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  1. Biochemical and Genetic Engineering of Diatoms for Polyunsaturated Fatty Acid Biosynthesis

    Directory of Open Access Journals (Sweden)

    Hong-Ye Li

    2014-01-01

    Full Text Available The role of diatoms as a source of bioactive compounds has been recently explored. Diatom cells store a high amount of fatty acids, especially certain polyunsaturated fatty acids (PUFAs. However, many aspects of diatom metabolism and the production of PUFAs remain unclear. This review describes a number of technical strategies, such as modulation of environmental factors (temperature, light, chemical composition of culture medium and culture methods, to influence the content of PUFAs in diatoms. Genetic engineering, a newly emerging field, also plays an important role in controlling the synthesis of fatty acids in marine microalgae. Several key points in the biosynthetic pathway of PUFAs in diatoms as well as recent progresses are also a critical part and are summarized here.

  2. Biochemical and genetic engineering of diatoms for polyunsaturated fatty acid biosynthesis.

    Science.gov (United States)

    Li, Hong-Ye; Lu, Yang; Zheng, Jian-Wei; Yang, Wei-Dong; Liu, Jie-Sheng

    2014-01-07

    The role of diatoms as a source of bioactive compounds has been recently explored. Diatom cells store a high amount of fatty acids, especially certain polyunsaturated fatty acids (PUFAs). However, many aspects of diatom metabolism and the production of PUFAs remain unclear. This review describes a number of technical strategies, such as modulation of environmental factors (temperature, light, chemical composition of culture medium) and culture methods, to influence the content of PUFAs in diatoms. Genetic engineering, a newly emerging field, also plays an important role in controlling the synthesis of fatty acids in marine microalgae. Several key points in the biosynthetic pathway of PUFAs in diatoms as well as recent progresses are also a critical part and are summarized here.

  3. Purification and biochemical characterization of mutacin I from the group I strain of Streptococcus mutans, CH43, and genetic analysis of mutacin I biosynthesis genes.

    Science.gov (United States)

    Qi, F; Chen, P; Caufield, P W

    2000-08-01

    Previously, we reported isolation and characterization of mutacin III and genetic analysis of mutacin III biosynthesis genes from the group III strain of Streptococcus mutans, UA787 (F. Qi, P. Chen, and P. W. Caufield, Appl. Environ. Microbiol. 65:3880-3887, 1999). During the same process of isolating the mutacin III structural gene, we also cloned the structural gene for mutacin I. In this report, we present purification and biochemical characterization of mutacin I from the group I strain CH43 and compare mutacin I and mutacin III biosynthesis genes. The mutacin I biosynthesis gene locus consists of 14 genes in the order mutR, -A, -A', -B, -C, -D, -P, -T, -F, -E, -G, orfX, orfY, orfZ. mutA is the structural gene for mutacin I, while mutA' is not required for mutacin I activity. DNA and protein sequence analysis revealed that mutacins I and III are homologous to each other, possibly arising from a common ancestor. The mature mutacin I is 24 amino acids in size and has a molecular mass of 2, 364 Da. Ethanethiol modification and peptide sequencing of mutacin I revealed that it contains six dehydrated serines, four of which are probably involved with thioether bridge formation. Comparison of the primary sequence of mutacin I with that of mutacin III and epidermin suggests that mutacin I likely has the same bridging pattern as epidermin.

  4. Chemical genetics to examine cellulose biosynthesis

    Directory of Open Access Journals (Sweden)

    Seth eDebolt

    2013-01-01

    Full Text Available Long-term efforts to decode plant cellulose biosynthesis via molecular genetics and biochemical strategies are being enhanced by the ever-expanding scale of omics technologies. An alternative approach to consider are the prospects for inducing change in plant metabolism using exogenously supplied chemical ligands. Cellulose biosynthesis inhibitors (CBI have been identified among known herbicides, during diverse combinatorial chemical libraries screens, and natural chemical screens from microbial agents. In this review, we summarize the current knowledge of the inhibitory effects of CBIs and further group them by how they influence fluorescently tagged cellulose synthase A (CESA proteins. Additional attention is paid to the continuing development of the CBI toolbox to explore the cell biology and genetic mechanisms underpinning effector molecule activity.

  5. (-)-Menthol biosynthesis and molecular genetics

    Science.gov (United States)

    Croteau, Rodney B.; Davis, Edward M.; Ringer, Kerry L.; Wildung, Mark R.

    2005-12-01

    (-)-Menthol is the most familiar of the monoterpenes as both a pure natural product and as the principal and characteristic constituent of the essential oil of peppermint ( Mentha x piperita). In this paper, we review the biosynthesis and molecular genetics of (-)-menthol production in peppermint. In Mentha species, essential oil biosynthesis and storage is restricted to the peltate glandular trichomes (oil glands) on the aerial surfaces of the plant. A mechanical method for the isolation of metabolically functional oil glands, has provided a system for precursor feeding studies to elucidate pathway steps, as well as a highly enriched source of the relevant biosynthetic enzymes and of their corresponding transcripts with which cDNA libraries have been constructed to permit cloning and characterization of key structural genes. The biosynthesis of (-)-menthol from primary metabolism requires eight enzymatic steps, and involves the formation and subsequent cyclization of the universal monoterpene precursor geranyl diphosphate to the parent olefin (-)-(4 S)-limonene as the first committed reaction of the sequence. Following hydroxylation at C3, a series of four redox transformations and an isomerization occur in a general “allylic oxidation-conjugate reduction” scheme that installs three chiral centers on the substituted cyclohexanoid ring to yield (-)-(1 R, 3 R, 4 S)-menthol. The properties of each enzyme and gene of menthol biosynthesis are described, as are their probable evolutionary origins in primary metabolism. The organization of menthol biosynthesis is complex in involving four subcellular compartments, and regulation of the pathway appears to reside largely at the level of gene expression. Genetic engineering to up-regulate a flux-limiting step and down-regulate a side route reaction has led to improvement in the composition and yield of peppermint oil.

  6. Biochemical genetics of some Indian fishes

    Digital Repository Service at National Institute of Oceanography (India)

    Menezes, M.R.; Qasim, S.Z.

    Studies on biochemical genetics of fishes, using electrophoretic methods, are relatively of recent origin. Earlier serum and eye lens protein were used to identify marine populations. This technique showed that closely related species have...

  7. Molecular and biochemical studies of fragrance biosynthesis in rose

    NARCIS (Netherlands)

    Sun, P.

    2017-01-01

    Roses are one of the most popular ornamental plants, whose floral volatiles are not only involved in environmental interactions but also widely used by industries. The biosynthesis of many of these volatiles in roses is not well understood. This thesis describes alternative pathways for the

  8. The Biochemical Mechanism of Auxin Biosynthesis by an Arabidopsis YUCCA Flavin-containing Monooxygenase*

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    Dai, Xinhua; Mashiguchi, Kiyoshi; Chen, Qingguo; Kasahara, Hiroyuki; Kamiya, Yuji; Ojha, Sunil; DuBois, Jennifer; Ballou, David; Zhao, Yunde

    2013-01-01

    Auxin regulates every aspect of plant growth and development. Previous genetic studies demonstrated that YUCCA (YUC) flavin-containing monooxygenases (FMOs) catalyze a rate-limiting step in auxin biosynthesis and that YUCs are essential for many developmental processes. We proposed that YUCs convert indole-3-pyruvate (IPA) to indole-3-acetate (IAA). However, the exact biochemical mechanism of YUCs has remained elusive. Here we present the biochemical characterization of recombinant Arabidopsis YUC6. Expressed in and purified from Escherichia coli, YUC6 contains FAD as a cofactor, which has peaks at 448 nm and 376 nm in the UV-visible spectrum. We show that YUC6 uses NADPH and oxygen to convert IPA to IAA. The first step of the YUC6-catalyzed reaction is the reduction of the FAD cofactor to FADH− by NADPH. Subsequently, FADH− reacts with oxygen to form a flavin-C4a-(hydro)peroxy intermediate, which we show has a maximum absorbance at 381 nm in its UV-visible spectrum. The final chemical step is the reaction of the C4a-intermediate with IPA to produce IAA. Although the sequences of the YUC enzymes are related to those of the mammalian FMOs, which oxygenate nucleophilic substrates, YUC6 oxygenates an electrophilic substrate (IPA). Nevertheless, both classes of enzymes form quasi-stable C4a-(hydro)peroxyl FAD intermediates. The YUC6 intermediate has a half-life of ∼20 s whereas that of some FMOs is >30 min. This work reveals the catalytic mechanism of the first known plant flavin monooxygenase and provides a foundation for further investigating how YUC activities are regulated in plants. PMID:23188833

  9. BIOCHEMICAL AND GENETIC CHARACTERIZATION OF AN EARLY STEP IN A NOVEL PATHWAY FOR THE BIOSYNTHESIS OF AROMATIC AMINO ACIDS AND P-AMINOBENZOIC ACID IN THE ARCHAEON METHANOCOCCUS MARIPALUDIS

    Science.gov (United States)

    Methanococcus maripaludis is a strictly anaerobic, methane-producing archaeon and facultative autotroph capable of biosynthesizing all the amino acids and vitamins required for growth. In this work, the novel 6-deoxy-5-ketofructose-1-phosphate (DKFP) pathway for the biosynthesis ...

  10. Muriel Wheldale Onslow and early biochemical genetics.

    Science.gov (United States)

    Richmond, Marsha L

    2007-01-01

    Muriel Whedale, a distinguished graduate of Newnham College, Cambridge, was a member of William Bateson's school of genetics at Cambridge University from 1903. Her investigation of flower color inheritance in snapdragons (Antirrhinum), a topic of particular interest to botanists, contributed to establishing Mendelism as a powerful new tool in studying heredity. Her understanding of the genetics of pigment formation led her to do cutting-edge work in biochemistry, culminating in the publication of her landmark work, The Anthocyanin Pigments of Plants (1916). In 1915, she joined Frederick Gowland Hopkin's Department of Biochemistry as assistant and in 1926 became one of the first women to be appointed university lecturer. In 1919 she married the biochemist Huia Onslow, with whom she collaborated until his death in 1922. This paper examines Whedale's work in genetics and especially focuses on the early linkage of Mendelian methodology with new techniques in biochemistry that eventually led to the founding of biochemical genetics. It highlights significant issues in the early history of women in genetics, including the critical role of mentors, funding opportunities, and career strategies.

  11. GENETIC ANALYSIS OF ABSCISIC ACID BIOSYNTHESIS

    Energy Technology Data Exchange (ETDEWEB)

    MCCARTY D R

    2012-01-10

    The carotenoid cleavage dioxygenases (CCD) catalyze synthesis of a variety of apo-carotenoid secondary metabolites in plants, animals and bacteria. In plants, the reaction catalyzed by the 11, 12, 9-cis-epoxy carotenoid dioxygenase (NCED) is the first committed and key regulated step in synthesis of the plant hormone, abscisic acid (ABA). ABA is a key regulator of plant stress responses and has critical functions in normal root and seed development. The molecular mechanisms responsible for developmental control of ABA synthesis in plant tissues are poorly understood. Five of the nine CCD genes present in the Arabidopsis genome encode NCED's involved in control of ABA synthesis in the plant. This project is focused on functional analysis of these five AtNCED genes as a key to understanding developmental regulation of ABA synthesis and dissecting the role of ABA in plant development. For this purpose, the project developed a comprehensive set of gene knockouts in the AtNCED genes that facilitate genetic dissection of ABA synthesis. These mutants were used in combination with key molecular tools to address the following specific objectives: (1) the role of ABA synthesis in root development; (2) developmental control of ABA synthesis in seeds; (3) analysis of ATNCED over-expressers; (4) preliminary crystallography of the maize VP14 protein.

  12. Biochemical genetic variation between four populations of ...

    African Journals Online (AJOL)

    system) to 0.093 in the Spekboom River population (Limpopo River system). The genetic distance, FST and NEM values, as well as pair-wise contingency c2 analyses indicate a lack of gene flow between populations, as expected for isolated fish. Evidence of foreign genetic material in one population was also observed.

  13. Genetics of Dothistromin Biosynthesis of Dothistroma septosporum: An Update

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    Rosie E. Bradshaw

    2010-11-01

    Full Text Available Dothistroma needle blight is one of the most devastating fungal pine diseases worldwide. The disease is characterized by accumulation in pine needles of a red toxin, dothistromin, that is chemically related to aflatoxin (AF and sterigmatocystin (ST. This review updates current knowledge of the genetics of dothistromin biosynthesis by the Dothistroma septosporum pathogen and highlights differences in gene organization and regulation that have been discovered between the dothistromin and AF/ST systems. Some previously reported genes are promoted or demoted as ‘dothistromin genes’ based on recent research. A new dothistromin gene, norB, is reported, and evidence of dothistromin gene homologs in other Dothideomycete fungi is presented. A hypothesis for the biological role of dothistromin is outlined. Finally, the impact that the availability of the D. septosporum genome sequence will have on dothistromin research is discussed.

  14. Appraisal of biochemical and genetic diversity of mango cultivars ...

    African Journals Online (AJOL)

    Mango (Mangifera indica L.) is one of the oldest fruit crops and is broadly cultivated worldwide. To determine the level of genetic diversity, a total of 13 mango genotypes have been collected from different farms of Fayoum oasis in Egypt and were analyzed using molecular (DNA) and biochemical (SDS-PAGE) markers ...

  15. Genetic and biochemical evidences reveal novel insights into the ...

    Indian Academy of Sciences (India)

    Home; Journals; Journal of Biosciences; Volume 41; Issue 4. Genetic and biochemical evidences reveal novel insights into the mechanism underlying Saccharomyces cerevisiae Sae2-mediated abrogation of DNA replication stress. INDRAJEET GHODKE K MUNIYAPPA. ARTICLE Volume 41 Issue 4 December 2016 pp ...

  16. Recombinant biosynthesis of bacterial cellulose in genetically modified Escherichia coli.

    Science.gov (United States)

    Buldum, Gizem; Bismarck, Alexander; Mantalaris, Athanasios

    2018-02-01

    Bacterial cellulose (BC) exhibits unique properties such as high purity compared to plant-based cellulose; however, commercial production of BC has remained a challenge, primarily due to the strain properties of cellulose-producing bacteria. Herein, we developed a functional and stable BC production system in genetically modified (GM) Escherichia coli by recombinant expression of both the BC synthase operon (bcsABCD) and the upstream operon (cmcax, ccp Ax). BC production was achieved in GM HMS174 (DE3) and in GM C41 (DE3) by optimization of the culture temperature (22 °C, 30 °C, and 37 °C) and IPTG concentration. BC biosynthesis was detected much earlier in GM C41 (DE3) cultures (3 h after IPTG induction) than those of Gluconacetobacter hansenii. GM HMS174 (DE3) produced dense fibres having a length of approximately 1000-3000 μm and a diameter of 10-20 μm, which were remarkably larger than the fibres of BC typically produced by G. hansenii.

  17. Biochemical, radiological, and genetic characterization of congenital hypothyroidism in Abu Dhabi, United Arab Emirates.

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    Deeb, Asma; Elkadry, Ihab; Attia, Salima; Al Suwaidi, Hana; Obaid, Laila; Schoenmakers, Nadia A

    2016-07-01

    Congenital hypothyroidism (CH) is caused by thyroid gland (TG) dysgenesis or inadequate thyroid hormone biosynthesis in a structurally normal gland. Different etiologies are known to be associated with various clinical, biochemical and imaging markers and a subset of cases have an underlying genetic basis. Despite the presence of neonatal screening programs in the UAE, there is a lack of data on the disease etiology in the area. We aim to study the etiology of CH in our center and examine its relationship with the clinical, biochemical, genetic and radiological features. Patients with CH who were followed in our center between 2011 and 2014 are enrolled in the study. Data collected included gender, gestational age, history of CH in a first-degree relative, initial thyroid stimulating hormone (TSH) and thyroxine (T4) levels, imaging findings, associated disease/malformation and treatment details. Selected patients with associated systemic disease or familial CH underwent genetic testing. Sixty-five patients were enrolled. Ten patients underwent genetic study: seven patients with associated congenital disease/malformation, one with a sibling and two with cousins with CH. Forty-nine subjects had technetium99 and/or ultrasound scans. Dyshormonogenesis was diagnosed in two-thirds of the patients. Three patients of 10 tested had likely causative genetic mutations; two homozygous thyroid peroxidase (TPO) and one heterozygous thyroid stimulating hormone receptor (TSHR) missense mutations. Dyshormonogenesis is the commonest etiology in CH in the studied group. It is expected that genetic mutations are more prevalent in our region due to the nature of the CH etiology and the rate of high consanguinity rate.

  18. Biochemical genetics in marine fisheries management and conservation

    Digital Repository Service at National Institute of Oceanography (India)

    Menezes, M.R.

    - 403004 NBDBlDBTSponsored Training on Taxonomy, GOIdia turd Gme Bturking o/Coastal and Marine Bloresources, CIFE, Mumbal BIOCHEMICAL GENETICS IN MARINE FISHERIES MANAGEMENT AND CONSERVATION Maria R. Menezes Introduction . Species of fish, like most... population structure may have evolved and been maintained in species of fish ofeconomic interest has led to the concept of 'stock'. The stock concept dominates much of marine fisheries management, theory and practice because the identification of discrete...

  19. Biochemical and genetic engineering strategies to enhance hydrogen production in photosynthetic algae and cyanobacteria.

    Science.gov (United States)

    Srirangan, Kajan; Pyne, Michael E; Perry Chou, C

    2011-09-01

    As an energy carrier, hydrogen gas is a promising substitute to carbonaceous fuels owing to its superb conversion efficiency, non-polluting nature, and high energy content. At present, hydrogen is predominately synthesized via chemical reformation of fossil fuels. While various biological methods have been extensively explored, none of them is justified as economically feasible. A sustainable platform for biological production of hydrogen will certainly impact the biofuel market. Among a selection of biological systems, algae and cyanobacteria have garnered major interests as potential cell factories for hydrogen production. In conjunction with photosynthesis, these organisms utilize inexpensive inorganic substrates and solar energy for simultaneous biosynthesis and hydrogen evolution. However, the hydrogen yield associated with these organisms remains far too low to compete with the existing chemical systems. This article reviews recent advances of biochemical, bioprocess, and genetic engineering strategies in circumventing technological limitations to hopefully improve the applicative potential of these photosynthetic hydrogen production systems. Copyright © 2011 Elsevier Ltd. All rights reserved.

  20. Genetic determinants of reutericyclin biosynthesis in Lactobacillus reuteri.

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    Lin, Xiaoxi B; Lohans, Christopher T; Duar, Rebbeca; Zheng, Jinshui; Vederas, John C; Walter, Jens; Gänzle, Michael

    2015-03-01

    Reutericyclin is a unique antimicrobial tetramic acid produced by some strains of Lactobacillus reuteri. This study aimed to identify the genetic determinants of reutericyclin biosynthesis. Comparisons of the genomes of reutericyclin-producing L. reuteri strains with those of non-reutericyclin-producing strains identified a genomic island of 14 open reading frames (ORFs) including genes coding for a nonribosomal peptide synthetase (NRPS), a polyketide synthase (PKS), homologues of PhlA, PhlB, and PhlC, and putative transport and regulatory proteins. The protein encoded by rtcN is composed of a condensation domain, an adenylation domain likely specific for d-leucine, and a thiolation domain. rtcK codes for a PKS that is composed of a ketosynthase domain, an acyl-carrier protein domain, and a thioesterase domain. The products of rtcA, rtcB, and rtcC are homologous to the diacetylphloroglucinol-biosynthetic proteins PhlABC and may acetylate the tetramic acid moiety produced by RtcN and RtcK, forming reutericyclin. Deletion of rtcN or rtcABC in L. reuteri TMW1.656 abrogated reutericyclin production but did not affect resistance to reutericyclin. Genes coding for transport and regulatory proteins could be deleted only in the reutericyclin-negative L. reuteri strain TMW1.656ΔrtcN, and these deletions eliminated reutericyclin resistance. The genomic analyses suggest that the reutericyclin genomic island was horizontally acquired from an unknown source during a unique event. The combination of PhlABC homologues with both an NRPS and a PKS has also been identified in the lactic acid bacteria Streptococcus mutans and Lactobacillus plantarum, suggesting that the genes in these organisms and those in L. reuteri share an evolutionary origin. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  1. Growth-based determination and biochemical confirmation of genetic requirements for protein degradation in Saccharomyces cerevisiae.

    Science.gov (United States)

    Watts, Sheldon G; Crowder, Justin J; Coffey, Samuel Z; Rubenstein, Eric M

    2015-02-16

    Regulated protein degradation is crucial for virtually every cellular function. Much of what is known about the molecular mechanisms and genetic requirements for eukaryotic protein degradation was initially established in Saccharomyces cerevisiae. Classical analyses of protein degradation have relied on biochemical pulse-chase and cycloheximide-chase methodologies. While these techniques provide sensitive means for observing protein degradation, they are laborious, time-consuming, and low-throughput. These approaches are not amenable to rapid or large-scale screening for mutations that prevent protein degradation. Here, a yeast growth-based assay for the facile identification of genetic requirements for protein degradation is described. In this assay, a reporter enzyme required for growth under specific selective conditions is fused to an unstable protein. Cells lacking the endogenous reporter enzyme but expressing the fusion protein can grow under selective conditions only when the fusion protein is stabilized (i.e. when protein degradation is compromised). In the growth assay described here, serial dilutions of wild-type and mutant yeast cells harboring a plasmid encoding a fusion protein are spotted onto selective and non-selective medium. Growth under selective conditions is consistent with degradation impairment by a given mutation. Increased protein abundance should be biochemically confirmed. A method for the rapid extraction of yeast proteins in a form suitable for electrophoresis and western blotting is also demonstrated. A growth-based readout for protein stability, combined with a simple protocol for protein extraction for biochemical analysis, facilitates rapid identification of genetic requirements for protein degradation. These techniques can be adapted to monitor degradation of a variety of short-lived proteins. In the example presented, the His3 enzyme, which is required for histidine biosynthesis, was fused to Deg1-Sec62. Deg1-Sec62 is targeted for

  2. Some aspects of genetic control of antibiotic biosynthesis in Streptomyces

    Directory of Open Access Journals (Sweden)

    М. P. Teplitskaya

    2005-12-01

    Full Text Available These work contain a review of basic hypotheses and experimental information in relation to the problem of antibiotic synthesis regulation by the bacteria of the Streptomyces family. Data on cluster organization of antibiotics biosynthesis genes in these microorganisms were generalized. The examples of the positive and negative specific control of antibiotic production genes were resulted. Except for it, proofs that confirm participation of a few genes of more high level in the process of initiation and expression of antibiotics biosynthesis genes also were found. In this connection А-factor role in the mechanism of cascade-organized process of streptomycin biosynthesis control, some other antibiotics and spore determinations is discussed in detail.

