obtained. Plant sterols are essential components of membrane lipids, which contributing to their fluidity and permeability. Besides their cholesterol-lowering properties, they also have anti-inflammatory, antidiabetic and anticancer activities. Squalene, which is phytosterol precursor, is widely used in medicine, foods and cosmetics due to its anti-tumor, antioxidant and anti-aging activities. Nowadays, vegetable oils constitute the main sources of phytosterols and squalene, but their isolation and purification involve complex extraction protocols and high costs. In this work, Daucus carota cell cultures were used to evaluate the effect of cyclodextrins and terbinafine on the production and accumulation of squalene and phytosterols as well as the expression levels of squalene synthase and cycloartenol synthase genes. D. carota cell cultures were able to produce high levels of extracellular being phytosterols in the presence of cyclodextrins (12 mg/L), these compounds able to increase both the secretion and accumulation of phytosterols in the culture medium. Moreover, terbinafine induced a significant increase in intracellular squalene production, as seen after 168 h of treatment (497.0 ± 23.5 µg g dry weight -1 ) while its extracellular production only increased in the presence of cyclodextrins.The analysis of sqs and cas gene expression revealed that cyclodextrins did not induce genes encoding enzymes involved in the phytosterol biosynthetic pathway since the expression levels of sqs and cas genes in cyclodextrin-treated cells were lower than in control cells. The results, therefore, suggest that cyclodextrins were only able to release phytosterols from the cells to the extracellular medium, thus contributing to their acumulation. To sum up, D. carota
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Integrating computational methods to retrofit enzymes to synthetic pathways.
Brunk, Elizabeth; Neri, Marilisa; Tavernelli, Ivano; Hatzimanikatis, Vassily; Rothlisberger, Ursula
2012-02-01
Microbial production of desired compounds provides an efficient framework for the development of renewable energy resources. To be competitive to traditional chemistry, one requirement is to utilize the full capacity of the microorganism to produce target compounds with high yields and turnover rates. We use integrated computational methods to generate and quantify the performance of novel biosynthetic routes that contain highly optimized catalysts. Engineering a novel reaction pathway entails addressing feasibility on multiple levels, which involves handling the complexity of large-scale biochemical networks while respecting the critical chemical phenomena at the atomistic scale. To pursue this multi-layer challenge, our strategy merges knowledge-based metabolic engineering methods with computational chemistry methods. By bridging multiple disciplines, we provide an integral computational framework that could accelerate the discovery and implementation of novel biosynthetic production routes. Using this approach, we have identified and optimized a novel biosynthetic route for the production of 3HP from pyruvate. Copyright © 2011 Wiley Periodicals, Inc.
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Extending the biosynthetic repertoires of cyanobacteria and chloroplasts.
Nielsen, Agnieszka Zygadlo; Mellor, Silas Busck; Vavitsas, Konstantinos; Wlodarczyk, Artur Jacek; Gnanasekaran, Thiyagarajan; Perestrello Ramos H de Jesus, Maria; King, Brian Christopher; Bakowski, Kamil; Jensen, Poul Erik
2016-07-01
Chloroplasts in plants and algae and photosynthetic microorganisms such as cyanobacteria are emerging hosts for sustainable production of valuable biochemicals, using only inorganic nutrients, water, CO2 and light as inputs. In the past decade, many bioengineering efforts have focused on metabolic engineering and synthetic biology in the chloroplast or in cyanobacteria for the production of fuels, chemicals and complex, high-value bioactive molecules. Biosynthesis of all these compounds can be performed in photosynthetic organelles/organisms by heterologous expression of the appropriate pathways, but this requires optimization of carbon flux and reducing power, and a thorough understanding of regulatory pathways. Secretion or storage of the compounds produced can be exploited for the isolation or confinement of the desired compounds. In this review, we explore the use of chloroplasts and cyanobacteria as biosynthetic compartments and hosts, and we estimate the levels of production to be expected from photosynthetic hosts in light of the fraction of electrons and carbon that can potentially be diverted from photosynthesis. The supply of reducing power, in the form of electrons derived from the photosynthetic light reactions, appears to be non-limiting, but redirection of the fixed carbon via precursor molecules presents a challenge. We also discuss the available synthetic biology tools and the need to expand the molecular toolbox to facilitate cellular reprogramming for increased production yields in both cyanobacteria and chloroplasts. © 2016 The Authors The Plant Journal © 2016 John Wiley & Sons Ltd.
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Evolutionary systems biology of amino acid biosynthetic cost in yeast.
Directory of Open Access Journals (Sweden)
Michael D Barton
2010-08-01
Full Text Available Every protein has a biosynthetic cost to the cell based on the synthesis of its constituent amino acids. In order to optimise growth and reproduction, natural selection is expected, where possible, to favour the use of proteins whose constituents are cheaper to produce, as reduced biosynthetic cost may confer a fitness advantage to the organism. Quantifying the cost of amino acid biosynthesis presents challenges, since energetic requirements may change across different cellular and environmental conditions. We developed a systems biology approach to estimate the cost of amino acid synthesis based on genome-scale metabolic models and investigated the effects of the cost of amino acid synthesis on Saccharomyces cerevisiae gene expression and protein evolution. First, we used our two new and six previously reported measures of amino acid cost in conjunction with codon usage bias, tRNA gene number and atomic composition to identify which of these factors best predict transcript and protein levels. Second, we compared amino acid cost with rates of amino acid substitution across four species in the genus Saccharomyces. Regardless of which cost measure is used, amino acid biosynthetic cost is weakly associated with transcript and protein levels. In contrast, we find that biosynthetic cost and amino acid substitution rates show a negative correlation, but for only a subset of cost measures. In the economy of the yeast cell, we find that the cost of amino acid synthesis plays a limited role in shaping transcript and protein expression levels compared to that of translational optimisation. Biosynthetic cost does, however, appear to affect rates of amino acid evolution in Saccharomyces, suggesting that expensive amino acids may only be used when they have specific structural or functional roles in protein sequences. However, as there appears to be no single currency to compute the cost of amino acid synthesis across all cellular and environmental
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Directory of Open Access Journals (Sweden)
Rosanna Mancinelli
2013-03-01
Full Text Available INTRODUCTION. Long term alcohol abuse is associated with deficiencies in essential nutrients and minerals that can cause a variety of medical consequences including accumulation of toxic metals. Aim. The aim of this research is to get evidence-based data to evaluate alcohol damage and to optimize treatment. Thiamine and thiamine diphosphate (T/TDP, zinc (Zn, selenium (Se, lead (Pb and oxidative stress in terms of reactive oxygen metabolites (ROMs were examined in blood samples from 58 alcohol dependent patients (17 females and 41 males. RESULTS. T/TDP concentration in alcoholics resulted significantly lower than controls (p < 0.005 for both sexes. Serum Zn and Se did not significantly differ from reference values. Levels of blood Pb in alcoholics resulted significantly higher (p < 0.0001 than Italian reference values and were higher in females than in males. ROMs concentration was significantly higher than healthy population only in female abusers (p = 0.005. CONCLUSION. Alcoholics show a significant increase in blood oxidative stress and Pb and decrease in thiamine. Impairment occurs mainly in female abusers confirming a gender specific vulnerability.
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Bakker, SJL; ter Maaten, JC; Gans, ROB
Conventional treatment for gestational diabetes mellitus increases the proportion of infants born with a low birth weight, a risk factor for cardiovascular disease and diabetes mellitus in later life. Thiamine supplementation during pregnancy may be shown to be a safe preventive measure. During
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Linking fungal secondary metabolites and pathways to their genes in Aspergillus
DEFF Research Database (Denmark)
Petersen, Lene Maj
. oryzae metabolites, however, revealed the chemical link between the two species. In two parallel projects, involving A. niger and A. aculeatus respectively, the polyketide 6-methyl salicylic acid (6-MSA), and corresponding biosynthetic pathways, were investigated. In A. niger, 6-MSA was converted...
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Directory of Open Access Journals (Sweden)
M. Salehi
2016-05-01
Full Text Available The improvement of growth and flowering of petunia as one of the most popular and cultivated bedding plants in Iran, is of significant importance. Thus, a CRD experiment with five replications was conducted at the Research Greenhouse of Shahid Bahonar University, Kerman, Iran. From 48 days after sowing, when the seedlings had 5-6 true leaves, the seedlings were sprayed with thiamine (0 and 100 mgL-1, ascorbic acid (0 and 100 mg L-1 and benzyl adenine (0 and 200 mg L-1 at 4 steps during growth and development. The results indicated that the treatment of ascorbic acid with thiamine and benzyl adenine led to 2.5 and 3.5-fold increases in the number and length of lateral shoots compared to control treatment. The greatest fresh weight was obtained with ascorbic acid with thiamine and benzyl adenine treatment which led to a 2.5-fold increase in this trait, compared to the control. The highest dry weight was achieved in benzyl adenine treatment. The greatest vase-life and flower diameter were found with ascorbic acid (100 mg L-1, thiamine (100 mg L-1 and benzyl adenine (200 mg L-1 treatments in an extent that the flower longevity and diameter were increased by 83% and 72%, respectively, in comparison to control. Furthermore, chlorophyll a, chlorophyll b, total chlorophyll, carotenoids and reduced sugars concentrations were significantly increased by the foliar-applied compounds compared to control.
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Directory of Open Access Journals (Sweden)
Nagaraja Reddy Rama Reddy
Full Text Available Senna (Cassia angustifolia Vahl. is a world's natural laxative medicinal plant. Laxative properties are due to sennosides (anthraquinone glycosides natural products. However, little genetic information is available for this species, especially concerning the biosynthetic pathways of sennosides. We present here the transcriptome sequencing of young and mature leaf tissue of Cassia angustifolia using Illumina MiSeq platform that resulted in a total of 6.34 Gb of raw nucleotide sequence. The sequence assembly resulted in 42230 and 37174 transcripts with an average length of 1119 bp and 1467 bp for young and mature leaf, respectively. The transcripts were annotated using NCBI BLAST with 'green plant database (txid 33090', Swiss Prot, Kyoto Encylcopedia of Genes & Genomes (KEGG, Cluster of Orthologous Gene (COG and Gene Ontology (GO. Out of the total transcripts, 40138 (95.0% and 36349 (97.7% from young and mature leaf, respectively, were annotated by BLASTX against green plant database of NCBI. We used InterProscan to see protein similarity at domain level, a total of 34031 (young leaf and 32077 (mature leaf transcripts were annotated against the Pfam domains. All transcripts from young and mature leaf were assigned to 191 KEGG pathways. There were 166 and 159 CDS, respectively, from young and mature leaf involved in metabolism of terpenoids and polyketides. Many CDS encoding enzymes leading to biosynthesis of sennosides were identified. A total of 10,763 CDS differentially expressing in both young and mature leaf libraries of which 2,343 (21.7% CDS were up-regulated in young compared to mature leaf. Several differentially expressed genes found functionally associated with sennoside biosynthesis. CDS encoding for many CYPs and TF families were identified having probable roles in metabolism of primary as well as secondary metabolites. We developed SSR markers for molecular breeding of senna. We have identified a set of putative genes involved in
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Rama Reddy, Nagaraja Reddy; Mehta, Rucha Harishbhai; Soni, Palak Harendrabhai; Makasana, Jayanti; Gajbhiye, Narendra Athamaram; Ponnuchamy, Manivel; Kumar, Jitendra
2015-01-01
Senna (Cassia angustifolia Vahl.) is a world's natural laxative medicinal plant. Laxative properties are due to sennosides (anthraquinone glycosides) natural products. However, little genetic information is available for this species, especially concerning the biosynthetic pathways of sennosides. We present here the transcriptome sequencing of young and mature leaf tissue of Cassia angustifolia using Illumina MiSeq platform that resulted in a total of 6.34 Gb of raw nucleotide sequence. The sequence assembly resulted in 42230 and 37174 transcripts with an average length of 1119 bp and 1467 bp for young and mature leaf, respectively. The transcripts were annotated using NCBI BLAST with 'green plant database (txid 33090)', Swiss Prot, Kyoto Encylcopedia of Genes & Genomes (KEGG), Cluster of Orthologous Gene (COG) and Gene Ontology (GO). Out of the total transcripts, 40138 (95.0%) and 36349 (97.7%) from young and mature leaf, respectively, were annotated by BLASTX against green plant database of NCBI. We used InterProscan to see protein similarity at domain level, a total of 34031 (young leaf) and 32077 (mature leaf) transcripts were annotated against the Pfam domains. All transcripts from young and mature leaf were assigned to 191 KEGG pathways. There were 166 and 159 CDS, respectively, from young and mature leaf involved in metabolism of terpenoids and polyketides. Many CDS encoding enzymes leading to biosynthesis of sennosides were identified. A total of 10,763 CDS differentially expressing in both young and mature leaf libraries of which 2,343 (21.7%) CDS were up-regulated in young compared to mature leaf. Several differentially expressed genes found functionally associated with sennoside biosynthesis. CDS encoding for many CYPs and TF families were identified having probable roles in metabolism of primary as well as secondary metabolites. We developed SSR markers for molecular breeding of senna. We have identified a set of putative genes involved in various
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Beltramo, Elena; Nizheradze, Konstantin; Berrone, Elena; Tarallo, Sonia; Porta, Massimo
2009-10-01
Early and selective loss of pericytes and thickening of the basement membrane are hallmarks of diabetic retinopathy. We reported reduced adhesion, but no changes in apoptosis, of bovine retinal pericytes cultured on extracellular matrix (ECM) produced by endothelial cells in high glucose (HG). Since human and bovine pericytes may behave differently in conditions mimicking the diabetic milieu, we verified the behaviour of human retinal pericytes cultured on HG-conditioned ECM. Pericytes were cultured in physiological/HG on ECM produced by human umbilical vein endothelial cells in physiological/HG, alone or in the presence of thiamine and benfotiamine. Adhesion, proliferation, apoptosis, p53 and Bcl-2/Bax ratio (mRNA levels and protein concentrations) were measured in wild-type and immortalized human pericytes. Both types of pericytes adhered less to HG-conditioned ECM and plastic than to physiological glucose-conditioned ECM. DNA synthesis was impaired in pericytes cultured in HG on the three different surfaces but there were no differences in proliferation. DNA fragmentation and Bcl-2/Bax ratio were greatly enhanced by HG-conditioned ECM in pericytes kept in both physiological and HG. Addition of thiamine and benfotiamine to HG during ECM production completely prevented these damaging effects. Apoptosis is strongly increased in pericytes cultured on ECM produced by endothelium in HG, probably due to impairment of the Bcl-2/Bax ratio. Thiamine and benfotiamine completely revert this effect. This behaviour is therefore completely different from that of bovine pericytes, underlining the importance of establishing species-specific cell models to study the mechanisms of diabetic retinopathy. (c) 2009 John Wiley & Sons, Ltd.
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Directory of Open Access Journals (Sweden)
Nuria Huertas-González
2015-01-01
Full Text Available Background. Wernicke’s encephalopathy (WE is an acute neurological disorder resulting from thiamine deficiency. It is mainly related to alcohol abuse but it can be associated with other conditions such as gastrointestinal disorders. This vitamin deficiency can also present with cardiovascular symptoms, called “wet beriberi.” Association with folate deficit worsens the clinical picture. Subject. A 70-year-old man with gastric phytobezoar presented with gait instability, dyspnoea, chest pain associated with right heart failure and pericarditis, and folate deficiency. Furosemide was administered and cardiac symptoms improved but he soon developed vertiginous syndrome, nystagmus, diplopia, dysmetria, and sensitive and motor deficit in all four limbs with areflexia. Results. A cerebral magnetic resonance imaging (MRI showed typical findings of WE. He was immediately treated with thiamine. Neurological symptoms improved in a few days and abnormal signals disappeared in a follow-up MRI two weeks later. Conclusion. Patients with malabsorption due to gastrointestinal disorders have an increased risk of thiamine deficiency, and folate deficiency can make this vitamin malabsorption worse. An established deficiency mainly shows neurological symptoms, WE, or rarely cardiovascular symptoms, wet beriberi. Early vitamin treatment in symptomatic patients improves prognosis. We recommend administration of prophylactic multivitamins supplements in patients at risk as routine clinical practice.
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Functional analysis of aromatic biosynthetic pathways in Pseudomonas putida KT2440
Molina‐Henares, M. Antonia; García‐Salamanca, Adela; Molina‐Henares, A. Jesús; De La Torre, Jesús; Herrera, M. Carmen; Ramos, Juan L.; Duque, Estrella
2009-01-01
Summary Pseudomonas putida KT2440 is a non‐pathogenic prototrophic bacterium with high potential for biotechnological applications. Despite all that is known about this strain, the biosynthesis of essential chemicals has not been fully analysed and auxotroph mutants are scarce. We carried out massive mini‐Tn5 random mutagenesis and screened for auxotrophs that require aromatic amino acids. The biosynthesis of aromatic amino acids was analysed in detail including physical and transcriptional organization of genes, complementation assays and feeding experiments to establish pathway intermediates. There is a single pathway from chorismate leading to the biosynthesis of tryptophan, whereas the biosynthesis of phenylalanine and tyrosine is achieved through multiple convergent pathways. Genes for tryptophan biosynthesis are grouped in unlinked regions with the trpBA and trpGDE genes organized as operons and the trpI, trpE and trpF genes organized as single transcriptional units. The pheA and tyrA gene‐encoding multifunctional enzymes for phenylalanine and tyrosine biosynthesis are linked in the chromosome and form an operon with the serC gene involved in serine biosynthesis. The last step in the biosynthesis of these two amino acids requires an amino transferase activity for which multiple tyrB‐like genes are present in the host chromosome. PMID:21261884
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Metformin Is a Substrate and Inhibitor of the Human Thiamine Transporter, THTR-2 (SLC19A3).
Liang, Xiaomin; Chien, Huan-Chieh; Yee, Sook Wah; Giacomini, Marilyn M; Chen, Eugene C; Piao, Meiling; Hao, Jia; Twelves, Jolyn; Lepist, Eve-Irene; Ray, Adrian S; Giacomini, Kathleen M
2015-12-07
The biguanide metformin is widely used as first-line therapy for the treatment of type 2 diabetes. Predominately a cation at physiological pH's, metformin is transported by membrane transporters, which play major roles in its absorption and disposition. Recently, our laboratory demonstrated that organic cation transporter 1, OCT1, the major hepatic uptake transporter for metformin, was also the primary hepatic uptake transporter for thiamine, vitamin B1. In this study, we tested the reverse, i.e., that metformin is a substrate of thiamine transporters (THTR-1, SLC19A2, and THTR-2, SLC19A3). Our study demonstrated that human THTR-2 (hTHTR-2), SLC19A3, which is highly expressed in the small intestine, but not hTHTR-1, transports metformin (Km = 1.15 ± 0.2 mM) and other cationic compounds (MPP(+) and famotidine). The uptake mechanism for hTHTR-2 was pH and electrochemical gradient sensitive. Furthermore, metformin as well as other drugs including phenformin, chloroquine, verapamil, famotidine, and amprolium inhibited hTHTR-2 mediated uptake of both thiamine and metformin. Species differences in the substrate specificity of THTR-2 between human and mouse orthologues were observed. Taken together, our data suggest that hTHTR-2 may play a role in the intestinal absorption and tissue distribution of metformin and other organic cations and that the transporter may be a target for drug-drug and drug-nutrient interactions.
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Chapter 7. Cloning and analysis of natural product pathways.
Gust, Bertolt
2009-01-01
The identification of gene clusters of natural products has lead to an enormous wealth of information about their biosynthesis and its regulation, and about self-resistance mechanisms. Well-established routine techniques are now available for the cloning and sequencing of gene clusters. The subsequent functional analysis of the complex biosynthetic machinery requires efficient genetic tools for manipulation. Until recently, techniques for the introduction of defined changes into Streptomyces chromosomes were very time-consuming. In particular, manipulation of large DNA fragments has been challenging due to the absence of suitable restriction sites for restriction- and ligation-based techniques. The homologous recombination approach called recombineering (referred to as Red/ET-mediated recombination in this chapter) has greatly facilitated targeted genetic modifications of complex biosynthetic pathways from actinomycetes by eliminating many of the time-consuming and labor-intensive steps. This chapter describes techniques for the cloning and identification of biosynthetic gene clusters, for the generation of gene replacements within such clusters, for the construction of integrative library clones and their expression in heterologous hosts, and for the assembly of entire biosynthetic gene clusters from the inserts of individual library clones. A systematic approach toward insertional mutation of a complete Streptomyces genome is shown by the use of an in vitro transposon mutagenesis procedure.
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Biosynthetic origin of the isoprene units in chromenes of Piper aduncum (Piperaceae)
Energy Technology Data Exchange (ETDEWEB)
Leite, Ana C.; Lopes, Adriana A.; Bolzani, Vanderlan da S.; Furlan, Maysa [UNESP, Araraquara, SP (Brazil). Inst. de Quimica]. E-mail: maysaf@iq.unesp.br; Kato, Massuo J. [Universidade de Sao Paulo (USP), SP (Brazil). Inst. de Quimica
2007-07-01
Metabolic studies involving the incorporation of [1-{sup 13}C]-D-glucose into intact leaves of Piper aduncum (Piperaceae) have indicated that both the mevalonate (MVA) and the pyruvate-triose (MEP) non-mevalonate pathways are implicated in the biosynthesis of isoprene moieties present in methyl 2,2-dimethyl-2H-1-chromene-6-carboxylate (1) and methyl 2,2-dimethyl-8-(3'-methyl- 2'-butenyl)-2H-1-chromene-6-carboxylate (2). The pattern of incorporation of label from [1- {sup 13}C]-D-glucose into these chromenes was determined by quantitative {sup 13}C NMR spectroscopy. The results confirmed that biosynthetic compartment of 1 and 2 could either be the plastid and/ or the cytosol or, possibly, an additional compartment such as the plastid inter-membrane space. (author)
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Rodriguez, Alberto; Martínez, Juan A; Millard, Pierre; Gosset, Guillermo; Portais, Jean-Charles; Létisse, Fabien; Bolivar, Francisco
2017-06-01
Metabolic engineering strategies applied over the last two decades to produce shikimate (SA) in Escherichia coli have resulted in a battery of strains bearing many expression systems. However, the effects that these systems have on the host physiology and how they impact the production of SA are still not well understood. In this work we utilized an engineered E. coli strain to determine the consequences of carrying a vector that promotes SA production from glucose with a high-yield but that is also expected to impose a significant cellular burden. Kinetic comparisons in fermentors showed that instead of exerting a negative effect, the sole presence of the plasmid increased glucose consumption without diminishing the growth rate. By constitutively expressing a biosynthetic operon from this vector, the more active glycolytic metabolism was exploited to redirect intermediates toward the production of SA, which further increased the glucose consumption rate and avoided excess acetate production. Fluxomics and metabolomics experiments revealed a global remodeling of the carbon and energy metabolism in the production strain, where the increased SA production reduced the carbon available for oxidative and fermentative pathways. Moreover, the results showed that the production of SA relies on a specific setup of the pentose phosphate pathway, where both its oxidative and non-oxidative branches are strongly activated to supply erythrose-4-phosphate and balance the NADPH requirements. This work improves our understanding of the metabolic reorganization observed in E. coli in response to the plasmid-based expression of the SA biosynthetic pathway. Biotechnol. Bioeng. 2017;114: 1319-1330. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.
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DEFF Research Database (Denmark)
Whitfield, Kyly C; Smith, Geoffry; Chamnan, Chhoun
2017-01-01
the thiamine status (measured as erythrocyte thiamine diphosphate concentrations; eThDP) among a representative sample of Cambodian women of childbearing age (15-49 y) and their young children (6-69 mo). METHODOLOGY/PRINCIPLE FINDINGS: Samples for this cross-sectional analysis were collected as part...... of a national micronutrient survey linked to the Cambodian Demographic and Health Survey (CDHS) 2014. One-sixth of households taking part in the CDHS were randomly selected and re-visited for additional blood sampling for eThDP analysis (719 women and 761 children). Thiamine status was assessed using different...... cut-offs from literature. Women were mean (SD) 30 (6) y, and children (46% girls) were 41 (17) mo. Women had lower mean (95% CI) eThDP of 150 nmol/L (146-153) compared to children, 174 nmol/L (171-179; P
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Energy Technology Data Exchange (ETDEWEB)
Chakraborty, Saumen; Reed, Julian; Sage, Timothy; Branagan, Nicole C.; Petrik, Igor D.; Miner, Kyle D.; Hu, Michael Y.; Zhao, Jiyong; Alp, E. Ercan; Lu, Yi
2015-10-05
This Forum Article focuses on recent advances in structural and spectroscopic studies of biosynthetic models of nitric oxide reductases (NORs). NORs are complex metalloenzymes found in the denitrification pathway of Earth's nitrogen cycle where they catalyze the proton-dependent twoelectron reduction of nitric oxide (NO) to nitrous oxide (N2O). While much progress has been made in biochemical and biophysical studies of native NORs and their variants, a. clear mechanistic understanding of this important metalloenzyme related to its function is still elusive. We report herein UV vis and nuclear resonance vibrational spectroscopy (NRVS) studies of mononitrosylated intermediates of the NOR reaction of a biosynthetic model. The ability to selectively substitute metals at either heme or nonheme metal sites allows the introduction of independent 57Fe probe atoms at either site, as well as allowing the preparation of analogues of stable reaction intermediates by replacing either metal with a redox inactive metal. Together with previous structural and spectroscopic results, we summarize insights gained from studying these biosynthetic models toward understanding structural features responsible for the NOR activity and its mechanism. As a result, the outlook on NOR modeling is also discussed, with an emphasis on the design of models capable of catalytic turnovers designed based on close mimics of the secondary coordination sphere of native NORs.
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Xie, Zhengzhi; Ma, Xiaoqiang; Gang, David R
2009-01-01
Turmeric is an excellent example of a plant that produces large numbers of metabolites from diverse metabolic pathways or networks. It is hypothesized that these metabolic pathways or networks contain biosynthetic modules, which lead to the formation of metabolite modules-groups of metabolites whose production is co-regulated and biosynthetically linked. To test whether such co-regulated metabolite modules do exist in this plant, metabolic profiling analysis was performed on turmeric rhizome samples that were collected from 16 different growth and development treatments, which had significant impacts on the levels of 249 volatile and non-volatile metabolites that were detected. Importantly, one of the many co-regulated metabolite modules that were indeed readily detected in this analysis contained the three major curcuminoids, whereas many other structurally related diarylheptanoids belonged to separate metabolite modules, as did groups of terpenoids. The existence of these co-regulated metabolite modules supported the hypothesis that the 3-methoxyl groups on the aromatic rings of the curcuminoids are formed before the formation of the heptanoid backbone during the biosynthesis of curcumin and also suggested the involvement of multiple polyketide synthases with different substrate selectivities in the formation of the array of diarylheptanoids detected in turmeric. Similar conclusions about terpenoid biosynthesis could also be made. Thus, discovery and analysis of metabolite modules can be a powerful predictive tool in efforts to understand metabolism in plants.
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Pan, Ya-Jie; Liu, Jia; Guo, Xiao-Rui; Zu, Yuan-Gang; Tang, Zhong-Hua
2015-05-01
Research on transcriptional regulation of terpenoid indole alkaloid (TIA) biosynthesis of the medicinal plant, Catharanthus roseus, has largely been focused on gene function and not clustering analysis of multiple genes at the transcript level. Here, more than ten key genes encoding key enzyme of alkaloid synthesis in TIA biosynthetic pathways were chosen to investigate the integrative responses to exogenous elicitor ethylene and copper (Cu) at both transcriptional and metabolic levels. The ethylene-induced gene transcripts in leaves and roots, respectively, were subjected to principal component analysis (PCA) and the results showed the overall expression of TIA pathway genes indicated as the Q value followed a standard normal distribution after ethylene treatments. Peak gene expression was at 15-30 μM of ethephon, and the pre-mature leaf had a higher Q value than the immature or mature leaf and root. Treatment with elicitor Cu found that Cu up-regulated overall TIA gene expression more in roots than in leaves. The combined effects of Cu and ethephon on TIA gene expression were stronger than their separate effects. It has been documented that TIA gene expression is tightly regulated by the transcriptional factor (TF) ethylene responsive factor (ERF) and mitogen-activated protein kinase (MAPK) cascade. The loading plot combination with correlation analysis for the genes of C. roseus showed that expression of the MPK gene correlated with strictosidine synthase (STR) and strictosidine b-D-glucosidase(SGD). In addition, ERF expression correlated with expression of secologanin synthase (SLS) and tryptophan decarboxylase (TDC), specifically in roots, whereas MPK and myelocytomatosis oncogene (MYC) correlated with STR and SGD genes. In conclusion, the ERF regulates the upstream pathway genes in response to heavy metal Cu mainly in C. roseus roots, while the MPK mainly participates in regulating the STR gene in response to ethylene in pre-mature leaf. Interestingly, the
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Responses of Synechocystis sp. PCC 6803 to heterologous biosynthetic pathways
DEFF Research Database (Denmark)
Vavitsas, Konstantinos; Rue, Emil Østergaard; Stefánsdóttir, Lára Kristín
2017-01-01
BACKGROUND: There are an increasing number of studies regarding genetic manipulation of cyanobacteria to produce commercially interesting compounds. The majority of these works study the expression and optimization of a selected heterologous pathway, largely ignoring the wholeness and complexity...... different compounds, the cyanogenic glucoside dhurrin and the diterpenoid 13R-manoyl oxide in Synechocystis PCC 6803. We used genome-scale metabolic modelling to study fluxes in individual reactions and pathways, and we determined the concentrations of key metabolites, such as amino acids, carotenoids...
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Structure, function and regulation of the enzymes in the starch biosynthetic pathway.
Energy Technology Data Exchange (ETDEWEB)
Geiger, Jim
2013-11-30
structure of ADP- Glucose pyrophosphorylase from potato in its inhibited conformation, and bound to both ATP and ADP-glucose. In addition, we have determined the first structure of glycogen synthase in its "closed", catalytically active conformation bound to ADP-glucose. We also determined the structure of glycogen synthase bound to malto-oligosaccharides, showing for the first time that an enzyme in the starch biosynthetic pathway recognizes glucans not just in its active site but on binding sites on the surface of the enzyme ten’s of Angstroms from the active site. In addition our structure of a glycogen branching enzyme bound to malto-oligosaccharides identified seven distinct binding sites distributed about the surface of the enzyme. We will now determine the function of these sites to get a molecular-level picture of exactly how these enzymes interact with their polymeric substrates and confer specificity leading to the complex structure of the starch granule. We will extend our studies to other isoforms of the enzymes, to understand how their structures give rise to their distinct function. Our goal is to understand what accounts for the various functional differences between SS and SBE isoforms at a molecular level.
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Determining Antifungal Target Sites in the Sterol Pathway of the Yeast Candida and Saccharomyces
National Research Council Canada - National Science Library
Bard, Martin
1998-01-01
... as in topical infections which lead to significant losses in work-place productivity. The work reported here seeks to identify new target sites in the sterol biosynthetic pathway against which new antifungal compounds might be developed...
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Protein design for pathway engineering.
Eriksen, Dawn T; Lian, Jiazhang; Zhao, Huimin
2014-02-01
Design and construction of biochemical pathways has increased the complexity of biosynthetically-produced compounds when compared to single enzyme biocatalysis. However, the coordination of multiple enzymes can introduce a complicated set of obstacles to overcome in order to achieve a high titer and yield of the desired compound. Metabolic engineering has made great strides in developing tools to optimize the flux through a target pathway, but the inherent characteristics of a particular enzyme within the pathway can still limit the productivity. Thus, judicious protein design is critical for metabolic and pathway engineering. This review will describe various strategies and examples of applying protein design to pathway engineering to optimize the flux through the pathway. The proteins can be engineered for altered substrate specificity/selectivity, increased catalytic activity, reduced mass transfer limitations through specific protein localization, and reduced substrate/product inhibition. Protein engineering can also be expanded to design biosensors to enable high through-put screening and to customize cell signaling networks. These strategies have successfully engineered pathways for significantly increased productivity of the desired product or in the production of novel compounds. Copyright © 2013 Elsevier Inc. All rights reserved.
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Natural Product Biosynthetic Diversity and Comparative Genomics of the Cyanobacteria.
Dittmann, Elke; Gugger, Muriel; Sivonen, Kaarina; Fewer, David P
2015-10-01
Cyanobacteria are an ancient lineage of slow-growing photosynthetic bacteria and a prolific source of natural products with intricate chemical structures and potent biological activities. The bulk of these natural products are known from just a handful of genera. Recent efforts have elucidated the mechanisms underpinning the biosynthesis of a diverse array of natural products from cyanobacteria. Many of the biosynthetic mechanisms are unique to cyanobacteria or rarely described from other organisms. Advances in genome sequence technology have precipitated a deluge of genome sequences for cyanobacteria. This makes it possible to link known natural products to biosynthetic gene clusters but also accelerates the discovery of new natural products through genome mining. These studies demonstrate that cyanobacteria encode a huge variety of cryptic gene clusters for the production of natural products, and the known chemical diversity is likely to be just a fraction of the true biosynthetic capabilities of this fascinating and ancient group of organisms. Copyright © 2015. Published by Elsevier Ltd.
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Key roles of Arf small G proteins and biosynthetic trafficking for animal development.
Rodrigues, Francisco F; Harris, Tony J C
2017-04-14
Although biosynthetic trafficking can function constitutively, it also functions specifically for certain developmental processes. These processes require either a large increase to biosynthesis or the biosynthesis and targeted trafficking of specific players. We review the conserved molecular mechanisms that direct biosynthetic trafficking, and discuss how their genetic disruption affects animal development. Specifically, we consider Arf small G proteins, such as Arf1 and Sar1, and their coat effectors, COPI and COPII, and how these proteins promote biosynthetic trafficking for cleavage of the Drosophila embryo, the growth of neuronal dendrites and synapses, extracellular matrix secretion for bone development, lumen development in epithelial tubes, notochord and neural tube development, and ciliogenesis. Specific need for the biosynthetic trafficking system is also evident from conserved CrebA/Creb3-like transcription factors increasing the expression of secretory machinery during several of these developmental processes. Moreover, dysfunctional trafficking leads to a range of developmental syndromes.
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Wang, Kui-Lin; Bolitho, Karen; Grafton, Karryn; Kortstee, A.J.; Karunairetnam, Sakuntala; McGhie, T.K.; Espley, R.V.; Hellens, R.P.; Allan, A.C.
2010-01-01
Background - The control of plant anthocyanin accumulation is via transcriptional regulation of the genes encoding the biosynthetic enzymes. A key activator appears to be an R2R3 MYB transcription factor. In apple fruit, skin anthocyanin levels are controlled by a gene called MYBA or MYB1, while the gene determining fruit flesh and foliage anthocyanin has been termed MYB10. In order to further understand tissue-specific anthocyanin regulation we have isolated orthologous MYB genes from all th...
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DEFF Research Database (Denmark)
Anyaogu, Diana Chinyere
in industrial applications for the productionof these bioactive compounds and other chemicals as well as for enzyme production. Especially Aspergillusniger and Aspergillus oryzae are used as industrial workhorses for the production of various enzymes. Manyof the secreted proteins are glycosylated, indicating...... aspharmaceuticals. Access to this unexploited reservoir is hampered as many of the clusters are silent orbarely expressed under laboratory conditions. Methods for activating these pathways are thereforeessential for pathway discovery and elucidation. Filamentous fungi and Aspergillus species in particular are used...... that glycosylation plays an important role in thesecretory pathway. Thus, understanding the role and process of glycosylation will enable directedglycoengineering in Aspergilli to improve protein production and expand the repertoire of proteins, whichcan be produced by these fungi. Aspergillus nidulans has been used...
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Yu, Dayu; Xu, Fuchao; Valiente, Jonathan; Wang, Siyuan; Zhan, Jixun
2013-01-01
A putative indigoidine biosynthetic gene cluster was located in the genome of Streptomyces chromofuscus ATCC 49982. The silent 9.4-kb gene cluster consists of five open reading frames, named orf1, Sc-indC, Sc-indA, Sc-indB, and orf2, respectively. Sc-IndC was functionally characterized as an indigoidine synthase through heterologous expression of the enzyme in both Streptomyces coelicolor CH999 and Escherichia coli BAP1. The yield of indigoidine in E. coli BAP1 reached 2.78 g/l under the optimized conditions. The predicted protein product of Sc-indB is unusual and much larger than any other reported IndB-like protein. The N-terminal portion of this enzyme resembles IdgB and the C-terminal portion is a hypothetical protein. Sc-IndA and/or Sc-IndB were co-expressed with Sc-IndC in E. coli BAP1, which demonstrated the involvement of Sc-IndB, but not Sc-IndA, in the biosynthetic pathway of indigoidine. The yield of indigoidine was dramatically increased by 41.4 % (3.93 g/l) when Sc-IndB was co-expressed with Sc-IndC in E. coli BAP1. Indigoidine is more stable at low temperatures.
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Giant virus Megavirus chilensis encodes the biosynthetic pathway for uncommon acetamido sugars.
Piacente, Francesco; De Castro, Cristina; Jeudy, Sandra; Molinaro, Antonio; Salis, Annalisa; Damonte, Gianluca; Bernardi, Cinzia; Abergel, Chantal; Tonetti, Michela G
2014-08-29
Giant viruses mimicking microbes, by the sizes of their particles and the heavily glycosylated fibrils surrounding their capsids, infect Acanthamoeba sp., which are ubiquitous unicellular eukaryotes. The glycans on fibrils are produced by virally encoded enzymes, organized in gene clusters. Like Mimivirus, Megavirus glycans are mainly composed of virally synthesized N-acetylglucosamine (GlcNAc). They also contain N-acetylrhamnosamine (RhaNAc), a rare sugar; the enzymes involved in its synthesis are encoded by a gene cluster specific to Megavirus close relatives. We combined activity assays on two enzymes of the pathway with mass spectrometry and NMR studies to characterize their specificities. Mg534 is a 4,6-dehydratase 5-epimerase; its three-dimensional structure suggests that it belongs to a third subfamily of inverting dehydratases. Mg535, next in the pathway, is a bifunctional 3-epimerase 4-reductase. The sequential activity of the two enzymes leads to the formation of UDP-l-RhaNAc. This study is another example of giant viruses performing their glycan synthesis using enzymes different from their cellular counterparts, raising again the question of the origin of these pathways. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.
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Anholt, H; Himsworth, C; Britton, A
2016-07-01
Two 4-year-old spayed female Siamese cats were seized by the British Columbia Society for the Prevention of Cruelty to Animals after confinement to an abandoned housing unit without food for 9 weeks. One cat was found dead, and the second was euthanized within 24 hours due to neurologic deterioration despite therapy. Polioencephalomalacia of the caudal colliculus, hepatic lipidosis, cachexia, and congestive heart failure with cardiomyocyte atrophy were identified in both cats through postmortem examination and attributed to a prolonged period of starvation. Brain lesions were likely the result of thiamine deficiency (Chastek paralysis), which can be associated with both malnutrition and liver disease. This case highlights the importance of thiamine supplementation during realimentation of cats with hepatic lipidosis. Heart failure resulting from cachexia may have contributed to the death of the first cat and the morbidity of the second cat. © The Author(s) 2016.
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Martinez-Villaluenga, C; Michalska, A; Frias, J; Piskula, M K; Vidal-Valverde, C; Zieliński, H
2009-01-01
The effect of rye flour extraction rates and baking on thiamine and riboflavin content, and antioxidant capacity of traditional rye bread were studied and compared with white wheat flour. The content of thiamine was higher (10.9%) in rye dough formulated with dark rye flour (F-100%; extraction rate of 100%) than in rye dough formulated with brown rye flour (F-92%; extraction rate of 92%) that was similar to dough made with wheat flour. The riboflavin content in rye dough made from flour F-100% was also higher (16%) than in dough formulated with flour F-92%, and both provided larger riboflavin content than wheat dough. Baking led to reductions in thiamine of 56% for wheat bread and of 20% for both rye breads; however, this process caused only a 10% decrease in riboflavin for wheat bread and a 30% decrease for rye breads. Trolox equivalent antioxidant capacity, peroxyl radical scavenging capacity, DPPH radical scavenging activity, and Folin-Ciocalteu reducing capacity were higher in rye than in wheat dough and bread. Baking process produced slight changes in antioxidant activity, except for Superoxide Dismutase-like activity where a sharp decrease was observed. Our findings showed that rye breads are an important source of B vitamins and rye breads formulated with dark and brown flours showed better antioxidant properties than wheat bread. Therefore, rye breads should be more widely recommended in human nutrition.
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Zahr, Natalie M; Sullivan, Edith V; Rohlfing, Torsten; Mayer, Dirk; Collins, Amy M; Luong, Richard; Pfefferbaum, Adolf
2016-07-01
Serious neurological concomitants of alcoholism include Wernicke's encephalopathy (WE), Korsakoff's syndrome (KS), and hepatic encephalopathy (HE). This study was conducted in animal models to determine neuroradiological signatures associated with liver damage caused by carbon tetrachloride (CCl4), thiamine deficiency caused by pyrithiamine treatment, and nonspecific nutritional deficiency caused by food deprivation. Magnetic resonance imaging (MRI) and magnetic resonance spectroscopy (MRS) were used to evaluate brains of wild-type Wistar rats at baseline and following treatment. Similar to observations in ethanol (EtOH) exposure models, thiamine deficiency caused enlargement of the lateral ventricles. Liver damage was not associated with effects on cerebrospinal fluid volumes, whereas food deprivation caused modest enlargement of the cisterns. In contrast to what has repeatedly been shown in EtOH exposure models, in which levels of choline-containing compounds (Cho) measured by MRS are elevated, Cho levels in treated animals in all three experiments (i.e., liver damage, thiamine deficiency, and food deprivation) were lower than those in baseline or controls. These results add to the growing body of literature suggesting that MRS-detectable Cho is labile and can depend on a number of variables that are not often considered in human experiments. These results also suggest that reductions in Cho observed in humans with alcohol use disorder (AUD) may well be due to mild manifestations of concomitants of AUD such as liver damage or nutritional deficiencies and not necessarily to alcohol consumption per se.
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Liseron-Monfils, Christophe; Lewis, Tim; Ashlock, Daniel; McNicholas, Paul D; Fauteux, François; Strömvik, Martina; Raizada, Manish N
2013-03-15
The discovery of genetic networks and cis-acting DNA motifs underlying their regulation is a major objective of transcriptome studies. The recent release of the maize genome (Zea mays L.) has facilitated in silico searches for regulatory motifs. Several algorithms exist to predict cis-acting elements, but none have been adapted for maize. A benchmark data set was used to evaluate the accuracy of three motif discovery programs: BioProspector, Weeder and MEME. Analysis showed that each motif discovery tool had limited accuracy and appeared to retrieve a distinct set of motifs. Therefore, using the benchmark, statistical filters were optimized to reduce the false discovery ratio, and then remaining motifs from all programs were combined to improve motif prediction. These principles were integrated into a user-friendly pipeline for motif discovery in maize called Promzea, available at http://www.promzea.org and on the Discovery Environment of the iPlant Collaborative website. Promzea was subsequently expanded to include rice and Arabidopsis. Within Promzea, a user enters cDNA sequences or gene IDs; corresponding upstream sequences are retrieved from the maize genome. Predicted motifs are filtered, combined and ranked. Promzea searches the chosen plant genome for genes containing each candidate motif, providing the user with the gene list and corresponding gene annotations. Promzea was validated in silico using a benchmark data set: the Promzea pipeline showed a 22% increase in nucleotide sensitivity compared to the best standalone program tool, Weeder, with equivalent nucleotide specificity. Promzea was also validated by its ability to retrieve the experimentally defined binding sites of transcription factors that regulate the maize anthocyanin and phlobaphene biosynthetic pathways. Promzea predicted additional promoter motifs, and genome-wide motif searches by Promzea identified 127 non-anthocyanin/phlobaphene genes that each contained all five predicted promoter
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Directory of Open Access Journals (Sweden)
Sarah J Fletcher
Full Text Available Tight junctions (TJs link adjacent cells and are critical for maintenance of apical-basolateral polarity in epithelial monolayers. The TJ protein occludin functions in disparate processes, including wound healing and Hepatitis C Virus infection. Little is known about steady-state occludin trafficking into and out of the plasma membrane. Therefore, we determined the mechanisms responsible for occludin turnover in confluent Madin-Darby canine kidney (MDCK epithelial monolayers. Using various biotin-based trafficking assays we observed continuous and rapid endocytosis of plasma membrane localised occludin (the majority internalised within 30 minutes. By 120 minutes a significant reduction in internalised occludin was observed. Inhibition of lysosomal function attenuated the reduction in occludin signal post-endocytosis and promoted co-localisation with the late endocytic system. Using a similar method we demonstrated that ∼20% of internalised occludin was transported back to the cell surface. Consistent with these findings, significant co-localisation between internalised occludin and recycling endosomal compartments was observed. We then quantified the extent to which occludin synthesis and transport to the plasma membrane contributes to plasma membrane occludin homeostasis, identifying inhibition of protein synthesis led to decreased plasma membrane localised occludin. Significant co-localisation between occludin and the biosynthetic secretory pathway was demonstrated. Thus, under steady-state conditions occludin undergoes turnover via a continuous cycle of endocytosis, recycling and degradation, with degradation compensated for by biosynthetic exocytic trafficking. We developed a mathematical model to describe the endocytosis, recycling and degradation of occludin, utilising experimental data to provide quantitative estimates for the rates of these processes.
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Directory of Open Access Journals (Sweden)
Audray Dugrand-Judek
Full Text Available Citrus plants are able to produce defense compounds such as coumarins and furanocoumarins to cope with herbivorous insects and pathogens. In humans, these chemical compounds are strong photosensitizers and can interact with medications, leading to the "grapefruit juice effect". Removing coumarins and furanocoumarins from food and cosmetics imply additional costs and might alter product quality. Thus, the selection of Citrus cultivars displaying low coumarin and furanocoumarin contents constitutes a valuable alternative. In this study, we performed ultra-performance liquid chromatography coupled with mass spectrometry analyses to determine the contents of these compounds within the peel and the pulp of 61 Citrus species representative of the genetic diversity all Citrus. Generally, Citrus peel contains larger diversity and higher concentrations of coumarin/furanocoumarin than the pulp of the same fruits. According to the chemotypes found in the peel, Citrus species can be separated into 4 groups that correspond to the 4 ancestral taxa (pummelos, mandarins, citrons and papedas and extended with their respective secondary species descendants. Three of the 4 ancestral taxa (pummelos, citrons and papedas synthesize high amounts of these compounds, whereas mandarins appear practically devoid of them. Additionally, all ancestral taxa and their hybrids are logically organized according to the coumarin and furanocoumarin pathways described in the literature. This organization allows hypotheses to be drawn regarding the biosynthetic origin of compounds for which the biogenesis remains unresolved. Determining coumarin and furanocoumarin contents is also helpful for hypothesizing the origin of Citrus species for which the phylogeny is presently not firmly established. Finally, this work also notes favorable hybridization schemes that will lead to low coumarin and furanocoumarin contents, and we propose to select mandarins and Ichang papeda as Citrus
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Horsman, Geoff P.; Chen, Yihua; Thorson, Jon S.; Shen, Ben
2010-01-01
Enediynes are potent antitumor antibiotics that are classified as 9- or 10-membered according to the size of the enediyne core structure. However, almost nothing is known about enediyne core biosynthesis, and the determinants of 9- versus 10-membered enediyne core biosynthetic divergence remain elusive. Previous work identified enediyne-specific polyketide synthases (PKSEs) that can be phylogenetically distinguished as being involved in 9- versus 10-membered enediyne biosynthesis, suggesting that biosynthetic divergence might originate from differing PKSE chemistries. Recent in vitro studies have identified several compounds produced by the PKSE and associated thioesterase (TE), but condition-dependent product profiles make it difficult to ascertain a true catalytic difference between 9- and 10-membered PKSE-TE systems. Here we report that PKSE chemistry does not direct 9- versus 10-membered enediyne core biosynthetic divergence as revealed by comparing the products from three 9-membered and two 10-membered PKSE-TE systems under identical conditions using robust in vivo assays. Three independent experiments support a common catalytic function for 9- and 10-membered PKSEs by the production of a heptaene metabolite from: (i) all five cognate PKSE-TE pairs in Escherichia coli; (ii) the C-1027 and calicheamicin cognate PKSE-TEs in Streptomyces lividans K4-114; and (iii) selected native producers of both 9- and 10-membered enediynes. Furthermore, PKSEs and TEs from different 9- and 10-membered enediyne biosynthetic machineries are freely interchangeable, revealing that 9- versus 10-membered enediyne core biosynthetic divergence occurs beyond the PKSE-TE level. These findings establish a starting point for determining the origins of this biosynthetic divergence. PMID:20534556
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International Nuclear Information System (INIS)
Moorthy, P.N.; Hayon, E.
1977-01-01
The technique of pulse radiolysis and kinetic absorption spectrophotometry was used to study the one-electron reduction of thiamin, thiazole, 4-aminopyrimidine, biotin, and pantothenic acid in aqueous solution. The acetone ketyl radical and e/sub aq/ - were used as the one-electron reducing agents. The reaction rate constants of e/sub aq/ - and (CH 3 ) 2 COH with these compounds were determined at different pH values, taking into account the dissociation constants of the substrates. The transient optical absorption spectra of the intermediates produced, their extinction coefficients, decay kinetics, and ionization constants were determined. One-electron reduction of the thiazolium ring of thiamin is suggested, based on the formation of dihydrothiamin as a final product. Other assignments for these radicals are suggested and discussed. The reaction of OH radicals with biotin and pantothenic acid leads, primarily, to H atom abstraction at various sites in the molecule. The formation and ionization of the -C(OH)CONH- radical from pantothenic acid, pK/sub a/ = 6.0 +- 0.3, is proposed
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Crosstalk of Autophagy and the Secretory Pathway and Its Role in Diseases.
Zahoor, Muhammad; Farhan, Hesso
2018-01-01
The secretory and autophagic pathways are two fundamental, evolutionary highly conserved endomembrane processes. Typically, secretion is associated with biosynthesis and delivery of proteins. In contrast, autophagy is usually considered as a degradative pathway. Thus, an analogy to metabolic pathways is evident. Anabolic (biosynthetic) and catabolic (degradative) pathways are usually intimately linked and intertwined, and likewise, the secretory and autophagy pathways are intertwined. Investigation of this link is an emerging area of research, and we will provide an overview of some of the major advances that have been made to contribute to understanding of how secretion regulates autophagy and vice versa. Finally, we will highlight evidence that supports a potential involvement of the autophagy-secretion crosstalk in human diseases. © 2018 Elsevier Inc. All rights reserved.
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Emergent biosynthetic capacity in simple microbial communities.
Directory of Open Access Journals (Sweden)
Hsuan-Chao Chiu
2014-07-01
Full Text Available Microbes have an astonishing capacity to transform their environments. Yet, the metabolic capacity of a single species is limited and the vast majority of microorganisms form complex communities and join forces to exhibit capabilities far exceeding those achieved by any single species. Such enhanced metabolic capacities represent a promising route to many medical, environmental, and industrial applications and call for the development of a predictive, systems-level understanding of synergistic microbial capacity. Here we present a comprehensive computational framework, integrating high-quality metabolic models of multiple species, temporal dynamics, and flux variability analysis, to study the metabolic capacity and dynamics of simple two-species microbial ecosystems. We specifically focus on detecting emergent biosynthetic capacity--instances in which a community growing on some medium produces and secretes metabolites that are not secreted by any member species when growing in isolation on that same medium. Using this framework to model a large collection of two-species communities on multiple media, we demonstrate that emergent biosynthetic capacity is highly prevalent. We identify commonly observed emergent metabolites and metabolic reprogramming patterns, characterizing typical mechanisms of emergent capacity. We further find that emergent secretion tends to occur in two waves, the first as soon as the two organisms are introduced, and the second when the medium is depleted and nutrients become limited. Finally, aiming to identify global community determinants of emergent capacity, we find a marked association between the level of emergent biosynthetic capacity and the functional/phylogenetic distance between community members. Specifically, we demonstrate a "Goldilocks" principle, where high levels of emergent capacity are observed when the species comprising the community are functionally neither too close, nor too distant. Taken together
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Evolutionary origins and functions of the carotenoid biosynthetic pathway in marine diatoms
Czech Academy of Sciences Publication Activity Database
Coesel, S.; Oborník, Miroslav; Varela, J.; Falciatore, A.; Bowler, C.