  3. Biochemical pathways supporting beta-lactam biosynthesis in the springtail Folsomia candida

    NARCIS (Netherlands)

    Suring, Wouter; Marien, Janine; Broekman, R.A.; van Straalen, N.M.; Roelofs, D.

    2016-01-01

    Recently, an active set of beta-lactam biosynthesis genes was reported in the genome of the arthropod springtail Folsomia candida (Collembola). Evidence was provided that these genes were acquired through horizontal gene transfer. However, successful integration of fungal- or bacterial-derived

  4. Genetic control and regulatory mechanisms of succinoglycan and curdlan biosynthesis in genus Agrobacterium.

    Science.gov (United States)

    Wu, Dan; Li, Ang; Ma, Fang; Yang, Jixian; Xie, Yutong

    2016-07-01

    Agrobacterium is a genus of gram-negative bacteria that can produce several typical exopolysaccharides with commercial uses in the food and pharmaceutical fields. In particular, succinoglycan and curdlan, due to their good quality in high yield, have been employed on an industrial scale comparatively early. Exopolysaccharide biosynthesis is a multiple-step process controlled by different functional genes, and various environmental factors cause changes in exopolysaccharide biosynthesis through regulatory mechanisms. In this mini-review, we focus on the genetic control and regulatory mechanisms of succinoglycan and curdlan produced by Agrobacterium. Some key functional genes and regulatory mechanisms for exopolysaccharide biosynthesis are described, possessing a high potential for application in metabolic engineering to modify exopolysaccharide production and physicochemical properties. This review may contribute to the understanding of exopolysaccharide biosynthesis and exopolysaccharide modification by metabolic engineering methods in Agrobacterium.

  5. Genetic Dissection of Tropodithietic Acid Biosynthesis by Marine Roseobacters

    DEFF Research Database (Denmark)

    Geng, Haifeng; Bruhn, Jesper Bartholin; Nielsen, Kristian Fog

    2008-01-01

    formation is coincident with the production of an antibiotic and a yellow-brown pigment. In this report, we demonstrate that the antibiotic is a sulfur-containing compound, tropodithietic acid (TDA). Using random transposon insertion mutagenesis, 12 genes were identified as critical for TDA biosynthesis...... by the bacteria, and mutation in any one of these results in a loss of antibiotic activity (Tda(-)) and pigment production. Unexpectedly, six of the genes, referred to as tdaA-F, could not be found on the annotated TM1040 genome and were instead located on a previously unidentified plasmid (ca. 130 kb; pSTM3...

  6. Polyphenol Oxidases in Crops: Biochemical, Physiological and Genetic Aspects.

    Science.gov (United States)

    Taranto, Francesca; Pasqualone, Antonella; Mangini, Giacomo; Tripodi, Pasquale; Miazzi, Monica Marilena; Pavan, Stefano; Montemurro, Cinzia

    2017-02-10

    Enzymatic browning is a colour reaction occurring in plants, including cereals, fruit and horticultural crops, due to oxidation during postharvest processing and storage. This has a negative impact on the colour, flavour, nutritional properties and shelf life of food products. Browning is usually caused by polyphenol oxidases (PPOs), following cell damage caused by senescence, wounding and the attack of pests and pathogens. Several studies indicated that PPOs play a role in plant immunity, and emerging evidence suggested that PPOs might also be involved in other physiological processes. Genomic investigations ultimately led to the isolation of PPO homologs in several crops, which will be possibly characterized at the functional level in the near future. Here, focusing on the botanic families of Poaceae and Solanaceae, we provide an overview on available scientific literature on PPOs, resulting in useful information on biochemical, physiological and genetic aspects.

  7. Genetics of Dothistromin Biosynthesis in the Peanut Pathogen Passalora arachidicola

    Directory of Open Access Journals (Sweden)

    Rosie E. Bradshaw

    2010-11-01

    Full Text Available The peanut leaf spot pathogen Passalora arachidicola (Mycosphaerella arachidis is known to produce dothistromin, a mycotoxin related to aflatoxin. This is a feature shared with the pine needle pathogen Dothistroma septosporum (Mycosphaerella pini. Dothistromin biosynthesis in D. septosporum commences at an unusually early stage of growth in culture compared to most other fungal secondary metabolites, and the biosynthetic genes are arranged in fragmented groups, in contrast to aflatoxin gene clusters. Dothistromin biosynthetic genes were identified and studied in P. arachidicola to determine if the attributes described in D. septosporum are shared by another dothistromin-producing species within the Class Dothideomycetes. It was shown that dothistromin biosynthesis is very similar in the two species with regard to gene sequence and gene synteny. Functional complementation of D. septosporum mutants with P. arachidicola dothistromin genes was also possible. These similarities support a vertical mode of dothistromin gene transmission. P. arachidicola also produced dothistromin at an early growth stage in culture, suggesting that this type of regulation pattern may be relevant to the biological role of dothistromin.

  8. Mycobacterium tuberculosis phosphoribosylpyrophosphate synthetase: biochemical features of a crucial enzyme for mycobacterial cell wall biosynthesis.

    Directory of Open Access Journals (Sweden)

    Anna P Lucarelli

    Full Text Available The selection and soaring spread of Mycobacterium tuberculosis multidrug-resistant (MDR-TB and extensively drug-resistant strains (XDR-TB is a severe public health problem. Currently, there is an urgent need for new drugs for tuberculosis treatment, with novel mechanisms of action and, moreover, the necessity to identify new drug targets. Mycobacterial phosphoribosylpyrophosphate synthetase (MtbPRPPase is a crucial enzyme involved in the biosynthesis of decaprenylphosphoryl-arabinose, an essential precursor for the mycobacterial cell wall biosynthesis. Moreover, phosphoribosylpyrophosphate, which is the product of the PRPPase catalyzed reaction, is the precursor for the biosynthesis of nucleotides and of some amino acids such as histidine and tryptophan. In this context, the elucidation of the molecular and functional features of MtbPRPPase is mandatory. MtbPRPPase was obtained as a recombinant form, purified to homogeneity and characterized. According to its hexameric form, substrate specificity and requirement of phosphate for activity, the enzyme proved to belong to the class I of PRPPases. Although the sulfate mimicked the phosphate, it was less effective and required higher concentrations for the enzyme activation. MtbPRPPase showed hyperbolic response to ribose 5-phosphate, but sigmoidal behaviour towards Mg-ATP. The enzyme resulted to be allosterically activated by Mg(2+ or Mn(2+ and inhibited by Ca(2+ and Cu(2+ but, differently from other characterized PRPPases, it showed a better affinity for the Mn(2+ and Cu(2+ ions, indicating a different cation binding site geometry. Moreover, the enzyme from M. tuberculosis was allosterically inhibited by ADP, but less sensitive to inhibition by GDP. The characterization of M. tuberculosis PRPPase provides the starting point for the development of inhibitors for antitubercular drug design.

  9. Genetic analysis of selenocysteine biosynthesis in the archaeon Methanococcus maripaludis.

    Science.gov (United States)

    Hohn, Michael J; Palioura, Sotiria; Su, Dan; Yuan, Jing; Söll, Dieter

    2011-07-01

    In Archaea selenocysteine (Sec) is synthesized in three steps. First seryl-tRNA synthetase acylates tRNA(Sec) with serine to generate Ser-tRNA(Sec). Then phosphoseryl-tRNA(Sec) kinase (PSTK) forms Sep-tRNA(Sec) , which is converted to Sec-tRNA(Sec) by Sep-tRNA:Sec-tRNA synthase (SepSecS) in the presence of selenophosphate produced by selenophosphate synthetase (SelD). A complete in vivo analysis of the archaeal Sec biosynthesis pathway is still unavailable, and the existence of a redundant pathway or of a rescue mechanism based on the conversion of Sep-tRNA(Sec) to Cys-tRNA(Sec) during selenium starvation, cannot be excluded. Here we present a mutational analysis of Sec biosynthesis in Methanococcus maripaludis strain Mm900. Sec formation is abolished upon individually deleting the genes encoding SelD, PSTK or SepSecS; the resulting mutant strains could no longer grow on formate while growth with H(2) + CO(2) remained unaffected. However, deletion of the PSTK and SepSecS genes was not possible unless the selenium-free [NiFe]-hydrogenases Frc and Vhc were expressed. This required the prior deletion of either the gene encoding SelD or that of HrsM, a LysR-type regulator suppressing transcription of the frc and vhc operons in the presence of selenium. These results show that M. maripaludis Mm900 is facultatively selenium-dependent with a single pathway of Sec-tRNA(Sec) formation. © 2011 Blackwell Publishing Ltd.

  10. Genetics, Biosynthesis, Structure, and Mode of Action of Lantibiotics

    Science.gov (United States)

    Kuipers, Anneke; Rink, Rick; Moll, Gert N.

    Lantibiotics are lanthionine-containing peptide antibiotics. They are characterized by having meso-lanthionine(s) and/or β-methyllanthionine(s) or both. These intramolecular monosulfide cross-links render the peptide resistant against breakdown by peptidases. Moreover, in several cases, the (methyl)lanthionines are essential for interaction with the so-called docking molecule lipid II. The best known lantibiotic, nisin, highly effectively inhibits growth of target cells via two mechanisms: (1) abduction of the cell wall precursor lipid II from the septum and (2) formation of pores composed of lipid II and nisin. (Methyl)lanthionines result from two enzyme-catalyzed posttranslational modifications: dehydration of serines/threonines and coupling of the resulting dehydro amino acids to cysteines. Besides the localization of the thioether bridges and dehydro amino acids in the lantibiotics, also the three-dimensional structure of some lantibiotics has been resolved by NMR. Genes encoding proteins involved in the biosynthesis of lantibiotics are present in clusters and may comprise combinations of the following genes in varying order: a structural gene that encodes a leader peptide and the lantibiotic propeptide, modification enzyme(s), a transporter responsible for the export of the lantibiotic and in some cases for cleavage of the leader peptide, a leader peptidase, a so-called immunity protein involved in self-protection of the host cell, components of a transporter also involved in self-protection, and two components of an autoinduction system.

  11. Biochemical and histochemical localization of monoterpene biosynthesis in the glandular trichomes of spearmint (Mentha spicata)

    Energy Technology Data Exchange (ETDEWEB)

    Gershenzon, J.; Maffei, M.; Croteau, R. (Washington State Univ., Pullman (USA))

    1989-04-01

    The primary monoterpene accumulated in the glandular trichomes of spearmint (Mentha spicata) is the ketone (-)-carvone which is formed by cyclization of the C{sub 10} isoprenoid intermediate geranyl pyrophosphate to the olefin (-)-limonene, hydroxylation to (-)-trans-carveol and subsequent dehydrogenation. Selective extraction of the contents of the glandular trichomes indicated that essentially all of the cyclase and hydroxylase activities resided in these structures, whereas only about 30% of the carveol dehydrogenase was located here with the remainder located in the rest of the leaf. This distribution of carveol dehydrogenase activity was confirmed by histochemical methods. Electrophoretic analysis of the partially purified carveol dehydrogenase from extracts of both the glands and the leaves following gland removal indicated the presence of a unique carveol dehydrogenase species in the glandular trichomes, suggesting that the other dehydrogenase found throughout the leaf probably utilizes carveol only as an adventitious substrate. These results demonstrate that carvone biosynthesis takes place exclusively in the glandular trichomes in which this natural product accumulates.

  12. Biochemical and Histochemical Localization of Monoterpene Biosynthesis in the Glandular Trichomes of Spearmint (Mentha spicata) 12

    Science.gov (United States)

    Gershenzon, Jonathan; Maffei, Massimo; Croteau, Rodney

    1989-01-01

    The primary monoterpene accumulated in the glandular trichomes of spearmint (Mentha spicata) is the ketone (−)-carvone which is formed by cyclization of the C10 isoprenoid intermediate geranyl pyrophosphate to the olefin (−)-limonene, hydroxylation to (−)-trans-carveol and subsequent dehydrogenation. Selective extraction of the contents of the glandular trichomes indicated that essentially all of the cyclase and hydroxylase activities resided in these structures, whereas only about 30% of the carveol dehydrogenase was located here with the remainder located in the rest of the leaf. This distribution of carveol dehydrogenase activity was confirmed by histochemical methods. Electrophoretic analysis of the partially purified carveol dehydrogenase from extracts of both the glands and the leaves following gland removal indicated the presence of a unique carveol dehydrogenase species in the glandular trichomes, suggesting that the other dehydrogenase found throughout the leaf probably utilizes carveol only as an adventitious substrate. These results demonstrate that carvone biosynthesis takes place exclusively in the glandular trichomes in which this natural product accumulates. Images Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 PMID:16666709

  13. Clinical, biochemical and molecular genetic correlations in adenylosuccinate lyase deficiency.

    Science.gov (United States)

    Race, V; Marie, S; Vincent, M F; Van den Berghe, G

    2000-09-01

    Adenylosuccinate lyase (ADSL) deficiency (MIM 103050) is an autosomal recessive inborn error of purine synthesis characterized by the accumulation in body fluids of succinylaminoimidazolecarboxamide (SAICA) riboside and succinyladenosine (S-Ado), the dephosphorylated derivatives of the two substrates of the enzyme. Because ADSL-deficient patients display widely variable degrees of psychomotor retardation, we have expressed eight mutated ADSL enzymes as thioredoxin fusions and compared their properties with the clinical and biochemical characteristics of 10 patients. Three expressed mutated ADSL enzymes (M26L, R426H and T450S) were thermolabile, four (A2V, R141W, R303C and S395R) were thermostable and one (del206-218), was inactive. Thermolabile mutations decreased activities with SAICA ribotide (SAICAR) and adenylosuccinate (S-AMP) in parallel, or more with SAICAR than with S-AMP. Patients homozygous for one of these mutations, R426H, displayed similarly decreased ADSL activities in their fibroblasts, S-Ado:SAICA riboside ratios of approximately 1 in their cerebrospinal fluid and were profoundly retarded. With the exception of A2V, thermostable mutations decreased activity with S-AMP to a much more marked extent than with SAICAR. Two unrelated patients homozygous for one of the thermostable mutations, R303C, also displayed a much more marked decrease in the activity of fibroblast ADSL with S-AMP than with SAICAR, had S-Ado:SAICA riboside ratios between 3 and 4 in their cerebrospinal fluid and were mildly retarded. These results suggest that, in some cases, the genetic lesion of ADSL determines the ratio of its activities with S-AMP versus SAICAR, which in turn defines the S-Ado:SAICA riboside ratio and the patients' mental status.

  14. The clinical, biochemical and genetic features associated with RMND1-related mitochondrial disease

    DEFF Research Database (Denmark)

    Ng, Yi Shiau; Alston, Charlotte L; Diodato, Daria

    2016-01-01

    BACKGROUND: Mutations in the RMND1 (Required for Meiotic Nuclear Division protein 1) gene have recently been linked to infantile onset mitochondrial disease characterised by multiple mitochondrial respiratory chain defects. METHODS: We summarised the clinical, biochemical and molecular genetic in...

  15. A Classical Genetic Solution to Enhance the Biosynthesis of Anticancer Phytochemicals in Andrographis paniculata Nees

    Science.gov (United States)

    Talei, Daryush; Abdul Kadir, Mihdzar; Rafii, Mohd Yusop; Sagineedu, Sreenivasa Rao

    2014-01-01

    Andrographolides, the diterpene lactones, are major bioactive phytochemicals which could be found in different parts of the medicinal herb Andrographis paniculata. A number of such compounds namely andrographolide (AG), neoandrographolide (NAG), and 14-deoxy-11,12-didehydroandrographolide (DDAG) have already attracted a great deal of attention due to their potential therapeutic effects in hard-to-treat diseases such as cancers and HIV. Recently, they have also been considered as substrates for the discovery of novel pharmaceutical compounds. Nevertheless, there is still a huge gap in knowledge on the genetic pattern of the biosynthesis of these bioactive compounds. Hence, the present study aimed to investigate the genetic mechanisms controlling the biosynthesis of these phytochemicals using a diallel analysis. The high performance liquid chromatography analysis of the three andrographolides in 210 F1 progenies confirmed that the biosynthesis of these andrographolides was considerably increased via intraspecific hybridization. The results revealed high, moderate and low heterosis for DDAG, AG and NAG, respectively. Furthermore, the preponderance of non-additive gene actions was affirmed in the enhancement of the three andrographolides contents. The consequence of this type of gene action was the occurrence of high broad-sense and low narrow-sense heritabilities for the above mentioned andrographolides. The prevalence of non-additive gene action suggests the suitability of heterosis breeding and hybrid seed production as a preferred option to produce new plant varieties with higher andrographolide contents using the wild accessions of A. paniculata. Moreover, from an evolutionary point of view, the occurrence of population bottlenecks in the Malaysian accessions of A. paniculata was unveiled by observing a low level of additive genetic variance (VA) for all the andrographolides. PMID:24586262

  16. A classical genetic solution to enhance the biosynthesis of anticancer phytochemicals in Andrographis paniculata Nees.

    Directory of Open Access Journals (Sweden)

    Alireza Valdiani

    Full Text Available Andrographolides, the diterpene lactones, are major bioactive phytochemicals which could be found in different parts of the medicinal herb Andrographis paniculata. A number of such compounds namely andrographolide (AG, neoandrographolide (NAG, and 14-deoxy-11,12-didehydroandrographolide (DDAG have already attracted a great deal of attention due to their potential therapeutic effects in hard-to-treat diseases such as cancers and HIV. Recently, they have also been considered as substrates for the discovery of novel pharmaceutical compounds. Nevertheless, there is still a huge gap in knowledge on the genetic pattern of the biosynthesis of these bioactive compounds. Hence, the present study aimed to investigate the genetic mechanisms controlling the biosynthesis of these phytochemicals using a diallel analysis. The high performance liquid chromatography analysis of the three andrographolides in 210 F1 progenies confirmed that the biosynthesis of these andrographolides was considerably increased via intraspecific hybridization. The results revealed high, moderate and low heterosis for DDAG, AG and NAG, respectively. Furthermore, the preponderance of non-additive gene actions was affirmed in the enhancement of the three andrographolides contents. The consequence of this type of gene action was the occurrence of high broad-sense and low narrow-sense heritabilities for the above mentioned andrographolides. The prevalence of non-additive gene action suggests the suitability of heterosis breeding and hybrid seed production as a preferred option to produce new plant varieties with higher andrographolide contents using the wild accessions of A. paniculata. Moreover, from an evolutionary point of view, the occurrence of population bottlenecks in the Malaysian accessions of A. paniculata was unveiled by observing a low level of additive genetic variance (VA for all the andrographolides.

  17. Filling gaps in bacterial amino acid biosynthesis pathways with high-throughput genetics.

    Directory of Open Access Journals (Sweden)

    Morgan N Price

    2018-01-01

    Full Text Available For many bacteria with sequenced genomes, we do not understand how they synthesize some amino acids. This makes it challenging to reconstruct their metabolism, and has led to speculation that bacteria might be cross-feeding amino acids. We studied heterotrophic bacteria from 10 different genera that grow without added amino acids even though an automated tool predicts that the bacteria have gaps in their amino acid synthesis pathways. Across these bacteria, there were 11 gaps in their amino acid biosynthesis pathways that we could not fill using current knowledge. Using genome-wide mutant fitness data, we identified novel enzymes that fill 9 of the 11 gaps and hence explain the biosynthesis of methionine, threonine, serine, or histidine by bacteria from six genera. We also found that the sulfate-reducing bacterium Desulfovibrio vulgaris synthesizes homocysteine (which is a precursor to methionine by using DUF39, NIL/ferredoxin, and COG2122 proteins, and that homoserine is not an intermediate in this pathway. Our results suggest that most free-living bacteria can likely make all 20 amino acids and illustrate how high-throughput genetics can uncover previously-unknown amino acid biosynthesis genes.

  18. A Single Gene Cluster for Chalcomycins and Aldgamycins: Genetic Basis for Bifurcation of Their Biosynthesis.

    Science.gov (United States)

    Tang, Xiao-Long; Dai, Ping; Gao, Hao; Wang, Chuan-Xi; Chen, Guo-Dong; Hong, Kui; Hu, Dan; Yao, Xin-Sheng

    2016-07-01

    Aldgamycins are 16-membered macrolide antibiotics with a rare branched-chain sugar d-aldgarose or decarboxylated d-aldgarose at C-5. In our efforts to clone the gene cluster for aldgamycins from a marine-derived Streptomyces sp. HK-2006-1 capable of producing both aldgamycins and chalcomycins, we found that both are biosynthesized from a single gene cluster. Whole-genome sequencing combined with gene disruption established the entire gene cluster of aldgamycins: nine new genes are incorporated with the previously identified chalcomycin gene cluster. Functional analysis of these genes revealed that almDI/almDII, (encoding α/β subunits of pyruvate dehydrogenase) triggers the biosynthesis of aldgamycins, whereas almCI (encoding an oxidoreductase) initiates chalcomycins biosynthesis. This is the first report that aldgamycins and chalcomycins are derived from a single gene cluster and of the genetic basis for bifurcation in their biosynthesis. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  19. Genetic control of ascorbic acid biosynthesis and recycling in horticultural crops

    Science.gov (United States)

    Mellidou, Ifigeneia; Kanellis, Angelos K.