2008-01-01
Roč. 3, č. 8 (2008), s. 1-16 E-ISSN 1932-6203 R&D Projects: GA AV ČR IAA500220502 Institutional research plan: CEZ:AV0Z60220518 Keywords : marine diatoms * carotenoid pathway * evolution Subject RIV: EB - Genetics ; Molecular Biology
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In silico tools for the analysis of antibiotic biosynthetic pathways
DEFF Research Database (Denmark)
Weber, Tilmann
2014-01-01
Natural products of bacteria and fungi are the most important source for antimicrobial drug leads. For decades, such compounds were exclusively found by chemical/bioactivity-guided screening approaches. The rapid progress in sequencing technologies only recently allowed the development of novel...... screening methods based on the genome sequences of potential producing organisms. The basic principle of such genome mining approaches is to identify genes, which are involved in the biosynthesis of such molecules, and to predict the products of the identified pathways. Thus, bioinformatics methods...... and tools are crucial for genome mining. In this review, a comprehensive overview is given on programs and databases for the identification and analysis of antibiotic biosynthesis gene clusters in genomic data....
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Rodas-Junco, Beatriz A; Cab-Guillén, Yahaira; Muñoz-Sánchez, J Armando; Vázquez-Flota, Felipe; Monforte-González, Miriam; Hernández-Sotomayor, S M Teresa
2013-10-01
Signal transduction via phospholipids is mediated by phospholipases such as phospholipase C (PLC) and D (PLD), which catalyze hydrolysis of plasma membrane structural phospholipids. Phospholipid signaling is also involved in plant responses to phytohormones such as salicylic acid (SA). The relationships between phospholipid signaling, SA, and secondary metabolism are not fully understood. Using a Capsicum chinense cell suspension as a model, we evaluated whether phospholipid signaling modulates SA-induced vanillin production through the activation of phenylalanine ammonia lyase (PAL), a key enzyme in the biosynthetic pathway. Salicylic acid was found to elicit PAL activity and consequently vanillin production, which was diminished or reversed upon exposure to the phosphoinositide-phospholipase C (PI-PLC) signaling inhibitors neomycin and U73122. Exposure to the phosphatidic acid inhibitor 1-butanol altered PLD activity and prevented SA-induced vanillin production. Our results suggest that PLC and PLD-generated secondary messengers may be modulating SA-induced vanillin production through the activation of key biosynthetic pathway enzymes.
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Monjas, Leticia; Swier, Lotteke J Y M; Setyawati, Inda; Slotboom, Dirk Jan; Hirsch, Anna Katharina Herta
2017-01-01
We applied dynamic combinatorial chemistry (DCC) to identify ligands of ThiT, the S-component of the energy-coupling factor (ECF) transporter for thiamine in Lactococcus lactis. We used a pre-equilibrated dynamic combinatorial library (DCL) and saturation-transfer difference (STD) NMR spectroscopy
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On the biosynthetic origin of carminic acid
DEFF Research Database (Denmark)
Rasmussen, Silas A.; Kongstad, Kenneth T; Khorsand-Jamal, Paiman
2018-01-01
provides solid evidence of a polyketide, rather than a shikimate, origin of coccid pigments. Based on the newly identified compounds, we present a detailed biosynthetic scheme that accounts for the formation of carminic acid (CA) in D. coccus and all described coccid pigments which share a flavokermesic...... distribution suggests a common evolutionary origin for the trait in all coccid dye producing insect species....
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Chapter 3: Omics Advances of Biosynthetic Pathways of Isoprenoid Production in Microalgae
Energy Technology Data Exchange (ETDEWEB)
Paniagua-Michel, J.; Subramanian, Venkataramanan
2017-01-01
In this chapter, the current status of microalgal isoprenoids and the role of omics technologies, or otherwise specified, in bioproducts optimization and applications are reviewed. Emphasis is focused in the metabolic pathways of microalgae involved in the production of commercially important products, namely, hydrocarbons and biofuels, nutraceuticals, and pharmaceuticals.
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Directory of Open Access Journals (Sweden)
Muhammad Tariq Saeed
2016-09-01
Full Text Available The alteration of glucose metabolism, through increased uptake of glucose and glutamine addiction, is essential to cancer cell growth and invasion. Increased flux of glucose through the Hexosamine Biosynthetic Pathway (HBP drives increased cellular O-GlcNAcylation (hyper-O-GlcNAcylation and contributes to cancer progression by regulating key oncogenes. However, the association between hyper-O-GlcNAcylation and activation of these oncogenes remains poorly characterized. Here, we implement a qualitative modeling framework to analyze the role of the Biological Regulatory Network in HBP activation and its potential effects on key oncogenes. Experimental observations are encoded in a temporal language format and model checking is applied to infer the model parameters and qualitative model construction. Using this model, we discover step-wise genetic alterations that promote cancer development and invasion due to an increase in glycolytic flux, and reveal critical trajectories involved in cancer progression. We compute delay constraints to reveal important associations between the production and degradation rates of proteins. O-linked N-acetylglucosamine transferase (OGT, an enzyme used for addition of O-GlcNAc during O-GlcNAcylation, is identified as a key regulator to promote oncogenesis in a feedback mechanism through the stabilization of c-Myc. Silencing of the OGT and c-Myc loop decreases glycolytic flux and leads to programmed cell death. Results of network analyses also identify a significant cycle that highlights the role of p53-Mdm2 circuit oscillations in cancer recovery and homeostasis. Together, our findings suggest that the OGT and c-Myc feedback loop is critical in tumor progression, and targeting these mediators may provide a mechanism-based therapeutic approach to regulate hyper-O-GlcNAcylation in human cancer.
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Zhang, Lihan; Hoshino, Shotaro; Awakawa, Takayoshi; Wakimoto, Toshiyuki; Abe, Ikuro
2016-08-03
Natural products have enormous structural diversity, yet little is known about how such diversity is achieved in nature. Here we report the structural diversification of a cyanotoxin-lyngbyatoxin A-and its biosynthetic intermediates by heterologous expression of the Streptomyces-derived tleABC biosynthetic gene cluster in three different Streptomyces hosts: S. lividans, S. albus, and S. avermitilis. Notably, the isolated lyngbyatoxin derivatives, including four new natural products, were biosynthesized by crosstalk between the heterologous tleABC gene cluster and the endogenous host enzymes. The simple strategy described here has expanded the structural diversity of lyngbyatoxin A and its biosynthetic intermediates, and provides opportunities for investigation of the currently underestimated hidden biosynthetic crosstalk. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
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Fabrication of biosynthetic vascular prostheses by 193-nm excimer laser radiation
Husinsky, Wolfgang; Csek, Ch.; Bartel, A.; Grabenwoeger, M.; Fitzal, F.; Wolner, Ernst
1998-05-01
This study was undertaken to investigate the feasibility of transmural capillary ingrowth into the inner surface of biosynthetic vascular prostheses (OmniflowTM) through perforations created by an excimer-laser, thus inducing an endothelial cell coverage. The biosynthetic vascular prostheses (10 cm length, 6 mm (phi) ) were perforated with an excimer laser ((phi) of the holes 50 - 100 micrometer, distance 4 mm) and implanted into the carotid arteries of 8 sheep. The laser tissue interaction process of 193 nm radiation ensures minimal thermal damage to the prostheses. They were compared to untreated OmniflowTM prostheses implanted at the contralateral side. Three months after implantation the prostheses were explanted and evaluated by gross morphology, histological examination and scanning electron microscopy. Scanning electron microscopy showed endothelial cells in the midgraft portion of all perforated prostheses, whereas collagen fibers, fibrin meshwork and activated platelets formed the inner layer in 6 out of 8 untreated OmniflowTM prostheses. It can be concluded, that spontaneous endothelialization of biosynthetic vascular prostheses can be achieved by transmural capillary ingrowth through perforations in the wall of the prostheses in an experimental sheep model.
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Directory of Open Access Journals (Sweden)
Dian Xiao
2016-04-01
Full Text Available Ampakine compounds have been shown to reverse opiate-induced respiratory depression by activation of amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA glutamate receptors. However, their pharmacological exploitations are hindered by low blood-brain barrier (BBB permeability and limited brain distribution. Here, we explored whether thiamine disulfide prodrugs with the ability of “lock-in” can be used to solve these problems. A series of thiamine disulfide prodrugs 7a–7f of ampakine compound LCX001 was synthesized and evaluated. The trials in vitro showed that prodrugs 7e, 7d, 7f possessed a certain stability in plasma and quickly decomposed in brain homogenate by the disulfide reductase. In vivo, prodrug 7e decreased the peripheral distribution of LCX001 and significantly increased brain distribution of LCX001 after i.v. administration. This compound showed 2.23- and 3.29-fold greater increases in the AUC0-t and MRT0-t of LCX001 in brain, respectively, than did LCX001 itself. A preliminary pharmacodynamic study indicated that the required molar dose of prodrug 7e was only one eighth that of LCX001 required to achieve the same effect in mice. These findings provide an important reference to evaluate the clinical outlook of ampakine compounds.
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Köckerling, F; Alam, N N; Antoniou, S A; Daniels, I R; Famiglietti, F; Fortelny, R H; Heiss, M M; Kallinowski, F; Kyle-Leinhase, I; Mayer, F; Miserez, M; Montgomery, A; Morales-Conde, S; Muysoms, F; Narang, S K; Petter-Puchner, A; Reinpold, W; Scheuerlein, H; Smietanski, M; Stechemesser, B; Strey, C; Woeste, G; Smart, N J
2018-04-01
Although many surgeons have adopted the use of biologic and biosynthetic meshes in complex abdominal wall hernia repair, others have questioned the use of these products. Criticism is addressed in several review articles on the poor standard of studies reporting on the use of biologic meshes for different abdominal wall repairs. The aim of this consensus review is to conduct an evidence-based analysis of the efficacy of biologic and biosynthetic meshes in predefined clinical situations. A European working group, "BioMesh Study Group", composed of invited surgeons with a special interest in surgical meshes, formulated key questions, and forwarded them for processing in subgroups. In January 2016, a workshop was held in Berlin where the findings were presented, discussed, and voted on for consensus. Findings were set out in writing by the subgroups followed by consensus being reached. For the review, 114 studies and background analyses were used. The cumulative data regarding biologic mesh under contaminated conditions do not support the claim that it is better than synthetic mesh. Biologic mesh use should be avoided when bridging is needed. In inguinal hernia repair biologic and biosynthetic meshes do not have a clear advantage over the synthetic meshes. For prevention of incisional or parastomal hernias, there is no evidence to support the use of biologic/biosynthetic meshes. In complex abdominal wall hernia repairs (incarcerated hernia, parastomal hernia, infected mesh, open abdomen, enterocutaneous fistula, and component separation technique), biologic and biosynthetic meshes do not provide a superior alternative to synthetic meshes. The routine use of biologic and biosynthetic meshes cannot be recommended.
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Directory of Open Access Journals (Sweden)
Maciej Pronicki
2017-06-01
Full Text Available Biotin-thiamine-responsive basal ganglia disease is a severe form of a rare neurogenetic disorder caused by pathogenic molecular variants in the thiamine transporter gene. Nowadays, a potentially effective treatment is known, therefore the early diagnosis is mandatory. The aim of the paper was to assess the contribution of neuropathological and magnetic resonance imaging (MRI studies to a proper diagnosis. We present the brain study of two Polish patients with SLC19A3 mutations, including (1 an infant with an intriguing “walnut” appearance of the brain autopsied many years before the discovery of the SLC19A3 defect, and (2 a one-year-old patient with clinical features of Leigh syndrome. In patient 2, biotin/thiamine responsiveness was not tested at the time of diagnosis and causal treatment started with one-year delay. The central nervous system lesions found in the patients displayed almost clearly a specific pattern for SLC19A3 defect, as previously proposed in diagnostic criteria. Our study presents a detailed description of neuropathological and MRI findings of both patients. We confirm that the autopsy and/or MRI of the brain is sufficient to qualify a patient with an unknown neuropathological disorder directly for SLC19A3 mutations testing and a prompt trial of specific treatment.
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Integrating nitric oxide into salicylic acid and jasmonic acid/ ethylene plant defense pathways.
Mur, Luis A J; Prats, Elena; Pierre, Sandra; Hall, Michael A; Hebelstrup, Kim H
2013-01-01
Plant defense against pests and pathogens is known to be conferred by either salicylic acid (SA) or jasmonic acid (JA)/ethylene (ET) pathways, depending on infection or herbivore-grazing strategy. It is well attested that SA and JA/ET pathways are mutually antagonistic allowing defense responses to be tailored to particular biotic stresses. Nitric oxide (NO) has emerged as a major signal influencing resistance mediated by both signaling pathways but no attempt has been made to integrate NO into established SA/JA/ET interactions. NO has been shown to act as an inducer or suppressor of signaling along each pathway. NO will initiate SA biosynthesis and nitrosylate key cysteines on TGA-class transcription factors to aid in the initiation of SA-dependent gene expression. Against this, S-nitrosylation of NONEXPRESSOR OF PATHOGENESIS-RELATED PROTEINS1 (NPR1) will promote the NPR1 oligomerization within the cytoplasm to reduce TGA activation. In JA biosynthesis, NO will initiate the expression of JA biosynthetic enzymes, presumably to over-come any antagonistic effects of SA on JA-mediated transcription. NO will also initiate the expression of ET biosynthetic genes but a suppressive role is also observed in the S-nitrosylation and inhibition of S-adenosylmethionine transferases which provides methyl groups for ET production. Based on these data a model for NO action is proposed but we have also highlighted the need to understand when and how inductive and suppressive steps are used.
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Wang, Jian; Wu, Yifei; Sun, Xinxiao; Yuan, Qipeng; Yan, Yajun
2017-10-20
Microbial based bioplastics are promising alternatives to petroleum based synthetic plastics due to their renewability and economic feasibility. Glutarate is one of the most potential building blocks for bioplastics. The recent biosynthetic routes for glutarate were mostly based on the l-lysine degradation pathway from Pseudomonas putida that required lysine either by feeding or lysine overproduction via genetic manipulations. Herein, we established a novel glutarate biosynthetic pathway by incorporation of a "+1" carbon chain extension pathway from α-ketoglutarate (α-KG) in combination with α-keto acid decarboxylation pathway in Escherichia coli. Introduction of homocitrate synthase (HCS), homoaconitase (HA) and homoisocitrate dehydrogenase (HICDH) from Saccharomyces cerevisiae into E. coli enabled "+1" carbon extension from α-KG to α-ketoadipate (α-KA), which was subsequently converted into glutarate by a promiscuous α-keto acid decarboxylase (KivD) and a succinate semialdehyde dehydrogenase (GabD). The recombinant E. coli coexpressing all five genes produced 0.3 g/L glutarate from glucose. To further improve the titers, α-KG was rechanneled into carbon chain extension pathway via the clustered regularly interspersed palindromic repeats system mediated interference (CRISPRi) of essential genes sucA and sucB in tricarboxylic acid (TCA) cycle. The final strain could produce 0.42 g/L glutarate, which was increased by 40% compared with the parental strain.
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Insulin Biosynthetic Interaction Network Component, TMEM24, Facilitates Insulin Reserve Pool Release
Directory of Open Access Journals (Sweden)
Anita Pottekat
2013-09-01
Full Text Available Insulin homeostasis in pancreatic β cells is now recognized as a critical element in the progression of obesity and type II diabetes (T2D. Proteins that interact with insulin to direct its sequential synthesis, folding, trafficking, and packaging into reserve granules in order to manage release in response to elevated glucose remain largely unknown. Using a conformation-based approach combined with mass spectrometry, we have generated the insulin biosynthetic interaction network (insulin BIN, a proteomic roadmap in the β cell that describes the sequential interacting partners of insulin along the secretory axis. The insulin BIN revealed an abundant C2 domain-containing transmembrane protein 24 (TMEM24 that manages glucose-stimulated insulin secretion from a reserve pool of granules, a critical event impaired in patients with T2D. The identification of TMEM24 in the context of a comprehensive set of sequential insulin-binding partners provides a molecular description of the insulin secretory pathway in β cells.
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Lemetre, Christophe; Maniko, Jeffrey; Charlop-Powers, Zachary; Sparrow, Ben; Lowe, Andrew J; Brady, Sean F
2017-10-31
Although bacterial bioactive metabolites have been one of the most prolific sources of lead structures for the development of small-molecule therapeutics, very little is known about the environmental factors associated with changes in secondary metabolism across natural environments. Large-scale sequencing of environmental microbiomes has the potential to shed light on the richness of bacterial biosynthetic diversity hidden in the environment, how it varies from one environment to the next, and what environmental factors correlate with changes in biosynthetic diversity. In this study, the sequencing of PCR amplicons generated using primers targeting either ketosynthase domains from polyketide biosynthesis or adenylation domains from nonribosomal peptide biosynthesis was used to assess biosynthetic domain composition and richness in soils collected across the Australian continent. Using environmental variables collected at each soil site, we looked for environmental factors that correlated with either high overall domain richness or changes in the domain composition. Among the environmental variables we measured, changes in biosynthetic domain composition correlate most closely with changes in latitude and to a lesser extent changes in pH. Although it is unclear at this time the exact mix of factors that may drive the relationship between biosynthetic domain composition and latitude, from a practical perspective the identification of a latitudinal basis for differences in soil metagenome biosynthetic domain compositions should help guide future natural product discovery efforts. Published under the PNAS license.
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Overlapping riboflavin supply pathways in bacteria.
García-Angulo, Víctor Antonio
2017-03-01
Riboflavin derivatives are essential cofactors for a myriad of flavoproteins. In bacteria, flavins importance extends beyond their role as intracellular protein cofactors, as secreted flavins are a key metabolite in a variety of physiological processes. Bacteria obtain riboflavin through the endogenous riboflavin biosynthetic pathway (RBP) or by the use of importer proteins. Bacteria frequently encode multiple paralogs of the RBP enzymes and as for other micronutrient supply pathways, biosynthesis and uptake functions largely coexist. It is proposed that bacteria shut down biosynthesis and would rather uptake riboflavin when the vitamin is environmentally available. Recently, the overlap of riboflavin provisioning elements has gained attention and the functions of duplicated paralogs of RBP enzymes started to be addressed. Results point towards the existence of a modular structure in the bacterial riboflavin supply pathways. Such structure uses subsets of RBP genes to supply riboflavin for specific functions. Given the importance of riboflavin in intra and extracellular bacterial physiology, this complex array of riboflavin provision pathways may have developed to contend with the various riboflavin requirements. In riboflavin-prototrophic bacteria, riboflavin transporters could represent a module for riboflavin provision for particular, yet unidentified processes, rather than substituting for the RBP as usually assumed.
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Enzyme organization in the proline biosynthetic pathway of Escherichia coli
Energy Technology Data Exchange (ETDEWEB)
Gamper, H; Moses, V
1974-01-01
The conversion of glutamic acid to proline by an Escherichia coli extract was studied. The activity was dependent upon the presence of ATP and NADPH and was largely unaffected by the presence of NH/sub 3/ or imidazole. The first two pathway enzymes appear to exist as a complex which stabilizes a labile intermediate postulated as ..gamma..-glutamyl phosphate. Attempted synthesis of this compound was unsuccessful due to its spontaneous cyclization to 2-pyrrolidone 5-carboxylate. Dissociation of the enzyme complex upon dilution of the extract is presumed responsible for an experimentally observed dilution effect. E. coli pro/sub A//sup -/ and pro/sub B//sup -/ auxotroph extracts failed to complement one another in the biosynthesis of proline. This is attributed to the lack of a dynamic equilibrium between the complex and its constituent enzymes. In vivo studies with E. coli showed no evidence for metabolic channeling in the final reaction of proline synthesis, the reduction of ..delta../sup 1/-pyrroline 5-carboxylate.
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Mitu, Shahida Akter; Bose, Utpal; Suwansa-Ard, Saowaros; Turner, Luke H; Zhao, Min; Elizur, Abigail; Ogbourne, Steven M; Shaw, Paul Nicholas; Cummins, Scott F
2017-11-07
The sea cucumber (phylum Echinodermata) body wall is the first line of defense and is well known for its production of secondary metabolites; including vitamins and triterpenoid glycoside saponins that have important ecological functions and potential benefits to human health. The genes involved in the various biosynthetic pathways are unknown. To gain insight into these pathways in an echinoderm, we performed a comparative transcriptome analysis and functional annotation of the body wall and the radial nerve of the sea cucumber Holothuria scabra ; to define genes associated with body wall metabolic functioning and secondary metabolite biosynthesis. We show that genes related to signal transduction mechanisms were more highly represented in the H. scabra body wall, including genes encoding enzymes involved in energy production. Eight of the core triterpenoid biosynthesis enzymes were found, however, the identity of the saponin specific biosynthetic pathway enzymes remains unknown. We confirm the body wall release of at least three different triterpenoid saponins using solid phase extraction followed by ultra-high-pressure liquid chromatography-quadrupole time of flight-mass spectrometry. The resource we have established will help to guide future research to explore secondary metabolite biosynthesis in the sea cucumber.
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Directory of Open Access Journals (Sweden)
Shahida Akter Mitu
2017-11-01
Full Text Available The sea cucumber (phylum Echinodermata body wall is the first line of defense and is well known for its production of secondary metabolites; including vitamins and triterpenoid glycoside saponins that have important ecological functions and potential benefits to human health. The genes involved in the various biosynthetic pathways are unknown. To gain insight into these pathways in an echinoderm, we performed a comparative transcriptome analysis and functional annotation of the body wall and the radial nerve of the sea cucumber Holothuria scabra; to define genes associated with body wall metabolic functioning and secondary metabolite biosynthesis. We show that genes related to signal transduction mechanisms were more highly represented in the H. scabra body wall, including genes encoding enzymes involved in energy production. Eight of the core triterpenoid biosynthesis enzymes were found, however, the identity of the saponin specific biosynthetic pathway enzymes remains unknown. We confirm the body wall release of at least three different triterpenoid saponins using solid phase extraction followed by ultra-high-pressure liquid chromatography-quadrupole time of flight-mass spectrometry. The resource we have established will help to guide future research to explore secondary metabolite biosynthesis in the sea cucumber.
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Directory of Open Access Journals (Sweden)
Wu Tian-Shung
2006-07-01
Full Text Available Abstract Background Floral scent is one of the important strategies for ensuring fertilization and for determining seed or fruit set. Research on plant scents has hampered mainly by the invisibility of this character, its dynamic nature, and complex mixtures of components that are present in very small quantities. Most progress in scent research, as in other areas of plant biology, has come from the use of molecular and biochemical techniques. Although volatile components have been identified in several orchid species, the biosynthetic pathways of orchid flower fragrance are far from understood. We investigated how flower fragrance was generated in certain Phalaenopsis orchids by determining the chemical components of the floral scent, identifying floral expressed-sequence-tags (ESTs, and deducing the pathways of floral scent biosynthesis in Phalaneopsis bellina by bioinformatics analysis. Results The main chemical components in the P. bellina flower were shown by gas chromatography-mass spectrometry to be monoterpenoids, benzenoids and phenylpropanoids. The set of floral scent producing enzymes in the biosynthetic pathway from glyceraldehyde-3-phosphate (G3P to geraniol and linalool were recognized through data mining of the P. bellina floral EST database (dbEST. Transcripts preferentially expressed in P. bellina were distinguished by comparing the scent floral dbEST to that of a scentless species, P. equestris, and included those encoding lipoxygenase, epimerase, diacylglycerol kinase and geranyl diphosphate synthase. In addition, EST filtering results showed that transcripts encoding signal transduction and Myb transcription factors and methyltransferase, in addition to those for scent biosynthesis, were detected by in silico hybridization of the P. bellina unigene database against those of the scentless species, rice and Arabidopsis. Altogether, we pinpointed 66% of the biosynthetic steps from G3P to geraniol, linalool and their derivatives
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Hsiao, Yu-Yun; Tsai, Wen-Chieh; Kuoh, Chang-Sheng; Huang, Tian-Hsiang; Wang, Hei-Chia; Wu, Tian-Shung; Leu, Yann-Lii; Chen, Wen-Huei; Chen, Hong-Hwa
2006-07-13
Floral scent is one of the important strategies for ensuring fertilization and for determining seed or fruit set. Research on plant scents has hampered mainly by the invisibility of this character, its dynamic nature, and complex mixtures of components that are present in very small quantities. Most progress in scent research, as in other areas of plant biology, has come from the use of molecular and biochemical techniques. Although volatile components have been identified in several orchid species, the biosynthetic pathways of orchid flower fragrance are far from understood. We investigated how flower fragrance was generated in certain Phalaenopsis orchids by determining the chemical components of the floral scent, identifying floral expressed-sequence-tags (ESTs), and deducing the pathways of floral scent biosynthesis in Phalaneopsis bellina by bioinformatics analysis. The main chemical components in the P. bellina flower were shown by gas chromatography-mass spectrometry to be monoterpenoids, benzenoids and phenylpropanoids. The set of floral scent producing enzymes in the biosynthetic pathway from glyceraldehyde-3-phosphate (G3P) to geraniol and linalool were recognized through data mining of the P. bellina floral EST database (dbEST). Transcripts preferentially expressed in P. bellina were distinguished by comparing the scent floral dbEST to that of a scentless species, P. equestris, and included those encoding lipoxygenase, epimerase, diacylglycerol kinase and geranyl diphosphate synthase. In addition, EST filtering results showed that transcripts encoding signal transduction and Myb transcription factors and methyltransferase, in addition to those for scent biosynthesis, were detected by in silico hybridization of the P. bellina unigene database against those of the scentless species, rice and Arabidopsis. Altogether, we pinpointed 66% of the biosynthetic steps from G3P to geraniol, linalool and their derivatives. This systems biology program combined
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Directory of Open Access Journals (Sweden)
Schulz Burkhard
2010-12-01
Full Text Available Abstract Background Plant growth depends on both cell division and cell expansion. Plant hormones, including brassinosteroids (BRs, are central to the control of these two cellular processes. Despite clear evidence that BRs regulate cell elongation, their roles in cell division have remained elusive. Results Here, we report results emphasizing the importance of BRs in cell division. An Arabidopsis BR biosynthetic mutant, dwarf7-1, displayed various characteristics attributable to slower cell division rates. We found that the DWARF4 gene which encodes for an enzyme catalyzing a rate-determining step in the BR biosynthetic pathways, is highly expressed in the actively dividing callus, suggesting that BR biosynthesis is necessary for dividing cells. Furthermore, dwf7-1 showed noticeably slower rates of callus growth and shoot induction relative to wild-type control. Flow cytometric analyses of the nuclei derived from either calli or intact roots revealed that the cell division index, which was represented as the ratio of cells at the G2/M vs. G1 phases, was smaller in dwf7-1 plants. Finally, we found that the expression levels of the genes involved in cell division and shoot induction, such as PROLIFERATING CELL NUCLEAR ANTIGEN2 (PCNA2 and ENHANCER OF SHOOT REGENERATION2 (ESR2, were also lower in dwf7-1 as compared with wild type. Conclusions Taken together, results of callus induction, shoot regeneration, flow cytometry, and semi-quantitative RT-PCR analysis suggest that BRs play important roles in both cell division and cell differentiation in Arabidopsis.
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International Nuclear Information System (INIS)
Leister, K.J.; Cutry, A.F.; Wenner, C.E.
1986-01-01
Recently, it has been shown that there exists a point in the cell cycle (approximately 2 h prior to S phase entry) when (Na + /K + )ATPase pump activity is no longer needed for progression through the cycle. These data suggests that pump activity is critical in the biosynthetic processes which enables the cell to proceed through the G 1 phase. A scheme is proposed which is currently being tested that (Na + /K + )ATPase pump activity serves as the driving force in the regulation of other membrane transport processes critical for cell proliferation. For example, in post-confluent quiescent C3H-10T1/2 fibroblasts, when [K + ]/sub o/ is lowered just below the K/sub m/ of the pump for K + there is a 10-fold increase in 3 H-uridine uptake into both acid soluble and insoluble cell fractions. By modulation of the pump in this manner, glucose utilization is enhanced whereas inhibition of the pump by ouabain suppresses glucose utilization. In both methods of affecting the pump, 3 H-leucine incorporation is inhibited. Electron acceptors that influence the redox state of the cell have been shown to both stimulate or inhibit cell cycle progression. Under conditions where [K + ]/sub o/ is lowered, the nucleoside uptake responses observed were modified by electron acceptors depending on the ability to oxidize NAD(P)H directly or to interact with a cytochrome-like component, (e.g. phenazine methosulfate) reversed the enhanced uridine uptake and p-phenylene diamine further enhanced the uridine uptake response. These findings suggest that a plasma membrane redox system (presumably cyt-c like) is linked to nucleoside transport which is subject to (Na + /K + )ATPase activity
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International Nuclear Information System (INIS)
Gonzalez-Noriega, Alfonso; Ortega Cuellar, Daniel D.; Michalak, Colette
2006-01-01
Recent work has shown that the cation-independent mannose 6-phosphate and the 78 kDa receptors for lysosomal enzyme targeting are located in different cell compartments. While the mannose 6-phosphate receptor is enriched in the Percoll fractions that contain Golgi apparatus, most of the 78 kDa receptor is localized in a heavy fraction at the bottom of the Percoll gradient. This report presents the biosynthetic transport of the 78 kDa receptor. Newly synthesized 78 kDa receptor was transported to Golgi from endoplasmic reticulum with a half life of 5 min. From the Golgi apparatus, the receptor takes two routes; about 15-25% is transported to the plasma membrane, and the rest migrates to late endosomes, subsequently to prelysosomes and finally to the dense vesicles. The 78 kDa receptor starts appearing at the dense vesicles 120 min after biosynthesis and reaches a maximum of 40-50% of the total receptor. Treatment of cells with NH 4 Cl causes depletion of the receptor from the dense vesicles and prelysosomes and corresponding augmentation in endosomes and plasma membrane. These results suggest that the 78 kDa receptor cycles between compartments and that the dense vesicles seem to represent the most distal compartment in the biosynthetic pathway of this receptor
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Koopen, NR; Post, SM; Wolters, H; Havinga, R; Stellaard, F; Boverhof, R; Kuipers, F; Princen, HMG
Effects of 17 alpha-ethinylestradiol (EE) on the neutral and acidic biosynthetic pathways of bile salt (BS) synthesis were evaluated in rats with an intact enterohepatic circulation and in rats with long-term bile diversion to induce BS synthesis, For this purpose, bile salt pool composition,
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Genes encoding enzymes of the lignin biosynthesis pathway in Eucalyptus
Directory of Open Access Journals (Sweden)
Ricardo Harakava
2005-01-01
Full Text Available Eucalyptus ESTs libraries were screened for genes involved in lignin biosynthesis. This search was performed under the perspective of recent revisions on the monolignols biosynthetic pathway. Eucalyptus orthologues of all genes of the phenylpropanoid pathway leading to lignin biosynthesis reported in other plant species were identified. A library made with mRNAs extracted from wood was enriched for genes involved in lignin biosynthesis and allowed to infer the isoforms of each gene family that play a major role in wood lignin formation. Analysis of the wood library suggests that, besides the enzymes of the phenylpropanoids pathway, chitinases, laccases, and dirigent proteins are also important for lignification. Colocalization of several enzymes on the endoplasmic reticulum membrane, as predicted by amino acid sequence analysis, supports the existence of metabolic channeling in the phenylpropanoid pathway. This study establishes a framework for future investigations on gene expression level, protein expression and enzymatic assays, sequence polymorphisms, and genetic engineering.
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Energy Technology Data Exchange (ETDEWEB)
Baumgartner, Jackson; Lee, Jesi; Halavaty, Andrei S.; Minasov, George; Anderson, Wayne F.; Kuhn, Misty L. (NWU); (SFSU)
2017-10-30
L-Rhamnose is a ubiquitous bacterial cell-wall component. The biosynthetic pathway for its precursor dTDP-L-rhamnose is not present in humans, which makes the enzymes of the pathway potential drug targets. In this study, the three-dimensional structure of the first protein of this pathway, glucose-1-phosphate thymidylyltransferase (RfbA), from
Bacillus anthracis was determined. In other organisms this enzyme is referred to as RmlA. RfbA was co-crystallized with the products of the enzymatic reaction, dTDP-α-D-glucose and pyrophosphate, and its structure was determined at 2.3 Å resolution. This is the first reported thymidylyltransferase structure from a Gram-positive bacterium. RfbA shares overall structural characteristics with known RmlA homologs. However, RfbA exhibits a shorter sequence at its C-terminus, which results in the absence of three α-helices involved in allosteric site formation. Consequently, RfbA was observed to exhibit a quaternary structure that is unique among currently reported glucose-1-phosphate thymidylyltransferase bacterial homologs. These structural analyses suggest that RfbA may not be allosterically regulated in some organisms and is structurally distinct from other RmlA homologs. -
Giant linear plasmids in Streptomyces: a treasure trove of antibiotic biosynthetic clusters.
Kinashi, Haruyasu
2011-01-01
Many giant linear plasmids have been isolated from Streptomyces by using pulsed-field gel electrophoresis and some of them were found to carry an antibiotic biosynthetic cluster(s); SCP1 carries biosynthetic genes for methylenomycin, pSLA2-L for lankacidin and lankamycin, and pKSL for lasalocid and echinomycin. Accumulated data suggest that giant linear plasmids have played critical roles in genome evolution and horizontal transfer of secondary metabolism. In this review, I summarize typical examples of giant linear plasmids whose involvement in antibiotic production has been studied in some detail, emphasizing their finding processes and interaction with the host chromosomes. A hypothesis on horizontal transfer of secondary metabolism involving giant linear plasmids is proposed at the end.
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Directory of Open Access Journals (Sweden)
Xia Wang
Full Text Available The plant Dioscorea composita has important applications in the medical and energy industries, and can be used for the extraction of steroidal sapogenins (important raw materials for the synthesis of steroidal drugs and bioethanol production. However, little is known at the genetic level about how sapogenins are biosynthesized in this plant. Using Illumina deep sequencing, 62,341 unigenes were obtained by assembling its transcriptome, and 27,720 unigenes were annotated. Of these, 8,022 unigenes were mapped to 243 specific pathways, and 531 unigenes were identified to be involved in 24 secondary metabolic pathways. 35 enzymes, which were encoded by 79 unigenes, were related to the biosynthesis of steroidal sapogenins in this transcriptome database, covering almost all the nodes in the steroidal pathway. The results of real-time PCR experiments on ten related transcripts (HMGR, MK, SQLE, FPPS, DXS, CAS, HMED, CYP51, DHCR7, and DHCR24 indicated that sapogenins were mainly biosynthesized by the mevalonate pathway. The expression of these ten transcripts in the tuber and leaves was found to be much higher than in the stem. Also, expression in the shoots was low. The nucleotide and protein sequences and conserved domains of four related genes (HMGR, CAS, SQS, and SMT1 were highly conserved between D. composita and D. zingiberensis; but expression of these four genes is greater in D. composita. However, there is no expression of these key enzymes in potato and no steroidal sapogenins are synthesized.
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Wilson, Edward C F; Stanley, George; Mirza, Zulfiquar
2016-04-01
Wernicke's encephalopathy (WE) is an acute neuropsychiatric condition caused by depleted intracellular thiamine, most commonly arising in chronic alcohol misusers, who may present to emergency departments (EDs) for a variety of reasons. Guidelines recommend a minimum 5-day course of intravenous (IV) thiamine in at-risk patients unless WE can be excluded. To estimate the cost impact on the UK public sector (NHS and social services) of a 5-day course of IV thiamine, vs a 2- and 10-day course, in harmful or dependent drinkers presenting to EDs. A Markov chain model compared expected prognosis of patients under alternative admission strategies over 35 years. Model inputs were derived from a prospective cohort study, expert opinion via structured elicitation and NHS costing databases. Costs (2012/2013 price year) were discounted at 3.5 %. Increasing treatment from 2 to 5 days increased acute care costs but reduced the probability of disease progression and thus reduced the expected net costs by GBP87,000 per patient (95 % confidence interval GBP19,300 to GBP172,300) over 35 years. Increasing length of stay to optimize IV thiamine replacement will place additional strain on acute care but has potential UK public sector cost savings. Social services and the NHS should explore collaborations to realise both the health benefits to patients and savings to the public purse.
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Unal, Meral; Palavan-Unsal, Narcin; Tufekci, M A
2008-03-01
The genotoxic and cytotoxic effects of exogenous polyamines (PAs), putrescine (Put), spermidine (Spd), spermine (Spm) and PA biosynthetic inhibitors, alpha-difluoromethylornithine (DFMO), cyclohexilamine (CHA), methylglioxal bis-(guanylhydrazone) (MGBG) were investigated in the root meristems of Allium cepa L. The reduction of mitotic index and the induction of chromosomal aberrations such as bridges, stickiness, c-mitotic anaphases, micronuclei, endoredupliction by PAs and PA biosynthetic inhibitors were observed and these were used as evidence of genotoxicity and cytotoxicity.
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International Nuclear Information System (INIS)
Hanis, T.; Pospisil, M.; Klir, P.; Bondy, R.
1985-01-01
Two diets for laboratory rat, based on cereals, were either irradiated with 25 or 50 kGy or untreated and stored under free access of air and day light at ambient temperature for 79 d. At the beginning of the experiment and at one month intervals the changes in moisture and vitamin A, thiamine and riboflavin content were evaluated. (author)
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Directory of Open Access Journals (Sweden)
Ceres Mattos Della Lucia
Full Text Available Abstract Fortification of food constitutes an important strategy for the control of micronutrient deficiency and has advantages such as high population coverage and maintenance of eating habits. This study aimed to assess the impact of using fortified rice (Ultra Rice® - UR® on the nutritional status of preschoolers. Ninety-nine children enrolled in two philanthropic preschools participated of the study. Children of one of the preschools were offered UR® mixed with polished rice, as part of school meals (test group and the children of another preschool were offered pure polished rice (control group. Biochemical evaluations were performed before and after 4 months of intervention. Dietary assessment and sensory evaluation of UR® mixed with polished rice were performed during the study. The fortified rice improved the concentrations of zinc (p < 0.001, thiamine (p < 0.001, folic acid (p = 0.003, mean corpuscular hemoglobin (p < 0.001 and mean corpuscular hemoglobin concentration (p < 0.001. The fortified rice showed good acceptability among preschoolers. This study demonstrated the effectiveness of using rice fortified with iron, zinc, thiamine and folic acid on the nutritional status of children.
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Distribution of secondary metabolite biosynthetic gene clusters in 343 Fusarium genomes
Fusarium consists of over 200 phylogenetically distinct species, many of which cause important crop diseases and/or produce mycotoxins and other secondary metabolites (SMs). Some fusaria also cause opportunistic infections in humans and other animals. To investigate the distribution of biosynthetic ...
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Takemura, Miho; Maoka, Takashi; Misawa, Norihiko
2015-03-01
MpBHY codes for a carotene β-ring 3(,3')-hydroxylase responsible for both zeaxanthin and lutein biosynthesis in liverwort. MpCYP97C functions as an ε-ring hydroxylase (zeinoxanthin 3'-hydroxylase) to produce lutein in liverwort. Xanthophylls are oxygenated or hydroxylated carotenes that are most abundant in the light-harvesting complexes of plants. The plant-type xanthophylls consist of α-xanthophyll (lutein) and β-xanthophylls (zeaxanthin, antheraxanthin, violaxanthin and neoxanthin). The α-xanthophyll and β-xanthophylls are derived from α-carotene and β-carotene by carotene hydroxylase activities, respectively. β-Ring 3,3'-hydroxylase that mediates the route of zeaxanthin from β-carotene via β-cryptoxanthin is present in higher plants and is encoded by the BHY (BCH) gene. On the other hand, CYP97A (or BHY) and CYP97C genes are responsible for β-ring 3-hydroxylation and ε-ring 3'-hydroxylation, respectively, in routes from α-carotene to lutein. To elucidate the evolution of the biosynthetic routes of such hydroxylated carotenoids from carotenes in land plants, we identified and functionally analyzed carotenoid hydroxylase genes of liverwort Marchantia polymorpha L. Three genes homologous to higher plants, BHY, CYP97A, and CYP97C, were isolated and named MpBHY, MpCYP97A, and MpCYP97C, respectively. MpBHY was found to code for β-ring hydroxylase, which is responsible for both routes starting from β-carotene and α-carotene. MpCYP97C functioned as an ε-ring hydroxylase not for α-carotene but for zeinoxanthin, while MpCYP97A showed no hydroxylation activity for β-carotene or α-carotene. These findings suggest the original functions of the hydroxylation enzymes of carotenes in land plants, which are thought to diversify in higher plants. In addition, we generated recombinant Escherichia coli cells, which produced rare and novel carotenoids such as α-echinenone and 4-ketozeinoxanthin, through pathway engineering using bacterial carotenogenic genes
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A chloroplast pathway for the de novo biosynthesis of triacylglycerol in Chlamydomonas reinhardtii
Energy Technology Data Exchange (ETDEWEB)
Fan, J.; Xu, C.; Andre, C.
2011-06-23
Neutral lipid metabolism has been extensively studied in yeast, plants and mammals. In contrast, little information is available regarding the biochemical pathway, enzymes and regulatory factors involved in the biosynthesis of triacylglycerol (TAG) in microalgae. In the conventional TAG biosynthetic pathway widely accepted for yeast, plants and mammals, TAG is assembled in the endoplasmic reticulum (ER) from its immediate precursor diacylglycerol (DAG) made by ER-specific acyltransferases, and is deposited exclusively in lipid droplets in the cytosol. Here, we demonstrated that the unicellular microalga Chlamydomonas reinhardtii employs a distinct pathway that uses DAG derived almost exclusively from the chloroplast to produce TAG. This unique TAG biosynthesis pathway is largely dependent on de novo fatty acid synthesis, and the TAG formed in this pathway is stored in lipid droplets in both the chloroplast and the cytosol. These findings have wide implications for understanding TAG biosynthesis and storage and other areas of lipid metabolism in microalgae and other organisms.
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Signal Transduction Pathways that Regulate CAB Gene Expression
Energy Technology Data Exchange (ETDEWEB)
Chory, Joanne
2004-12-31
The process of chloroplast differentiation, involves the coordinate regulation of many nuclear and chloroplast genes. The cues for the initiation of this developmental program are both extrinsic (e.g., light) and intrinsic (cell-type and plastid signals). During this project period, we utilized a molecular genetic approach to select for Arabidopsis mutants that did not respond properly to environmental light conditions, as well as mutants that were unable to perceive plastid damage. These latter mutants, called gun mutants, define two retrograde signaling pathways that regulate nuclear gene expression in response to chloroplasts. A major finding was to identify a signal from chloroplasts that regulates nuclear gene transcription. This signal is the build-up of Mg-Protoporphyrin IX, a key intermediate of the chlorophyll biosynthetic pathway. The signaling pathways downstream of this signal are currently being studied. Completion of this project has provided an increased understanding of the input signals and retrograde signaling pathways that control nuclear gene expression in response to the functional state of chloroplasts. These studies should ultimately influence our abilities to manipulate plant growth and development, and will aid in the understanding of the developmental control of photosynthesis.
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Signal Transduction Pathways that Regulate CAB Gene Expression
Energy Technology Data Exchange (ETDEWEB)
Chory, Joanne
2006-01-16
The process of chloroplast differentiation, involves the coordinate regulation of many nuclear and chloroplast genes. The cues for the initiation of this developmental program are both extrinsic (e.g., light) and intrinsic (cell-type and plastid signals). During this project period, we utilized a molecular genetic approach to select for Arabidopsis mutants that did not respond properly to environmental light conditions, as well as mutants that were unable to perceive plastid damage. These latter mutants, called gun mutants, define two retrograde signaling pathways that regulate nuclear gene expression in response to chloroplasts. A major finding was to identify a signal from chloroplasts that regulates nuclear gene transcription. This signal is the build-up of Mg-Protoporphyrin IX, a key intermediate of the chlorophyll biosynthetic pathway. The signaling pathways downstream of this signal are currently being studied. Completion of this project has provided an increased understanding of the input signals and retrograde signaling pathways that control nuclear gene expression in response to the functional state of chloroplasts. These studies should ultimately influence our abilities to manipulate plant growth and development, and will aid in the understanding of the developmental control of photosynthesis.
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Biosynthetic multitasking facilitates thalassospiramide structural diversity in marine bacteria
Ross, Avena C.
2013-01-23
Thalassospiramides A and B are immunosuppressant cyclic lipopeptides first reported from the marine α-proteobacterium Thalassospira sp. CNJ-328. We describe here the discovery and characterization of an extended family of 14 new analogues from four Tistrella and Thalassospira isolates. These potent calpain 1 protease inhibitors belong to six structure classes in which the length and composition of the acylpeptide side chain varies extensively. Genomic sequence analysis of the thalassospiramide-producing microbes revealed related, genus-specific biosynthetic loci encoding hybrid nonribosomal peptide synthetase/polyketide synthases consistent with thalassospiramide assembly. The bioinformatics analysis of the gene clusters suggests that structural diversity, which ranges from the 803.4 Da thalassospiramide C to the 1291.7 Da thalassospiramide F, results from a complex sequence of reactions involving amino acid substrate channeling and enzymatic multimodule skipping and iteration. Preliminary biochemical analysis of the N-terminal nonribosomal peptide synthetase module from the Thalassospira TtcA megasynthase supports a biosynthetic model in which in cis amino acid activation competes with in trans activation to increase the range of amino acid substrates incorporated at the N terminus. © 2012 American Chemical Society.
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Biosynthetic multitasking facilitates thalassospiramide structural diversity in marine bacteria
Ross, Avena C.; Xü , Ying; Lu, Liang; Kersten, Roland D.; Shao, Zongze; Al-Suwailem, Abdulaziz M.; Dorrestein, Pieter C.; Qian, Peiyuan; Moore, Bradley S.
2013-01-01
Thalassospiramides A and B are immunosuppressant cyclic lipopeptides first reported from the marine α-proteobacterium Thalassospira sp. CNJ-328. We describe here the discovery and characterization of an extended family of 14 new analogues from four Tistrella and Thalassospira isolates. These potent calpain 1 protease inhibitors belong to six structure classes in which the length and composition of the acylpeptide side chain varies extensively. Genomic sequence analysis of the thalassospiramide-producing microbes revealed related, genus-specific biosynthetic loci encoding hybrid nonribosomal peptide synthetase/polyketide synthases consistent with thalassospiramide assembly. The bioinformatics analysis of the gene clusters suggests that structural diversity, which ranges from the 803.4 Da thalassospiramide C to the 1291.7 Da thalassospiramide F, results from a complex sequence of reactions involving amino acid substrate channeling and enzymatic multimodule skipping and iteration. Preliminary biochemical analysis of the N-terminal nonribosomal peptide synthetase module from the Thalassospira TtcA megasynthase supports a biosynthetic model in which in cis amino acid activation competes with in trans activation to increase the range of amino acid substrates incorporated at the N terminus. © 2012 American Chemical Society.
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Wang, Yonglin; Hu, Xiaoping; Fang, Yulin; Anchieta, Amy; Goldman, Polly H; Hernandez, Gustavo; Klosterman, Steven J
2018-04-01
Verticillium dahliae is a soilborne fungus that causes vascular wilt diseases on numerous plant species worldwide. The production of darkly melanized microsclerotia is crucial in the disease cycle of V. dahliae, as these structures allow for long-term survival in soil. Previously, transcriptomic and genomic analysis identified a cluster of genes in V. dahliae that encodes some dihydroxynaphthalene (DHN) melanin biosynthetic pathway homologues found in related fungi. In this study, we explored the roles of cluster-specific transcription factor VdCmr1, as well as two other genes within the cluster encoding a polyketide synthase (VdPKS1) and a laccase (VdLac1), enzymes at initial and endpoint steps in DHN melanin production. The results revealed that VdCmr1 and VdPKS1 are required for melanin production, but neither is required for microsclerotia production. None of the three genes were required for pathogenesis on tobacco and lettuce. Exposure of ΔVdCmr1 and wild-type strains to UV irradiation, or to high temperature (40 °C), revealed an approx. 50 % reduction of survival in the ΔVdCmr1 strain, relative to the wild-type strain, in response to either condition. Expression profiles revealed that expression of some melanin biosynthetic genes are in part dependent on VdCmr1. Combined data indicate VdCmr1 is a key regulator of melanin biosynthesis, and that via regulation of melanogenesis, VdCmr1 affects survival of V. dahliae in response to abiotic threats. We conclude with a model showing regulation of VdCmr1 by a high osmolarity glycerol response (Hog)-type MAP kinase pathway.
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Directory of Open Access Journals (Sweden)
Karen M Vernau
Full Text Available Alaskan Husky Encephalopathy (AHE has been previously proposed as a mitochondrial encephalopathy based on neuropathological similarities with human Leigh Syndrome (LS. We studied 11 Alaskan Husky dogs with AHE, but found no abnormalities in respiratory chain enzyme activities in muscle and liver, or mutations in mitochondrial or nuclear genes that cause LS in people. A genome wide association study was performed using eight of the affected dogs and 20 related but unaffected control AHs using the Illumina canine HD array. SLC19A3 was identified as a positional candidate gene. This gene controls the uptake of thiamine in the CNS via expression of the thiamine transporter protein THTR2. Dogs have two copies of this gene located within the candidate interval (SLC19A3.2 - 43.36-43.38 Mb and SLC19A3.1 - 43.411-43.419 Mb on chromosome 25. Expression analysis in a normal dog revealed that one of the paralogs, SLC19A3.1, was expressed in the brain and spinal cord while the other was not. Subsequent exon sequencing of SLC19A3.1 revealed a 4bp insertion and SNP in the second exon that is predicted to result in a functional protein truncation of 279 amino acids (c.624 insTTGC, c.625 C>A. All dogs with AHE were homozygous for this mutation, 15/41 healthy AH control dogs were heterozygous carriers while 26/41 normal healthy AH dogs were wild type. Furthermore, this mutation was not detected in another 187 dogs of different breeds. These results suggest that this mutation in SLC19A3.1, encoding a thiamine transporter protein, plays a critical role in the pathogenesis of AHE.
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Transcriptome analysis of bitter acid biosynthesis and precursor pathways in hop (Humulus lupulus
Directory of Open Access Journals (Sweden)
Clark Shawn M
2013-01-01
Full Text Available Abstract Background Bitter acids (e.g. humulone are prenylated polyketides synthesized in lupulin glands of the hop plant (Humulus lupulus which are important contributors to the bitter flavour and stability of beer. Bitter acids are formed from acyl-CoA precursors derived from branched-chain amino acid (BCAA degradation and C5 prenyl diphosphates from the methyl-D-erythritol 4-phosphate (MEP pathway. We used RNA sequencing (RNA-seq to obtain the transcriptomes of isolated lupulin glands, cones with glands removed and leaves from high α-acid hop cultivars, and analyzed these datasets for genes involved in bitter acid biosynthesis including the supply of major precursors. We also measured the levels of BCAAs, acyl-CoA intermediates, and bitter acids in glands, cones and leaves. Results Transcripts encoding all the enzymes of BCAA metabolism were significantly more abundant in lupulin glands, indicating that BCAA biosynthesis and subsequent degradation occurs in these specialized cells. Branched-chain acyl-CoAs and bitter acids were present at higher levels in glands compared with leaves and cones. RNA-seq analysis showed the gland-specific expression of the MEP pathway, enzymes of sucrose degradation and several transcription factors that may regulate bitter acid biosynthesis in glands. Two branched-chain aminotransferase (BCAT enzymes, HlBCAT1 and HlBCAT2, were abundant, with gene expression quantification by RNA-seq and qRT-PCR indicating that HlBCAT1 was specific to glands while HlBCAT2 was present in glands, cones and leaves. Recombinant HlBCAT1 and HlBCAT2 catalyzed forward (biosynthetic and reverse (catabolic reactions with similar kinetic parameters. HlBCAT1 is targeted to mitochondria where it likely plays a role in BCAA catabolism. HlBCAT2 is a plastidial enzyme likely involved in BCAA biosynthesis. Phylogenetic analysis of the hop BCATs and those from other plants showed that they group into distinct biosynthetic (plastidial and
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Ancient horizontal gene transfer from bacteria enhances biosynthetic capabilities of fungi.
Directory of Open Access Journals (Sweden)
Imke Schmitt
Full Text Available Polyketides are natural products with a wide range of biological functions and pharmaceutical applications. Discovery and utilization of polyketides can be facilitated by understanding the evolutionary processes that gave rise to the biosynthetic machinery and the natural product potential of extant organisms. Gene duplication and subfunctionalization, as well as horizontal gene transfer are proposed mechanisms in the evolution of biosynthetic gene clusters. To explain the amount of homology in some polyketide synthases in unrelated organisms such as bacteria and fungi, interkingdom horizontal gene transfer has been evoked as the most likely evolutionary scenario. However, the origin of the genes and the direction of the transfer remained elusive.We used comparative phylogenetics to infer the ancestor of a group of polyketide synthase genes involved in antibiotic and mycotoxin production. We aligned keto synthase domain sequences of all available fungal 6-methylsalicylic acid (6-MSA-type PKSs and their closest bacterial relatives. To assess the role of symbiotic fungi in the evolution of this gene we generated 24 6-MSA synthase sequence tags from lichen-forming fungi. Our results support an ancient horizontal gene transfer event from an actinobacterial source into ascomycete fungi, followed by gene duplication.Given that actinobacteria are unrivaled producers of biologically active compounds, such as antibiotics, it appears particularly promising to study biosynthetic genes of actinobacterial origin in fungi. The large number of 6-MSA-type PKS sequences found in lichen-forming fungi leads us hypothesize that the evolution of typical lichen compounds, such as orsellinic acid derivatives, was facilitated by the gain of this bacterial polyketide synthase.