    2017-07-01

    Ascorbic acid (AsA) is an essential compound present in almost all living organisms that has important functions in several aspects of plant growth and development, hormone signalling, as well as stress defense networks. In recent years, the genetic regulation of AsA metabolic pathways has received much attention due to its beneficial role in human diet. Despite the great variability within species, genotypes, tissues and developmental stages, AsA accumulation is considered to be controlled by the fine orchestration of net biosynthesis, recycling, degradation/oxidation, and/or intercellular and intracellular transport. To date, several structural genes from the AsA metabolic pathways and transcription factors are considered to significantly affect AsA in plant tissues, either at the level of activity, transcription or translation via feedback inhibition. Yet, all the emerging studies support the notion that the steps proceeding through GDP-L-galactose phosphorylase and to a lesser extent through GDP-D-mannose-3,5-epimerase are control points in governing AsA pool size in several species. In this mini review, we discuss the current consensus of the genetic regulation of AsA biosynthesis and recycling, with a focus on horticultural crops. The aspects of AsA degradation and transport are not discussed herein. Novel insights of how this multifaceted trait is regulated are critical to prioritize candidate genes for follow-up studies towards improving the nutritional value of fruits and vegetables.

  20. A preliminary biochemical genetic survey of four South African ...

    African Journals Online (AJOL)

    Allozyme electrophoresis was used to evaluate genetic variability in painted reed frog (Hyperolius marmoratus) populations from the Northern Province and the Eastern Cape. Sixteen protein-encoding loci were resolved, of which seven displayed allelic polymorphism. Average heterozygosity (H) values in two permanent H.

  1. Genetic Investigations Using Immuno-biochemical Markers in a Maramureş Brown Cattle Population

    Directory of Open Access Journals (Sweden)

    Nicoleta Isfan

    2011-05-01

    Full Text Available The study of the genetic markers and identifying new markers involves an increasing number of research projects in the fields of genetics of immunology, biochemical genetics, molecular genetics, quantity genetics and the genetic improvement of animals. Some studies on genes frequency determining the red cells specificity and for whey hemoglobin are approached in the present report. In this way, some blood factors, most of them belonging to B system (the most complex system in cattle have been evidenced. The lowest gene frequency was present in K factor (7%, and highest one in, O1, G’ , W and F1 (100%. In addition to basic importance on knowledge and determination of cattle population genetic structure for studied protein loci, another theme proposed to correlate hemoglobin type with some traits of economical importance: milk yield, fat and protein content, fat and protein yield. Higher performance was recorded by HbA/HbA individuals.

  2. Metabologenomics of Phaeochromocytoma and Paraganglioma: An Integrated Approach for Personalised Biochemical and Genetic Testing.

    Science.gov (United States)

    Eisenhofer, Graeme; Klink, Barbara; Richter, Susan; Lenders, Jacques Wm; Robledo, Mercedes

    2017-04-01

    The tremendous advances over the past two decades in both clinical genetics and biochemical testing of chromaffin cell tumours have led to new considerations about how these aspects of laboratory medicine can be integrated to improve diagnosis and management of affected patients. With germline mutations in 15 genes now identified to be responsible for over a third of all cases of phaeochromocytomas and paragangliomas, these tumours are recognised to have one of the richest hereditary backgrounds among all neoplasms. Depending on the mutation, tumours show distinct differences in metabolic pathways that relate to or even directly impact clinical presentation. At the same time, there has been improved understanding about how catecholamines are synthesised, stored, secreted and metabolised by chromaffin cell tumours. Although the tumours may not always secrete catecholamines it has become clear that almost all continuously produce and metabolise catecholamines. This has not only fuelled changes in laboratory medicine, but has also assisted in recognition of genotype-biochemical phenotype relationships important for diagnostics and clinical care. In particular, differences in catecholamine and energy pathway metabolomes can guide genetic testing, assist with test interpretation and provide predictions about the nature, behaviour and imaging characteristics of the tumours. Conversely, results of genetic testing are important for guiding how routine biochemical testing should be employed and interpreted in surveillance programmes for at-risk patients. In these ways there are emerging needs for modern laboratory medicine to seamlessly integrate biochemical and genetic testing into the diagnosis and management of patients with chromaffin cell tumours.

  3. Aspirin resistance: effect of clinical, biochemical and genetic factors.

    Science.gov (United States)

    Fitzgerald, Richard; Pirmohamed, Munir

    2011-05-01

    Aspirin is one of the cornerstones of treatment for cardiovascular disease. However, some patients may be 'resistant' to its effect: this is associated with adverse cardiovascular outcomes and increased mortality. Measuring response to aspirin is often difficult and there is no accepted definition of aspirin resistance. Many assays are available to test aspirin sensitivity but most are not specific to aspirin and the degree of agreement between different assays is poor. Each assay has its own advantages and disadvantages, and there is currently no one assay that can be recommended for routine clinical practise. There are also many potential modifiers of aspirin response including aspirin dose, non-compliance, disease severity, genetic factors, inflammation, diabetes mellitus, hyperlipidaemia, smoking and interacting drugs. Treating the underlying cause may improve aspirin sensitivity but current data are contradictory with no large clinical trials that have addressed this. Further work is required in this area to determine whether and how aspirin resistance is important clinically, what the best measurement is phenotypically and how this should be used in clinical practise, and whether there are any genetic predisposing factors. This will require well designed prospective studies which take into account the numerous confounding factors that can modify aspirin resistance. Copyright © 2011 Elsevier Inc. All rights reserved.

  4. Genetic and metabolomic analysis of AdeD and AdeI mutants of de novo purine biosynthesis: cellular models of de novo purine biosynthesis deficiency disorders.

    Science.gov (United States)

    Duval, Nathan; Luhrs, Kyleen; Wilkinson, Terry G; Baresova, Veronika; Skopova, Vaclava; Kmoch, Stanislav; Vacano, Guido N; Zikanova, Marie; Patterson, David

    2013-03-01

    Purines are molecules essential for many cell processes, including RNA and DNA synthesis, regulation of enzyme activity, protein synthesis and function, energy metabolism and transfer, essential coenzyme function, and cell signaling. Purines are produced via the de novo purine biosynthesis pathway. Mutations in purine biosynthetic genes, for example phosphoribosylaminoimidazole carboxylase/phosphoribosylaminoimidazole succinocarboxamide synthetase (PAICS, E.C. 6.3.2.6/E.C. 4.1.1.21), can lead to developmental anomalies in lower vertebrates. Alterations in PAICS expression in humans have been associated with various types of cancer. Mutations in adenylosuccinate lyase (ADSL, E.C. 4.3.2.2) or 5-aminoimidazole-4-carboxamide ribonucleotide formyltransferase/IMP cyclohydrolase (ATIC, E.C. 2.1.2.3/E.C. 3.5.4.10) lead to inborn errors of metabolism with a range of clinical symptoms, including developmental delay, severe neurological symptoms, and autistic features. The pathogenetic mechanism is unknown for these conditions, and no effective treatments exist. The study of cells carrying mutations in the various de novo purine biosynthesis pathway genes provides one approach to analysis of purine disorders. Here we report the characterization of AdeD Chinese hamster ovary (CHO) cells, which carry genetic mutations encoding p.E177K and p.W363* variants of PAICS. Both mutations impact PAICS structure and completely abolish its biosynthesis. Additionally, we describe a sensitive and rapid analytical method for detection of purine de novo biosynthesis intermediates based on high performance liquid chromatography with electrochemical detection. Using this technique we detected accumulation of AIR in AdeD cells. In AdeI cells, mutant for the ADSL gene, we detected accumulation of SAICAR and SAMP and, somewhat unexpectedly, accumulation of AIR. This method has great potential for metabolite profiling of de novo purine biosynthesis pathway mutants, identification of novel genetic

  5. Sucrose accumulation in watermelon fruits: genetic variation and biochemical analysis.

    Science.gov (United States)

    Yativ, Merav; Harary, Idan; Wolf, Shmuel

    2010-05-15

    Sugar accumulation, the key process determining fruit quality, is controlled by both the translocation of sugars and their metabolism in developing fruits. Sugar composition in watermelon, as in all cucurbit fruits, includes sucrose, fructose and glucose. The proportions of these three sugars are determined primarily by three enzyme families: invertases, sucrose synthases (SuSys) and sucrose phosphate synthases (SPSs). The goal of the present research was to explore the process of sugar metabolism in watermelon fruits. Crosses between the domestic watermelon (Citrullus lanatus) and three wild species provided a wide germplasm to explore genetic variability in sugar composition and metabolism. This survey demonstrated great genetic variability in sugar content and in the proportions of sucrose, glucose and fructose in mature fruits. Genotypes accumulating high and low percentage of sucrose provided an experimental system to study sugar metabolism in developing fruits. Insoluble invertase activity was high and constant throughout fruit development in control lines and in genotypes accumulating low levels of sucrose, while in genotypes accumulating high levels of sucrose, activity declined sharply 4 weeks after pollination. Soluble acid invertase activity was significantly lower in genotypes accumulating high levels of sucrose than in low-sucrose-accumulating genotypes. Conversely, activities of SuSy and SPS were higher in the high-sucrose-accumulating genotypes. The present results establish that, within the genus Citrullus, there are genotypes that accumulate a high percentage of sucrose in the fruit, while others accumulate high percentages of glucose and fructose. The significant negative correlation between insoluble invertase activity and fruit sucrose level suggests that sucrose accumulation is affected by both phloem unloading and sugar metabolism. (c) 2009 Elsevier GmbH. All rights reserved.

  6. Recommendations for reporting results of diagnostic genetic testing (biochemical, cytogenetic and molecular genetic)

    NARCIS (Netherlands)

    Claustres, Mireille; Kozich, Viktor; Dequeker, Els; Fowler, Brain; Hehir-Kwa, Jayne Y.; Miller, Konstantin; Oosterwijk, Cor; Peterlin, Borut; van Ravenswaaij-Arts, Conny; Zimmermann, Uwe; Zuffardi, Orsetta; Hastings, Ros J.; Barton, David E.

    Genetic test results can have considerable importance for patients, their parents and more remote family members. Clinical therapy and surveillance, reproductive decisions and genetic diagnostics in family members, including prenatal diagnosis, are based on these results. The genetic test report

  7. The Genetic Determinism of Biochemical Systems Polymorphous From the Blood Serum in Pigs

    Directory of Open Access Journals (Sweden)

    Nicoleta Işfan

    2010-05-01

    Full Text Available The study of genetic markers and identification of new markers make the subject of an increasing number of research projects in various fields such as genetics of immunology, biochemical genetics, molecular genetics, quantitative genetics and the genetic amelioration of animals. The information provided by electrophoresis graphs has been used to determine the frequency of various categories of alleles (for the loci of pre-albumin, transferines and serum amylases, the frequency of various phenotypes and the genetic structure for each and every locus and, simultaneously, for the loci being studied. The discussion over the varieties of serum proteins was carried on for the purpose of using them as genetic markers, in order to appreciate the levels of genetic unity or diversity within the stock of swine that has been studied. A pair of simple alleles has been determined for each of the three loci. When the three loci were studied simultaneously, out of the 27 possible combinations, only 15 have been found. The sample studied has found to be genetically balanced for every of the three loci. However, when the simultaneous study has been applied, the same sample has not been found genetically balanced.

  8. Genetic and biochemical analysis of peptide transport in Escherichia coli

    International Nuclear Information System (INIS)

    Andrews, J.C.

    1986-01-01

    E. coli peptide transport mutants have been isolated based on their resistance to toxic tripeptides. These genetic defects were found to map in two distinct chromosomal locations. The transport systems which require expression of the trp-linked opp genes and the oppE gene(s) for activity were shown to have different substrate preferences. Growth of E. coli in medium containing leucine results in increased entry of exogenously supplied tripeptides into the bacterial cell. This leucine-mediated elevation of peptide transport required expression of the trp-linked opp operon and was accompanied by increased sensitivity to toxic tripeptides, by an enhanced capacity to utilize nutritional peptides, and by an increase in both the velocity and apparent steady-state level of L-(U- 14 C)alanyl-L-alanyl-L-alanine accumulation for E. coli grown in leucine-containing medium relative to these parameters of peptide transport measured with bacteria grown in media lacking leucine. Direct measurement of opp operon expression by pulse-labeling experiments demonstrated that growth of E. coli in the presence of leucine resulted in increased synthesis of the oppA-encoded periplasmic binding protein. The transcriptional regulation of the trp-linked opp operon of E. coli was investigated using λ placMu51-generated lac operon fusions. Synthesis of β-galactosidase by strains harboring oppA-lac, oppB-lac, and oppD-lac fusions occurred at a basal level when the fusion-containing strains were grown in minimal medium

  9. CRISPR-Cas9-enabled genetic disruptions for understanding ethanol and ethyl acetate biosynthesis in Kluyveromyces marxianus.

    Science.gov (United States)

    Löbs, Ann-Kathrin; Engel, Ronja; Schwartz, Cory; Flores, Andrew; Wheeldon, Ian

    2017-01-01

    The thermotolerant yeast Kluyveromyces marxianus shows promise as an industrial host for the biochemical production of fuels and chemicals. Wild-type strains are known to ferment high titers of ethanol and can effectively convert a wide range of C 5 , C 6 , and C 12 sugars into the volatile short-chain ester ethyl acetate. Strain engineering, however, has been limited due to a lack of advanced genome-editing tools and an incomplete understanding of ester and ethanol biosynthesis. Enabled by the design of hybrid RNA polymerase III promoters, this work adapts the CRISPR-Cas9 system from Streptococcus pyogenes for use in K. marxianus . The system was used to rapidly create functional disruptions to alcohol dehydrogenase (ADH) and alcohol- O -acetyltransferase (ATF) genes with putative function in ethyl acetate and ethanol biosynthesis. Screening of the Km ATF disrupted strain revealed that Atf activity contributes to ethyl acetate biosynthesis, but the knockout reduced ethyl acetate titers by only ~15%. Overexpression experiments revealed that Km Adh7 can catalyze the oxidation of hemiacetal to ethyl acetate. Finally, analysis of the Km ADH2 disrupted strain showed that the knockout almost completely eliminated ethanol production and resulted in the accumulation of acetaldehyde. Newly designed RNA polymerase III promoters for sgRNA expression in K. marxianus enable a CRISPR-Cas9 genome-editing system for the thermotolerant yeast. This system was used to disrupt genes involved in ethyl acetate biosynthesis, specifically Km ADH1-7 and Km ATF. Km Adh2 was found to be critical for aerobic and anaerobic ethanol production. Aerobically produced ethanol supplies the biosynthesis of ethyl acetate catalyzed by Km Atf. Km Adh7 was found to exhibit activity toward the oxidation of hemiacetal, a possible alternative route for the synthesis of ethyl acetate.

  10. Tramadol Induced Adrenal Insufficiency: Histological, Immunohistochemical, Ultrastructural, and Biochemical Genetic Experimental Study

    OpenAIRE

    Abdelaleem, Shereen Abdelhakim; Hassan, Osama A.; Ahmed, Rasha F.; Zenhom, Nagwa M.; Rifaai, Rehab A.; El-Tahawy, Nashwa F.

    2017-01-01

    Tramadol is a synthetic, centrally acting analgesic. It is the most consumed narcotic drug that is prescribed in the world. Tramadol abuse has dramatically increased in Egypt. Long term use of tramadol can induce endocrinopathy. So, the aim of this study was to analyze the adrenal insufficiency induced by long term use of tramadol in experimental animals and also to assess its withdrawal effects through histopathological and biochemical genetic study. Forty male albino rats were used in this ...

  11. [Genetic and biochemical mechanisms of involvement of antioxidant defense enzymes in the development of bronchial asthma].

    Science.gov (United States)

    Polonikov, A V; Ivanov, V P; Bogomazov, A D; Solodilova, M A

    2015-01-01

    In the present review we have analyzed and summarized recent literature data on genetic and biochemical mechanisms responsible for involvement of antioxidant defense enzymes in the etiology and pathogenesis of bronchial asthma. It has been shown that the mechanisms of asthma development are linked with genetically determined abnormalities in the functioning of antioxidant defense enzymes. These alterations are accompanied by a systemic imbalance between oxidative and anti-oxidative reactions with the shift of the redox state toward increased free radical production and oxidative stress, a key element in the pathogenesis of bronchial asthma.

  12. Genetic analysis of pathway regulation for enhancing branched-chain amino acid biosynthesis in plants

    KAUST Repository

    Chen, Hao

    2010-08-01

    The branched-chain amino acids (BCAAs) valine, leucine and isoleucine are essential amino acids that play critical roles in animal growth and development. Animals cannot synthesize these amino acids and must obtain them from their diet. Plants are the ultimate source of these essential nutrients, and they synthesize BCAAs through a conserved pathway that is inhibited by its end products. This feedback inhibition has prevented scientists from engineering plants that accumulate high levels of BCAAs by simply over-expressing the respective biosynthetic genes. To identify components critical for this feedback regulation, we performed a genetic screen for Arabidopsis mutants that exhibit enhanced resistance to BCAAs. Multiple dominant allelic mutations in the VALINE-TOLERANT 1 (VAT1) gene were identified that conferred plant resistance to valine inhibition. Map-based cloning revealed that VAT1 encodes a regulatory subunit of acetohydroxy acid synthase (AHAS), the first committed enzyme in the BCAA biosynthesis pathway. The VAT1 gene is highly expressed in young, rapidly growing tissues. When reconstituted with the catalytic subunit in vitro, the vat1 mutant-containing AHAS holoenzyme exhibits increased resistance to valine. Importantly, transgenic plants expressing the mutated vat1 gene exhibit valine tolerance and accumulate higher levels of BCAAs. Our studies not only uncovered regulatory characteristics of plant AHAS, but also identified a method to enhance BCAA accumulation in crop plants that will significantly enhance the nutritional value of food and feed. © 2010 Blackwell Publishing Ltd.

  13. PR Toxin - Biosynthesis, Genetic Regulation, Toxicological Potential, Prevention and Control Measures: Overview and Challenges.

    Science.gov (United States)

    Dubey, Manish K; Aamir, Mohd; Kaushik, Manish S; Khare, Saumya; Meena, Mukesh; Singh, Surendra; Upadhyay, Ram S

    2018-01-01

    Out of the various mycotoxigenic food and feed contaminant, the fungal species belonging to Penicillium genera, particularly Penicillium roqueforti is of great economic importance, and well known for its crucial role in the manufacturing of Roquefort and Gorgonzola cheese. The mycotoxicosis effect of this mold is due to secretion of several metabolites, of which PR toxin is of considerable importance, with regard to food quality and safety challenges issues. The food products and silages enriched with PR toxin could lead into damage to vital internal organs, gastrointestinal perturbations, carcinogenicity, immunotoxicity, necrosis, and enzyme inhibition. Moreover, it also has the significant mutagenic potential to disrupt/alter the crucial processes like DNA replication, transcription, and translation at the molecular level. The high genetic diversities in between the various strains of P. roqueforti persuaded their nominations with Protected Geographical Indication (PGI), accordingly to the cheese type, they have been employed. Recently, the biosynthetic mechanism and toxicogenetic studies unraveled the role of ari1 and prx gene clusters that cross-talk with the synthesis of other metabolites or involve other cross-regulatory pathways to negatively regulate/inhibit the other biosynthetic route targeted for production of a strain-specific metabolites. Interestingly, the chemical conversion that imparts toxic properties to PR toxin is the substitution/oxidation of functional hydroxyl group (-OH) to aldehyde group (-CHO). The rapid conversion of PR toxin to the other derivatives such as PR imine, PR amide, and PR acid, based on conditions available reflects their unstability and degradative aspects. Since the PR toxin-induced toxicity could not be eliminated safely, the assessment of dose-response and other pharmacological aspects for its safe consumption is indispensable. The present review describes the natural occurrences, diversity, biosynthesis, genetics

  14. PR Toxin – Biosynthesis, Genetic Regulation, Toxicological Potential, Prevention and Control Measures: Overview and Challenges

    Science.gov (United States)

    Dubey, Manish K.; Aamir, Mohd; Kaushik, Manish S.; Khare, Saumya; Meena, Mukesh; Singh, Surendra; Upadhyay, Ram S.

    2018-01-01

    Out of the various mycotoxigenic food and feed contaminant, the fungal species belonging to Penicillium genera, particularly Penicillium roqueforti is of great economic importance, and well known for its crucial role in the manufacturing of Roquefort and Gorgonzola cheese. The mycotoxicosis effect of this mold is due to secretion of several metabolites, of which PR toxin is of considerable importance, with regard to food quality and safety challenges issues. The food products and silages enriched with PR toxin could lead into damage to vital internal organs, gastrointestinal perturbations, carcinogenicity, immunotoxicity, necrosis, and enzyme inhibition. Moreover, it also has the significant mutagenic potential to disrupt/alter the crucial processes like DNA replication, transcription, and translation at the molecular level. The high genetic diversities in between the various strains of P. roqueforti persuaded their nominations with Protected Geographical Indication (PGI), accordingly to the cheese type, they have been employed. Recently, the biosynthetic mechanism and toxicogenetic studies unraveled the role of ari1 and prx gene clusters that cross-talk with the synthesis of other metabolites or involve other cross-regulatory pathways to negatively regulate/inhibit the other biosynthetic route targeted for production of a strain-specific metabolites. Interestingly, the chemical conversion that imparts toxic properties to PR toxin is the substitution/oxidation of functional hydroxyl group (-OH) to aldehyde group (-CHO). The rapid conversion of PR toxin to the other derivatives such as PR imine, PR amide, and PR acid, based on conditions available reflects their unstability and degradative aspects. Since the PR toxin-induced toxicity could not be eliminated safely, the assessment of dose-response and other pharmacological aspects for its safe consumption is indispensable. The present review describes the natural occurrences, diversity, biosynthesis, genetics

  15. Nucleoside antibiotics: biosynthesis, regulation, and biotechnology.

    Science.gov (United States)

    Niu, Guoqing; Tan, Huarong

    2015-02-01

    The alarming rise in antibiotic-resistant pathogens has coincided with a decline in the supply of new antibiotics. It is therefore of great importance to find and create new antibiotics. Nucleoside antibiotics are a large family of natural products with diverse biological functions. Their biosynthesis is a complex process through multistep enzymatic reactions and is subject to hierarchical regulation. Genetic and biochemical studies of the biosynthetic machinery have provided the basis for pathway engineering and combinatorial biosynthesis to create new or hybrid nucleoside antibiotics. Dissection of regulatory mechanisms is leading to strategies to increase the titer of bioactive nucleoside antibiotics. Copyright © 2014. Published by Elsevier Ltd.