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Nkembo, Marguerite Kasidimoko; Lee, Jung-Bum; Nakagiri, Takeshi; Hayashi, Toshimitsu
2006-05-01
Specific inhibitors of the MVA pathway (pravastatin) and the MEP pathway (fosmidomycin) were used to interfere with the biosynthetic flux which leads to the production of aphidicolin-like diterpene in leaf organ cultures of Scoparia dulcis. Treatment of leaf organs with fosmidomycin resulted in dose dependent inhibition of chlorophylls, carotenoids, scopadulcic acid B (SDB) and phytol production, and no effect on sterol production was observed. In response to the pravastatin treatment, a significant decrease in sterol and perturbation of SDB production was observed.
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Rational synthetic pathway refactoring of natural products biosynthesis in actinobacteria.
Tan, Gao-Yi; Liu, Tiangang
2017-01-01
Natural products (NPs) and their derivatives are widely used as frontline treatments for many diseases. Actinobacteria spp. are used to produce most of NP antibiotics and have also been intensively investigated for NP production, derivatization, and discovery. However, due to the complicated transcriptional and metabolic regulation of NP biosynthesis in Actinobacteria, especially in the cases of genome mining and heterologous expression, it is often difficult to rationally and systematically engineer synthetic pathways to maximize biosynthetic efficiency. With the emergence of new tools and methods in metabolic engineering, the synthetic pathways of many chemicals, such as fatty acids and biofuels, in model organisms (e.g. Escherichia coli ), have been refactored to realize precise and flexible control of production. These studies also offer a promising approach for synthetic pathway refactoring in Actinobacteria. In this review, the great potential of Actinobacteria as a microbial cell factory for biosynthesis of NPs is discussed. To this end, recent progress in metabolic engineering of NP synthetic pathways in Actinobacteria are summarized and strategies and perspectives to rationally and systematically refactor synthetic pathways in Actinobacteria are highlighted. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.
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Energy Technology Data Exchange (ETDEWEB)
Yaeger, E.; Siegel, B.Z.; Siegel, S.M.; Lasconia, M.; Correal, T.
1986-01-01
The positive photoatactic behavior of Artemia was disturbed when larvae cultured in 0.25 M NaCl were introduced into 10/sup -7/ to 10/sup -5/ M HgCl/sub 2/. At 10/sup -8/ M, and in controls, Hg had no effect either on orientation or mortality, and 65% of the larvae were collected at the light source in 30 min or less. In contrast, at 10/sup -6/ M, only 29% were able to do so. The presence of 10/sup -4/ M thiamine hydrochloride in the excystment medium prevented fully the loss of light-sensitivity.
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Directory of Open Access Journals (Sweden)
Jungsuwadee Paiboon
2011-02-01
Full Text Available Abstract Background Lactation increases energy demands four- to five-fold, leading to a two- to three-fold increase in food consumption, requiring a proportional adjustment in the ability of the lactating dam to absorb nutrients and to synthesize critical biomolecules, such as cholesterol, to meet the dietary needs of both the offspring and the dam. The size and hydrophobicity of the bile acid pool increases during lactation, implying an increased absorption and disposition of lipids, sterols, nutrients, and xenobiotics. In order to investigate changes at the transcriptomics level, we utilized an exon array and calculated expression levels to investigate changes in gene expression in the liver, duodenum, jejunum, and ileum of lactating dams when compared against age-matched virgin controls. Results A two-way mixed models ANOVA was applied to detect differentially expressed genes. Significance calls were defined as a p Cyp7a1, which catalyzes the rate limiting step in the bile acid biosynthetic pathway, was also significantly increased in liver. In addition, decreased levels of mRNA associated with T-cell signaling were found in the jejunum and ileum. Several members of the Solute Carrier (SLC and Adenosine Triphosphate Binding Cassette (ABC superfamilies of membrane transporters were found to be differentially expressed; these genes may play a role in differences in nutrient and xenobiotic absorption and disposition. mRNA expression of SLC39a4_predicted, a zinc transporter, was increased in all tissues, suggesting that it is involved in increased zinc uptake during lactation. Microarray data are available through GEO under GSE19175. Conclusions We detected differential expression of mRNA from several pathways in lactating dams, including upregulation of the cholesterol biosynthetic pathway in liver and intestine, consistent with Srebp activation. Differential T-Cell signaling in the two most distal regions of the small intestine (ileum and
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Diurnal and circadian expression profiles of glycerolipid biosynthetic genes in Arabidopsis.
Nakamura, Yuki; Andrés, Fernando; Kanehara, Kazue; Liu, Yu-chi; Coupland, George; Dörmann, Peter
2014-01-01
Glycerolipid composition in plant membranes oscillates in response to diurnal change. However, its functional significance remained unclear. A recent discovery that Arabidopsis florigen FT binds diurnally oscillating phosphatidylcholine molecules to promote flowering suggests that diurnal oscillation of glycerolipid composition is an important input in flowering time control. Taking advantage of public microarray data, we globally analyzed the expression pattern of glycerolipid biosynthetic genes in Arabidopsis under long-day, short-day, and continuous light conditions. The results revealed that 12 genes associated with glycerolipid metabolism showed significant oscillatory profiles. Interestingly, expression of most of these genes followed circadian profiles, suggesting that glycerolipid biosynthesis is partially under clock regulation. The oscillating expression profile of one representative gene, PECT1, was analyzed in detail. Expression of PECT1 showed a circadian pattern highly correlated with that of the clock-regulated gene GIGANTEA. Thus, our study suggests that a considerable number of glycerolipid biosynthetic genes are under circadian control.
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Arjunan, Palaniappa; Chandrasekhar, Krishnamoorthy; Sax, Martin; Brunskill, Andrew; Nemeria, Natalia; Jordan, Frank; Furey, William
2004-03-09
Thiamin thiazolone diphosphate (ThTDP), a potent inhibitor of the E1 component from the Escherichia coli pyruvate dehydrogenase multienzyme complex (PDHc), binds to the enzyme with greater affinity than does the cofactor thiamin diphosphate (ThDP). To identify what determines this difference, the crystal structure of the apo PDHc E1 component complex with ThTDP and Mg(2+) has been determined at 2.1 A and compared to the known structure of the native holoenzyme, PDHc E1-ThDP-Mg(2+) complex. When ThTDP replaces ThDP, reorganization occurs in the protein structure in the vicinity of the active site involving positional and conformational changes in some amino acid residues, a change in the V coenzyme conformation, addition of new hydration sites, and elimination of others. These changes culminate in an increase in the number of hydrogen bonds to the protein, explaining the greater affinity of the apoenzyme for ThTDP. The observed hydrogen bonding pattern is not an invariant feature of ThDP-dependent enzymes but rather specific to this enzyme since the extra hydrogen bonds are made with nonconserved residues. Accordingly, these sequence-related hydrogen bonding differences likewise explain the wide variation in the affinities of different thiamin-dependent enzymes for ThTDP and ThDP. The sequence of each enzyme determines its ability to form hydrogen bonds to the inhibitor or cofactor. Mechanistic roles are suggested for the aforementioned reorganization and its reversal in PDHc E1 catalysis: to promote substrate binding and product release. This study also provides additional insight into the role of water in enzyme inhibition and catalysis.
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A nuclear-receptor-dependent phosphatidylcholine pathway with antidiabetic effects.
Lee, Jae Man; Lee, Yoon Kwang; Mamrosh, Jennifer L; Busby, Scott A; Griffin, Patrick R; Pathak, Manish C; Ortlund, Eric A; Moore, David D
2011-05-25
Nuclear hormone receptors regulate diverse metabolic pathways and the orphan nuclear receptor LRH-1 (also known as NR5A2) regulates bile acid biosynthesis. Structural studies have identified phospholipids as potential LRH-1 ligands, but their functional relevance is unclear. Here we show that an unusual phosphatidylcholine species with two saturated 12 carbon fatty acid acyl side chains (dilauroyl phosphatidylcholine (DLPC)) is an LRH-1 agonist ligand in vitro. DLPC treatment induces bile acid biosynthetic enzymes in mouse liver, increases bile acid levels, and lowers hepatic triglycerides and serum glucose. DLPC treatment also decreases hepatic steatosis and improves glucose homeostasis in two mouse models of insulin resistance. Both the antidiabetic and lipotropic effects are lost in liver-specific Lrh-1 knockouts. These findings identify an LRH-1 dependent phosphatidylcholine signalling pathway that regulates bile acid metabolism and glucose homeostasis.
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Directory of Open Access Journals (Sweden)
Chengjian Li
2018-01-01
Full Text Available L-Theanine is a nonprotein amino acid in tea, and its immunomodulatory function has been confirmed. This study aimed to investigate the effect of L-theanine addition on cytokines secretion in rat splenic lymphocytes and explore its potential immunomodulatory effects on the mevalonate biosynthetic pathway. Our results showed that L-theanine treatment did not influence the proliferation and division indexes of the splenic lymphocytes subsets. Interestingly, L-theanine treatment had regulated the contents of IFN-γ, IL-2, IL-4, IL-10, IL-12, and TNF-α (P<0.001 except IL-6 and upregulated the mRNA and protein expression of Ras-related protein Rap-1A (Rap1A, 3-hydroxy-3-methylglutaryl-CoA reductase (HMGCR, and farnesyl diphosphate synthase (FDPs (P<0.001. Additionally, there was a positive correlation between Rap1A and HMGCR proteins expression and IFN-γ, IL-4, and IL-6 levels. In conclusion, L-theanine regulated the secretion of cytokines probably by activating expression of Rap1A and HMGCR proteins involved in the mevalonate biosynthetic pathway in rat splenic lymphocytes. Therefore, L-theanine might be a promising potential drug candidate as immunopotentiator.
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Directory of Open Access Journals (Sweden)
Jine Li
Full Text Available The paulomycins are a group of glycosylated compounds featuring a unique paulic acid moiety. To locate their biosynthetic gene clusters, the genomes of two paulomycin producers, Streptomyces paulus NRRL 8115 and Streptomyces sp. YN86, were sequenced. The paulomycin biosynthetic gene clusters were defined by comparative analyses of the two genomes together with the genome of the third paulomycin producer Streptomyces albus J1074. Subsequently, the identity of the paulomycin biosynthetic gene cluster was confirmed by inactivation of two genes involved in biosynthesis of the paulomycose branched chain (pau11 and the ring A moiety (pau18 in Streptomyces paulus NRRL 8115. After determining the gene cluster boundaries, a convergent biosynthetic model was proposed for paulomycin based on the deduced functions of the pau genes. Finally, a paulomycin high-producing strain was constructed by expressing an activator-encoding gene (pau13 in S. paulus, setting the stage for future investigations.
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Expression of Xanthophyll Biosynthetic Genes during Light-Dependent Chloroplast Differentiation1
Woitsch, Sonja; Römer, Susanne
2003-01-01
In higher plants, etioplast to chloroplast differentiation is characterized by dramatic ultrastructural changes of the plastid and a concomitant increase in chlorophylls and carotenoids. Whereas the formation and function of carotenes and their oxygenated derivatives, the xanthophylls, have been well studied, little is known about the regulation of the genes involved in xanthophyll biosynthesis. Here, we analyze the expression of three xanthophyll biosynthetic genes (i.e. β-carotene hydroxylase [bhy], zeaxanthin epoxidase [zep], and violaxanthin de-epoxidase [vde]) during de-etiolation of seedlings of tobacco (Nicotiana tabacum L. cv Samsun) under different light conditions. White-light illumination caused an increase in the amount of all corresponding mRNAs. The expression profiles of bhy and zep not only resembled each other but were also similar to the pattern of a gene encoding a major light-harvesting protein of photosystem II. This finding indicates a coordinated synthesis during formation of the antenna complex. In contrast, the expression pattern of vde was clearly different. Furthermore, the gene expression of bhy was shown to be modulated after illumination with different white-light intensities. The expression of all xanthophyll biosynthetic genes under examination was up-regulated upon exposure to red, blue, and white light. Gene expression of bhy and vde but not of zep was more pronounced under red-light illumination, pointing at an involvement of the phytochrome system. Expression analysis in the presence of the photosynthetic electron transport inhibitors 3-(3,4-dichlorophenyl)-1,1-dimethyl-urea and 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone indicated a redox control of transcription of two of the xanthophyll biosynthetic genes (bhy and zep). PMID:12857831
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Electron density reactivity indexes of the tautomeric/ionization forms of thiamin diphosphate.
Jaña, Gonzalo A; Delgado, Eduardo J
2013-09-01
The generation of the highly reactive ylide in thiamin diphosphate catalysis is analyzed in terms of the nucleophilicity of key atoms, by means of density functional calculations at X3LYP/6-31++G(d,p) level of theory. The Fukui functions of all tautomeric/ionization forms are calculated in order to assess their reactivity. The results allow to conclude that the highly conserved glutamic residue does not protonate the N1' atom of the pyrimidyl ring, but it participates in a strong hydrogen bonding, stabilizing the eventual negative charge on the nitrogen, in all forms involved in the ylide generation. This condition provides the necessary reactivity on key atoms, N4' and C2, to carry out the formation of the ylide required to initiate the catalytic cycle of ThDP-dependent enzymes. This study represents a new approach for the ylide formation in ThDP catalysis.
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Yamada, Tetsuya; Matsuda, Fumio; Kasai, Koji; Fukuoka, Shuichi; Kitamura, Keisuke; Tozawa, Yuzuru; Miyagawa, Hisashi; Wakasa, Kyo
2008-01-01
Two distinct biosynthetic pathways for Phe in plants have been proposed: conversion of prephenate to Phe via phenylpyruvate or arogenate. The reactions catalyzed by prephenate dehydratase (PDT) and arogenate dehydratase (ADT) contribute to these respective pathways. The Mtr1 mutant of rice (Oryza sativa) manifests accumulation of Phe, Trp, and several phenylpropanoids, suggesting a link between the synthesis of Phe and Trp. Here, we show that the Mtr1 mutant gene (mtr1-D) encodes a form of rice PDT with a point mutation in the putative allosteric regulatory region of the protein. Transformed callus lines expressing mtr1-D exhibited all the characteristics of Mtr1 callus tissue. Biochemical analysis revealed that rice PDT possesses both PDT and ADT activities, with a preference for arogenate as substrate, suggesting that it functions primarily as an ADT. The wild-type enzyme is feedback regulated by Phe, whereas the mutant enzyme showed a reduced feedback sensitivity, resulting in Phe accumulation. In addition, these observations indicate that rice PDT is critical for regulating the size of the Phe pool in plant cells. Feeding external Phe to wild-type callus tissue and seedlings resulted in Trp accumulation, demonstrating a connection between Phe accumulation and Trp pool size. PMID:18487352
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Asymmetric Stetter reactions catalyzed by thiamine diphosphate-dependent enzymes.
Kasparyan, Elena; Richter, Michael; Dresen, Carola; Walter, Lydia S; Fuchs, Georg; Leeper, Finian J; Wacker, Tobias; Andrade, Susana L A; Kolter, Geraldine; Pohl, Martina; Müller, Michael
2014-12-01
The intermolecular asymmetric Stetter reaction is an almost unexplored transformation for biocatalysts. Previously reported thiamine diphosphate (ThDP)-dependent PigD from Serratia marcescens is the first enzyme identified to catalyze the Stetter reaction of α,β-unsaturated ketones (Michael acceptor substrates) and α-keto acids. PigD is involved in the biosynthesis of the potent cytotoxic agent prodigiosin. Here, we describe the investigation of two new ThDP-dependent enzymes, SeAAS from Saccharopolyspora erythraea and HapD from Hahella chejuensis. Both show a high degree of homology to the amino acid sequence of PigD (39 and 51 %, respectively). The new enzymes were heterologously overproduced in Escherichia coli, and the yield of soluble protein was enhanced by co-expression of the chaperone genes groEL/ES. SeAAS and HapD catalyze intermolecular Stetter reactions in vitro with high enantioselectivity. The enzymes possess a characteristic substrate range with respect to Michael acceptor substrates. This provides support for a new type of ThDP-dependent enzymatic activity, which is abundant in various species and not restricted to prodigiosin biosynthesis in different strains. Moreover, PigD, SeAAS, and HapD are also able to catalyze asymmetric carbon-carbon bond formation reactions of aldehydes and α-keto acids, resulting in 2-hydroxy ketones.
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Evolutionary rate patterns of the Gibberellin pathway genes
Directory of Open Access Journals (Sweden)
Zhang Fu-min
2009-08-01
Full Text Available Abstract Background Analysis of molecular evolutionary patterns of different genes within metabolic pathways allows us to determine whether these genes are subject to equivalent evolutionary forces and how natural selection shapes the evolution of proteins in an interacting system. Although previous studies found that upstream genes in the pathway evolved more slowly than downstream genes, the correlation between evolutionary rate and position of the genes in metabolic pathways as well as its implications in molecular evolution are still less understood. Results We sequenced and characterized 7 core structural genes of the gibberellin biosynthetic pathway from 8 representative species of the rice tribe (Oryzeae to address alternative hypotheses regarding evolutionary rates and patterns of metabolic pathway genes. We have detected significant rate heterogeneity among 7 GA pathway genes for both synonymous and nonsynonymous sites. Such rate variation is mostly likely attributed to differences of selection intensity rather than differential mutation pressures on the genes. Unlike previous argument that downstream genes in metabolic pathways would evolve more slowly than upstream genes, the downstream genes in the GA pathway did not exhibited the elevated substitution rate and instead, the genes that encode either the enzyme at the branch point (GA20ox or enzymes catalyzing multiple steps (KO, KAO and GA3ox in the pathway had the lowest evolutionary rates due to strong purifying selection. Our branch and codon models failed to detect signature of positive selection for any lineage and codon of the GA pathway genes. Conclusion This study suggests that significant heterogeneity of evolutionary rate of the GA pathway genes is mainly ascribed to differential constraint relaxation rather than the positive selection and supports the pathway flux theory that predicts that natural selection primarily targets enzymes that have the greatest control on fluxes.
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Nicotinamide riboside kinase structures reveal new pathways to NAD+.
Directory of Open Access Journals (Sweden)
Wolfram Tempel
2007-10-01
Full Text Available The eukaryotic nicotinamide riboside kinase (Nrk pathway, which is induced in response to nerve damage and promotes replicative life span in yeast, converts nicotinamide riboside to nicotinamide adenine dinucleotide (NAD+ by phosphorylation and adenylylation. Crystal structures of human Nrk1 bound to nucleoside and nucleotide substrates and products revealed an enzyme structurally similar to Rossmann fold metabolite kinases and allowed the identification of active site residues, which were shown to be essential for human Nrk1 and Nrk2 activity in vivo. Although the structures account for the 500-fold discrimination between nicotinamide riboside and pyrimidine nucleosides, no enzyme feature was identified to recognize the distinctive carboxamide group of nicotinamide riboside. Indeed, nicotinic acid riboside is a specific substrate of human Nrk enzymes and is utilized in yeast in a novel biosynthetic pathway that depends on Nrk and NAD+ synthetase. Additionally, nicotinic acid riboside is utilized in vivo by Urh1, Pnp1, and Preiss-Handler salvage. Thus, crystal structures of Nrk1 led to the identification of new pathways to NAD+.
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Prokaryotic Heme Biosynthesis: Multiple Pathways to a Common Essential Product.
Dailey, Harry A; Dailey, Tamara A; Gerdes, Svetlana; Jahn, Dieter; Jahn, Martina; O'Brian, Mark R; Warren, Martin J
2017-03-01
The advent of heme during evolution allowed organisms possessing this compound to safely and efficiently carry out a variety of chemical reactions that otherwise were difficult or impossible. While it was long assumed that a single heme biosynthetic pathway existed in nature, over the past decade, it has become clear that there are three distinct pathways among prokaryotes, although all three pathways utilize a common initial core of three enzymes to produce the intermediate uroporphyrinogen III. The most ancient pathway and the only one found in the Archaea converts siroheme to protoheme via an oxygen-independent four-enzyme-step process. Bacteria utilize the initial core pathway but then add one additional common step to produce coproporphyrinogen III. Following this step, Gram-positive organisms oxidize coproporphyrinogen III to coproporphyrin III, insert iron to make coproheme, and finally decarboxylate coproheme to protoheme, whereas Gram-negative bacteria first decarboxylate coproporphyrinogen III to protoporphyrinogen IX and then oxidize this to protoporphyrin IX prior to metal insertion to make protoheme. In order to adapt to oxygen-deficient conditions, two steps in the bacterial pathways have multiple forms to accommodate oxidative reactions in an anaerobic environment. The regulation of these pathways reflects the diversity of bacterial metabolism. This diversity, along with the late recognition that three pathways exist, has significantly slowed advances in this field such that no single organism's heme synthesis pathway regulation is currently completely characterized. Copyright © 2017 American Society for Microbiology.
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DEFF Research Database (Denmark)
Henriques de Jesus, Maria Perestrello Ramos; Nielsen, Agnieszka Janina Zygadlo; Mellor, Silas Busck
2017-01-01
to their complex structures. Some of the crucial enzymes catalyzing their biosynthesis are the cytochromes P450 (P450s) situated in the endoplasmic reticulum (ER), powered by electron transfers from NADPH. Dhurrin is a cyanogenic glucoside and its biosynthesis involves a dynamic metabolon formed by two P450s....... Nevertheless, translocation of the pathway from the ER to the chloroplast creates other difficulties, such as the loss of metabolon formation and intermediate diversion into other metabolic pathways. We show here that co-localization of these enzymes in the thylakoid membrane leads to a significant increase...... in product formation, with a concomitant decrease in off-pathway intermediates. This was achieved by exchanging the membrane anchors of the dhurrin pathway enzymes to components of the Twin-arginine translocation pathway, TatB and TatC, which have self-assembly properties. Consequently, we show 5-fold...
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A novel, rapid and sensitive Ultra Performance Liquid-Chromatography tandem Mass-Spectrometry (UPLC-MS/MS) method for the simultaneous determination of several B-vitamins in human milk was developed. Resolution by retention time or multiple reaction monitoring (MRM) for thiamin, riboflavin, flavin a...
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Ach, Thomas; Kardorff, Rüdiger; Rohrschneider, Klaus
2015-01-01
To report ophthalmologic fundus autofluorescence and spectral domain optical coherence tomography findings in a patient with thiamine responsive megaloblastic anemia (TRMA). A 13-year-old girl with genetically proven TRMA was ophthalmologically (visual acuity, funduscopy, perimetry, electroretinogram) followed up over >5 years. Fundus imaging also included autofluorescence and spectral domain optical coherence tomography. During a 5-year follow-up, visual acuity and visual field decreased, despite a special TRMA diet. Funduscopy revealed bull's eye appearance, whereas fundus autofluorescence showed central and peripheral hyperfluorescence and perifoveal hypofluorescence. Spectral domain optical coherence tomography revealed affected inner segment ellipsoid band and irregularities in the retinal pigment epithelium and choroidea. Autofluorescence and spectral domain optical coherence tomography findings in a patient with TRMA show retinitis pigmentosa-like retina, retinal pigment epithelium, and choroid alterations. These findings might progress even under special TRMA diet, indispensable to life. Ophthalmologist should consider TRMA in patients with deafness and ophthalmologic disorders.
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DEFF Research Database (Denmark)
Pissarra, P.D.; Nielsen, Jens Bredal
1997-01-01
This paper describes the thermodynamic analysis of pathways related to penicillin production in Penicillium chrysogenum. First a thermodynamic feasibility analysis is performed of the L-lysine pathway of which one of the precursors for penicillin biosynthesis (alpha-aminoadipic acid......) is an intermediate. It is found that the L-lysine pathway in P. chrysogenum is thermodynamically feasible and that the calculated standard Gibbs free energy values of the two enzymes controlling the pathway flux indicate that they operate far from equilibrium. It is therefore proposed that the regulation of alpha......-aminoadipate reductase by lysine is important to maintain a high concentration of alpha-aminoadipate in order to direct the carbon flux to penicillin production. Secondly the changes in Gibbs free energy in the penicillin biosynthetic pathway during fed-batch cultivation were studied. The analysis showed that all...
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Genetic analysis of the capsular biosynthetic locus from all 90 pneumococcal serotypes.
Directory of Open Access Journals (Sweden)
Stephen D Bentley
2006-03-01
Full Text Available Several major invasive bacterial pathogens are encapsulated. Expression of a polysaccharide capsule is essential for survival in the blood, and thus for virulence, but also is a target for host antibodies and the basis for effective vaccines. Encapsulated species typically exhibit antigenic variation and express one of a number of immunochemically distinct capsular polysaccharides that define serotypes. We provide the sequences of the capsular biosynthetic genes of all 90 serotypes of Streptococcus pneumoniae and relate these to the known polysaccharide structures and patterns of immunological reactivity of typing sera, thereby providing the most complete understanding of the genetics and origins of bacterial polysaccharide diversity, laying the foundations for molecular serotyping. This is the first time, to our knowledge, that a complete repertoire of capsular biosynthetic genes has been available, enabling a holistic analysis of a bacterial polysaccharide biosynthesis system. Remarkably, the total size of alternative coding DNA at this one locus exceeds 1.8 Mbp, almost equivalent to the entire S. pneumoniae chromosomal complement.