  16. Genetic manipulation of putrescine biosynthesis reprograms the cellular transcriptome and the metabolome.

    Science.gov (United States)

    Page, Andrew F; Cseke, Leland J; Minocha, Rakesh; Turlapati, Swathi A; Podila, Gopi K; Ulanov, Alexander; Li, Zhong; Minocha, Subhash C

    2016-05-18

    With the increasing interest in metabolic engineering of plants using genetic manipulation and gene editing technologies to enhance growth, nutritional value and environmental adaptation, a major concern is the potential of undesirable broad and distant effects of manipulating the target gene or metabolic step in the resulting plant. A comprehensive transcriptomic and metabolomic analysis of the product may shed some useful light in this regard. The present study used these two techniques with plant cell cultures to analyze the effects of genetic manipulation of a single step in the biosynthesis of polyamines because of their well-known roles in plant growth, development and stress responses. The transcriptomes and metabolomes of a control and a high putrescine (HP) producing cell line of poplar (Populus nigra x maximowiczii) were compared using microarrays and GC/MS. The HP cells expressed an ornithine decarboxylase transgene and accumulated several-fold higher concentrations of putrescine, with only small changes in spermidine and spermine. The results show that up-regulation of a single step in the polyamine biosynthetic pathway (i.e. ornithine → putrescine) altered the expression of a broad spectrum of genes; many of which were involved in transcription, translation, membrane transport, osmoregulation, shock/stress/wounding, and cell wall metabolism. More than half of the 200 detected metabolites were significantly altered (p ≤ 0.05) in the HP cells irrespective of sampling date. The most noteworthy differences were in organic acids, carbohydrates and nitrogen-containing metabolites. The results provide valuable information about the role of polyamines in regulating nitrogen and carbon use pathways in cell cultures of high putrescine producing transgenic cells of poplar vs. their low putrescine counterparts. The results underscore the complexity of cellular responses to genetic perturbation of a single metabolic step related to nitrogen metabolism

  17. Genetic Profiling of the Isoprenoid and Sterol Biosynthesis Pathway Genes of Trypanosoma cruzi

    Science.gov (United States)

    Cosentino, Raúl O.; Agüero, Fernán

    2014-01-01

    In Trypanosoma cruzi the isoprenoid and sterol biosynthesis pathways are validated targets for chemotherapeutic intervention. In this work we present a study of the genetic diversity observed in genes from these pathways. Using a number of bioinformatic strategies, we first identified genes that were missing and/or were truncated in the T. cruzi genome. Based on this analysis we obtained the complete sequence of the ortholog of the yeast ERG26 gene and identified a non-orthologous homolog of the yeast ERG25 gene (sterol methyl oxidase, SMO), and we propose that the orthologs of ERG25 have been lost in trypanosomes (but not in Leishmanias). Next, starting from a set of 16 T. cruzi strains representative of all extant evolutionary lineages, we amplified and sequenced ∼24 Kbp from 22 genes, identifying a total of 975 SNPs or fixed differences, of which 28% represent non-synonymous changes. We observed genes with a density of substitutions ranging from those close to the average (∼2.5/100 bp) to some showing a high number of changes (11.4/100 bp, for the putative lathosterol oxidase gene). All the genes of the pathway are under apparent purifying selection, but genes coding for the sterol C14-demethylase, the HMG-CoA synthase, and the HMG-CoA reductase have the lowest density of missense SNPs in the panel. Other genes (TcPMK, TcSMO-like) have a relatively high density of non-synonymous SNPs (2.5 and 1.9 every 100 bp, respectively). However, none of the non-synonymous changes identified affect a catalytic or ligand binding site residue. A comparative analysis of the corresponding genes from African trypanosomes and Leishmania shows similar levels of apparent selection for each gene. This information will be essential for future drug development studies focused on this pathway. PMID:24828104

  18. Genetic basis of coaggregation receptor polysaccharide biosynthesis in Streptococcus sanguinis and related species.

    Science.gov (United States)

    Yang, J; Yoshida, Y; Cisar, J O

    2014-02-01

    Interbacterial adhesion between streptococci and actinomyces promotes early dental plaque biofilm development. Recognition of coaggregation receptor polysaccharides (RPS) on strains of Streptococcus sanguinis, Streptococcus gordonii and Streptococcus oralis by Actinomyces spp. type 2 fimbriae is the principal mechanism of these interactions. Previous studies of genetic loci for synthesis of RPS (rps) and RPS precursors (rml, galE1 and galE2) in S. gordonii 38 and S. oralis 34 revealed differences between these strains. To determine whether these differences are strain-specific or species-specific, we identified and compared loci for polysaccharide biosynthesis in additional strains of these species and in several strains of the previously unstudied species, S. sanguinis. Genes for synthesis of RPS precursors distinguished the rps loci of different streptococci. Hence, rml genes for synthesis of TDP-L-Rha were in rps loci of S. oralis strains but at other loci in S. gordonii and S. sanguinis. Genes for two distinct galactose epimerases were also distributed differently. Hence, galE1 for epimerization of UDP-Glc and UDP-Gal was in galactose operons of S. gordonii and S. sanguinis strains but surprisingly, this gene was not present in S. oralis. Moreover, galE2 for epimerization of both UDP-Glc and UDP-Gal and UDP-GlcNAc and UDP-GalNAc was at a different locus in each species, including rps operons of S. sanguinis. The findings provide insight into cell surface properties that distinguish different RPS-producing streptococci and open an approach for identifying these bacteria based on the arrangement of genes for synthesis of polysaccharide precursors. Published 2013. This article is a U.S. Government work and is in the public domain in the USA.

  19. Plant responses to UV and blue light: biochemical and genetic approaches

    International Nuclear Information System (INIS)

    Jenkins, G.I.; Christie, J.M.; Fuglevand, G.; Long, J.C.; Jackson, J.A.

    1995-01-01

    UV and blue light control many aspects of plant growth and development. It is evident that several different photoreceptors mediate responses to UV and blue light, and there are reports of the functional and biochemical characterisation of a putative photoreceptor for phototropism and of the functional and molecular characterisation of the CRY1 photoreceptor, encoded by the Arabidopsis HY4 gene. The CRY1 photoreceptor mediates extension growth and gene expression responses to UV-A/blue light presumably through different or branching signal transduction pathways. Progress has been made in cell physiological and biochemical studies of UV/blue light signal transduction, but much remains to be done to relate candidate UV/blue signal transduction events to particular photoreceptors and responses. The application of a genetic approach in Arabidopsis has been responsible for many advances in understanding UV/blue responses, but further UV-B, UV-A and blue light response mutants need to be isolated. (author)

  20. Acetylcholine receptors and cholinergic ligands: biochemical and genetic aspects in Torpedo californica and Drosophila melanogaster

    International Nuclear Information System (INIS)

    Rosenthal, L.S.

    1987-01-01

    This study evaluates the biochemical and genetic aspects of the acetylcholine receptor proteins and cholinergic ligands in Drosophila melanogaster and Torpedo californica. Included are (1) a comparative study of nicotinic ligand-induced cation release from acetylcholine receptors isolated from Torpedo californica and from Drosophila melanogaster, (2) solution studies of the cholinergic ligands, nikethamide and ethamivan, aimed at measuring internal molecular rotational barriers in solvents of different polarity; and (3) the isolation and characterization of the gene(s) for the acetylcholine receptor in Drosophila melasogaster. Acetylcholine receptor proteins isolated from Drosphila melanogaster heads were found to behave kinetically similar (with regards to cholinergic ligand-induced 155 Eu: 3+ displacement from prelabeled proteins) to receptor proteins isolated from Torpedo californica electric tissue, providing additional biochemical evidence for the existence of a Drosophila acetylcholine receptor

  1. Sterol biosynthesis via cycloartenol and other biochemical features related to photosynthetic phyla in the amoeba Naegleria lovaniensis and Naegleria gruberi.

    Science.gov (United States)

    Raederstorff, D; Rohmer, M

    1987-04-15

    The sterols and sterol precursors of two amoebae of the genus Naegleria, Naegleria lovaniensis and Naegleria gruberi were investigated. Cycloartenol, the sterol precursor in photosynthetic organisms, is present in both amoebae. In N. lovaniesis, it is accompanied by lanosterol and parkeol, as well as by the 24,25-dihydro derivatives of these triterpenes. One of the most striking features of these amoebae is the accumulation of 4 alpha-methylsterols which are present in similar amounts as those of 4,4-desmethylsterols (3-5 mg/g, dry weight). 4 alpha-Methylergosta-7,22-dienol was identified as a new compound. Ergosterol was the major 4,4-desmethylsterol, accompanied by small amounts of C27 and other C28 sterols. Treatment of N. lovaniensis with fenpropimorph modified the sterol pattern of this amoeba and inhibited its growth. This fungicide, known to inhibit steps of sterol biosynthesis in fungi and plants, induced the disappearance of 4 alpha-methyl-delta 7-sterols and the appearance of the unusual delta 6,8,22-ergostatrienol as in A. polyphaga. These results might be explained by a partial inhibition of the delta 8----delta 7 isomerase, the small amounts of delta 7-sterols formed being converted into ergosterol which is still present in fenpropimorph-exposed cells. De novo sterol biosynthesis in N. lovaniensis was shown by incorporation of [1-14C]acetate into sterols and sterol precursors, especially cycloartenol. Lanosterol and parkeol were not significantly labelled. Furthermore, [3-3H]squalene epoxide was efficiently cyclized by a cell-free system of this amoeba into cycloartenol, and again no significant radioactivity was detected in lanosterol and parkeol. This shows that cycloartenol, the sterol precursor in plants and algae, is also the sterol precursor in Naegleria species, and that these amoebae, like A. polyphaga, are related by some biosynthetic pathways to photosynthetic phyla. Lanosterol, the sterol precursor in non-photosynthetic phyla (animal and

  2. Overexpression, purification, and biochemical characterization of GumC, an enzyme involved in the biosynthesis of exopolysaccharide by Xylella fastidiosa.

    Science.gov (United States)

    de Pieri, Celina; Beltramini, Leila M; Selistre-de-Araújo, Heloisa S; Vettore, André L; da Silva, Felipe R; Arruda, Paulo; Oliva, Glaucius; de Souza, Dulce H F

    2004-04-01

    GumC is one of nine enzymes involved in the biosynthesis of fastidian gum, an exopolysaccharide produced by Xylella fastidiosa that may be linked directly to the pathogenicity of the microorganism. GumC may be responsible for gum polymerization or secretion through the membrane of X. fastidiosa. To perform structure and functions studies, we developed an expression system for the production of GumC as a fusion protein with maltose binding protein (MBP) using pMAL-c2x vector. The GumC-MBP fusion protein was expressed as a 94 kDa protein, which strongly reacts with anti-MBP antibodies. GumC-MBP was isolated by affinity chromatography through an amylose column and used to produce antibodies against the fusion protein. After the enzymatic cleavage of MBP, GumC was purified on a Q Sepharose Fast Flow column. GumC showed a molecular weight corresponding to the expected one (52 kDa) and its N-terminal sequence was identical to that deduced from the DNA. The shape of the circular dichroism spectrum was compatible with a folded protein that contains alpha-helical regions in its structure. Therefore, in this study we describe, for the first time, the production of GumC recombinant protein.

  3. Lilium floral fragrance: A biochemical and genetic resource for aroma and flavor.

    Science.gov (United States)

    Johnson, Timothy S; Schwieterman, Michael L; Kim, Joo Young; Cho, Keun H; Clark, David G; Colquhoun, Thomas A

    2016-02-01

    Hybrid Lilium (common name lily) cultivars are among the top produced domestic fresh cut flowers and potted plants in the US today. Many hybrid Lilium cultivars produce large and showy flowers that emit copious amounts of volatile molecules, which can negatively affect a consumer's appreciation or limit use of the plant product. There are few publications focused on the biochemistry, genetics, and/or molecular regulation of floral volatile biosynthesis for Lilium cultivars. In an initial pursuit to provide breeders with molecular markers for floral volatile biosynthesis, a total of five commercially available oriental and oriental-trumpet hybrid Lilium cultivars were selected for analytical characterization of floral volatile emission. In total, 66 volatile molecules were qualified and quantitated among all cultivars. Chemical classes of identified volatiles include monoterpene hydrocarbons, monoterpene alcohols and aldehydes, phenylpropanoids, benzenoids, fatty-acid-derived, nitrogen-containing, and amino-acid-derived compounds. In general, the floral volatile profiles of the three oriental-trumpet hybrids were dominated by monoterpene hydrocarbons, monoterpene alcohols and aldehydes, while the two oriental hybrids were dominated by monoterpene alcohols and aldehydes and phenylpropanoids, respectively. Tepal tissues (two petal whirls) emitted the vast majority of total volatile molecules compared to the reproductive organs of the flowers. Tepal volatile profiles were cultivar specific with a high degree of distinction, which indicates the five cultivars chosen will provide an excellent differential genetic environment for gene discovery through comparative transcriptomics in the future. Cloning and assaying transcript accumulation from four floral volatile biosynthetic candidates provided few immediate or obvious trends with floral volatile emission. Copyright © 2015 Elsevier Ltd. All rights reserved.

  4. Understanding the Constitutive and Induced Biosynthesis of Mono- and Sesquiterpenes in Grapes (Vitis vinifera): A Key to Unlocking the Biochemical Secrets of Unique Grape Aroma Profiles.

    Science.gov (United States)

    Schwab, Wilfried; Wüst, Matthias

    2015-12-16

    The present review integrates current knowledge on mono- and sesquiterpenes in grapes with a special focus on biochemical and physiological aspects. Recent research has impressively shown the prominence of terpenoid metabolism in grapevine (Vitis sp). The 69 putatively functional mono- and sesquiterpene synthases that were identified by the analysis of the updated 12-fold sequencing and assembly of the grapevine genome deliver the scaffolds for structural diversity and display a surprising expansion of the terpene synthase (TPS) gene family in grapevine when compared to other plants like Arabidopsis thaliana (32 TPS). While monoterpenes occur as highly functionalized compounds and are stored as their corresponding glycoconjugates in berry tissues, sesquiterpenes are mainly present as unsaturated hydrocarbons and accumulate in the epicuticular wax layer of intact berries. Interestingly, both groups of terpenes appear to be involved as volatile organic compounds in plant defense and their biosynthesis is enhanced via the jasmonic acid signaling pathway. These novel aspects will help to understand how environmental cues affect the genes and enzymes of various metabolic pathways of relevant wine aroma compounds with numerous links to enology and wine flavor chemistry.

  5. Genetic effects in the leukotriene biosynthesis pathway and association with atherosclerosis

    Science.gov (United States)

    Crosslin, David R.; Shah, Svati H.; Nelson, Sarah C.; Haynes, Carol S.; Connelly, Jessica J.; Gadson, Shera; Goldschmidt-Clermont, Pascal J.; Vance, Jeffery M.; Rose, Jason; Granger, Chris B.; Seo, David; Gregory, Simon G.; Kraus, William E.

    2009-01-01

    Leukotrienes are arachidonic acid derivatives long known for their inflammatory properties and their involvement with a number of human diseases, most particularly asthma. Recently, leukotriene-based inflammation has also been shown to play an important role in atherosclerosis: ALOX5AP and LTA4H, both genes in the leukotriene biosynthesis pathway, have individually been shown to be associated with various cardiovascular disease (CVD) phenotypes. To assess the role of the leukotriene pathway in CVD pathogenesis, we performed genetic association studies of ALOX5AP and LTA4H in a family based study of early onset coronary artery disease (EOCAD) (GENECARD, 1,101 families) and in a non-familial dataset of EOCAD (CATHGEN, 656 cases and 405 controls). We found weak to moderate association between single nucleotide polymorphisms (SNPs) in ALOX5AP and LTA4H with EOCAD. The previously reported four-SNP haplotype (HapA) in ALOX5AP showed association with EOCAD in CATHGEN (P = 0.02), while controlling for age, race and CVD risk factors. HapK, the previously reported ten-SNP haplotype in LTA4H was associated with EOCAD in CATHGEN (P = 0.04). Another previously reported four-SNP haplotype in ALOX5AP (HapB) was not significant in our sample (P = 0.39). The overall lack of (or weak) association of single SNPs as compared with the haplotype results demonstrates the need for analyzing multiple SNPs within each gene in such studies. Interestingly, we detected an association of SNPs in ALOX5 (P < 0.05), the target of ALOX5AP, with CVD. Using a pathway-based approach, we also detected statistical evidence for interactions among ALOX5, ALOX5AP and LTA4H using RNA expression data from a collection of freshly harvested human aortas with varying degrees of atherosclerosis. The GENECARD families did not demonstrate evidence for linkage or association with ALOX5, ALOX5AP or LTA4H. Our results support a modest role for the leukotriene pathway in atherosclerosis pathogenesis, reveal important

  6. Transcriptome analysis of Panax vietnamensis var. fuscidicus discovers putative ocotillol-type ginsenosides biosynthesis genes and genetic markers.

    Science.gov (United States)

    Zhang, Guang-Hui; Ma, Chun-Hua; Zhang, Jia-Jin; Chen, Jun-Wen; Tang, Qing-Yan; He, Mu-Han; Xu, Xiang-Zeng; Jiang, Ni-Hao; Yang, Sheng-Chao

    2015-03-08

    liquid chromatography (HPLC) and evaporative light scattering detector (ELSD). The genomic resources generated from P. vietnamensis var. fuscidiscus provide new insights into the identification of putative genes involved in triterpenoid saponins biosynthesis pathway. This will facilitate our understanding of the biosynthesis of triterpenoid saponins at molecular level. The SSR markers identified and developed in this study show genetic diversity for this important crop and will contribute to marker-assisted breeding for P. vietnamensis var. fuscidiscus.

  7. Genetic and biochemical diversity among Valeriana jatamansi populations from Himachal Pradesh.

    Science.gov (United States)

    Singh, Sunil Kumar; Katoch, Rajan; Kapila, Rakesh Kumar

    2015-01-01

    Valeriana jatamansi Jones is an important medicinal plant that grows wild in Himachal Pradesh, India. Molecular and biochemical diversity among 13 natural populations from Himachal Pradesh was assessed using RAPD and GC-MS to know the extent of existing variation. A total of seven genetically diverse groups have been identified based on RAPD analysis which corroborated well with the analysis based on chemical constituents. The essential oil yield ranged from 0.6% to 1.66% (v/w). A negative correlation between patchouli alcohol and viridiflorol, the two major valued constituents, limits the scope of their simultaneous improvement. However, other few populations like Chamba-II and Kandi-I were found promising for viridiflorol and patchouli alcohol, respectively. The analysis of chemical constitution of oil of the populations from a specific region revealed predominance of specific constituents indicating possibility of their collection/selection for specific end uses like phytomedicines. The prevalence of genetically diverse groups along with sufficient chemical diversity in a defined region clearly indicates the role of ecology in the maintenance of evolution of this species. Sufficient molecular and biochemical diversity detected among natural populations of this species will form basis for the future improvement.

  8. Isolation, biochemical and genetic characterizations of alcohol-producing yeasts from the flowers of Woodfordia fruticosa.

    Science.gov (United States)

    Manwar, Jagdish; Mahadik, Kakasaheb; Paradkar, Anant; Sathiyanarayanan, Lohidasan; Vohra, Mustafa; Patil, Sanjay

    2013-12-01

    To isolate and characterize the alcohol-producing yeasts from Woodfordia fruticosa flowers, which are used for the induction and maintenance of fermentation in the making of Ayurvedic formulations. Initially twenty four yeasts strains were isolated on MGYP agar plate. Among them, four strains were selected for further studies on the basis of their alcohol generation capacity using jaggery media (50% w/v). Physiological, biochemical and genetic characterization (18S rRNA sequencing) of selected strains were carried out. Physiological, biochemical and genetic characterization (18S rRNA sequencing) confirmed the strains as Saccharomycopsis fibuligera Jm.8, S. fibuligera Jm.10, S. fibuligera Jm.16 and Saccharomyces cerevisiae Jm.20. Under the controlled conditions, S. cerevisiae Jm.20 produced 69.57 g/l of alcohol, whereas remaining strains produced the alcohol in the range of 6.04-7.32 g/l. Among selected strains, strains S. fibuligera are a newer in the flowers. Kinetic study of alcohol generation revealed the strain S. cerevisiae Jm.20 can be efficiently used in making of fermented Ayurvedic formulations instead of use W. fruticosa flowers.

  9. Genetic and biochemical differences in populations bred for extremes in maize grain methionine concentration.

    Science.gov (United States)

    Newell, Mark A; Vogel, Karla E; Adams, Marie; Aydin, Nevzat; Bodnar, Anastasia L; Ali, Muhammad; Lauter, Adrienne N Moran; Scott, M Paul

    2014-02-19

    Methionine is an important nutrient in animal feed and several approaches have been developed to increase methionine concentration in maize (Zea mays L.) grain. One approach is through traditional breeding using recurrent selection. Using divergent selection, genetically related populations with extreme differences in grain methionine content were produced. In order to better understand the molecular mechanisms controlling grain methionine content, we examined seed proteins, transcript levels of candidate genes, and genotypes of these populations. Two populations were selected for high or low methionine concentration for eight generations and 40 and 56% differences between the high and low populations in grain methionine concentration were observed. Mean values between the high and low methionine populations differed by greater than 1.5 standard deviations in some cycles of selection. Other amino acids and total protein concentration exhibited much smaller changes. In an effort to understand the molecular mechanisms that contribute to these differences, we compared transcript levels of candidate genes encoding high methionine seed storage proteins involved in sulfur assimilation or methionine biosynthesis. In combination, we also explored the genetic mechanisms at the SNP level through implementation of an association analysis. Significant differences in methionine-rich seed storage protein genes were observed in comparisons of high and low methionine populations, while transcripts of seed storage proteins lacking high levels of methionine were unchanged. Seed storage protein levels were consistent with transcript levels. Two genes involved in sulfur assimilation, Cys2 and CgS1 showed substantial differences in allele frequencies when two selected populations were compared to the starting populations. Major genes identified across cycles of selection by a high-stringency association analysis included dzs18, wx, dzs10, and zp27. We hypothesize that transcriptional

  10. Biochemical traits useful for the determination of genetic variation in a natural population of Myracrodruon urundeuva

    Directory of Open Access Journals (Sweden)

    Abdala Ludmila

    2002-01-01

    Full Text Available The objectives of this work were to analyze seeds from 20 trees of aroeira (Myracrodruon urundeuva Fr. All. of a natural population located in the region of Selvíria, State of Mato Grosso do Sul, Brazil, in order to evaluate their protein, lipid and carbohydrate contents, and to estimate their genetic variation. A completely randomized experimental design consisting of 20 treatments (families was set up, with two replications. Four types of proteins were detected: albumin (35.0 to 107.3 mg/g seed, globulin (3.4 to 9.3 mg/g, prolamin (60.0 to 135.2 mg/g and glutelin (118.0 to 286.0 mg/g. The lipid content varied between 200 and 334 mg/g seed. The total sugars also varied (26.5 to 46.3 mg/g seed, with a predominance of polyols (arabinitol, mannitol, glucitol and xylitol. The main monosaccharides detected were glucose and arabinose. Total hydrolysis of the sugars indicated the presence of neutral arabinan and xylan oligosaccharides. The starch content varied from 0.35 to 1.58 mg/g seed. These biochemical traits showed considerable genetic variability, indicating that only the collection of seeds from many different trees can provide a representative sample of the population for conservation and genetic improvement.

  11. Genetic and biochemical markers of hydroxyurea therapeutic response in sickle cell anemia.

    Science.gov (United States)

    Silva, Danilo Grunig Humberto; Belini Junior, Edis; Carrocini, Gisele Cristine de Souza; Torres, Lidiane de Souza; Ricci Júnior, Octávio; Lobo, Clarisse Lopes de Castro; Bonini-Domingos, Claudia Regina; de Almeida, Eduardo Alves

    2013-10-09

    Sickle cell anemia (SCA) presents a complex pathophysiology which can be affected by a number of modifying factors, including genetic and biochemical ones. In Brazil, there have been no studies verifying βS-haplotypes effect on oxidative stress parameters. This study evaluated βS-haplotypes and Hb F levels effects on oxidative stress markers and their relationship with hydroxyurea (HU) treatment in SCA patients. The studied group was composed by 28 SCA patients. Thirteen of these patients were treated with HU and 15 of them were not. We used molecular methodology (PCR-RFLP) for hemoglobin S genotype confirmation and haplotypes identification. Biochemical parameters were measured using spectrophotometric methods (Thiobarbituric-acid-reactive substances and Trolox equivalent antioxidant capacity levels, catalase and GST activities) and plasma glutathione levels by High-performance liquid chromatography coupled to electrochemical detection. We found the highest frequency of Bantu haplotype (48.2%) which was followed by Benin (32.1%). We observed also the presence of Cameroon haplotype, rare in Brazilian population and 19.7% of atypical haplotypes. The protective Hb F effect was confirmed in SCA patients because these patients showed an increase in Hb F levels that resulted in a 41.3% decrease on the lipid peroxidation levels (r =-0.74, p=0.01). Other biochemical parameters have not shown differential expression according to patient's haplotypes. Bantu haplotype presence was related to the highest lipid peroxidation levels in patients (p < 0,01), but it also conferred a differential response to HU treatment, raising Hb F levels in 52.6% (p = 0.03) when compared with the group with the same molecular profile without HU usage. SCA patients with Bantu haplotype showed the worst oxidative status. However these patients also demonstrated a better response to the treatment with HU. Such treatment seems to have presented a "haplotype-dependent" pharmacological

  12. A comparative clinical, pathological, biochemical and genetic study of fused in sarcoma proteinopathies

    DEFF Research Database (Denmark)

    Lashley, Tammaryn; Rohrer, Jonathan D; Bandopadhyay, Rina

    2011-01-01

    Neuronal intermediate filament inclusion disease and atypical frontotemporal lobar degeneration are rare diseases characterized by ubiquitin-positive inclusions lacking transactive response DNA-binding protein-43 and tau. Recently, mutations in the fused in sarcoma gene have been shown to cause...... familial amyotrophic lateral sclerosis and fused in sarcoma-positive neuronal inclusions have subsequently been demonstrated in neuronal intermediate filament inclusion disease and atypical frontotemporal lobar degeneration with ubiquitinated inclusions. Here we provide clinical, imaging, morphological...... findings, as well as genetic and biochemical data in 14 fused in sarcoma proteinopathy cases. In this cohort, the age of onset was variable but included cases of young-onset disease. Patients with atypical frontotemporal lobar degeneration with ubiquitinated inclusions all presented with behavioural...

  13. Genetic Confirmation of the Role of Sulfopyruvate Decarboxylase in Coenzyme M Biosynthesis in Methanococcus maripaludis

    Directory of Open Access Journals (Sweden)

    Felipe Sarmiento

    2013-01-01

    Full Text Available Coenzyme M is an essential coenzyme for methanogenesis. The proposed biosynthetic pathway consists of five steps, of which the fourth step is catalyzed by sulfopyruvate decarboxylase (ComDE. Disruption of the gene comE by transposon mutagenesis resulted in a partial coenzyme M auxotroph, which grew poorly in the absence of coenzyme M and retained less than 3% of the wild type level of coenzyme M biosynthesis. Upon coenzyme M addition, normal growth of the mutant was restored. Moreover, complementation of the mutation with the wild type comE gene in trans restored full growth in the absence of coenzyme M. These results confirm that ComE plays an important role in coenzyme M biosynthesis. The inability to yield a complete CoM auxotroph suggests that either the transposon insertion failed to completely inactivate the gene or M. maripaludis possesses a promiscuous activity that partially complemented the mutation.

  14. Genetic confirmation of the role of sulfopyruvate decarboxylase in coenzyme M biosynthesis in Methanococcus maripaludis.

    Science.gov (United States)

    Sarmiento, Felipe; Ellison, Courtney K; Whitman, William B

    2013-01-01

    Coenzyme M is an essential coenzyme for methanogenesis. The proposed biosynthetic pathway consists of five steps, of which the fourth step is catalyzed by sulfopyruvate decarboxylase (ComDE). Disruption of the gene comE by transposon mutagenesis resulted in a partial coenzyme M auxotroph, which grew poorly in the absence of coenzyme M and retained less than 3% of the wild type level of coenzyme M biosynthesis. Upon coenzyme M addition, normal growth of the mutant was restored. Moreover, complementation of the mutation with the wild type comE gene in trans restored full growth in the absence of coenzyme M. These results confirm that ComE plays an important role in coenzyme M biosynthesis. The inability to yield a complete CoM auxotroph suggests that either the transposon insertion failed to completely inactivate the gene or M. maripaludis possesses a promiscuous activity that partially complemented the mutation.

  15. Genetic and biochemical changes of the serotonergic system in migraine pathobiology.

    Science.gov (United States)

    Gasparini, Claudia Francesca; Smith, Robert Anthony; Griffiths, Lyn Robyn

    2017-12-01

    Migraine is a brain disorder characterized by a piercing headache which affects one side of the head, located mainly at the temples and in the area around the eye. Migraine imparts substantial suffering to the family in addition to the sufferer, particularly as it affects three times more women than men and is most prevalent between the ages of 25 and 45, the years of child rearing. Migraine typically occurs in individuals with a genetic predisposition and is aggravated by specific environmental triggers. Attempts to study the biochemistry of migraine began as early as the 1960s and were primarily directed at serotonin metabolism after an increase of 5-hydroxyindoleacetic acid (5-HIAA), the main metabolite of serotonin was observed in urine of migraineurs. Genetic and biochemical studies have primarily focused on the neurotransmitter serotonin, considering receptor binding, transport and synthesis of serotonin and have investigated serotonergic mediators including enzymes, receptors as well as intermediary metabolites. These studies have been mainly assayed in blood, CSF and urine as the most accessible fluids. More recently PET imaging technology integrated with a metabolomics and a systems biology platform are being applied to study serotonergic biology. The general trend observed is that migraine patients have alterations of neurotransmitter metabolism detected in biological fluids with different biochemistry from controls, however the interpretation of the biological significance of these peripheral changes is unresolved. In this review we present the biology of the serotonergic system and metabolic routes for serotonin and discuss results of biochemical studies with regard to alterations in serotonin in brain, cerebrospinal fluid, saliva, platelets, plasma and urine of migraine patients.

  16. Increased cell hydration promotes both tumor growth and metastasis: a biochemical mechanism consistent with genetic signatures.

    Science.gov (United States)

    McIntyre, G I

    2007-01-01

    It was postulated previously that a progressive increase in cell hydration, induced by successive genetic or epigenetic changes, is the basic mechanism of multistep carcinogenesis, and also that the degree of malignancy increases with the degree of cell hydration. These hypotheses implied that increased cell hydration is a common factor promoting both tumor growth and metastasis, and that metastatic potential increases with the degree of cell hydration. This paper discusses these implications in relation to current concepts of genetic mechanisms determining the acquisition of metastatic potential. It was also postulated previously that the enhancement of metabolic activity by increased cell hydration will increase the ability of tumor cells to compete for nutrients with their normal counterparts. This effect may favor the preferential selection of cells whose genotypes confer the greatest increase in cell hydration and which, on the present hypothesis, would be those with the greatest capacity for metastasis. An important feature of this "common factor" hypothesis is that it suggests a biochemical explanation for DNA-microarray data showing a similarity between the gene expression patterns associated with both tumor growth and metastasis, while the postulated role of genes causing increased cell hydration might explain the apparent acquisition of metastatic potential at an early stage of tumorigenesis. Previous investigations were consistent with the hypothesis that various factors promoting carcinogenesis may do so by increasing cell hydration. A survey of the literature showed that all of these factors also promote cell motility, migration or metastasis, and provided evidence that these effects could be attributed to the associated increase in cell hydration. Methods are suggested for testing the hypothesis, and the paper concludes by emphasizing the need for more research on the biochemistry of cancer, and on the role of water as a biochemical factor of

  17. Haploid Mammalian Genetic Screen Identifies UBXD8 as a Key Determinant of HMGCR Degradation and Cholesterol Biosynthesis.

    Science.gov (United States)

    Loregger, Anke; Raaben, Matthijs; Tan, Josephine; Scheij, Saskia; Moeton, Martina; van den Berg, Marlene; Gelberg-Etel, Hila; Stickel, Elmer; Roitelman, Joseph; Brummelkamp, Thijn; Zelcer, Noam

    2017-11-01

    The cellular demand for cholesterol requires control of its biosynthesis by the mevalonate pathway. Regulation of HMGCR (3-hydroxy-3-methylglutaryl coenzyme A reductase), a rate-limiting enzyme in this pathway and the target of statins, is a key control point herein. Accordingly, HMGCR is subject to negative and positive regulation. In particular, the ability of oxysterols and intermediates of the mevalonate pathway to stimulate its proteasomal degradation is an exquisite example of metabolically controlled feedback regulation. To define the genetic determinants that govern this process, we conducted an unbiased haploid mammalian genetic screen. We generated human haploid cells with mNeon fused to endogenous HMGCR using CRISPR/Cas9 and used these cells to interrogate regulation of HMGCR abundance in live cells. This resulted in identification of known and new regulators of HMGCR, and among the latter, UBXD8 (ubiquitin regulatory X domain-containing protein 8), a gene that has not been previously implicated in this process. We demonstrate that UBXD8 is an essential determinant of metabolically stimulated degradation of HMGCR and of cholesterol biosynthesis in multiple cell types. Accordingly, UBXD8 ablation leads to aberrant cholesterol synthesis due to loss of feedback control. Mechanistically, we show that UBXD8 is necessary for sterol-stimulated dislocation of ubiquitylated HMGCR from the endoplasmic reticulum membrane en route to proteasomal degradation, a function dependent on its UBX domain. We establish UBXD8 as a previously unrecognized determinant that couples flux across the mevalonate pathway to control of cholesterol synthesis and demonstrate the feasibility of applying mammalian haploid genetics to study metabolic traits. © 2017 The Authors.

  18. The genetic origins of biosynthesis and light-responsive control of the chemical UV screen of land plants

    International Nuclear Information System (INIS)

    Jorgensen, R.

    1994-01-01

    Most land plants possess the capacity to protect themselves from UV light, and do so by producing pigments that absorb efficiently in the UV-A and UV-B regions of the spectrum while allowing transmission of nearly all photosynthetically useful wavelengths. These UV-absorbing pigments are mainly phenylpropanoids and flavonoids. This chapter summarizes current understanding of the mechanism of UV protection in higher land plants, evaluates the information available from lower land plants and their green-algal relatives, and then considers the possible evolutionary origins of this use of chemical filters for selectively screening UV light from solar radiation. It is proposed that photo control over the biosynthesis of UV-absorbing phenylpropanoids and flavonoids may have evolved in concert with the evolution of the high biosynthetic activity necessary for UV protection. The toxicity of phenylpropanoids and flavonoids has been postulated to have been a barrier to the evolution of an effective chemical UV screen, and that some means for sequestering these compounds and/or for controlling their synthesis probably evolved prior to, or in concert with, the evolution of high rates of biosynthesis. The original photoreceptor and signal transduction system is speculated to have been based on photo isomerization of a phenylpropanoid ester and a pre-existing product feedback mechanism for controlling phenylpropanoid biosynthesis. Understanding the original mechanism for photo control of the chemical UV screen of land plants could be valuable for understanding the adaptability of extant land plants to rising levels of solar UV-B radiation and may suggest genetic strategies for engineering improved UV tolerance in crop plants. (author)

  19. Genetic and Biochemical Alterations in Non-Small Cell Lung Cancer

    Directory of Open Access Journals (Sweden)

    Jackie L. Johnson

    2012-01-01

    Full Text Available Despite significant advances in the detection and treatment of lung cancer, it causes the highest number of cancer-related mortality. Recent advances in the detection of genetic alterations in patient samples along with physiologically relevant animal models has yielded a new understanding of the molecular etiology of lung cancer. This has facilitated the development of potent and specific targeted therapies, based on the genetic and biochemical alterations present in the tumor, especially non-small-cell lung cancer (NSCLC. It is now clear that heterogeneous cell signaling pathways are disrupted to promote NSCLC, including mutations in critical growth regulatory proteins (K-Ras, EGFR, B-RAF, MEK-1, HER2, MET, EML-4-ALK, KIF5B-RET, and NKX2.1 and inactivation of growth inhibitory pathways (TP53, PTEN, p16, and LKB-1. How these pathways differ between smokers and non-smokers is also important for clinical treatment strategies and development of targeted therapies. This paper describes these molecular targets in NSCLC, and describes the biological significance of each mutation and their potential to act as a therapeutic target.

  20. Tramadol Induced Adrenal Insufficiency: Histological, Immunohistochemical, Ultrastructural, and Biochemical Genetic Experimental Study

    Directory of Open Access Journals (Sweden)

    Shereen Abdelhakim Abdelaleem

    2017-01-01

    Full Text Available Tramadol is a synthetic, centrally acting analgesic. It is the most consumed narcotic drug that is prescribed in the world. Tramadol abuse has dramatically increased in Egypt. Long term use of tramadol can induce endocrinopathy. So, the aim of this study was to analyze the adrenal insufficiency induced by long term use of tramadol in experimental animals and also to assess its withdrawal effects through histopathological and biochemical genetic study. Forty male albino rats were used in this study. The rats were divided into 4 groups (control group, tramadol-treated group, and withdrawal groups. Tramadol was given to albino rats at a dose of 80 mg/kg body weight for 3 months and after withdrawal periods (7–15 days rats were sacrificed. Long term use of tramadol induced severe histopathological changes in adrenal glands. Tramadol decreased the levels of serum cortisol and DHEAS hormones. In addition, it increased the level of adrenal MDA and decreased the genetic expression of glutathione peroxidase and thioredoxin reductase in adrenal gland tissues. All these changes started to return to normal after withdrawal of tramadol. Thus, it was confirmed that long term use of tramadol can induce severe adrenal insufficiency.

  1. Tramadol Induced Adrenal Insufficiency: Histological, Immunohistochemical, Ultrastructural, and Biochemical Genetic Experimental Study.

    Science.gov (United States)

    Abdelaleem, Shereen Abdelhakim; Hassan, Osama A; Ahmed, Rasha F; Zenhom, Nagwa M; Rifaai, Rehab A; El-Tahawy, Nashwa F

    2017-01-01

    Tramadol is a synthetic, centrally acting analgesic. It is the most consumed narcotic drug that is prescribed in the world. Tramadol abuse has dramatically increased in Egypt. Long term use of tramadol can induce endocrinopathy. So, the aim of this study was to analyze the adrenal insufficiency induced by long term use of tramadol in experimental animals and also to assess its withdrawal effects through histopathological and biochemical genetic study. Forty male albino rats were used in this study. The rats were divided into 4 groups (control group, tramadol-treated group, and withdrawal groups). Tramadol was given to albino rats at a dose of 80 mg/kg body weight for 3 months and after withdrawal periods (7-15 days) rats were sacrificed. Long term use of tramadol induced severe histopathological changes in adrenal glands. Tramadol decreased the levels of serum cortisol and DHEAS hormones. In addition, it increased the level of adrenal MDA and decreased the genetic expression of glutathione peroxidase and thioredoxin reductase in adrenal gland tissues. All these changes started to return to normal after withdrawal of tramadol. Thus, it was confirmed that long term use of tramadol can induce severe adrenal insufficiency.

  2. [Medico-genetic study of isolates in Uzbekistan. IV. Clinico-biochemical diagnosis of hereditary diseases].

    Science.gov (United States)

    Kozlova, S I; Diachenko, S S; Khannanova, F K; Kuleshov, N P; Khodzhaeva, G K

    1976-01-01

    An exhaustive clinico-biochemical examination of the population of two kishlaks of the Samarkand Region, viz. Karakent (210 persons) and Ishan (248 persons) was carried out. The program of this examination permitted to exclude over 160 forms of hereditary pathology. A total of 45 persons affected with diseases belonging to 12 nosological forms were revealed in the course of the examination. Among the diseases observed only 5 are hereditary sensu stricto, viz. myoclonus-epilepsy, Bonevi-Ulrich's syndrome, imperfect osteogenesis, pigment choreoretinite and Down's syndrome, others belong to diseases with a pronounced hereditary predisposition. The main part of this group comprises neuro-psychic diseases, such as non-differentiated olygophreny (5.0%), epilepsy (1.3%), schizophreny; many of these cases have a familial character, particularly in Karakent. Besides the persons suffering from diseases, 20 heterozygous carriers of beta-thalassemia and 17 heterozygous carriers of G6PD-deficiency were discovered in the kishlaks examined. On the whole the frequency of the diseases revealed did not exceed the level in the general population. Despite the different degree of isolation of the kishlaks examined (Karakent is isolated on a religious basis, F = 0.0064; while Ishan is a desintagrated isolate, F = = 0.0014), no substantial differences between them in the distribution of pathological phenomena were observed. On the basis of the experience of this expedition recomendations are proposed concerning the origination and accomplishment of medico-genetic expeditions. A scheme is proposed for the performance of medico-genetic examination through several stages. The first stage in the composition of tentative maps of the distribution of hereditary diseases within a region on the basis of the information obtained from the medical personnel and from the examination of the documents of district and regional hospitals. Subsequently the primary information is specified, the regions to

  3. Genetic and biochemical heterogeneity in patients with the rhizomelic form of chondrodysplasia punctata--a complementation study

    NARCIS (Netherlands)

    Heikoop, J. C.; Wanders, R. J.; Strijland, A.; Purvis, R.; Schutgens, R. B.; Tager, J. M.

    1992-01-01

    The genetic relationship between 10 patients with clinical manifestations of rhizomelic chondrodysplasia punctata (RCDP) was studied by complementation analysis after somatic cell fusion. Biochemically, 9 out of the 10 patients were characterized by a partial deficiency of acyl-CoA: dihydroxyacetone

  4. Behavioral, biochemical, and genetic analysis of iron metabolism in high-intensity blood donors.

    Science.gov (United States)

    Mast, Alan E; Foster, Tisha M; Pinder, Holly L; Beczkiewicz, Craig A; Bellissimo, Daniel B; Murphy, Anthony T; Kovacevic, Steve; Wroblewski, Victor J; Witcher, Derrick R

    2008-10-01

    Individuals donating whole blood 13 times in a 2-year period without development of iron deficiency anemia (superdonors) are a self-selected population that is deferred for low hematocrit (Hct) level less frequently than other donors. Iron metabolism was assessed in 138 superdonors through a questionnaire and measurement of Hct, serum ferritin, serum hepcidin, and serum growth differentiation factor 15 (GDF15). Genetic testing for HFE and JAK-2 mutations was also performed. Iron deficiency (ferritin level, <30 microg/L) is present in more than 60 percent of superdonors. Behaviors altering iron status included casual use of iron supplements in males, but not in females, and cigarette smoking that produced increased Hct associated with decreased ferritin. The striking biochemical characteristic of superdonors is greatly decreased serum hepcidin, consistent with their need to absorb maximal amounts of dietary iron to replace that lost from blood donation. GDF15 is normal in most superdonors, indicating that GDF15 overexpression arising from the expanded erythroid pool necessary to replace donated red cells is not the biochemical mechanism for the decreased serum hepcidin. Mutations in JAK-2 were not found, indicating that undiagnosed polycythemia vera is not a common cause for successful repeated blood donation by superdonors. Mutations in HFE associated with hemochromatosis were present in superdonors at the same frequency as the normal population. However, superdonors heterozygous for the H63D mutation in HFE had significantly decreased hepcidin : ferritin ratios demonstrating for the first time that the heterozygous state for HFE mutations is associated with alterations in hepcidin expression.