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Liu, Lan; Salam, Nimaichand; Jiao, Jian-Yu; Jiang, Hong-Chen; Zhou, En-Min; Yin, Yi-Rui; Ming, Hong; Li, Wen-Jun
2016-07-01
The class Actinobacteria has been a goldmine for the discovery of antibiotics and has attracted interest from both academics and industries. However, an absence of novel approaches during the last few decades has limited the discovery of new microbial natural products useful for industries. Scientists are now focusing on the ecological aspects of diverse environments including unexplored or underexplored habitats and extreme environments in the search for new metabolites. This paper reports on the diversity of culturable actinobacteria associated with hot springs located in Tengchong County, Yunnan Province, southwestern China. A total of 58 thermophilic actinobacterial strains were isolated from the samples collected from ten hot springs distributed over three geothermal fields (e.g., Hehua, Rehai, and Ruidian). Phylogenetic positions and their biosynthetic profiles were analyzed by sequencing 16S rRNA gene and three biosynthetic gene clusters (KS domain of PKS-I, KSα domain of PKS-II and A domain of NRPS). On the basis of 16S rRNA gene phylogenetic analysis, the 58 strains were affiliated with 12 actinobacterial genera: Actinomadura Micromonospora, Microbispora, Micrococcus, Nocardiopsis, Nonomuraea, Promicromonospora, Pseudonocardia, Streptomyces, Thermoactinospora, Thermocatellispora, and Verrucosispora, of which the two novel genera Thermoactinospora and Thermocatellisopora were recently described from among these strains. Considering the biosynthetic potential of these actinobacterial strains, 22 were positive for PCR amplification of at least one of the three biosynthetic gene clusters (PKS-I, PKS-II, and NRPS). These actinobacteria were further subjected to antimicrobial assay against five opportunistic human pathogens (Acinetobacter baumannii, Escherichia coli, Micrococcus luteus, Staphylococcus aureus and Streptococcus faecalis). All of the 22 strains that were positive for PCR amplification of at least one of the biosynthetic gene domains exhibited
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Zhou, Yikang; Li, Gang; Dong, Junkai; Xing, Xin-Hui; Dai, Junbiao; Zhang, Chong
2018-05-01
Facing boosting ability to construct combinatorial metabolic pathways, how to search the metabolic sweet spot has become the rate-limiting step. We here reported an efficient Machine-learning workflow in conjunction with YeastFab Assembly strategy (MiYA) for combinatorial optimizing the large biosynthetic genotypic space of heterologous metabolic pathways in Saccharomyces cerevisiae. Using β-carotene biosynthetic pathway as example, we first demonstrated that MiYA has the power to search only a small fraction (2-5%) of combinatorial space to precisely tune the expression level of each gene with a machine-learning algorithm of an artificial neural network (ANN) ensemble to avoid over-fitting problem when dealing with a small number of training samples. We then applied MiYA to improve the biosynthesis of violacein. Feed with initial data from a colorimetric plate-based, pre-screened pool of 24 strains producing violacein, MiYA successfully predicted, and verified experimentally, the existence of a strain that showed a 2.42-fold titer improvement in violacein production among 3125 possible designs. Furthermore, MiYA was able to largely avoid the branch pathway of violacein biosynthesis that makes deoxyviolacein, and produces very pure violacein. Together, MiYA combines the advantages of standardized building blocks and machine learning to accelerate the Design-Build-Test-Learn (DBTL) cycle for combinatorial optimization of metabolic pathways, which could significantly accelerate the development of microbial cell factories. Copyright © 2018 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.
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Directory of Open Access Journals (Sweden)
Deniz Sabahattin
2017-01-01
Full Text Available The existence of heavy metal ions in aqueous media is one of the biggest environmental pollution problems and thus the removal of heavy metals is a very important procedure. In this work, a new adsorbent was synthesized by modifying 3-aminopropyl-functionalized silica gel with thiamine (vitamin B1 and characterized. The influence of the uptake conditions, such as pH, contact time, initial feed concentration and foreign metal ions, on the binding capacity of thiamine-functionalized silica gel sorbent (M3APS were investigated. Maximum obtained adsorption capacities for Pb(II, Hg(II and Cd(II were 39.4±0.2, 30.9±0.5 and 9.54±0.4 mg g-1 M3APS, respectively, at pH 5.0. The observed selectivity of M3APS for these metal ions was the following: Pb(II > Hg(II > Cd(II. Adsorption isotherm models were also applied to the adsorption process. As a result, the Langmuir isotherm model gave the best fit for the adsorption of metal ions on M3APS. The Gibbs energy change (ΔG for the adsorption of Pb(II, Hg(II and Cd(II were calculated to predict the nature of adsorption process. Having such satisfactory adsorption results, M3APS is a potential candidate adsorbent for Pb(II and Hg(II removal from aqueous media.
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International Nuclear Information System (INIS)
Ayranci, Guler; Sahin, Melike; Ayranci, Erol
2007-01-01
Apparent molar volumes and apparent molar isentropic compressibilities of ascorbic acid (vitamin C) and thiamine hydrochloride (vitamin B 1 ) were determined from accurately measured density and sound velocity data in water and in aqueous NaCl solutions at (283.15, 293.15, 298.15, 303.15, 308.15, and 313.15) K. These volume and compressibility data were extrapolated to zero concentration using suitable empirical or theoretical equations to determine the corresponding infinite dilution values. Apparent molar expansibilities at infinite dilution were determined from slopes of apparent molar volume vs. temperature plots. Ionization of both ascorbic acid and thiamine hydrochloride were suppressed using sufficiently acidic solutions. Apparent molar volumes at infinite dilution for ascorbic acid and thiamine hydrochloride were found to increase with temperature in acidic solutions and in the presence of co-solute, NaCl. Apparent molar expansibility at infinite dilution were found to be constant over the temperature range studied and were all positive, indicating the hydrophilic character of the two vitamins studied in water and in the presence of co-solute, NaCl. Apparent molar isentropic compressibilities of ascorbic acid at infinite dilution were positive in water and in the presence of co-solute, NaCl, at low molalities. Those of thiamine hydrochloride at infinitive dilution were all negative, consistent with its ionic nature. Transfer apparent molar volumes of vitamins at infinite dilution from water solutions to NaCl solutions at various temperatures were determined. The results were interpreted in terms of complex vitamin-water-co-solute (NaCl) interactions
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Insights into secondary metabolism from a global analysis of prokaryotic biosynthetic gene clusters
Cimermancic, P.; Medema, Marnix; Claesen, J.; Kurika, K.; Wieland Brown, L.C.; Mavrommatis, K.; Pati, A.; Godfrey, P.A.; Koehrsen, M.; Clardy, J.; Birren, B. W.; Takano, Eriko; Sali, A.; Linington, R.G.; Fischbach, M.A.
2014-01-01
Although biosynthetic gene clusters (BGCs) have been discovered for hundreds of bacterial metabolites, our knowledge of their diversity remains limited. Here, we used a novel algorithm to systematically identify BGCs in the extensive extant microbial sequencing data. Network analysis of the
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Cerneus, D. P.; Ueffing, E.; Posthuma, G.; Strous, G. J.; van der Ende, A.
1993-01-01
Alkaline phosphatase is anchored to the outer leaflet of the plasma membrane by a covalently attached glycosyl-phosphatidylinositol anchor. We have studied the biosynthetic transport and endocytosis of alkaline phosphatase in the choriocarcinoma cell line BeWo, which endogenously expresses this
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Chromatin landscaping in algae reveals novel regulation pathway for biofuels production
Energy Technology Data Exchange (ETDEWEB)
Ngan, Chew Yee; Wong, Chee-Hong; Choi, Cindy; Pratap, Abhishek; Han, James; Wei, Chia-Lin
2013-02-19
The diminishing reserve of fossil fuels calls for the development of biofuels. Biofuels are produced from renewable resources, including photosynthetic organisms, generating clean energy. Microalgae is one of the potential feedstock for biofuels production. It grows easily even in waste water, and poses no competition to agricultural crops for arable land. However, little is known about the algae lipid biosynthetic regulatory mechanisms. Most studies relied on the homology to other plant model organisms, in particular Arabidopsis or through low coverage expression analysis to identify key enzymes. This limits the discovery of new components in the biosynthetic pathways, particularly the genetic regulators and effort to maximize the production efficiency of algal biofuels. Here we report an unprecedented and de novo approach to dissect the algal lipid pathways through disclosing the temporal regulations of chromatin states during lipid biosynthesis. We have generated genome wide chromatin maps in chlamydomonas genome using ChIP-seq targeting 7 histone modifications and RNA polymerase II in a time-series manner throughout conditions activating lipid biosynthesis. To our surprise, the combinatory profiles of histone codes uncovered new regulatory mechanism in gene expression in algae. Coupled with matched RNA-seq data, chromatin changes revealed potential novel regulators and candidate genes involved in the activation of lipid accumulations. Genetic perturbation on these candidate regulators further demonstrated the potential to manipulate the regulatory cascade for lipid synthesis efficiency. Exploring epigenetic landscape in microalgae shown here provides powerful tools needed in improving biofuel production and new technology platform for renewable energy generation, global carbon management, and environmental survey.
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The synthesis of chlorophyll-a biosynthetic precursors and methyl substituted iron porphyrins
International Nuclear Information System (INIS)
Matera, K.M.
1988-01-01
The biosynthetic intermediates were incubated in a plant system. The activity levels calculated show that magnesium 6-acrylate porphyrins and one of the magnesium 6-β-hydroxypropionate porphyrins are not intermediates. In addition, plant systems incubated with 18 O 2 were found to synthesize magnesium 2,4-divinyl pheoporphyrin-a 5 incorporated with 18 O at the 9-carbonyl oxygen. Mass spectroscopy confirmed the presence of the oxygen label, thus eliminating one of two hypothesized pathways to chlorophyll-a. An overall description is given of iron porphyrins and iron porphyrin containing proteins. The function of the propionic side chains of the heme prosthetic group during electron transport reactions will be investigated. The synthesis of a series of iron(III) hexamethyl porphyrins with increasingly longer substituents in the remaining two peripheral positions of the porphyrin is described. Models for NMR studies of iron chlorin containing enzymes are discussed. Iron(III) pyropheophorbide-a and methyl pyropheophorbide-a were synthesized in addition to 5-CD 3 , 10-CD 2 iron(III) pyropheophorbide-a and methyl pyropheophorbide-a. Together, these pyropheophorbides were used to assign NMR resonances and ultimately provide a model for other iron chlorins. The synthesis of nickel(II) anhydro-mesorhodoporphyrin from zinc(III) anhydromesorhodochlorin is described; this nickel porphyrin was used as a standard for ring current calculations of reduced nickel analogs of anhydromesorhodoporphyrin
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New insights into the organization and regulation of trichothecene biosynthetic genes in Trichoderma
Collectively, species of the genus Trichoderma can produce numerous structurally diverse secondary metabolites (SM). This ability is conferred by the presence of SM biosynthetic gene clusters in their genomes. Species of Trichoderma in the Brevicompactum clade are able to produce trichothecenes, a f...
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Use of [75Se]selenomethionine in immunoglobulin biosynthetic studies
International Nuclear Information System (INIS)
Gutman, G.A.; Warner, N.L.; Harris, A.W.; Bowles, A.
1978-01-01
The gamma-emitting amino acid analog, [ 75 Se] selenomethionine, has been used as a biosynthetic label for immunoglobulins secreted by plasmacytomas in tissue culture. The secreted products are structurally intact with respect to their antibody combining sites and their class and allotype antigenic specificities. A component of [ 75 Se] selenomethionine preparations was found to bind to fetal calf serum proteins, in a manner releasable by mercaptoethanol, but not by sodium dodecyl sulfate and urea. Methods for circumventing the problems caused by this binding are described. (Auth.)
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Pearce, Stephen; Huttly, Alison K; Prosser, Ian M; Li, Yi-dan; Vaughan, Simon P; Gallova, Barbora; Patil, Archana; Coghill, Jane A; Dubcovsky, Jorge; Hedden, Peter; Phillips, Andrew L
2015-06-05
The gibberellin (GA) pathway plays a central role in the regulation of plant development, with the 2-oxoglutarate-dependent dioxygenases (2-ODDs: GA20ox, GA3ox, GA2ox) that catalyse the later steps in the biosynthetic pathway of particularly importance in regulating bioactive GA levels. Although GA has important impacts on crop yield and quality, our understanding of the regulation of GA biosynthesis during wheat and barley development remains limited. In this study we identified or assembled genes encoding the GA 2-ODDs of wheat, barley and Brachypodium distachyon and characterised the wheat genes by heterologous expression and transcript analysis. The wheat, barley and Brachypodium genomes each contain orthologous copies of the GA20ox, GA3ox and GA2ox genes identified in rice, with the exception of OsGA3ox1 and OsGA2ox5 which are absent in these species. Some additional paralogs of 2-ODD genes were identified: notably, a novel gene in the wheat B genome related to GA3ox2 was shown to encode a GA 1-oxidase, named as TaGA1ox-B1. This enzyme is likely to be responsible for the abundant 1β-hydroxylated GAs present in developing wheat grains. We also identified a related gene in barley, located in a syntenic position to TaGA1ox-B1, that encodes a GA 3,18-dihydroxylase which similarly accounts for the accumulation of unusual GAs in barley grains. Transcript analysis showed that some paralogs of the different classes of 2-ODD were expressed mainly in a single tissue or at specific developmental stages. In particular, TaGA20ox3, TaGA1ox1, TaGA3ox3 and TaGA2ox7 were predominantly expressed in developing grain. More detailed analysis of grain-specific gene expression showed that while the transcripts of biosynthetic genes were most abundant in the endosperm, genes encoding inactivation and signalling components were more highly expressed in the seed coat and pericarp. The comprehensive expression and functional characterisation of the multigene families encoding the 2-ODD
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Redox Impact on Starch Biosynthetic Enzymes in Arabidopsis thaliana
DEFF Research Database (Denmark)
Skryhan, Katsiaryna
Summary The thesis provides new insight into the influence of the plant cell redox state on the transient starch metabolism in Arabidopsis thaliana with a focus on starch biosynthetic enzymes. Two main hypotheses forms the basis of this thesis: 1) photosynthesis and starch metabolism are coordina......Summary The thesis provides new insight into the influence of the plant cell redox state on the transient starch metabolism in Arabidopsis thaliana with a focus on starch biosynthetic enzymes. Two main hypotheses forms the basis of this thesis: 1) photosynthesis and starch metabolism...... are coordinated by the redox state of the cell via post-translational modification of the starch metabolic enzymes containing redox active cysteine residues and these cysteine residues became cross-linked upon oxidation providing a conformational change leading to activity loss; 2) cysteine residues...... of chloroplast enzymes can play a role not only in enzyme activity and redox sensitivity but also in protein folding and stability upon oxidation. Several redox sensitive enzymes identified in this study can serve as potential targets to control the carbon flux to and from starch during the day and night...
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Directory of Open Access Journals (Sweden)
Ximena Escalera-Fanjul
2017-06-01
Full Text Available Gene duplication is one of the major evolutionary mechanisms providing raw material for the generation of genes with new or modified functions. The yeast Saccharomyces cerevisiae originated after an allopolyploidization event, which involved mating between two different ancestral yeast species. ScALT1 and ScALT2 codify proteins with 65% identity, which were proposed to be paralogous alanine transaminases. Further analysis of their physiological role showed that while ScALT1 encodes an alanine transaminase which constitutes the main pathway for alanine biosynthesis and the sole pathway for alanine catabolism, ScAlt2 does not display alanine transaminase activity and is not involved in alanine metabolism. Moreover, phylogenetic studies have suggested that ScALT1 and ScALT2 come from each one of the two parental strains which gave rise to the ancestral hybrid. The present work has been aimed to the understanding of the properties of the ancestral type Lacchancea kluyveri LkALT1 and Kluyveromyces lactis KlALT1, alanine transaminases in order to better understand the ScALT1 and ScALT2 evolutionary history. These ancestral -type species were chosen since they harbor ALT1 genes, which are related to ScALT2. Presented results show that, although LkALT1 and KlALT1 constitute ScALT1 orthologous genes, encoding alanine transaminases, both yeasts display LkAlt1 and KlAlt1 independent alanine transaminase activity and additional unidentified alanine biosynthetic and catabolic pathway(s. Furthermore, phenotypic analysis of null mutants uncovered the fact that KlAlt1 and LkAlt1 have an additional role, not related to alanine metabolism but is necessary to achieve wild type growth rate. Our study shows that the ancestral alanine transaminase function has been retained by the ScALT1 encoded enzyme, which has specialized its catabolic character, while losing the alanine independent role observed in the ancestral type enzymes. The fact that ScAlt2 conserves 64
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Hoang, Van L T; Innes, David J; Shaw, P Nicholas; Monteith, Gregory R; Gidley, Michael J; Dietzgen, Ralf G
2015-07-30
Mango fruits contain a broad spectrum of phenolic compounds which impart potential health benefits; their biosynthesis is catalysed by enzymes in the phenylpropanoid-flavonoid (PF) pathway. The aim of this study was to reveal the variability in genes involved in the PF pathway in three different mango varieties Mangifera indica L., a member of the family Anacardiaceae: Kensington Pride (KP), Irwin (IW) and Nam Doc Mai (NDM) and to determine associations with gene expression and mango flavonoid profiles. A close evolutionary relationship between mango genes and those from the woody species poplar of the Salicaceae family (Populus trichocarpa) and grape of the Vitaceae family (Vitis vinifera), was revealed through phylogenetic analysis of PF pathway genes. We discovered 145 SNPs in total within coding sequences with an average frequency of one SNP every 316 bp. Variety IW had the highest SNP frequency (one SNP every 258 bp) while KP and NDM had similar frequencies (one SNP every 369 bp and 360 bp, respectively). The position in the PF pathway appeared to influence the extent of genetic diversity of the encoded enzymes. The entry point enzymes phenylalanine lyase (PAL), cinnamate 4-mono-oxygenase (C4H) and chalcone synthase (CHS) had low levels of SNP diversity in their coding sequences, whereas anthocyanidin reductase (ANR) showed the highest SNP frequency followed by flavonoid 3'-hydroxylase (F3'H). Quantitative PCR revealed characteristic patterns of gene expression that differed between mango peel and flesh, and between varieties. The combination of mango expressed sequence tags and availability of well-established reference PF biosynthetic genes from other plant species allowed the identification of coding sequences of genes that may lead to the formation of important flavonoid compounds in mango fruits and facilitated characterisation of single nucleotide polymorphisms between varieties. We discovered an association between the extent of sequence variation and
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Analytical determination of thiamine (vitamin B1) in irradiated and stored Brazilian beans
International Nuclear Information System (INIS)
Villavicencio, Anna L.C.H.; Mancini-Filho, Jorge; Bognar, A.
1997-01-01
Thiamine (vitamin B 1 ) content in two varieties of Brazilian beans, Phaselus vulgaris L., var. carioca and Vigna unguiculata (L) Walp, var. macacar , irradiated with doses ranging from 0,05, 1.0, 2.5, 5.0 and 10.0 kGy was investigated. After a 6 months storage period, the optimum cooking time was established for each dose and variety. Sensorial evaluation tests were carried out by a panel of six people trained to this specific task. Our purpose to work with these beans is because conflicting results have appeared in studies about vitamin loss after low-dose irradiation. In our case, after a 6-month storage period of this two kinds of beans, in addition to the analysis of cooking time and sensory properties the vitamin B 1 content was evaluated. No significant vitamin losses were observed in Macacar beans until 10.0 kGy. Carioca beans showed small losses after 2.5 kGy. (author). 23 refs., 2 figs., 5 tabs
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Ren, Hengqian; Hu, Pingfan; Zhao, Huimin
2017-08-01
Pathway refactoring serves as an invaluable synthetic biology tool for natural product discovery, characterization, and engineering. However, the complicated and laborious molecular biology techniques largely hinder its application in natural product research, especially in a high-throughput manner. Here we report a plug-and-play pathway refactoring workflow for high-throughput, flexible pathway construction, and expression in both Escherichia coli and Saccharomyces cerevisiae. Biosynthetic genes were firstly cloned into pre-assembled helper plasmids with promoters and terminators, resulting in a series of expression cassettes. These expression cassettes were further assembled using Golden Gate reaction to generate fully refactored pathways. The inclusion of spacer plasmids in this system would not only increase the flexibility for refactoring pathways with different number of genes, but also facilitate gene deletion and replacement. As proof of concept, a total of 96 pathways for combinatorial carotenoid biosynthesis were built successfully. This workflow should be generally applicable to different classes of natural products produced by various organisms. Biotechnol. Bioeng. 2017;114: 1847-1854. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.
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Hudson, André O; Singh, Bijay K; Leustek, Thomas; Gilvarg, Charles
2006-01-01
Although lysine (Lys) biosynthesis in plants is known to occur by way of a pathway that utilizes diaminopimelic acid (DAP) as a central intermediate, the available evidence suggests that none of the known DAP-pathway variants found in nature occur in plants. A new Lys biosynthesis pathway has been identified in Arabidopsis (Arabidopsis thaliana) that utilizes a novel transaminase that specifically catalyzes the interconversion of tetrahydrodipicolinate and LL-diaminopimelate, a reaction requiring three enzymes in the DAP-pathway variant found in Escherichia coli. The LL-DAP aminotransferase encoded by locus At4g33680 was able to complement the dapD and dapE mutants of E. coli. This result, in conjunction with the kinetic properties and substrate specificity of the enzyme, indicated that LL-DAP aminotransferase functions in the Lys biosynthetic direction under in vivo conditions. Orthologs of At4g33680 were identified in all the cyanobacterial species whose genomes have been sequenced. The Synechocystis sp. ortholog encoded by locus sll0480 showed the same functional properties as At4g33680. These results demonstrate that the Lys biosynthesis pathway in plants and cyanobacteria is distinct from the pathways that have so far been defined in microorganisms.
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Directory of Open Access Journals (Sweden)
Shicheng Zhao
2014-10-01
Full Text Available Kenaf (Hibiscus cannabinus is cultivated worldwide for its fiber; however, the medicinal properties of this plant are currently attracting increasing attention. In this study, we investigated the expression levels of genes involved in the biosynthesis of kaempferitrin, a compound with many biological functions, in different kenaf organs. We found that phenylalanine ammonia lyase (HcPAL was more highly expressed in stems than in other organs. Expression levels of cinnamate 4-hydroxylase (HcC4H and 4-coumarate-CoA ligase (Hc4CL were highest in mature leaves, followed by stems and young leaves, and lowest in roots and mature flowers. The expression of chalcone synthase (HcCHS, chalcone isomerase (HcCHI, and flavone 3-hydroxylase (HcF3H was highest in young flowers, whereas that of flavone synthase (HcFLS was highest in leaves. An analysis of kaempferitrin accumulation in the different organs of kenaf revealed that the accumulation of this compound was considerably higher (>10-fold in leaves than in other organs. On the basis of a comparison of kaempferitrin contents with the expression levels of different genes in different organs, we speculate that HcFLS plays an important regulatory role in the kaempferitrin biosynthetic pathway in kenaf.
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Zhao, Shicheng; Li, Xiaohua; Cho, Dong Ha; Arasu, Mariadhas Valan; Al-Dhabi, Naif Abdullah; Park, Sang Un
2014-10-23
Kenaf (Hibiscus cannabinus) is cultivated worldwide for its fiber; however, the medicinal properties of this plant are currently attracting increasing attention. In this study, we investigated the expression levels of genes involved in the biosynthesis of kaempferitrin, a compound with many biological functions, in different kenaf organs. We found that phenylalanine ammonia lyase (HcPAL) was more highly expressed in stems than in other organs. Expression levels of cinnamate 4-hydroxylase (HcC4H) and 4-coumarate-CoA ligase (Hc4CL) were highest in mature leaves, followed by stems and young leaves, and lowest in roots and mature flowers. The expression of chalcone synthase (HcCHS), chalcone isomerase (HcCHI), and flavone 3-hydroxylase (HcF3H) was highest in young flowers, whereas that of flavone synthase (HcFLS) was highest in leaves. An analysis of kaempferitrin accumulation in the different organs of kenaf revealed that the accumulation of this compound was considerably higher (>10-fold) in leaves than in other organs. On the basis of a comparison of kaempferitrin contents with the expression levels of different genes in different organs, we speculate that HcFLS plays an important regulatory role in the kaempferitrin biosynthetic pathway in kenaf.
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Hadjipieri, Margarita; Georgiadou, Egli C; Marin, Alicia; Diaz-Mula, Huertas M; Goulas, Vlasios; Fotopoulos, Vasileios; Tomás-Barberán, Francisco A; Manganaris, George A
2017-06-14
Carotenoids are the main colouring substances found in orange-fleshed loquat fruits. The aim of this study was to unravel the carotenoid biosynthetic pathway of loquat fruit (cv. 'Obusa') in peel and flesh tissue during distinct on-tree developmental stages through a targeted analytical and molecular approach. Substantial changes regarding colour parameters, both between peel and flesh and among the different developmental stages, were monitored, concomitant with a significant increment in carotenoid content. Key genes and individual compounds that are implicated in the carotenoid biosynthetic pathway were further dissected with the employment of molecular (RT-qPCR) and advanced analytical techniques (LC-MS). Results revealed significant differences in carotenoid composition between peel and flesh. Thirty-two carotenoids were found in the peel, while only eighteen carotenoids were identified in the flesh. Trans-lutein and trans-β-carotene were the major carotenoids in the peel; the content of the former decreased with the progress of ripening, while the latter registered a 7.2-fold increase. However, carotenoid profiling of loquat flesh indicated trans-β-cryptoxanthin, followed by trans-β-carotene and 5,8-epoxy-β-carotene to be the most predominant carotenoids. High amounts of trans-β-carotene in both tissues were supported by significant induction in a chromoplast-specific lycopene β-cyclase (CYCB) transcript levels. PSY1, ZDS, CYCB and BCH were up-regulated and CRTISO, LCYE, ECH and VDE were down-regulated in most of the developmental stages compared with the immature stage in both peel and flesh tissue. Overall, differential regulation of expression levels with the progress of on-tree fruit development was more evident in the middle and downstream genes of carotenoid biosynthetic pathway. Carotenoid composition is greatly affected during on-tree loquat development with striking differences between peel and flesh tissue. A link between gene up- or down
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IMG-ABC: An Atlas of Biosynthetic Gene Clusters to Fuel the Discovery of Novel Secondary Metabolites
Energy Technology Data Exchange (ETDEWEB)
Chen, I-Min; Chu, Ken; Ratner, Anna; Palaniappan, Krishna; Huang, Jinghua; Reddy, T. B.K.; Cimermancic, Peter; Fischbach, Michael; Ivanova, Natalia; Markowitz, Victor; Kyrpides, Nikos; Pati, Amrita
2014-10-28
In the discovery of secondary metabolites (SMs), large-scale analysis of sequence data is a promising exploration path that remains largely underutilized due to the lack of relevant computational resources. We present IMG-ABC (https://img.jgi.doe.gov/abc/) -- An Atlas of Biosynthetic gene Clusters within the Integrated Microbial Genomes (IMG) system1. IMG-ABC is a rich repository of both validated and predicted biosynthetic clusters (BCs) in cultured isolates, single-cells and metagenomes linked with the SM chemicals they produce and enhanced with focused analysis tools within IMG. The underlying scalable framework enables traversal of phylogenetic dark matter and chemical structure space -- serving as a doorway to a new era in the discovery of novel molecules.