  5. Clinical, biochemical, and genetic spectrum of seven new patients with NFU1 deficiency

    Directory of Open Access Journals (Sweden)

    Uwe eAhting

    2015-04-01

    Full Text Available Disorders of the mitochondrial energy metabolism are clinically and genetically heterogeneous. An increasingly recognized subgroup is caused by defective mitochondrial iron-sulfur (Fe-S cluster biosynthesis, with defects in 13 genes being linked to human disease to date. Mutations in three of them, NFU1, BOLA3, and IBA57 affect the assembly of mitochondrial [4Fe-4S] proteins leading to an impairment of diverse mitochondrial metabolic pathways and ATP production. Patients with defects in these three genes present with lactic acidosis, hyperglycinemia, and reduced activities of respiratory chain complexes I and II, the four lipoic acid-dependent 2-oxoacid dehydrogenases, and the glycine cleavage system (GCS. To date, 5 different NFU1 pathogenic variants have been reported in 15 patients from 12 families. We report on 7 new patients from 5 families carrying compound heterozygous or homozygous pathogenic NFU1 mutations identified by candidate gene screening and exome sequencing. 6 out of 8 different disease alleles were novel and functional studies were performed to support the pathogenicity of 5 of them. Characteristic clinical features included fatal infantile encephalopathy and pulmonary hypertension leading to death within the first 6 months of life in 6 out of 7 patients. Laboratory investigations revealed combined defects of PDHc (5 out of 5 and respiratory chain complexes I and II+III (4 out of 5 in skeletal muscle and/or fibroblasts as well as elevated lactate (5 out of 6 and glycine levels (7 out of 7. Our study adds to the definition of the phenotypic spectrum associated with NFU1 mutations and might contribute to the diagnostic workup of future patients

  6. New function for Escherichia coli xanthosine phophorylase (xapA): genetic and biochemical evidences on its participation in NAD+ salvage from nicotinamide

    Science.gov (United States)

    2014-01-01

    Background In an effort to reconstitute the NAD+ synthetic pathway in Escherichia coli (E. coli), we produced a set of gene knockout mutants with deficiencies in previously well-defined NAD+de novo and salvage pathways. Unexpectedly, the mutant deficient in NAD+de novo and salvage pathway I could grow in M9/nicotinamide medium, which was contradictory to the proposed classic NAD+ metabolism of E. coli. Such E. coli mutagenesis assay suggested the presence of an undefined machinery to feed nicotinamide into the NAD+ biosynthesis. We wanted to verify whether xanthosine phophorylase (xapA) contributed to a new NAD+ salvage pathway from nicotinamide. Results Additional knockout of xapA further slowed down the bacterial growth in M9/nicotinamide medium, whereas the complementation of xapA restored the growth phenotype. To further validate the new function of xapA, we cloned and expressed E. coli xapA as a recombinant soluble protein. Biochemical assay confirmed that xapA was capable of using nicotinamide as a substrate for nicotinamide riboside formation. Conclusions Both the genetic and biochemical evidences indicated that xapA could convert nicotinamide to nicotinamide riboside in E. coli, albeit with relatively weak activity, indicating that xapA may contribute to a second NAD+ salvage pathway from nicotinamide. We speculate that this xapA-mediated NAD+ salvage pathway might be significant in some bacteria lacking NAD+de novo and NAD+ salvage pathway I or II, to not only use nicotinamide riboside, but also nicotinamide as precursors to synthesize NAD+. However, this speculation needs to be experimentally tested. PMID:24506841

  7. New function for Escherichia coli xanthosine phophorylase (xapA): genetic and biochemical evidences on its participation in NAD(+) salvage from nicotinamide.

    Science.gov (United States)

    Dong, Wei-Ren; Sun, Cen-Cen; Zhu, Guan; Hu, Shi-Hua; Xiang, Li-Xin; Shao, Jian-Zhong

    2014-02-08

    In an effort to reconstitute the NAD(+) synthetic pathway in Escherichia coli (E. coli), we produced a set of gene knockout mutants with deficiencies in previously well-defined NAD(+)de novo and salvage pathways. Unexpectedly, the mutant deficient in NAD(+) de novo and salvage pathway I could grow in M9/nicotinamide medium, which was contradictory to the proposed classic NAD(+) metabolism of E. coli. Such E. coli mutagenesis assay suggested the presence of an undefined machinery to feed nicotinamide into the NAD(+) biosynthesis. We wanted to verify whether xanthosine phophorylase (xapA) contributed to a new NAD(+) salvage pathway from nicotinamide. Additional knockout of xapA further slowed down the bacterial growth in M9/nicotinamide medium, whereas the complementation of xapA restored the growth phenotype. To further validate the new function of xapA, we cloned and expressed E. coli xapA as a recombinant soluble protein. Biochemical assay confirmed that xapA was capable of using nicotinamide as a substrate for nicotinamide riboside formation. Both the genetic and biochemical evidences indicated that xapA could convert nicotinamide to nicotinamide riboside in E. coli, albeit with relatively weak activity, indicating that xapA may contribute to a second NAD(+) salvage pathway from nicotinamide. We speculate that this xapA-mediated NAD(+) salvage pathway might be significant in some bacteria lacking NAD(+) de novo and NAD(+) salvage pathway I or II, to not only use nicotinamide riboside, but also nicotinamide as precursors to synthesize NAD(+). However, this speculation needs to be experimentally tested.

  8. A genetic anomaly of oriented collagen biosynthesis and cross-linking: Keratoconus.

    Science.gov (United States)

    Bourges, J L; Robert, A M; Robert, L

    2015-02-01

    Oriented collagen biosynthesis is one of the major mechanisms involved in tissue and organ formation during development. Corneal biogenesis is one example. Defects in this process lead to anomalies in tissue structure and function. The transparency of cornea and its achievement are a good example as well as its pathological modifications. Keratoconus is one example of this type of pathologies, involving also inappropriate cross-linking of collagen fibers. Among the tentatives to correct this anomaly, the riboflavin-potentiated UV-cross-linking (CXL) of keratoconus corneas appears clinically satisfactory, although none of the experiments and clinical results published prove effective cross-linking. The published results are reviewed in this article. Copyright © 2014. Published by Elsevier SAS.

  9. Genetic manipulation of carotenoid biosynthesis in the green sulfur bacterium Chlorobium tepidum

    DEFF Research Database (Denmark)

    Frigaard, Niels-Ulrik; Maresca, Julia A; Yunker, Colleen E

    2004-01-01

    The green sulfur bacterium Chlorobium tepidum is a strict anaerobe and an obligate photoautotroph. On the basis of sequence similarity with known enzymes or sequence motifs, nine open reading frames encoding putative enzymes of carotenoid biosynthesis were identified in the genome sequence of C....... tepidum, and all nine genes were inactivated. Analysis of the carotenoid composition in the resulting mutants allowed the genes encoding the following six enzymes to be identified: phytoene synthase (crtB/CT1386), phytoene desaturase (crtP/CT0807), zeta-carotene desaturase (crtQ/CT1414), gamma......-carotene desaturase (crtU/CT0323), carotenoid 1',2'-hydratase (crtC/CT0301), and carotenoid cis-trans isomerase (crtH/CT0649). Three mutants (CT0180, CT1357, and CT1416 mutants) did not exhibit a discernible phenotype. The carotenoid biosynthetic pathway in C. tepidum is similar to that in cyanobacteria and plants...

  10. Environmental, biochemical and genetic drivers of DMSP degradation and DMS production in the Sargasso Sea.

    Science.gov (United States)

    Levine, Naomi Marcil; Varaljay, Vanessa A; Toole, Dierdre A; Dacey, John W H; Doney, Scott C; Moran, Mary Ann

    2012-05-01

    Dimethylsulfide (DMS) is a climatically relevant trace gas produced and cycled by the surface ocean food web. Mechanisms driving intraannual variability in DMS production and dimethylsulfoniopropionate (DMSP) degradation in open-ocean, oligotrophic regions were investigated during a 10-month time-series at the Bermuda Atlantic Time-series Study site in the Sargasso Sea. Abundance and transcription of bacterial DMSP degradation genes, DMSP lyase enzyme activity, and DMS and DMSP concentrations, consumption rates and production rates were quantified over time and depth. This interdisciplinary data set was used to test current hypotheses of the role of light and carbon supply in regulating upper-ocean sulfur cycling. Findings supported UV-A-dependent phytoplankton DMS production. Bacterial DMSP degraders may also contribute significantly to DMS production when temperatures are elevated and UV-A dose is moderate, but may favour DMSP demethylation under low UV-A doses. Three groups of bacterial DMSP degraders with distinct intraannual variability were identified and niche differentiation was indicated. The combination of genetic and biochemical data suggest a modified 'bacterial switch' hypothesis where the prevalence of different bacterial DMSP degradation pathways is regulated by a complex set of factors including carbon supply, temperature and UV-A dose. © 2012 Society for Applied Microbiology and Blackwell Publishing Ltd.

  11. Purification of mutacin III from group III Streptococcus mutans UA787 and genetic analyses of mutacin III biosynthesis genes.

    Science.gov (United States)

    Qi, F; Chen, P; Caufield, P W

    1999-09-01

    Previously, members of our group reported the isolation and characterization of mutacin II from Streptococcus mutans T8 and the genetic analyses of the mutacin II biosynthesis genes (J. Novak, P. W. Caufield, and E. J. Miller, J. Bacteriol. 176:4316-4320, 1994; F. Qi, P. Chen, and P. W. Caufield, Appl. Environ. Microbiol. 65:652-658, 1999; P. Chen, F. Qi, J. Novak, and P. W. Caufield, Appl. Environ. Microbiol. 65:1356-1360, 1999). In this study, we cloned and sequenced the mutacin III biosynthesis gene locus from a group III strain of S. mutans, UA787. DNA sequence analysis revealed eight open reading frames, which we designated mutR, -A, -A', -B, -C, -D, -P, and -T. MutR bears strong homology with MutR of mutacin II, while MutA, -B, -C, -D, -P, and -T are counterparts of proteins in the lantibiotic epidermin group. MutA' has 60% amino acid identity with MutA and therefore appears to be a duplicate of MutA. Insertional inactivation demonstrated that mutA is an essential gene for mutacin III production, while mutA' is not required. Mutacin III was purified to homogeneity by using reverse-phase high-pressure liquid chromatography. N-terminal peptide sequencing of the purified mutacin III determined mutA to be the structural gene for prepromutacin III. The molecular mass of the purified peptide was measured by laser disorption mass spectrophotometry and found to be 2,266.43 Da, consistent with our supposition that mutacin III has posttranslational modifications similar to those of the lantibiotic epidermin.

  12. Biochemical and genetic characterization of three molybdenum cofactor hydroxylases in Arabidopsis thaliana

    DEFF Research Database (Denmark)

    Hoff, Tine; Frandsen, Gitte Inselmann; Rocher, Anne

    1998-01-01

    Aldehyde oxidases and xanthine dehydrogenases/oxidases belong to the molybdenum cofactor dependent hydroxylase class of enzymes. Zymograms show that Arabidopsis thaliana has at least three different aldehyde oxidases and one xanthine oxidase. Three different cDNA clones encoding putative aldehyde...... oxidases (AtAO1, 2, 3) were isolated. An aldehyde oxidase is the last step in abscisic acid (ABA) biosynthesis. AtAO1 is mainly expressed in seeds and roots which might reflect that it is involved in ABA biosynthesis....

  13. β-lactamase-mediated resistance: a biochemical, epidemiological and genetic overview.

    Science.gov (United States)

    Gutkind, Gabriel O; Di Conza, Jose; Power, Pablo; Radice, Marcela

    2013-01-01

    Early after the introduction of the first (narrow spectrum) penicillins into clinical use, penicillinase-producing staphylococci replaced (worldwide) the previously susceptible microorganisms. Similarly, the extensive use of broad-spectrum, orally administered β- lactams (like ampicillin, amoxicillin or cefalexin) provided a favorable scenario for the selection of gram-negative microorganisms producing broad spectrum β-lactamases almost 45 years ago. These microorganisms could be controlled by the introduction of the so called "extended spectrum cephalosporins". However, overuse of these drugs resulted, after a few years, in the emergence of extended-spectrum β-lactamases (ESBLs) through point mutations in the existing broad-spectrum β-lactamases, such as TEM and SHV enzymes. Overuse of extended-spectrum β-lactams also gave rise to chromosomal mutations in regulatory genes which resulted in the overproduction of chromosomal AmpC genes, and, in other regions of the world, in the explosive emergence of other ESBL families, like the CTX-Ms. Carbapenems remained active on microorganisms harboring these extended-spectrum β-lactamases, while both carbapenems and fourth generation cephalosporins remained active towards those with derepressed (or the more recent plasmidic) AmpCs. However, microorganisms countered this assault by the emergence of the so called carbapenemases (both serine- and metallo- enzymes) which, in some cases, are actually capable of hydrolyzing almost all β-lactams including the carbapenems. Although all these enzyme families (some of them represented by hundreds of members) are for sure pre-dating the antibiotic era in environmental and clinically significant microorganisms, it was the misuse of these antibiotics that drove their evolution. This paper describes in detail each major class of β-lactamase including epidemiology, genetic, and biochemical evaluations.

  14. Biochemical and Genetical Responses of Phoenix dactylifera L. to Cadmium Stress

    Science.gov (United States)

    Al-Qurainy, Fahad; Alshameri, Aref

    2017-01-01

    The cadmium (Cd), a heavy metal, causes toxicity, which leads to hampering the growth and development of the plant. The molecular and biochemical approaches were used for the investigation of antioxidant system response and genotoxicity in date palm (Phoenix dactylifera L.) cv. Sagai in pot experiment having Cd. The root length was more affected than the shoot length as more accumulation of Cd occurs in roots. Fresh weights of root and shoot were reduced significantly in treated plants as compared to the control. The proline content was increased at low concentration of Cd (300 µM-CdCl2) than the medium and high concentrations (600 and 900 µM-CdCl2), respectively. The thiobarbituric acid reactive substances (TBARS) content was increased at 600 and 900 µM-CdCl2 compared to the plants treated at 300 µM-CdCl2 and controls. Antioxidant enzymatic assay was performed under Cd stress and compared with control plants. The catalase (CAT) and superoxide dismutase (SOD) activities were found to be high in plants treated with CdCl2 at 300 µM compared to at 600 and 900 µM-CdCl2, respectively. The genotoxicity of Cd was assessed using the inter-simple sequence repeat (ISSR) marker where all treated and control plants were clustered into three main groups based on genetic similarity. P. dactylifera plants were found to be more divergent at high Cd stress as compared to control and plants treated at low concentration of Cd. PMID:29201916

  15. Biochemical and Genetical Responses of Phoenix dactylifera L. to Cadmium Stress

    Directory of Open Access Journals (Sweden)

    Fahad Al-Qurainy

    2017-01-01

    Full Text Available The cadmium (Cd, a heavy metal, causes toxicity, which leads to hampering the growth and development of the plant. The molecular and biochemical approaches were used for the investigation of antioxidant system response and genotoxicity in date palm (Phoenix dactylifera L. cv. Sagai in pot experiment having Cd. The root length was more affected than the shoot length as more accumulation of Cd occurs in roots. Fresh weights of root and shoot were reduced significantly in treated plants as compared to the control. The proline content was increased at low concentration of Cd (300 µM-CdCl2 than the medium and high concentrations (600 and 900 µM-CdCl2, respectively. The thiobarbituric acid reactive substances (TBARS content was increased at 600 and 900 µM-CdCl2 compared to the plants treated at 300 µM-CdCl2 and controls. Antioxidant enzymatic assay was performed under Cd stress and compared with control plants. The catalase (CAT and superoxide dismutase (SOD activities were found to be high in plants treated with CdCl2 at 300 µM compared to at 600 and 900 µM-CdCl2, respectively. The genotoxicity of Cd was assessed using the inter-simple sequence repeat (ISSR marker where all treated and control plants were clustered into three main groups based on genetic similarity. P. dactylifera plants were found to be more divergent at high Cd stress as compared to control and plants treated at low concentration of Cd.

  16. Diagnostic approach to neurotransmitter monoamine disorders: experience from clinical, biochemical, and genetic profiles.

    Science.gov (United States)

    Kuster, Alice; Arnoux, Jean-Baptiste; Barth, Magalie; Lamireau, Delphine; Houcinat, Nada; Goizet, Cyril; Doray, Bérénice; Gobin, Stéphanie; Schiff, Manuel; Cano, Aline; Amsallem, Daniel; Barnerias, Christine; Chaumette, Boris; Plaze, Marion; Slama, Abdelhamid; Ioos, Christine; Desguerre, Isabelle; Lebre, Anne-Sophie; de Lonlay, Pascale; Christa, Laurence

    2018-01-01

    To improve the diagnostic work-up of patients with diverse neurological diseases, we have elaborated specific clinical and CSF neurotransmitter patterns. Neurotransmitter determinations in CSF from 1200 patients revealed abnormal values in 228 (19%) cases. In 54/228 (24%) patients, a final diagnosis was identified. We have reported primary (30/54, 56%) and secondary (24/54, 44%) monoamine neurotransmitter disorders. For primary deficiencies, the most frequently mutated gene was DDC (n = 9), and the others included PAH with neuropsychiatric features (n = 4), PTS (n = 5), QDPR (n = 3), SR (n = 1), and TH (n = 1). We have also identified mutations in SLC6A3, FOXG1 (n = 1 of each), MTHFR (n = 3), FOLR1, and MTHFD (n = 1 of each), for dopamine transporter, neuronal development, and folate metabolism disorders, respectively. For secondary deficiencies, we have identified POLG (n = 3), ACSF3 (n = 1), NFU1, and SDHD (n = 1 of each), playing a role in mitochondrial function. Other mutated genes included: ADAR, RNASEH2B, RNASET2, SLC7A2-IT1 A/B lncRNA, and EXOSC3 involved in nuclear and cytoplasmic metabolism; RanBP2 and CASK implicated in post-traductional and scaffolding modifications; SLC6A19 regulating amino acid transport; MTM1, KCNQ2 (n = 2), and ATP1A3 playing a role in nerve cell electrophysiological state. Chromosome abnormalities, del(8)(p23)/dup(12) (p23) (n = 1), del(6)(q21) (n = 1), dup(17)(p13.3) (n = 1), and non-genetic etiologies (n = 3) were also identified. We have classified the final 54 diagnoses in 11 distinctive biochemical profiles and described them through 20 clinical features. To identify the specific molecular cause of abnormal NT profiles, (targeted) genomics might be used, to improve diagnosis and allow early treatment of complex and rare neurological genetic diseases.

  17. Genetic and biochemical identification of a novel single-stranded DNA binding complex in Haloferax volcanii

    Directory of Open Access Journals (Sweden)

    Amy eStroud

    2012-06-01

    Full Text Available Single-stranded DNA binding proteins play an essential role in DNA replication and repair. They use oligosaccharide-binding folds, a five-stranded ß-sheet coiled into a closed barrel, to bind to single-stranded DNA thereby protecting and stabilizing the DNA. In eukaryotes the single-stranded DNA binding protein is known as replication protein A (RPA and consists of three distinct subunits that function as a heterotrimer. The bacterial homolog is termed single-stranded DNA-binding protein (SSB and functions as a homotetramer. In the archaeon Haloferax volcanii there are three genes encoding homologs of RPA. Two of the rpa genes (rpa1 and rpa3 exist in operons with a novel gene specific to Euryarchaeota, this gene encodes a protein that we have termed rpa-associated protein (RPAP. The rpap genes encode proteins belonging to COG3390 group and feature oligosaccharide-binding folds, suggesting that they might cooperate with RPA in binding to single-stranded DNA. Our genetic analysis showed that rpa1 and rpa3 deletion mutants have differing phenotypes; only ∆rpa3 strains are hypersensitive to DNA damaging agents. Deletion of the rpa3-associated gene rpap3 led to similar levels of DNA damage sensitivity, as did deletion of the rpa3 operon, suggesting that RPA3 and RPAP3 function in the same pathway. Protein pull-downs involving recombinant hexahistidine-tagged RPAs showed that RPA3 co-purifies with RPAP3, and RPA1 co-purifies with RPAP1. This indicates that the RPAs interact only with their respective associated proteins; this was corroborated by the inability to construct rpa1 rpap3 and rpa3 rpap1 double mutants. This is the first report investigating the individual function of the archaeal COG3390 RPA-associated proteins. We have shown genetically and biochemically that the RPAPs interact with their respective RPAs, and have uncovered a novel single-stranded DNA binding complex that is unique to Euryarchaeota.

  18. Social stratification in the Sikh population of Punjab (India) has a genetic basis: evidence from serological and biochemical markers.

    Science.gov (United States)

    Chahal, Sukh Mohinder Singh; Virk, Rupinder Kaur; Kaur, Sukhvir; Bansal, Rupinder

    2011-01-01

    The present study was planned to assess whether social stratification in the Sikh population inhabiting the northwest border Indian state of Punjab has any genetic basis. Blood samples were collected randomly from a total of 2851 unrelated subjects belonging to 21 groups of two low-ranking Sikh scheduled caste populations, viz. Mazhabi and Ramdasi, and a high-ranking Jat Sikh caste population of Punjab. The genetic profile of Sikh groups was investigated using a total of nine serobiochemical genetic markers, comprising two blood groups (ABO, RH(D)) and a battery of seven red cell enzyme polymorphisms (ADA, AK1, ESD, PGM1, GLO1, ACP1, GPI), following standard serological and biochemical laboratory protocols. Genetic structure was studied using original allele frequency data and statistical measures of heterozygosity, genic differentiation, genetic distance, and genetic admixture. Great heterogeneity was observed between Sikh scheduled caste and Jat Sikh populations, especially in the RH(D) blood group system, and distribution of ESD, ACP1, and PGM1 enzyme markers was also found to be significantly different between many of their groups. Genetic distance trees demonstrated little or no genetic affinities between Sikh scheduled caste and Jat Sikh populations; the Mazhabi and Ramdasi also showed little genetic relationship. Genetic admixture analysis suggested a higher element of autochthonous tribal extraction in the Ramdasi. The present study revealed much genetic heterogeneity in differently ranking Sikh caste populations of Punjab, mainly attributable to their different ethnic backgrounds, and provided a genetic basis to social stratification present in this religious community of Punjab, India.