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Heterologous stable expression of terpenoid biosynthetic genes using the moss Physcomitrella patens
DEFF Research Database (Denmark)
Bach, Søren Spanner; King, Brian Christopher; Zhan, Xin
2014-01-01
Heterologous and stable expression of genes encoding terpenoid biosynthetic enzymes in planta is an important tool for functional characterization and is an attractive alternative to expression in microbial hosts for biotechnological production. Despite improvements to the procedure, such as stre...
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Phytochemical and Biosynthetic Studies of Lignans, with a Focus on Indonesian Medicinal Plants
Elfahmi, [No Value
2006-01-01
In this thesis phytochemical and biosynthetic studies of lignans are described. The focus is on the Indonesian medicinal plants Phyllanthus niruri and Piper cubeba and on two Linum species, Linum flavum and L. leonii, native to European countries. Both Indonesian plants are used in jamu. Jamu is the
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Foulon, V; Croes, K; Waelkens, E
1999-01-01
Purification, molecular cloning, and expression of 2-hydroxyphytanoyl- CoA lyase, a peroxisomal thiamine pyrophosphate-dependent enzyme that catalyzes the carbon-carbon bond cleavage during à-oxidation of 3- methyl-branched fatty acids
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Viral exploitation of the MEK/ERK pathway - A tale of vaccinia virus and other viruses.
Bonjardim, Cláudio A
2017-07-01
The VACV replication cycle is remarkable in the sense that it is performed entirely in the cytoplasmic compartment of vertebrate cells, due to its capability to encode enzymes required either for regulating the macromolecular precursor pool or the biosynthetic processes. Although remarkable, this gene repertoire is not sufficient to confer the status of a free-living microorganism to the virus, and, consequently, the virus relies heavily on the host to successfully generate its progeny. During the complex virus-host interaction, viruses must deal not only with the host pathways to accomplish their temporal demands but also with pathways that counteract viral infection, including the inflammatory, innate and acquired immune responses. This review focuses on VACV and other DNA or RNA viruses that stimulate the MEK (MAPK - Mitogen Activated Protein Kinase)/ERK- Extracellular signal-Regulated Kinase) pathway as part of their replication cycle. Copyright © 2016 Elsevier Inc. All rights reserved.
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Shu, Qunfeng; Xu, Meijuan; Li, Jing; Yang, Taowei; Zhang, Xian; Xu, Zhenghong; Rao, Zhiming
2018-05-04
L-Ornithine is a non-protein amino acid with extensive applications in the food and pharmaceutical industries. In this study, we performed metabolic pathway engineering of an L-arginine hyper-producing strain of Corynebacterium crenatum for L-ornithine production. First, we amplified the L-ornithine biosynthetic pathway flux by blocking the competing branch of the pathway. To enhance L-ornithine synthesis, we performed site-directed mutagenesis of the ornithine-binding sites to solve the problem of L-ornithine feedback inhibition for ornithine acetyltransferase. Alternatively, the genes argA from Escherichia coli and argE from Serratia marcescens, encoding the enzymes N-acetyl glutamate synthase and N-acetyl-L-ornithine deacetylase, respectively, were introduced into Corynebacterium crenatum to mimic the linear pathway of L-ornithine biosynthesis. Fermentation of the resulting strain in a 5-L bioreactor allowed a dramatically increased production of L-ornithine, 40.4 g/L, with an overall productivity of 0.673 g/L/h over 60 h. This demonstrates that an increased level of transacetylation is beneficial for L-ornithine biosynthesis.
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Zimmermann, Katrin; Engeser, Marianne; Blunt, John W; Munro, Murray H G; Piel, Jörn
2009-03-04
The complex polyketide pederin is a potent antitumor agent isolated from Paederus spp. rove beetles. We have previously isolated a set of genes from a bacterial endosymbiont that are good candidates for pederin biosynthesis. To biochemically study this pathway, we expressed three methyltransferases from the putative pederin pathway and used the partially unmethylated analogue mycalamide A from the marine sponge Mycale hentscheli as test substrate. Analysis by high-resolution MS/MS and NMR revealed that PedO regiospecifically methylates the marine compound to generate the nonnatural hybrid compound 18-O-methylmycalamide A with increased cytotoxicity. To our knowledge, this is the first biochemical evidence that invertebrates can obtain defensive complex polyketides from bacterial symbionts.
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Reiterative Recombination for the in vivo assembly of libraries of multigene pathways.
Wingler, Laura M; Cornish, Virginia W
2011-09-13
The increasing sophistication of synthetic biology is creating a demand for robust, broadly accessible methodology for constructing multigene pathways inside of the cell. Due to the difficulty of rationally designing pathways that function as desired in vivo, there is a further need to assemble libraries of pathways in parallel, in order to facilitate the combinatorial optimization of performance. While some in vitro DNA assembly methods can theoretically make libraries of pathways, these techniques are resource intensive and inherently require additional techniques to move the DNA back into cells. All previously reported in vivo assembly techniques have been low yielding, generating only tens to hundreds of constructs at a time. Here, we develop "Reiterative Recombination," a robust method for building multigene pathways directly in the yeast chromosome. Due to its use of endonuclease-induced homologous recombination in conjunction with recyclable markers, Reiterative Recombination provides a highly efficient, technically simple strategy for sequentially assembling an indefinite number of DNA constructs at a defined locus. In this work, we describe the design and construction of the first Reiterative Recombination system in Saccharomyces cerevisiae, and we show that it can be used to assemble multigene constructs. We further demonstrate that Reiterative Recombination can construct large mock libraries of at least 10(4) biosynthetic pathways. We anticipate that our system's simplicity and high efficiency will make it a broadly accessible technology for pathway construction and render it a valuable tool for optimizing pathways in vivo.
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DEFF Research Database (Denmark)
Andersen-Ranberg, Johan; Kongstad, Kenneth Thermann; Nafisi, Majse
2017-01-01
unsuccessful. Herein, a novel combination of enzymes derived from a plant (Aloe arborescens, Aa), a bacterium (Streptomyces sp. R1128, St), and an insect (Dactylopius coccus, Dc) that allows for the biosynthesis of the C-glucosylated anthraquinone, dcII, a precursor for carminic acid, is reported. The pathway...
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Ketol-acid reductoisomerase enzymes and methods of use
Govindarajan, Sridhar; Li, Yougen; Liao, Der-Ing; O'Keefe, Daniel P.; Minshull, Jeremy Stephen; Rothman, Steven Cary; Tobias, Alexander Vincent
2015-10-27
Provided herein are polypeptides having ketol-aid reductoisomerase activity as well as microbial host cells comprising such polypeptides. Polypeptides provided herein may be used in biosynthetic pathways, including, but not limited to, isobutanol biosynthetic pathways.
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The oxalic acid biosynthetic activity of Burkholderia mallei is encoded by a single locus
Although it is known that oxalic acid provides a selective advantage to the secreting microbe, our understanding of how this acid is biosynthesized remains incomplete. This study reports the identification, cloning, and partial characterization of the oxalic acid biosynthetic enzyme from the animal ...
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DEFF Research Database (Denmark)
Månsson, Maria; Vynne, Nikolaj Grønnegaard; Klitgaard, Andreas
2016-01-01
Microorganisms are a rich source of bioactives; however, chemical identification is a major bottleneck. Strategies that can prioritize the most prolific microbial strains and novel compounds are of great interest. Here, we present an integrated approach to evaluate the biosynthetic richness in ba...
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Rabatel, Andréane; Febvay, Gérard; Gaget, Karen; Duport, Gabrielle; Baa-Puyoulet, Patrice; Sapountzis, Panagiotis; Bendridi, Nadia; Rey, Marjolaine; Rahbé, Yvan; Charles, Hubert; Calevro, Federica; Colella, Stefano
2013-04-10
Nutritional symbioses play a central role in insects' adaptation to specialized diets and in their evolutionary success. The obligatory symbiosis between the pea aphid, Acyrthosiphon pisum, and the bacterium, Buchnera aphidicola, is no exception as it enables this important agricultural pest insect to develop on a diet exclusively based on plant phloem sap. The symbiotic bacteria provide the host with essential amino acids lacking in its diet but necessary for the rapid embryonic growth seen in the parthenogenetic viviparous reproduction of aphids. The aphid furnishes, in exchange, non-essential amino acids and other important metabolites. Understanding the regulations acting on this integrated metabolic system during the development of this insect is essential in elucidating aphid biology. We used a microarray-based approach to analyse gene expression in the late embryonic and the early larval stages of the pea aphid, characterizing, for the first time, the transcriptional profiles in these developmental phases. Our analyses allowed us to identify key genes in the phenylalanine, tyrosine and dopamine pathways and we identified ACYPI004243, one of the four genes encoding for the aspartate transaminase (E.C. 2.6.1.1), as specifically regulated during development. Indeed, the tyrosine biosynthetic pathway is crucial for the symbiotic metabolism as it is shared between the two partners, all the precursors being produced by B. aphidicola. Our microarray data are supported by HPLC amino acid analyses demonstrating an accumulation of tyrosine at the same developmental stages, with an up-regulation of the tyrosine biosynthetic genes. Tyrosine is also essential for the synthesis of cuticular proteins and it is an important precursor for cuticle maturation: together with the up-regulation of tyrosine biosynthesis, we observed an up-regulation of cuticular genes expression. We were also able to identify some amino acid transporter genes which are essential for the switch
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Directory of Open Access Journals (Sweden)
Gladiol Zenunaj
Full Text Available Introduction: Biosynthetic prosthesis has become the trend to carry out arterial reconstruction in infected sites since considered to be resistant to infection. Late graft occlusion is the only complication reported in literature so far. We report a case of biosynthetic graft infection which led to early detachment of the femoral anastomosis of a femoral-popliteal above-knee bypass. Material: A 76-year-old man developed groin infection 3 months later after performing an ePTFE femoral-popliteal above-knee bypass for critical limb ischemia. He was re-admitted for groin infection involving the vascular structures. Explantation of the existing bypass and its replacement with a biosynthetic graft (omniflow II was performed. Detachment of the proximal anastomosis occurred 6 days later leading to groin haematoma. Consequently, retroperitoneal access was performed for clamping the external iliac artery so as to control haemorrhage followed by explantation of the biosynthetic graft. An external iliac-popliteal above-knee bypass was tailored in order to save the limb and it was performed using a transobturator approach avoiding the infected site. In both cases bacterial cultures resulted positive for Morganella Morganii. The groin wound was treated separately with negative pressure medication healing definitively within 20 days and after 3-month follow-up the bypass was still patent. Conclusion: This is the first report of biosynthetic graft infection used for infrainguinal reconstruction leading to haemorrhage due to anastomosis disrupture. Using an extra-anatomical access for providing blood inflow to the leg avoiding the infected site and treating safely the groin wound with VAC therapy revealed to be a valid approach. Keywords: Infrainguinal bypass, Graft infection, Biosynthetic material, Graft occlusion, Negative pressure medication, Morganella morgani
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Wang, Yaya; Deng, Zixin; Qu, Xudong
2014-01-01
Fluorination has been widely used in chemical synthesis, but is rare in nature. The only known biological fluorination scope is represented by the fl pathway from Streptomyces cattleya that produces fluoroacetate (FAc) and 4-fluorothreonine (4-FT). Here we report the identification of a novel pathway for FAc and 4-FT biosynthesis from the actinomycetoma-causing pathogen Nocardia brasiliensis ATCC 700358. The new pathway shares overall conservation with the fl pathway in S. cattleya. Biochemical characterization of the conserved domains revealed a novel fluorinase NobA that can biosynthesize 5'-fluoro-5'-deoxyadenosine (5'-FDA) from inorganic fluoride and S-adenosyl-l-methionine (SAM). The NobA shows similar halide specificity and characteristics to the fluorination enzyme FlA of the fl pathway. Kinetic parameters for fluoride ( K m 4153 μM, k cat 0.073 min (-1)) and SAM ( K m 416 μM, k cat 0.139 min (-1)) have been determined, revealing that NobA is slightly (2.3 fold) slower than FlA. Upon sequence comparison, we finally identified a distinct loop region in the fluorinases that probably accounts for the disparity of fluorination activity.
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Guo, Qianqian; Ma, Xiaojun; Wei, Shugen; Qiu, Deyou; Wilson, Iain W; Wu, Peng; Tang, Qi; Liu, Lijun; Dong, Shoukun; Zu, Wei
2014-08-12
The major medicinal alkaloids isolated from Uncaria rhynchophylla (gouteng in chinese) capsules are rhynchophylline (RIN) and isorhynchophylline (IRN). Extracts containing these terpene indole alkaloids (TIAs) can inhibit the formation and destabilize preformed fibrils of amyloid β protein (a pathological marker of Alzheimer's disease), and have been shown to improve the cognitive function of mice with Alzheimer-like symptoms. The biosynthetic pathways of RIN and IRN are largely unknown. In this study, RNA-sequencing of pooled Uncaria capsules RNA samples taken at three developmental stages that accumulate different amount of RIN and IRN was performed. More than 50 million high-quality reads from a cDNA library were generated and de novo assembled. Sequences for all of the known enzymes involved in TIAs synthesis were identified. Additionally, 193 cytochrome P450 (CYP450), 280 methyltransferase and 144 isomerase genes were identified, that are potential candidates for enzymes involved in RIN and IRN synthesis. Digital gene expression profile (DGE) analysis was performed on the three capsule developmental stages, and based on genes possessing expression profiles consistent with RIN and IRN levels; four CYP450s, three methyltransferases and three isomerases were identified as the candidates most likely to be involved in the later steps of RIN and IRN biosynthesis. A combination of de novo transcriptome assembly and DGE analysis was shown to be a powerful method for identifying genes encoding enzymes potentially involved in the biosynthesis of important secondary metabolites in a non-model plant. The transcriptome data from this study provides an important resource for understanding the formation of major bioactive constituents in the capsule extract from Uncaria, and provides information that may aid in metabolic engineering to increase yields of these important alkaloids.
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Pandey, Ramesh Prasad; Parajuli, Prakash; Koffas, Mattheos A G; Sohng, Jae Kyung
2016-01-01
In this review, we address recent advances made in pathway engineering, directed evolution, and systems/synthetic biology approaches employed in the production and modification of flavonoids from microbial cells. The review is divided into two major parts. In the first, various metabolic engineering and system/synthetic biology approaches used for production of flavonoids and derivatives are discussed broadly. All the manipulations/engineering accomplished on the microorganisms since 2000 are described in detail along with the biosynthetic pathway enzymes, their sources, structures of the compounds, and yield of each product. In the second part of the review, post-modifications of flavonoids by four major reactions, namely glycosylations, methylations, hydroxylations and prenylations using recombinant strains are described. Copyright © 2016 Elsevier Inc. All rights reserved.
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Use of (/sup 75/Se)selenomethionine in immunoglobulin biosynthetic studies
Energy Technology Data Exchange (ETDEWEB)
Gutman, G A; Warner, N L; Harris, A W; Bowles, A [Walter and Elisa Hall Institute of Medical Research, Victoria (Australia). Genetics Unit; Royal Melbourne Hospital, Victoria (Australia))
1978-05-01
The gamma-emitting amino acid analog, (/sup 75/Se) selenomethionine, has been used as a biosynthetic label for immunoglobulins secreted by plasmacytomas in tissue culture. The secreted products are structurally intact with respect to their antibody combining sites and their class and allotype antigenic specificities. A component of (/sup 75/Se) selenomethionine preparations was found to bind to fetal calf serum proteins, in a manner releasable by mercaptoethanol, but not by sodium dodecyl sulfate and urea. Methods for circumventing the problems caused by this binding are described.
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Directory of Open Access Journals (Sweden)
Carlos Eugenio Daudt
1995-01-01
Full Text Available O objetivo do trabalho foi estudar o comportamento das vitaminas,tiamina (vitamina B1 e riboflavina (vitamina B2,durante a maturação de Vitis vinifera cultivar Cabemet Sauvignon e durante a fermentação do mosto, usando diferentes quantidades de SO2. Os valores mínimos encontrados para tiamina e riboflavina foram respectivamente (ug/100nd: 7,67 e 6,85 com 5,5 °Brix e os máximos foram, respectivamente, 19,35 (na colheita e 15,75 com 12,8 °Brix. A tiamina foi quase totalmente consumida antes da fermentação inicial, aparecendo novamente ao final da mesma. A riboflavina, ao contrário aumentou durante a fermentação. Ambas as vitaminas aumentaram durante a maturação de Cabemet Sauvignon, estando a tiamina presente em maior quantidade. O aumento das quantidades de SO2 adicionado ao mosto, afetou a tiamina numa proporção direta, mas não teve nenhum efeito sobre a riboflavina. A tiamina diminuiu durante a fermentação, ao mesmo tempo em que ocorreu um aumento da riboflavina.The aim of the work was to follow the behaviour of both vitamins, thiamine and riboflavin, during ripening of Vitis vinifera Cabemet Sauvignon and during fermentation of the must using different amounts of SO2. Minimum values for thiamine and riboflavin were, respectivelly, (ug/100ml: 7.67 and 6.85 at 5.5 °Brix and the maximum values were, respectivelly, 19.35 (at harvest and 15.75 at 12.87 °Brix. Thiamine was almost completelly consumed before the start of fermentation showing up again at the end of the process; riboflavin, on the contrary, increased during fermentation. Both vitamins increased during ripening of Cabemet Sauvignon but -in grapes- the amount of thiamine was greater than riboflavin. Fermentation procedures, as increasing levels of SO2, affected thiamine in a direct proportion but did not have any effect upon riboflavin. Thiamine decreased during fermentation and riboflavin increased.
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Production of manganese peroxidase by white rot fungi from potato ...
African Journals Online (AJOL)
phenylalanine and L-tyrosine blocked the MnP biosynthetic pathway. Ammonium ion released from the L-phenylalanine by the L-phenylalanine ammonia-lyase participated in the repression of the MnP biosynthetic pathway of the strain L-25.
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Wang, Cheng; Zeng, Jian; Li, Yin; Hu, Wei; Chen, Ling; Miao, Yingjie; Deng, Pengyi; Yuan, Cuihong; Ma, Cheng; Chen, Xi; Zang, Mingli; Wang, Qiong; Li, Kexiu; Chang, Junli; Wang, Yuesheng; Yang, Guangxiao; He, Guangyuan
2014-06-01
Carotenoid content is a primary determinant of wheat nutritional value and affects its end-use quality. Wheat grains contain very low carotenoid levels and trace amounts of provitamin A content. In order to enrich the carotenoid content in wheat grains, the bacterial phytoene synthase gene (CrtB) and carotene desaturase gene (CrtI) were transformed into the common wheat cultivar Bobwhite. Expression of CrtB or CrtI alone slightly increased the carotenoid content in the grains of transgenic wheat, while co-expression of both genes resulted in a darker red/yellow grain phenotype, accompanied by a total carotenoid content increase of approximately 8-fold achieving 4.76 μg g(-1) of seed dry weight, a β-carotene increase of 65-fold to 3.21 μg g(-1) of seed dry weight, and a provitamin A content (sum of α-carotene, β-carotene, and β-cryptoxanthin) increase of 76-fold to 3.82 μg g(-1) of seed dry weight. The high provitamin A content in the transgenic wheat was stably inherited over four generations. Quantitative PCR analysis revealed that enhancement of provitamin A content in transgenic wheat was also a result of the highly coordinated regulation of endogenous carotenoid biosynthetic genes, suggesting a metabolic feedback regulation in the wheat carotenoid biosynthetic pathway. These transgenic wheat lines are not only valuable for breeding wheat varieties with nutritional benefits for human health but also for understanding the mechanism regulating carotenoid biosynthesis in wheat endosperm. © The Author 2014. Published by Oxford University Press on behalf of the Society for Experimental Biology.
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Directory of Open Access Journals (Sweden)
Yun Ji Park
2016-05-01
Full Text Available Valeriana fauriei (V. fauriei, which emits a characteristic and unpleasant odor, is important in traditional medicine. In this study, the expression of terpenoid biosynthetic genes was investigated in different organs that were also screened for volatile compounds including valerenic acid and its derivatives. Specific expression patterns from different parts of V. fauriei were observed using quantitative real-time PCR (qRT-PCR. The highest transcript levels of biosynthetic genes involved in mevalonic acid (MVA and methylerythritol phosphate (MEP production were found in the stem. Although the amounts of volatile compounds were varied by organ, most of the volatile terpenoids were accumulated in the root. Gas chromatography mass spectrometry (GC-MS analysis identified 128 volatile compounds, which represented 65.33% to 95.66% of total volatiles. Certain compounds were only found in specific organs. For example, isovalerenic acid and valerenic acid and its derivatives were restricted to the root. Organs with high transcript levels did not necessarily have high levels of the corresponding chemical constituents. According to these results, we hypothesize that translocation may occur between different organs in V. fauriei.
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Shaw-Reid, C A; McCormick, M M; Sinskey, A J; Stephanopoulos, G
1999-03-01
The N-succinyl-LL-diaminopimelate desuccinylase gene (dapE) in the four-step succinylase branch of the L-lysine biosynthetic pathway of Corynebacterium glutamicum was disrupted via marker-exchange mutagenesis to create a mutant strain that uses only the one-step meso-diaminopimelate dehydrogenase branch to overproduce lysine. This mutant strain grew and utilized glucose from minimal medium at the same rate as the parental strain. In addition, the dapE- strain produced lysine at the same rate as its parent strain. Transformation of the parental and dapE- strains with the amplified meso-diaminopimelate dehydrogenase gene (ddh) on a plasmid did not affect lysine production in either strain, despite an eightfold amplification of the activity of the enzyme. These results indicate that the four-step succinylase pathway is dispensable for lysine overproduction in shake-flask culture. In addition, the one-step meso-diaminopimelate dehydrogenase pathway does not limit lysine flux in Corynebacterium under these conditions.
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Role of Heme and Heme-Proteins in Trypanosomatid Essential Metabolic Pathways
Directory of Open Access Journals (Sweden)
Karina E. J. Tripodi
2011-01-01
Full Text Available Around the world, trypanosomatids are known for being etiological agents of several highly disabling and often fatal diseases like Chagas disease (Trypanosoma cruzi, leishmaniasis (Leishmania spp., and African trypanosomiasis (Trypanosoma brucei. Throughout their life cycle, they must cope with diverse environmental conditions, and the mechanisms involved in these processes are crucial for their survival. In this review, we describe the role of heme in several essential metabolic pathways of these protozoans. Notwithstanding trypanosomatids lack of the complete heme biosynthetic pathway, we focus our discussion in the metabolic role played for important heme-proteins, like cytochromes. Although several genes for different types of cytochromes, involved in mitochondrial respiration, polyunsaturated fatty acid metabolism, and sterol biosynthesis, are annotated at the Tritryp Genome Project, the encoded proteins have not yet been deeply studied. We pointed our attention into relevant aspects of these protein functions that are amenable to be considered for rational design of trypanocidal agents.
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International Nuclear Information System (INIS)
Klein, O.; Senger, H.
1978-01-01
The X-ray induced mutant C-2A' of Scenedesmus obliquus grows heterotrophically but forms only traces of chlorophyll in the dark. Upon illumination, delta-aminolevulinic acid (ALA) is synthesized and chlorophyll is formed. These processes are blue light dependent and ceased immediately when the cells were transferred back into darkness. Addition of levulinic acid (LA) inhibited the light-dependent formation of chlorophyll and caused accumulation of ALA by competitive inhibition of the ALA dehydratase (EC. 4.2.1.24). By feeding specifically labelled 14 C precursors to the pigment mutant, inhibiting the ALA dehydratase with LA, accumulating, extracting and analyzing the ALA, two pathways leading towards ALA could be established: glycine and succinyl CoA can be condensed to ALA and the 5 carbon skeleton of glutamate can completely be incorporated into ALA via a second pathway. The glycine-succinyl CoA pathway dominated over the glutamate pathway, but both led to chlorophyll formation. (author)
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El Khattabi, Mohamed; van Roosmalen, Maarten L.; Jager, Dennis; Metselaar, Heidi; Permentier, Hjalmar; Leenhouts, Kees; Broos, Jaap
2008-01-01
Incorporation of Trp (tryptophan) analogues into a protein may facilitate its structural analysis by spectroscopic techniques. Development of a biological system for the biosynthetic incorporation of such analogues into proteins is of considerable importance. The Gram-negative Escherichia coli is
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How to prove the existence of metabolons?
DEFF Research Database (Denmark)
Bassard, Jean-Étienne André; Halkier, Barbara Ann
2017-01-01
Sequential enzymes in biosynthetic pathways are organized in metabolons. It is challenging to provide experimental evidence for the existence of metabolons as biosynthetic pathways are composed of highly dynamic protein–protein interactions. Many different methods are being applied, each with str...
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Biosynthetic Studies on Water-Soluble Derivative 5c (DTX5c
Directory of Open Access Journals (Sweden)
José J. Fernández
2012-10-01
Full Text Available The dinoflagellate Prorocentrum belizeanum is responsible for the production of several toxins involved in the red tide phenomenon known as Diarrhetic Shellfish Poisoning (DSP. In this paper we report on the biosynthetic origin of an okadaic acid water-soluble ester derivative, DTX5c, on the basis of the spectroscopical analysis of 13C enriched samples obtained by addition of labelled sodium [l-13C], [2-13C] acetate to artificial cultures of this dinoflagellate.
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Hadjithomas, Michalis; Chen, I-Min Amy; Chu, Ken; Ratner, Anna; Palaniappan, Krishna; Szeto, Ernest; Huang, Jinghua; Reddy, T B K; Cimermančič, Peter; Fischbach, Michael A; Ivanova, Natalia N; Markowitz, Victor M; Kyrpides, Nikos C; Pati, Amrita
2015-07-14
In the discovery of secondary metabolites, analysis of sequence data is a promising exploration path that remains largely underutilized due to the lack of computational platforms that enable such a systematic approach on a large scale. In this work, we present IMG-ABC (https://img.jgi.doe.gov/abc), an atlas of biosynthetic gene clusters within the Integrated Microbial Genomes (IMG) system, which is aimed at harnessing the power of "big" genomic data for discovering small molecules. IMG-ABC relies on IMG's comprehensive integrated structural and functional genomic data for the analysis of biosynthetic gene clusters (BCs) and associated secondary metabolites (SMs). SMs and BCs serve as the two main classes of objects in IMG-ABC, each with a rich collection of attributes. A unique feature of IMG-ABC is the incorporation of both experimentally validated and computationally predicted BCs in genomes as well as metagenomes, thus identifying BCs in uncultured populations and rare taxa. We demonstrate the strength of IMG-ABC's focused integrated analysis tools in enabling the exploration of microbial secondary metabolism on a global scale, through the discovery of phenazine-producing clusters for the first time in Alphaproteobacteria. IMG-ABC strives to fill the long-existent void of resources for computational exploration of the secondary metabolism universe; its underlying scalable framework enables traversal of uncovered phylogenetic and chemical structure space, serving as a doorway to a new era in the discovery of novel molecules. IMG-ABC is the largest publicly available database of predicted and experimental biosynthetic gene clusters and the secondary metabolites they produce. The system also includes powerful search and analysis tools that are integrated with IMG's extensive genomic/metagenomic data and analysis tool kits. As new research on biosynthetic gene clusters and secondary metabolites is published and more genomes are sequenced, IMG-ABC will continue to
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Biochemical characterization of GDP-L-fucose de novo synthesis pathway in fungus Mortierella alpina
International Nuclear Information System (INIS)
Ren, Yan; Perepelov, Andrei V.; Wang, Haiyan; Zhang, Hao; Knirel, Yuriy A.; Wang, Lei; Chen, Wei
2010-01-01
Mortierella alpina is a filamentous fungus commonly found in soil, which is able to produce large amount of polyunsaturated fatty acids. L-Fucose is an important sugar found in a diverse range of organisms, playing a variety of biological roles. In this study, we characterized the de novo biosynthetic pathway of GDP-L-fucose (the nucleotide-activated form of L-fucose) in M. alpina. Genes encoding GDP-D-mannose 4,6-dehydratase (GMD) and GDP-keto-6-deoxymannose 3,5-epimerase/4-reductase (GMER) were expressed heterologously in Escherichia coli. The recombinant enzymes were produced as His-tagged fusion proteins. Conversion of GDP-mannose to GDP-4-keto-6-deoxy mannose by GMD and GDP-4-keto-6-deoxy mannose to GDP-L-fucose by GMER were analyzed by capillary electrophoresis, electro-spray ionization-mass spectrometry, and nuclear magnetic resonance spectroscopy. The k m values of GMD for GDP-mannose and GMER for GDP-4-keto-6-deoxy mannose were determined to be 0.77 mM and 1.047 mM, respectively. Both NADH and NADPH may be used by GMER as the coenzyme. The optimum temperature and pH were determined to be 37 o C and pH 9.0 (GMD) or pH 7.0 (GMER). Divalent cations are not required for GMD and GMER activity, and the activities of both enzymes may be enhanced by DTT. To our knowledge this is the first report on the characterization of GDP-L-fucose biosynthetic pathway in fungi.
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Current therapeutic interventions in the glycation pathway: evidence from clinical studies.
Engelen, L; Stehouwer, C D A; Schalkwijk, C G
2013-08-01
The increased formation of advanced glycation endproducts (AGEs) constitutes a potential mechanism of hyperglycaemia-induced micro- and macrovascular disease in diabetes. In vitro and animal experiments have shown that various interventions can inhibit formation and/or actions of AGEs, in particular the specific AGE inhibitor aminoguanidine and the AGEs crosslink breaker alagebrium, and the B vitamins pyridoxamine and thiamine, and the latter's synthetic derivative, benfotiamine. The potential clinical value of these interventions, however, remains to be established. The present review provides, from the clinical point of view, an overview of current evidence on interventions in the glycation pathway relating to (i) the clinical benefits of specific AGE inhibitors and AGE breakers and (ii) the potential AGE-inhibiting effects of therapies developed for purposes unrelated to the glycation pathway. We found that safety and/or efficacy in clinical studies with the specific AGE inhibitor, aminoguanidine and the AGE breaker, alagebrium, appeared to be a concern. The clinical evidence on the potential AGE-inhibiting effects of B vitamins is still limited. Finally, current evidence for AGE inhibition by therapies developed for purposes unrelated to glycation is limited due to a large heterogeneity in study designs and/or measurement techniques, which have often been sub-optimal. We conclude that, clinical evidence on interventions to inhibit formation and/or action of AGEs is currently weak and unconvincing. © 2012 Blackwell Publishing Ltd.
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Jiao, Jiao; Gai, Qing-Yan; Wang, Wei; Luo, Meng; Gu, Cheng-Bo; Fu, Yu-Jie; Ma, Wei
2015-09-23
In this work, Astragalus membranaceus hairy root cultures (AMHRCs) were exposed to ultraviolet radiation (UV-A, UV-B, and UV-C) for promoting isoflavonoid accumulation. The optimum enhancement for isoflavonoid production was achieved in 34-day-old AMHRCs elicited by 86.4 kJ/m(2) of UV-B. The resulting isoflavonoid yield was 533.54 ± 13.61 μg/g dry weight (DW), which was 2.29-fold higher relative to control (232.93 ± 3.08 μg/g DW). UV-B up-regulated the transcriptional expressions of all investigated genes involved in isoflavonoid biosynthetic pathway. PAL and C4H were found to be two potential key genes that controlled isoflavonoid biosynthesis. Moreover, a significant increase was noted in antioxidant activity of extracts from UV-B-elicited AMHRCs (IC50 values = 0.85 and 1.08 mg/mL) in comparison with control (1.38 and 1.71 mg/mL). Overall, this study offered a feasible elicitation strategy to enhance isoflavonoid accumulation in AMHRCs and also provided a basis for metabolic engineering of isoflavonoid biosynthesis in the future.
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Bernstein, Jeffrey R; Bulter, Thomas; Liao, James C
2008-01-01
This work demonstrates the transfer of the five-gene cyclohexane carboxylate (CHC) degradation pathway from the high-GC alphaproteobacterium Rhodopseudomonas palustris to Escherichia coli, a gammaproteobacterium. The degradation product of this pathway is pimeloyl-CoA, a key metabolite in E. coli's biotin biosynthetic pathway. This pathway is useful for biotin overproduction in E. coli; however, the expression of GC-rich genes is troublesome in this host. When the native R. palustris CHC degradation pathway is transferred to a DeltabioH pimeloyl-CoA auxotroph of E. coli, it is unable to complement growth in the presence of CHC. To overcome this expression problem we redesigned the operon with decreased GC content and removed stretches of high-GC intergenic DNA which comprise the 5' untranslated region of each gene, replacing these features with shorter low-GC sequences. We show this synthetic construct enables growth of the DeltabioH strain in the presence of CHC. When the synthetic degradation pathway is overexpressed in conjunction with the downstream genes for biotin biosynthesis, we measured significant accumulation of biotin in the growth medium, showing that the pathway transfer is successfully integrated with the host metabolism.
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Liu, Yong; Wei, Wen-Ping; Ye, Bang-Ce
2018-05-18
The overexpression of bacterial secondary metabolite biosynthetic enzymes is the basis for industrial overproducing strains. Genome editing tools can be used to further improve gene expression and yield. Saccharopolyspora erythraea produces erythromycin, which has extensive clinical applications. In this study, the CRISPR-Cas9 system was used to edit genes in the S. erythraea genome. A temperature-sensitive plasmid containing the PermE promoter, to drive Cas9 expression, and the Pj23119 and PkasO promoters, to drive sgRNAs, was designed. Erythromycin esterase, encoded by S. erythraea SACE_1765, inactivates erythromycin by hydrolyzing the macrolactone ring. Sequencing and qRT-PCR confirmed that reporter genes were successfully inserted into the SACE_1765 gene. Deletion of SACE_1765 in a high-producing strain resulted in a 12.7% increase in erythromycin levels. Subsequent PermE- egfp knock-in at the SACE_0712 locus resulted in an 80.3% increase in erythromycin production compared with that of wild type. Further investigation showed that PermE promoter knock-in activated the erythromycin biosynthetic gene clusters at the SACE_0712 locus. Additionally, deletion of indA (SACE_1229) using dual sgRNA targeting without markers increased the editing efficiency to 65%. In summary, we have successfully applied Cas9-based genome editing to a bacterial strain, S. erythraea, with a high GC content. This system has potential application for both genome-editing and biosynthetic gene cluster activation in Actinobacteria.
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Characterization of the fumonisin B2 biosynthetic gene cluster in Aspergillus niger and A. awamori.
Aspergillus niger and A. awamori strains isolated from grapes cultivated in Mediterranean basin were examined for fumonisin B2 (FB2) production and presence/absence of sequences within the fumonisin biosynthetic gene (fum) cluster. Presence of 13 regions in the fum cluster was evaluated by PCR assay...
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Extending the biosynthetic repertoires of cyanobacteria and chloroplasts
DEFF Research Database (Denmark)
Nielsen, Agnieszka Janina Zygadlo; Mellor, Silas Busck; Vavitsas, Konstantinos
2016-01-01
The chloroplasts found in plants and algae, and photosynthetic microorganisms such as cyanobacteria, are emerging hosts for sustainable production of valuable biochemicals, using only inorganic nutrients, water, CO2 and light as inputs. In the past decade, many bioengineering efforts have focused...... on metabolic engineering and synthetic biology in the chloroplast or in cyanobacteria for the production of fuels, chemicals, as well as complex, high-value bioactive molecules. Biosynthesis of all these compounds can be performed in photosynthetic organelles/organisms by heterologous expression...... of chloroplasts and cyanobacteria as biosynthetic compartments and hosts, and we estimate the production levels to be expected from photosynthetic hosts in light of the fraction of electrons and carbon that can potentially be diverted from photosynthesis. The supply of reducing power, in the form of electrons...
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Mei, Xianyi; Yan, Xiaoli; Zhang, Hui; Yu, Mingjia; Shen, Guangqing; Zhou, Linjun; Deng, Zixin; Lei, Chun; Qu, Xudong
2018-01-19
Anthrabenzoxocinones (ABXs) including (-)-ABXs and (+)-ABXs are a group of bacterial FabF-specific inhibitors with potent antimicrobial activity of resistant strains. Optimization of their chemical structures is a promising method to develop potent antibiotics. Through biosynthetic investigation, we herein identified and characterized two highly promiscuous enzymes involved in the (-)-ABX structural modification. The promiscuous halogenase and methyltransferase can respectively introduce halogen-modifications into various positions of the ABX scaffolds and methylation to highly diverse substrates. Manipulation of their activity in both of the (-)-ABXs and (+)-ABXs biosyntheses led to the generation of 14 novel ABX analogues of both enantiomers. Bioactivity assessment revealed that a few of the analogues showed significantly improved antimicrobial activity, with the C3-hydroxyl and chlorine substitutions critical for their activity. This study enormously expands the bioactive chemical space of the ABX family and FabF-specific inhibitors. The disclosed broad-selective biosynthetic machineries and structure-activity relationship provide a solid basis for further generation of potent antimicrobial agents.
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An Improved in Vivo Deuterium Labeling Method for Measuring the Biosynthetic Rate of Cytokinins
Directory of Open Access Journals (Sweden)
Petr Tarkowski
2010-12-01
Full Text Available An improved method for determining the relative biosynthetic rate of isoprenoid cytokinins has been developed. A set of 11 relevant isoprenoid cytokinins, including zeatin isomers, was separated by ultra performance liquid chromatography in less than 6 min. The iP-type cytokinins were observed to give rise to a previously-unknown fragment at m/z 69; we suggest that the diagnostic (204-69 transition can be used to monitor the biosynthetic rate of isopentenyladenine. Furthermore, we found that by treating the cytokinin nucleotides with alkaline phosphatase prior to analysis, the sensitivity of the detection process could be increased. In addition, derivatization (propionylation improved the ESI-MS response by increasing the analytes' hydrophobicity. Indeed, the ESI-MS response of propionylated isopentenyladenosine was about 34% higher than that of its underivatized counterpart. Moreover, the response of the derivatized zeatin ribosides was about 75% higher than that of underivatized zeatin ribosides. Finally, we created a web-based calculator (IZOTOP that facilitates MS/MS data processing and offer it freely to the research community.
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Controlling cell-free metabolism through physiochemical perturbations.
Karim, Ashty S; Heggestad, Jacob T; Crowe, Samantha A; Jewett, Michael C
2018-01-01
Building biosynthetic pathways and engineering metabolic reactions in cells can be time-consuming due to complexities in cellular metabolism. These complexities often convolute the combinatorial testing of biosynthetic pathway designs needed to define an optimal biosynthetic system. To simplify the optimization of biosynthetic systems, we recently reported a new cell-free framework for pathway construction and testing. In this framework, multiple crude-cell extracts are selectively enriched with individual pathway enzymes, which are then mixed to construct full biosynthetic pathways on the time scale of a day. This rapid approach to building pathways aids in the study of metabolic pathway performance by providing a unique freedom of design to modify and control biological systems for both fundamental and applied biotechnology. The goal of this work was to demonstrate the ability to probe biosynthetic pathway performance in our cell-free framework by perturbing physiochemical conditions, using n-butanol synthesis as a model. We carried out three unique case studies. First, we demonstrated the power of our cell-free approach to maximize biosynthesis yields by mapping physiochemical landscapes using a robotic liquid-handler. This allowed us to determine that NAD and CoA are the most important factors that govern cell-free n-butanol metabolism. Second, we compared metabolic profile differences between two different approaches for building pathways from enriched lysates, heterologous expression and cell-free protein synthesis. We discover that phosphate from PEP utilization, along with other physiochemical reagents, during cell-free protein synthesis-coupled, crude-lysate metabolic system operation inhibits optimal cell-free n-butanol metabolism. Third, we show that non-phosphorylated secondary energy substrates can be used to fuel cell-free protein synthesis and n-butanol biosynthesis. Taken together, our work highlights the ease of using cell-free systems to explore
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International Nuclear Information System (INIS)
Ozkaya, B.; Koksel, H.; Ozkaya, H.; Tutluer, H.
1994-01-01
Effects of gamma irradiation at the doses of 0.25,0.50,0.75,1.00 and 1.25 kGy on two flour samples with distinct technological properties was investigated in this study.The sedimentation values of the irradiated samples showed a steady decrease but falling number and wet gluten content did not change to an important extend as the radiation level increased. Thiamine and riboflavin contents also decreased with irradiation. Dough development time and stability values decreased but no apparent changes were observed in other farinogram and extensogram parameters as radiation levels increased.The baking quality of the samples irradiated at higher doses(over 1.00 kGy) deteriorated in both flours as radiation levels increased
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Guntaka, Naga Sandhya; Healy, Alan R; Crawford, Jason M; Herzon, Seth B; Bruner, Steven D
2017-10-20
Colibactin is a genotoxic hybrid nonribosomal peptide/polyketide secondary metabolite produced by various pathogenic and probiotic bacteria residing in the human gut. The presence of colibactin metabolites has been correlated to colorectal cancer formation in several studies. The specific function of many gene products in the colibactin gene cluster can be predicted. However, the role of ClbQ, a type II editing thioesterase, has not been established. The importance of ClbQ has been demonstrated by genetic deletions that abolish colibactin cytotoxic activity, and recent studies suggest an atypical role in releasing pathway intermediates from the assembly line. Here we report the 2.0 Å crystal structure and biochemical characterization of ClbQ. Our data reveal that ClbQ exhibits greater catalytic efficiency toward acyl-thioester substrates as compared to precolibactin intermediates and does not discriminate among carrier proteins. Cyclized pyridone-containing colibactins, which are off-pathway derivatives, are not viable substrates for ClbQ, while linear precursors are, supporting a role of ClbQ in facilitating the promiscuous off-loading of premature precolibactin metabolites and novel insights into colibactin biosynthesis.
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Energy Technology Data Exchange (ETDEWEB)
Guntaka, Naga Sandhya; Healy, Alan R.; Crawford, Jason M.; Herzon, Seth B.; Bruner, Steven D. (Yale); (Florida); (Yale-MED)
2017-09-08
Colibactin is a genotoxic hybrid nonribosomal peptide/polyketide secondary metabolite produced by various pathogenic and probiotic bacteria residing in the human gut. The presence of colibactin metabolites has been correlated to colorectal cancer formation in several studies. The specific function of many gene products in the colibactin gene cluster can be predicted. However, the role of ClbQ, a type II editing thioesterase, has not been established. The importance of ClbQ has been demonstrated by genetic deletions that abolish colibactin cytotoxic activity, and recent studies suggest an atypical role in releasing pathway intermediates from the assembly line. Here we report the 2.0 Å crystal structure and biochemical characterization of ClbQ. Our data reveal that ClbQ exhibits greater catalytic efficiency toward acyl-thioester substrates as compared to precolibactin intermediates and does not discriminate among carrier proteins. Cyclized pyridone-containing colibactins, which are off-pathway derivatives, are not viable substrates for ClbQ, while linear precursors are, supporting a role of ClbQ in facilitating the promiscuous off-loading of premature precolibactin metabolites and novel insights into colibactin biosynthesis.
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Directory of Open Access Journals (Sweden)
Jian Qiu
Full Text Available The rare wild species of snow lotus Saussurea involucrata is a commonly used medicinal herb with great pharmacological value for human health, resulting from its uniquely high level of phenylpropanoid compound production. To gain information on the phenylpropanid biosynthetic pathway genes in this critically important medicinal plant, global transcriptome sequencing was performed. It revealed that the phenylpropanoid pathway genes were well represented in S. involucrata. In addition, we introduced two key phenylpropanoid pathway inducing transcription factors (PAP1 and Lc into this medicinal plant. Transgenic S. involucrata co-expressing PAP1 and Lc exhibited purple pigments due to a massive accumulation of anthocyanins. The over-expression of PAP1 and Lc largely activated most of the phenylpropanoid pathway genes, and increased accumulation of several phenylpropanoid compounds significantly, including chlorogenic acid, syringin, cyanrine and rutin. Both ABTS (2,2'-azinobis-3-ethylbenzotiazo-line-6-sulfonic acid and FRAP (ferric reducing anti-oxidant power assays revealed that the antioxidant capacity of transgenic S. involucrata lines was greatly enhanced over controls. In addition to providing a deeper understanding of the molecular basis of phenylpropanoid metabolism, our results potentially enable an alternation of bioactive compound production in S. involucrata through metabolic engineering.
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Directory of Open Access Journals (Sweden)
William B Ludington
Full Text Available Climate change produces extremes in both temperature and precipitation causing increased drought severity and increased reliance on groundwater resources. Agricultural practices, which rely on groundwater, are sensitive to but also sources of contaminants, including nitrate. How agricultural contamination drives groundwater geochemistry through microbial metabolism is poorly understood.On an active cow dairy in the Central Valley of California, we sampled groundwater from three wells at depths of 4.3 m (two wells and 100 m (one well below ground surface (bgs as well as an effluent surface water lagoon that fertilizes surrounding corn fields. We analyzed the samples for concentrations of solutes, heavy metals, and USDA pathogenic bacteria of the Escherichia coli and Enterococcus groups as part of a long term groundwater monitoring study. Whole metagenome shotgun sequencing and assembly revealed taxonomic composition and metabolic potential of the community.Elevated nitrate and dissolved organic carbon occurred at 4.3m but not at 100m bgs. Metagenomics confirmed chemical observations and revealed several Planctomycete genomes, including a new Brocadiaceae lineage and a likely Planctomycetes OM190, as well novel diversity and high abundance of nano-prokaryotes from the Candidate Phyla Radiation (CPR, the Diapherotrites, Parvarchaeota, Aenigmarchaeota, Nanoarchaeota, Nanohaloarchaea (DPANN and the Thaumarchaeota, Aigarchaeota, Crenarchaeota, Korarchaeota (TACK superphyla. Pathway analysis suggests community interactions based on complimentary primary metabolic pathways and abundant secondary metabolite operons encoding antimicrobials and quorum sensing systems.The metagenomes show strong resemblance to activated sludge communities from a nitrogen removal reactor at a wastewater treatment plant, suggesting that natural bioremediation occurs through microbial metabolism. Elevated nitrate and rich secondary metabolite biosynthetic capacity suggest
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Genome Engineering of the 2,3-Butanediol Biosynthetic Pathway for Tight Regulation in Cyanobacteria.
Nozzi, Nicole E; Atsumi, Shota
2015-11-20
Cyanobacteria have gained popularity among the metabolic engineering community as a tractable photosynthetic host for renewable chemical production. However, though a number of successfully engineered production systems have been reported, long-term genetic stability remains an issue for cyanobacterial systems. The genetic engineering toolbox for cyanobacteria is largely lacking inducible systems for expression control. The characterization of tight regulation systems for use in cyanobacteria may help to alleviate this problem. In this work we explore the function of the IPTG inducible promoter P(L)lacO1 in the model cyanobacterium Synechococcus elongatus PCC 7942 as well as the effect of gene order within an operon on pathway expression. According to our experiments, P(L)lacO1 functions well as an inducible promoter in S. elongatus. Additionally, we found that gene order within an operon can strongly influence control of expression of each gene.
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Arioglu-Tuncil, Seda; Bhardwaj, Vivekanand; Taylor, Lynne S; Mauer, Lisa J
2017-09-01
Amorphous solid dispersions of thiamine chloride hydrochloride (THCl) were created using a variety of polymers with different physicochemical properties in order to investigate how effective the various polymers were as THCl crystallization inhibitors. THCl:polymer dispersions were prepared by lyophilizing solutions of THCl and amorphous polymers (guar gum, pectin, κ-carrageenan, gelatin, and polyvinylpyrrolidone (PVP)). These dispersions were stored at select temperature (25 and 40°C) and relative humidity (0, 23, 32, 54, 75, and 85% RH) conditions and monitored at different time points using powder X-ray diffraction (PXRD), Fourier transform infrared spectroscopy (FTIR), and differential scanning calorimetry (DSC). Moisture sorption isotherms of all samples were also obtained. Initially amorphous THCl was produced in the presence of ≥40% w/w pectin, κ-carrageenan, gelatin, and guar gum or ≥60% w/w PVP. Trends in polymer THCl crystallization inhibition (pectin≥κ-carrageenan>gelatin>guar gum≫PVP) were primarily based on the ability of the polymer to interact with THCl via hydrogen bonding and/or ionic interactions. The onset of THCl crystallization from the amorphous dispersions was also related to storage conditions. THCl remained amorphous at low RH conditions (0 and 23% RH) in all 1:1 dispersions except THCl:PVP. THCl crystallized in some dispersions below the glass transition temperature (T g ) but remained amorphous in others at T~T g . At high RHs (75 and 85% RH), THCl crystallized within one day in all samples. Given the ease of THCl amorphization in the presence of a variety of polymers, even at higher vitamin concentrations than would be found in foods, it is likely that THCl is amorphous in many low moisture foods. Copyright © 2017 Elsevier Ltd. All rights reserved.
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Paraptosis cell death induction by the thiamine analog benfotiamine in leukemia cells.
Directory of Open Access Journals (Sweden)
Naomi Sugimori
Full Text Available Benfotiamine is a synthetic thiamine analogue that stimulates transketolase, a cellular enzyme essential for glucose metabolism. Currently, benfotiamine is used to treat diabetic neuropathy. We recently reported that oral benfotiamine induced a temporary but remarkable recovery from acute myeloid leukemia in an elderly patient who was ineligible for standard chemotherapy due to dementia and renal failure. In the present study we present evidences that benfotiamine possess antitumor activity against leukemia cells. In a panel of nine myeloid leukemia cell lines benfotiamine impaired the viability of HL-60, NB4, K562 and KG1 cells and also inhibited the growing of primary leukemic blasts. The antitumor activity of benfotiamine is not mediated by apoptosis, necrosis or autophagy, but rather occurs though paraptosis cell death induction. Mechanistic studies revealed that benfotiamine inhibited the activity of constitutively active ERK1/2 and concomitantly increased the phosphorylation of JNK1/2 kinase in leukemic cells. In addition, benfotiamine induced the down regulation of the cell cycle regulator CDK3 which resulted in G1 cell cycle arrest in the sensitive leukemic cells. Moreover, combination index studies showed that benfotiamine enhanced the antiproliferative activities of cytarabine against leukemia cells. These findings suggest that benfotiamine has antitumor therapeutic potential.
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Paraptosis cell death induction by the thiamine analog benfotiamine in leukemia cells.
Sugimori, Naomi; Espinoza, J Luis; Trung, Ly Quoc; Takami, Akiyoshi; Kondo, Yukio; An, Dao Thi; Sasaki, Motoko; Wakayama, Tomohiko; Nakao, Shinji
2015-01-01
Benfotiamine is a synthetic thiamine analogue that stimulates transketolase, a cellular enzyme essential for glucose metabolism. Currently, benfotiamine is used to treat diabetic neuropathy. We recently reported that oral benfotiamine induced a temporary but remarkable recovery from acute myeloid leukemia in an elderly patient who was ineligible for standard chemotherapy due to dementia and renal failure. In the present study we present evidences that benfotiamine possess antitumor activity against leukemia cells. In a panel of nine myeloid leukemia cell lines benfotiamine impaired the viability of HL-60, NB4, K562 and KG1 cells and also inhibited the growing of primary leukemic blasts. The antitumor activity of benfotiamine is not mediated by apoptosis, necrosis or autophagy, but rather occurs though paraptosis cell death induction. Mechanistic studies revealed that benfotiamine inhibited the activity of constitutively active ERK1/2 and concomitantly increased the phosphorylation of JNK1/2 kinase in leukemic cells. In addition, benfotiamine induced the down regulation of the cell cycle regulator CDK3 which resulted in G1 cell cycle arrest in the sensitive leukemic cells. Moreover, combination index studies showed that benfotiamine enhanced the antiproliferative activities of cytarabine against leukemia cells. These findings suggest that benfotiamine has antitumor therapeutic potential.
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Modifications of the metabolic pathways of lipid and triacylglycerol production in microalgae
Directory of Open Access Journals (Sweden)
Yu Wei-Luen
2011-11-01
Full Text Available Abstract Microalgae have presented themselves as a strong candidate to replace diminishing oil reserves as a source of lipids for biofuels. Here we describe successful modifications of terrestrial plant lipid content which increase overall lipid production or shift the balance of lipid production towards lipid varieties more useful for biofuel production. Our discussion ranges from the biosynthetic pathways and rate limiting steps of triacylglycerol formation to enzymes required for the formation of triacylglycerol containing exotic lipids. Secondarily, we discuss techniques for genetic engineering and modification of various microalgae which can be combined with insights gained from research in higher plants to aid in the creation of production strains of microalgae.