  19. Biosynthesis and characterization of CdS quantum dots in genetically engineered Escherichia coli

    Science.gov (United States)

    Mi, Congcong; Wang, Yanyan; Zhang, Jingpu; Huang, Huaiqing; Xu, Linru; Wang, Shuo; Fang, Xuexun; Fang, Jin; Mao, Chuanbin; Xu, Shukun

    2011-01-01

    Quantum dots (QDs) were prepared in genetically engineered Escherichia coli (E. coli) through the introduction of foreign genes encoding a CdS binding peptide. The CdS QDs were successfully separated from the bacteria through two methods, lysis and freezing–thawing of cells, and purified with an anion-exchange resin. High-resolution transmission electron microscopy, X-ray diffraction, luminescence spectroscopy, and energy dispersive X-ray spectroscopy were applied to characterize the as-prepared CdS QDs. The effects of reactant concentrations, bacteria incubation times, and reaction times on QD growth were systematically investigated. Our work demonstrates that genetically engineered bacteria can be used to synthesize QDs. The biologically synthesized QDs are expected to be more biocompatible probes in bio-labeling and imaging. PMID:21458508

  20. Genetic Basis for Variation of Metalloproteinase-Associated Biochemical Activity in Venom of the Mojave Rattlesnake (Crotalus scutulatus scutulatus

    Directory of Open Access Journals (Sweden)

    Ruben K. Dagda

    2013-01-01

    Full Text Available The metalloproteinase composition and biochemical profiles of rattlesnake venom can be highly variable among rattlesnakes of the same species. We have previously shown that the neurotoxic properties of the Mojave rattlesnake (Crotalus scutulatus scutulatus are associated with the presence of the Mojave toxin A subunit suggesting the existence of a genetic basis for rattlesnake venom composition. In this report, we hypothesized the existence of a genetic basis for intraspecies variation in metalloproteinase-associated biochemical properties of rattlesnake venom of the Mojave rattlesnake. To address this question, we PCR-amplified and compared the genomic DNA nucleotide sequences that code for the mature metalloproteinase domain of fourteen Mojave rattlesnakes captured from different geographical locations across the southwest region of the United States. In addition, the venoms from the same rattlesnakes were tested for their ability to hydrolyze fibrinogen, fibrin, casein, and hide powder azure and for induction of hemorrhage in mice. Overall, based on genomic sequencing and biochemical data, we classified Mojave rattlesnake venom into four distinct groups of metalloproteinases. These findings indicate that differences in nucleotide sequences encoding the mature proteinase domain and noncoding regions contribute to differences in venom metalloproteinase activities among rattlesnakes of the same species.

  1. GM1-gangliosidosis in American black bears: clinical, pathological, biochemical and molecular genetic characterization.

    Science.gov (United States)

    Muthupalani, Sureshkumar; Torres, Paola A; Wang, Betty C; Zeng, Bai Jin; Eaton, Samuel; Erdelyi, Ildiko; Ducore, Rebecca; Maganti, Rajanikarath; Keating, John; Perry, Bain J; Tseng, Florina S; Waliszewski, Nicole; Pokras, Mark; Causey, Robert; Seger, Rita; March, Philip; Tidwell, Amy; Pfannl, Rolf; Seyfried, Thomas; Kolodny, Edwin H; Alroy, Joseph

    2014-04-01

    G(M1)-gangliosidosis is a rare progressive neurodegenerative disorder due to an autosomal recessively inherited deficiency of lysosomal β-galactosidase. We have identified seven American black bears (Ursus americanus) found in the Northeast United States suffering from G(M1)-gangliosidosis. This report describes the clinical features, brain MRI, and morphologic, biochemical and molecular genetic findings in the affected bears. Brain lipids were compared with those in the brain of a G(M1)-mouse. The bears presented at ages 10-14 months in poor clinical condition, lethargic, tremulous and ataxic. They continued to decline and were humanely euthanized. The T(2)-weighted MR images of the brain of one bear disclosed white matter hyperintensity. Morphological studies of the brain from five of the bears revealed enlarged neurons with foamy cytoplasm containing granules. Axonal spheroids were present in white matter. Electron microscopic examination revealed lamellated membrane structures within neurons. Cytoplasmic vacuoles were found in the liver, kidneys and chondrocytes and foamy macrophages within the lungs. Acid β-galactosidase activity in cultured skin fibroblasts was only 1-2% of control values. In the brain, ganglioside-bound sialic acid was increased more than 2-fold with G(M1)-ganglioside predominating. G(A1) content was also increased whereas cerebrosides and sulfatides were markedly decreased. The distribution of gangliosides was similar to that in the G(M1)-mouse brain, but the loss of myelin lipids was greater in the brain of the affected bear than in the brain of the G(M1) mouse. Isolated full-length cDNA of the black bear GLB1 gene revealed 86% homology to its human counterpart in nucleotide sequence and 82% in amino acid sequence. GLB1 cDNA from liver tissue of an affected bear contained a homozygous recessive T(1042) to C transition inducing a Tyr348 to His mutation (Y348H) within a highly conserved region of the GLB1 gene. The coincidence of several

  2. A spontaneous mutant of microcystin biosynthesis: genetic characterization and effect on Daphnia

    DEFF Research Database (Denmark)

    Kabernick, M.; Rohrlack, T.; Christoffersen, K.

    2001-01-01

    Microcystis aeruginosa strain MRC is unique in its' possession of the mcyA-J gene cluster, which encodes microcystin synthetase, but its' inability to produce microcystins. M. aeruginosa strain MRD is genetically identical to MRC at numerous genomic loci examined, but produces a variety...... of microcystins, mainly with the amino acid tyrosine in the molecule. Zooplankton studies with Daphnia galeata and D. pulicaria, using the mutant (MRC) and its' wild type (MRD), showed for the first time that microcystins other than microcystin-LR can be responsible for the poisoning of Daphnia by Microcystis....... Regardless of microcystin content, both Daphnia exhibited significantly reduced ingestion rates when fed with either strain of M. aeruginosa compared with the green alga Scenedesmus acutus. A disruption of the molting process in both Daphnia spp. was noted when these species were fed with MRC cells...

  3. Biochemical and molecular study of genetic stability in tomatoes plants rom seeds treated with low doses of X-ray

    International Nuclear Information System (INIS)

    Ramirez, R; Gonzalez, LM; Chavez, Licet; Camejo, Yanelis; Gonzalez, Maria C; Fernandez, Arais

    2008-01-01

    For the extensive agricultural exploitation of vegetable radio stimulation, it is indispensable to study the genetic stability of treated varieties, having in mind X ray potentialities of inducing not only physiological but genetic changes as well. Therefore, biochemical and molecular markers were employed in tomato plants derived from irradiated seeds at low doses of X rays. For the biochemical analysis, peroxidases, polyphenoloxidases and dismutase superoxide isoenzymes were determined whereas the Random Amplification of Polymorphic DNA (RAPD) method based on Polymerase Chain Reaction (PCR) was used for the molecular analysis. When comparing the electrophoretic patterns from the control and irradiated treatments applied to the three enzymatic systems, there were not appreciable variations on the number of bands and their intensities, indicating the little variability induced in these systems by the low X ray doses. Also, from the molecular viewpoint, electrophoretic patterns showed a clear amplification of DNA by generating a total of 155 bands in all varieties studied. This molecular marker showed a high monomorphism independently of the treatments applied, with values ranging between 86 and 97 %, indicating that irradiation at low doses did not induce an important genetic variability and confirming its possible practical usefulness for stimulating some physiological processes without causing. (Author)

  4. Evidence of Mitochondrial Dysfunction in Autism: Biochemical Links, Genetic-Based Associations, and Non-Energy-Related Mechanisms

    Directory of Open Access Journals (Sweden)

    Keren K. Griffiths

    2017-01-01

    Full Text Available Autism spectrum disorder (ASD, the fastest growing developmental disability in the United States, represents a group of neurodevelopmental disorders characterized by impaired social interaction and communication as well as restricted and repetitive behavior. The underlying cause of autism is unknown and therapy is currently limited to targeting behavioral abnormalities. Emerging studies suggest a link between mitochondrial dysfunction and ASD. Here, we review the evidence demonstrating this potential connection. We focus specifically on biochemical links, genetic-based associations, non-energy related mechanisms, and novel therapeutic strategies.

  5. Biochemical and genetic characterization of a carbamyl phosphate synthetase mutant of Escherichia coli K12.

    Science.gov (United States)

    Bolivar, F; Galván, M; Martuscelli, J

    1976-05-01

    An unusual Escherichia coli K12 mutant for carbamyl phosphate synthetase is described. The mutation was generated by bacteriophage MUI insertion and left a 5% residual activity of the enzyme using either ammonia or glutamine as donors. The mutation is recessive to the wild-type allele and maps at or near the pyrA gene, but the mutant requires only arginine and not uracil for growth. By a second block in the pyrB gene it was possible to shift the accumulated carbamyl phosphate to arginine biosynthesis. The Km values and the levels of ornithine activation and inhibition by UMP were normal in the mutant enzyme.

  6. Biochemical and Phylogenetic Characterization of a Novel Diaminopimelate Biosynthesis Pathway in Prokaryotes Identifies a Diverged Form of ll-Diaminopimelate Aminotransferase▿ †

    OpenAIRE

    Hudson, André O.; Gilvarg, Charles; Leustek, Thomas

    2008-01-01

    A variant of the diaminopimelate (DAP)-lysine biosynthesis pathway uses an ll-DAP aminotransferase (DapL, EC 2.6.1.83) to catalyze the direct conversion of l-2,3,4,5-tetrahydrodipicolinate to ll-DAP. Comparative genomic analysis and experimental verification of DapL candidates revealed the existence of two diverged forms of DapL (DapL1 and DapL2). DapL orthologs were identified in eubacteria and archaea. In some species the corresponding dapL gene was found to lie in genomic contiguity with o...

  7. Biochemical and phylogenetic characterization of a novel diaminopimelate biosynthesis pathway in prokaryotes identifies a diverged form of LL-diaminopimelate aminotransferase.

    Science.gov (United States)

    Hudson, André O; Gilvarg, Charles; Leustek, Thomas

    2008-05-01

    A variant of the diaminopimelate (DAP)-lysine biosynthesis pathway uses an LL-DAP aminotransferase (DapL, EC 2.6.1.83) to catalyze the direct conversion of L-2,3,4,5-tetrahydrodipicolinate to LL-DAP. Comparative genomic analysis and experimental verification of DapL candidates revealed the existence of two diverged forms of DapL (DapL1 and DapL2). DapL orthologs were identified in eubacteria and archaea. In some species the corresponding dapL gene was found to lie in genomic contiguity with other dap genes, suggestive of a polycistronic structure. The DapL candidate enzymes were found to cluster into two classes sharing approximately 30% amino acid identity. The function of selected enzymes from each class was studied. Both classes were able to functionally complement Escherichia coli dapD and dapE mutants and to catalyze LL-DAP transamination, providing functional evidence for a role in DAP/lysine biosynthesis. In all cases the occurrence of dapL in a species correlated with the absence of genes for dapD and dapE representing the acyl DAP pathway variants, and only in a few cases was dapL coincident with ddh encoding meso-DAP dehydrogenase. The results indicate that the DapL pathway is restricted to specific lineages of eubacteria including the Cyanobacteria, Desulfuromonadales, Firmicutes, Bacteroidetes, Chlamydiae, Spirochaeta, and Chloroflexi and two archaeal groups, the Methanobacteriaceae and Archaeoglobaceae.

  8. Heritability and genetic advance studies for biochemical traits in F2-3 introgressed families of Brassica

    International Nuclear Information System (INIS)

    Farhatullah, N.K.; Khalil, I.H.; Nahed, H.

    2015-01-01

    Higher heritability estimates along with high genetic advance values are effective in envisaging gain under selection in developing genotypes. The objective of the present study was to evaluate variability, heritability and genetic advance in 10 interspecific F2-3 families of Brassica species (B. napus * B. juncea, B. napus * B. rapa). These families were studied for heterospecific introgression of biochemical traits. Low to high heritability estimates were recorded for seed quality traits. Considerable variations within F2-3 families were observed for biochemical traits. Most of the F2-3 families for oil content and erucic showed moderate to high heritability indicating the slightest influence of environment thus modification of trait by selection would be more effective. Among F2-3 introgressed families Bn-510 x Bj-109 produced high oil i.e., 49.5% while Bn-532 x Br-118 (24.4%), Bn-533 x Bj-109 (24.1%) and high protein percentage in terms of mean performance. In the present research, individual segregating progenies of interspecific cross populations i.e., which possessed combination of desirable traits, were identified which could be incorporated in the future Breeding programs and it may facilitate varietal development. (author)

  9. Biochemical and Phylogenetic Characterization of a Novel Diaminopimelate Biosynthesis Pathway in Prokaryotes Identifies a Diverged Form of ll-Diaminopimelate Aminotransferase▿ †

    Science.gov (United States)

    Hudson, André O.; Gilvarg, Charles; Leustek, Thomas

    2008-01-01

    A variant of the diaminopimelate (DAP)-lysine biosynthesis pathway uses an ll-DAP aminotransferase (DapL, EC 2.6.1.83) to catalyze the direct conversion of l-2,3,4,5-tetrahydrodipicolinate to ll-DAP. Comparative genomic analysis and experimental verification of DapL candidates revealed the existence of two diverged forms of DapL (DapL1 and DapL2). DapL orthologs were identified in eubacteria and archaea. In some species the corresponding dapL gene was found to lie in genomic contiguity with other dap genes, suggestive of a polycistronic structure. The DapL candidate enzymes were found to cluster into two classes sharing approximately 30% amino acid identity. The function of selected enzymes from each class was studied. Both classes were able to functionally complement Escherichia coli dapD and dapE mutants and to catalyze ll-DAP transamination, providing functional evidence for a role in DAP/lysine biosynthesis. In all cases the occurrence of dapL in a species correlated with the absence of genes for dapD and dapE representing the acyl DAP pathway variants, and only in a few cases was dapL coincident with ddh encoding meso-DAP dehydrogenase. The results indicate that the DapL pathway is restricted to specific lineages of eubacteria including the Cyanobacteria, Desulfuromonadales, Firmicutes, Bacteroidetes, Chlamydiae, Spirochaeta, and Chloroflexi and two archaeal groups, the Methanobacteriaceae and Archaeoglobaceae. PMID:18310350

  10. The effects of genetic manipulation of putrescine biosynthesis on transcription and activities of the other polyamine biosynthetic enzymes

    Science.gov (United States)

    Andrew F. Page; Sridev Mohapatra; Rakesh Minocha; Subhash C. Minocha

    2007-01-01

    We have studied the effects of overproduction of putrescine (Put) via transgenic expression of a mouse ornithine decarboxylase (ODC) gene on the expression of native genes for four enzymes involved in polyamine biosynthesis in hybrid poplar (Populus nigra x maximowiczii) cells. An examination of the transcript levels of arginine...

  11. Haploid Mammalian Genetic Screen Identifies UBXD8 as a Key Determinant of HMGCR Degradation and Cholesterol Biosynthesis

    NARCIS (Netherlands)

    Loregger, Anke; Raaben, Matthijs; Tan, Josephine; Scheij, Saskia; Moeton, Martina; van den Berg, Marlene; Gelberg-Etel, Hila; Stickel, Elmer; Roitelman, Joseph; Brummelkamp, Thijn; Zelcer, Noam

    2017-01-01

    Objective-The cellular demand for cholesterol requires control of its biosynthesis by the mevalonate pathway. Regulation of HMGCR (3-hydroxy-3-methylglutaryl coenzyme A reductase), a rate-limiting enzyme in this pathway and the target of statins, is a key control point herein. Accordingly, HMGCR is

  12. Benzylisoquinoline alkaloid biosynthesis in opium poppy.

    Science.gov (United States)

    Beaudoin, Guillaume A W; Facchini, Peter J

    2014-07-01

    Opium poppy (Papaver somniferum) is one of the world's oldest medicinal plants and remains the only commercial source for the narcotic analgesics morphine, codeine and semi-synthetic derivatives such as oxycodone and naltrexone. The plant also produces several other benzylisoquinoline alkaloids with potent pharmacological properties including the vasodilator papaverine, the cough suppressant and potential anticancer drug noscapine and the antimicrobial agent sanguinarine. Opium poppy has served as a model system to investigate the biosynthesis of benzylisoquinoline alkaloids in plants. The application of biochemical and functional genomics has resulted in a recent surge in the discovery of biosynthetic genes involved in the formation of major benzylisoquinoline alkaloids in opium poppy. The availability of extensive biochemical genetic tools and information pertaining to benzylisoquinoline alkaloid metabolism is facilitating the study of a wide range of phenomena including the structural biology of novel catalysts, the genomic organization of biosynthetic genes, the cellular and sub-cellular localization of biosynthetic enzymes and a variety of biotechnological applications. In this review, we highlight recent developments and summarize the frontiers of knowledge regarding the biochemistry, cellular biology and biotechnology of benzylisoquinoline alkaloid biosynthesis in opium poppy.

  13. Clinical, genetic, biochemical, and testicular biopsy findings among 1,213 men evaluated for infertility

    DEFF Research Database (Denmark)

    Olesen, Inge Ahlmann; Andersson, Anna-Maria; Aksglaede, Lise

    2017-01-01

    work-up from 2005 to 2009. INTERVENTIONS(S): None. MAIN OUTCOME MEASURE(S): Health history, clinical findings, chromosome/genetic aberrations, semen quality, reproductive hormones. RESULT(S): In total, 64.4% of the infertile men had one or more reproductive disorders or factors influencing fertility...

  14. Genetic, Biochemical and Environmental Factors Associated with Pregnancy Outcomes in Newborns from the Czech Republic

    Czech Academy of Sciences Publication Activity Database

    Rössner ml., Pavel; Tabashidze, Nana; Dostál, Miroslav; Nováková, Zuzana; Chvátalová, Irena; Špátová, Milada; Šrám, Radim

    2011-01-01

    Roč. 119, č. 2 (2011), s. 265-271 ISSN 0091-6765 R&D Projects: GA MŽP(CZ) SP/1B3/50/07 Institutional research plan: CEZ:AV0Z50390512 Keywords : air pollution * biomarkers * genetic polymorphisms Subject RIV: DN - Health Impact of the Environment Quality Impact factor: 7.036, year: 2011

  15. The clinical, biochemical and genetic features associated with RMND1-related mitochondrial disease

    Czech Academy of Sciences Publication Activity Database

    Ng, Y. S.; Alston, Ch. L.; Diodato, D.; Morris, A. A.; Ulrick, N.; Kmoch, S.; Houštěk, Josef; Martinelli, D.; Haghighi, A.; Atiq, M.; Gamero, M. A.; Garcia-Martinez, E.; Kratochvílová, H.; Santra, S.; Brown, R. M.; Brown, G. K.; Ragge, N.; Monavari, A.; Pysden, K.; Ravn, K.; Casey, J. P.; Khan, A.; Chakrapani, A.; Vassallo, G.; Simons, C.; McKeever, K.; O´Sullivan, S.; Childs, A.-M.; Ostergaard, E.; Vanderver, A.; Goldstein, A.; Vogt, J.; Taylor, R. W.; McFarland, R.

    2016-01-01

    Roč. 53, č. 11 (2016), s. 768-775 ISSN 0022-2593 R&D Projects: GA ČR(CZ) GB14-36804G Institutional support: RVO:67985823 Keywords : congenital sensorineural deafness * lactic acidosis * mitochondrial respiratory chain deficiencies * prognosis * renal disease Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 5.451, year: 2016

  16. Biochemical, mechanical, and spectroscopic analyses of genetically engineered flax fibers producing bioplastic (poly-beta-hydroxybutyrate).

    Science.gov (United States)

    Wróbel-Kwiatkowska, Magdalena; Skórkowska-Telichowska, Katarzyna; Dymińska, Lucyna; Maczka, Mirosław; Hanuza, Jerzy; Szopa, Jan

    2009-01-01

    The interest in biofibers has grown in recent years due to their expanding range of applications in fields as diverse as biomedical science and the automotive industry. Their low production costs, biodegradability, physical properties, and perceived eco-friendliness allow for their extensive use as composite components, a role in which they could replace petroleum-based synthetic polymers. We performed biochemical, mechanical, and structural analyses of flax stems and fibers derived from field-grown transgenic flax enriched with PHB (poly-beta-hydroxybutyrate). The analyses of the plant stems revealed an increase in the cellulose content and a decrease in the lignin and pectin contents relative to the control plants. However, the contents of the fibers' major components (cellulose, lignin, pectin) remain unchanged. An FT-IR study confirmed the results of the biochemical analyses of the flax fibers. However, the arrangement of the cellulose polymer in the transgenic fibers differed from that in the control, and a significant increase in the number of hydrogen bonds was detected. The mechanical properties of the transgenic flax stems were significantly improved, reflecting the cellulose content increase. However, the mechanical properties of the fibers did not change in comparison with the control, with the exception of the fibers from transgenic line M13. The generated transgenic flax plants, which produce both components of the flax/PHB composites (i.e., fibers and thermoplastic matrix in the same plant organ) are a source of an attractive and ecologically safe material for industry and medicine. 2009 American Institute of Chemical Engineers Biotechnol.

  17. Genetic and biochemical studies of the lipid-containing bacteriophage PR4

    International Nuclear Information System (INIS)

    Vanden Boom, T.J.

    1989-01-01

    Bacteriophage PR4 is a lipid-containing bacterial virus able to infect Escherichia coli and Salmonella typhimurium. The icosahedral virion consists of an external protein capsid layer which surrounds a membrane vesicle enclosed ds DNA genome. The author has analyzed the time course of phage PR4 protein synthesis and have identified at least 34 proteins present in phage infected cells not detected in uninfected control cultures. In addition, he has isolated a more extensive set of conditional-lethal nonsense mutants of this virus. This collection of mutants permitted the identification of seven additional phage PR4 gene products, including the terminal genome protein and an accessory lytic factor. The present collection of phage PR4 mutants has been assigned to 19 distinct genetic groups on the basis of genetic complementation tests and sodium dodecyl sulfate polyacrylamide gel electrophoresis analysis of the proteins produced in mutant-infected UV-irradiated cells. A restriction endonuclease map of the phage PR4 genome was constructed which includes 59 sites for ten restriction endonucleases. In addition, he has constructed a collection of recombinant plasmids containing subgenomic DNA fragments of bacteriophage PR4. He has used this collection of plasmids to generate a physical-genetic map of the PR4 genome. The physical-genetic map localizes mutations in 13 phage PR4 genetic groups on the viral DNA molecule. To investigate the role of phosphatidylglycerol (PG) in phage assembly and infectivity, he propagated PR4 on an E. coli mutant defective in PG synthesis. The PG content of phage PR4 grown on the mutant host accounted for 0.4% of the total viral phospholipids, representing a 90-fold decrease in PG relative to the PG content of phage grown on a wild type host

  18. Genetic and Biochemical Basis for the Transformation of Energetic Materials (RDX, TNT, DNTs) by Plants

    Science.gov (United States)

    2007-04-01

    Determining the Genetics and Biochemistry of the Phytoremediation Transformation Pathways of RDX and TNT,” Food, Pharmaceutical and Bioengineering...Organic toxicants and plants. Ecotoxicology and Environmental Safety 47:1-26 Larson SL, Jones RP, Escalon L, Parker D (1999) Classification of explosives...dollars, compared to the pharmaceutical industry at 109 billion US dollars (McCutcheon and Schnoor, 2003). Since phytoremediation has minimal capital

  19. Attention-deficit hyperactivity disorder in adults: A systematic review and meta-analysis of genetic, pharmacogenetic and biochemical studies

    Science.gov (United States)

    Bonvicini, C; Faraone, S V; Scassellati, C

    2016-01-01

    The adult form of attention-deficit/hyperactivity disorder has a prevalence of up to 5% and is the most severe long-term outcome of this common disorder. Family studies in clinical samples as well as twin studies suggest a familial liability and consequently different genes were investigated in association studies. Pharmacotherapy with methylphenidate (MPH) seems to be the first-line treatment of choice in adults with attention-deficit hyperactive disorder (ADHD) and some studies were conducted on the genes influencing the response to this drug. Finally some peripheral biomarkers were identified in ADHD adult patients. We believe this work is the first systematic review and meta-analysis of candidate gene association studies, pharmacogenetic and biochemical (metabolomics) studies performed in adults with ADHD to identify potential genetic, predictive and peripheral markers linked specifically to ADHD in adults. After screening 5129 records, we selected 87 studies of which 61 were available for candidate gene association studies, 5 for pharmacogenetics and 21 for biochemical studies. Of these, 15 genetic, 2 pharmacogenetic and 6 biochemical studies were included in the meta-analyses. We obtained an association between adult ADHD and the gene BAIAP2 (brain-specific angiogenesis inhibitor 1-associated protein 2), even after Bonferroni correction, with any heterogeneity in effect size and no publication bias. If we did not apply the Bonferroni correction, a trend was found for the carriers allele 9R of dopamine transporter SLC6A3 40 bp variable tandem repeat polymorphism (VNTR) and for 6/6 homozygotes of SLC6A3 30 bp VNTR. Negative results were obtained for the 9-6 haplotype, the dopamine receptor DRD4 48 bp VNTR, and the enzyme COMT SNP rs4680. Concerning pharmacogenetic studies, no association was found for the SLC6A3 40 bp and response to MPH with only two studies selected. For the metabolomics studies, no differences between ADHD adults and controls were

  20. Molecular genetic and biochemical analyses of a DNA repair gene from Serratia marcescens

    International Nuclear Information System (INIS)

    Murphy, K.E.

    1989-01-01

    In Escherichia coli, the SOS response and two 3-methyladenine DNA glycosylases (TagI and TagII) are required for repair of DNA damaged by alkylating agents such as methyl methanesulfonate (MMS). Mutations of the recA gene eliminate the SOS response. TagI and TagII are encoded by the tag and alkA genes, respectively. A gene (rpr) encoding 3-methyladenine DNA glycosylase activity was isolated from the Gram-negative bacterium Serratia marcescens. The gene, localized to a 1.5-kilobase pair SmaI-HindIII restriction fragment, was cloned into plasmid pUC18. The clone complemented E. coli tag alkA and recA mutations for MMS resistance. The rpr gene did not, however, complement recA mutations for resistance to ultraviolet light or the ability to perform homologous recombination reactions, nor did it complement E. coli ada or alkB mutations. Two proteins of molecular weights 42,000 and 16,000 were produced from the rpr locus. Analysis of deletion and insertion mutants of rpr suggested that the 42kD molecule is the active protein. The 16kD protein may either be a breakdown product of the 42kD species or may be encoded by another gene overlapping the reading frame of the rpr gene. Biochemical assays showed that the rpr gene product (Rpr) possesses 3-methyladenine DNA glycosylase activity

  1. Biochemical and genetic functional dissection of the P38 viral suppressor of RNA silencing.

    Science.gov (United States)

    Iki, Taichiro; Tschopp, Marie-Aude; Voinnet, Olivier

    2017-05-01

    Phytoviruses encode viral suppressors of RNA silencing (VSRs) to counteract the plant antiviral silencing response, which relies on virus-derived small interfering (si)RNAs processed by Dicer RNaseIII enzymes and subsequently loaded into ARGONAUTE (AGO) effector proteins. Here, a tobacco cell-free system was engineered to recapitulate the key steps of antiviral RNA silencing and, in particular, the most upstream double-stranded (ds)RNA processing reaction, not kinetically investigated thus far in the context of plant VSR studies. Comparative biochemical analyses of distinct VSRs in the reconstituted assay showed that in all cases tested, VSR interactions with siRNA duplexes inhibited the loading, but not the activity, of antiviral AGO1 and AGO2. Turnip crinkle virus P38 displayed the additional and unique property to bind both synthetic and RNA-dependent-RNA-polymerase-generated long dsRNAs, and inhibited the processing into siRNAs. Single amino acid substitutions in P38 could dissociate dsRNA-processing from AGO-loading inhibition in vitro and in vivo, illustrating dual-inhibitory strategies discriminatively deployed within a single viral protein, which, we further show, are bona fide suppressor functions that evolved independently of the conserved coat protein function of P38. © 2017 Iki et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.

  2. Genetic Mapping and Biochemical Basis of Yellow Feather Pigmentation in Budgerigars.

    Science.gov (United States)

    Cooke, Thomas F; Fischer, Curt R; Wu, Ping; Jiang, Ting-Xin; Xie, Kathleen T; Kuo, James; Doctorov, Elizabeth; Zehnder, Ashley; Khosla, Chaitan; Chuong, Cheng-Ming; Bustamante, Carlos D

    2017-10-05

    Parrot feathers contain red, orange, and yellow polyene pigments called psittacofulvins. Budgerigars are parrots that have been extensively bred for plumage traits during the last century, but the underlying genes are unknown. Here we use genome-wide association mapping and gene-expression analysis to map the Mendelian blue locus, which abolishes yellow pigmentation in the budgerigar. We find that the blue trait maps to a single amino acid substitution (R644W) in an uncharacterized polyketide synthase (MuPKS). When we expressed MuPKS heterologously in yeast, yellow pigments accumulated. Mass spectrometry confirmed that these yellow pigments match those found in feathers. The R644W substitution abolished MuPKS activity. Furthermore, gene-expression data from feathers of different bird species suggest that parrots acquired their colors through regulatory changes that drive high expression of MuPKS in feather epithelia. Our data also help formulate biochemical models that may explain natural color variation in parrots. VIDEO ABSTRACT. Copyright © 2017 Elsevier Inc. All rights reserved.

  3. Potential biochemical and genetic toxicity of triclosan as an emerging pollutant on earthworms (Eisenia fetida).

    Science.gov (United States)

    Lin, Dasong; Zhou, Qixing; Xie, Xiujie; Liu, Yao

    2010-11-01

    Triclosan as an important antimicrobial agent is increasingly detected in the terrestrial environment as sewage sludge and reclaimed water are applied on land, but little is known about its effect on non-target soil organisms. In this study, biochemical responses including changes in the activity of catalase (CAT), superoxide dismutase (SOD), glutathione-S-transferase (GST), and malondialdehyde (MDA) of the earthworm Eisenia fetida were examined in order to assess ecological toxicity of the chemical. The single-cell gel electrophoresis (SCGE) was also used to measure the potential genotoxicity of the chemical. The results showed that the activity of CAT and GST at the highest tested dose could be stimulated after a 2-d exposure, reaching 148% and 123% of that in the control, respectively. However, with prolonged exposure, the activity of CAT and GST at the highest tested dose was inhibited, falling to 47% and 33% of that in the control, respectively. Triclosan induced an increase in the activity of SOD, but no significant (p>0.05) changes were observed. The content of MDA was dependent both on the dose of triclosan and on the exposure duration. The comet assay demonstrated that triclosan treatments led to a dose-dependent DNA damage of E. fetida after exposures of 7 and 14 d. Our findings can suggest that triclosan has sublethal effects on E. fetida. Copyright © 2010 Elsevier Ltd. All rights reserved.

  4. Seed Biochemical Analysis Based Profiling of Diverse Wheat Genetic Resource from Pakistan

    Science.gov (United States)

    Khalid, Anam; Hameed, Amjad

    2017-01-01

    Wheat is the major nutrient source worldwide. In Pakistan, it has a crucial place in agriculture as well as in national economy. For seed biochemical compositional analysis, wheat germplasm (77 genotypes) was collected from different agro-climatic zones of Pakistan. Significant variation (p sugar was found in Saleem-2000 (29.86 mg/g s. wt.), reducing sugars in Punjab-96 (12.68 mg/g s. wt.), non-reducing sugars in Saleem-2000 (27.33 mg/g s. wt.). However, highest albumins was identified in TC-4928 (352.89 mg/g s. wt.) and globulins in MEXI PAK (252.67 mg/g s. wt.), salt soluble proteins in Faisalabad-2008 (162.44 mg/g s. wt.), and total soluble proteins in Punjab-96 (487.33 mg/g s. wt.) indicating good quality of wheat genotypes as well as good nutritional status. Genotypes which have been ranked high in respective parameter can be employed in breeding to enhance the nutritional quality of wheat. PMID:28775731

  5. Development of enhanced radioprotectors - Biochemical and molecular genetical approaches on the radioprotective mechanism of natural products

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Jeong Hee; Lee, Eun Ju; Hong, Jung A [Kyunghee University, Seoul (Korea)

    2000-04-01

    To identify radio-protective agent candidate among medicinal plants and to elucidate the mechanism of action of the candidate material by using modern biochemical and molecular biological methods, we screened radio-protective activity among 48 medicinal plants. Seven samples showed above 20% protective activities against oxidative cell damage: Euryale ferox, Glycyrrhiza uralensis, Salvia miltiorrhiza, Eucomia ulmoides, Paeonia suffruticosa, Spirodela polyrrhiza, and Nelumbo nucifera. We also screened for oxidative stress sensitizing activity among other 51 medicinal plants. Among those samples, 11 samples showed good sensitizing effect; Melia azedarach, Agastache rugosa, Catalpa ovata, Prunus persica, Sinomenium acutum, Pulsatilla koreana, Oldenlandia diffusa, Anthriscus sylvestris, Schizandra chinensis, Gleditsia sinensis, and Cridium officinale. We also reported the radio-protective effect of DTT. The treatment of DTT increased cell survival after gamma-irradiation, decreased in the frequencies of micronucleus, and reduction in DNA fragmentation and apoptotic cells. Induction of apoptosis after UV-C irradiation was revealed by the changes in the relative cell death, increase in the relative amount of apoptotic cells, and the induction of DNA fragmentation. 165 refs., 9 figs., 8 tabs. (Author)

  6. Genetic and biochemical analysis of a laboratory-selected spirodiclofen-resistant strain of Tetranychus urticae Koch (Acari: Tetranychidae).

    Science.gov (United States)

    Van Pottelberge, Steven; Van Leeuwen, Thomas; Khajehali, Jahangir; Tirry, Luc

    2009-04-01

    Spirodiclofen is a selective, non-systemic acaricide from the new chemical class of tetronic acid derivatives. In order to develop strategies to minimise resistance in the field, a laboratory-selected spirodiclofen-resistant strain of the two-spotted spider mite, Tetranychus urticae Koch, was used to determine genetic, toxicological, biochemical and cross-resistance data. Selecting for spirodiclofen resistance in the laboratory yielded a strain (SR-VP) with a resistance ratio of 274, determined on the larval stage. The egg stage remained far more susceptible. No cross-resistance was found against other established acaricides, except for spiromesifen. Based on synergist experiments and enzyme assays, it appeared that especially P450 monooxygenases, but also esterases and glutathione-S-transferases, could be involved in the metabolic detoxification of spirodiclofen. Genetic analysis showed that the resistance is inherited as an intermediate trait under control of more than one gene. Resistance to spirodiclofen exceeded by far the recommended field rate. A good acaricide resistance management programme is necessary to prevent fast resistance build-up in the field. Spirodiclofen can be used in alternation with most established acaricides, except for other tetronic acid derivatives. Without selection pressure, resistance tends to be unstable and can decrease in the presence of susceptible individuals owing to the intermediate, polygenic inheritance mode. Copyright (c) 2009 Society of Chemical Industry.

  7. Genetic polymorphisms, Biochemical Factors, and Conventional Risk Factors in Young and Elderly North Indian Patients With Acute Myocardial Infarction.

    Science.gov (United States)

    Kaur, Rupinder; Das, Reena; Ahluwalia, Jasmina; Kumar, Rohit Manoj; Talwar, K K

    2016-03-01

    This study compared genetic polymorphisms (factor V Leiden [FVL] 1691G/A, factor VII [FVII] 10976G/A, FVII HVR4, platelet membrane glycoproteins GP1BA 1018C/T, GP1BA VNTR, integrin ITGB3 1565T/C, ITGA2 807C/T and methylenetetrahydrofolate reductase [MTHFR] 677C/T), biochemical (fibrinogen and homocysteine), and conventional risk factors in 184 young and 166 elderly north Indian patients with acute myocardial infarction (AMI). Univariate analysis revealed higher prevalence of hypertension and obesity in elderly patients while smoking, alcohol intake, and low socioeconomic status in young patients (P genetic polymorphisms were equally prevalent in young and elderly patients. Multiple logistic regression analysis showed smoking (P < .001), alcohol intake (P = .046), and hyperhomocysteinemia (P = .001) to be associated with AMI in the young patients while hypertension (P = .006) in elderly patients. To conclude, smoking, alcohol intake, and elevated homocysteine are the risk factors for AMI among young while hypertension among elderly patients. © The Author(s) 2014.

  8. Biochemical and genetic characterization of a novel metallo-β-lactamase from marine bacterium Erythrobacter litoralis HTCC 2594.

    Science.gov (United States)

    Jiang, Xia-Wei; Cheng, Hong; Huo, Ying-Yi; Xu, Lin; Wu, Yue-Hong; Liu, Wen-Hong; Tao, Fang-Fang; Cui, Xin-Jie; Zheng, Bei-Wen

    2018-01-16

    Metallo-β-lactamases (MBLs) are a group of enzymes that can inactivate most commonly used β-lactam-based antibiotics. Among MBLs, New Delhi metallo-β-lactamase-1 (NDM-1) constitutes an urgent threat to public health as evidenced by its success in rapidly disseminating worldwide since its first discovery. Here we report the biochemical and genetic characteristics of a novel MBL, ElBla2, from the marine bacterium Erythrobacter litoralis HTCC 2594. This enzyme has a higher amino acid sequence similarity to NDM-1 (56%) than any previously reported MBL. Enzymatic assays and secondary structure alignment also confirmed the high similarity between these two enzymes. Whole genome comparison of four Erythrobacter species showed that genes located upstream and downstream of elbla2 were highly conserved, which may indicate that elbla2 was lost during evolution. Furthermore, we predicted two prophages, 13 genomic islands and 25 open reading frames related to insertion sequences in the genome of E. litoralis HTCC 2594. However, unlike NDM-1, the chromosome encoded ElBla2 did not locate in or near these mobile genetic elements, indicating that it cannot transfer between strains. Finally, following our phylogenetic analysis, we suggest a reclassification of E. litoralis HTCC 2594 as a novel species: Erythrobacter sp. HTCC 2594.

  9. [Molecular mechanisms of protein biosynthesis initiation--biochemical and biomedical implications of a new model of translation enhanced by the RNA hypoxia response element (rHRE)].

    Science.gov (United States)

    Master, Adam; Nauman, Alicja

    2014-01-01

    Translation initiation is a key rate-limiting step in cellular protein synthesis. A cap-dependent initiation is the most effective mechanism of the translation. However, some physiological (mitosis) and pathological (oxidative stress) processes may switch the classic mechanism to an alternative one that is regulated by an mRNA element such as IRES, uORF, IRE, CPE, DICE, AURE or CITE. A recently discovered mechanism of RNA hypoxia response element (rHRE)-dependent translation initiation, may change the view of oxygen-regulated translation and give a new insight into unexplained biochemical processes. Hypoxia is one of the better-known factors that may trigger an alternative mechanism of the translation initiation. Temporal events of oxygen deficiency within tissues and organs may activate processes such as angiogenesis, myogenesis, regeneration, wound healing, and may promote an adaptive response in cardiovascular and neurodegenerative diseases. On the other hand, growth of solid tumors may be accompanied by cyclic hypoxia, allowing for synthesis of proteins required for further progression of cancer cells. This paper provides a review of current knowledge on translational control in the context of alternative models of translation initiation.

  10. Biochemical and functional characterization of AcUFGT3a, a galactosyltransferase involved in anthocyanin biosynthesis in the red-fleshed kiwifruit (Actinidia chinensis).

    Science.gov (United States)

    Liu, Yanfei; Zhou, Bin; Qi, Yingwei; Liu, Cuihua; Liu, Zhande; Ren, Xiaolin

    2018-04-01

    Much of the diversity of anthocyanin pigmentation in plant tissues is due to the action of glycosyltransferases, which attach sugar moieties to the anthocyanin aglycone. This step can increase both their solubility and stability. We investigated the pigmentation of the outer and inner pericarps of developing fruits of the red-fleshed kiwifruit Actinidia chinensis cv. 'Hongyang'. The results show that the red color of the inner pericarp is due to anthocyanin. Based on expression analyses of structural genes, AcUFGT was shown to be the key gene involved in the anthocyanin biosynthetic pathway. Expression of AcUFGT in developing fruit paralleled changes in anthocyanin concentration. Thirteen putative UFGT genes, including different transcripts, were identified in the genome of 'Hongyang'. Among these, only the expression of AcUFGT3a was found to be highly consistent with anthocyanin accumulation. Fruit infiltrated with virus-induced gene silencing showed delayed red colorations, lower anthocyanin contents and lower expressions of AcUFGT3a. At the same time, transient overexpression of AcUFGT3a in both Actinidia arguta and green apple fruit resulted in higher anthocyanin contents and deeper red coloration. In vitro biochemical assays revealed that recombinant AcUFGT3a recognized only anthocyanidins as substrate but not flavonols. Also, UDP-galactose was used preferentially as the sugar donor. These results indicate AcUFGT3a is the key enzyme regulating anthocyanin accumulation in red-fleshed kiwifruit. © 2017 Scandinavian Plant Physiology Society.

  11. Genetic, Biochemical, Nutritional and Antimicrobial Characteristics of Pomegranate (Punica granatum L. Grown in Istria

    Directory of Open Access Journals (Sweden)

    Ana Miklavčič Višnjevec

    2017-01-01

    Full Text Available This study characterises the genetic variability of local pomegranate (Punica granatum L. germplasm from the Slovenian and Croatian areas of Istria. The bioactive components and antioxidant and antimicrobial properties of ethanol and water extracts of different parts of pomegranate fruit were also determined, along with their preliminary nutritional characterisation. Twenty-six different genotypes identified with microsatellite analysis indicate the great diversity of pomegranate in Istria. The pomegranate fruit ethanol extracts represent rich sources of phenolic compounds (mean value of the mass fraction in exocarp and mesocarp expressed as gallic acid is 23 and 16 mg/g, respectively. The ethanol extracts of pomegranate exocarp and mesocarp showed the greatest antimicrobial activity against Candida albicans, Candida parapsilosis, Rhodotorula mucilaginosa, Exophiala dermatitidis and Staphylococcus aureus, and the same water extracts against S. aureus and Escherichia coli. To the best of our knowledge, this study represents the first report of the characterisation of pomegranate genetic resources from Istria at different levels, including the molecular, chemical, antimicrobial and nutritional properties.

  12. Genetic, Biochemical, Nutritional and Antimicrobial Characteristics of Pomegranate (Punica granatum L.)
Grown in Istria

    Science.gov (United States)

    2017-01-01

    Summary This study characterises the genetic variability of local pomegranate (Punica granatum L.) germplasm from the Slovenian and Croatian areas of Istria. The bioactive components and antioxidant and antimicrobial properties of ethanol and water extracts of different parts of pomegranate fruit were also determined, along with their preliminary nutritional characterisation. Twenty-six different genotypes identified with microsatellite analysis indicate the great diversity of pomegranate in Istria. The pomegranate fruit ethanol extracts represent rich sources of phenolic compounds (mean value of the mass fraction in exocarp and mesocarp expressed as gallic acid is 23 and 16 mg/g, respectively). The ethanol extracts of pomegranate exocarp and mesocarp showed the greatest antimicrobial activity against Candida albicans, Candida parapsilosis, Rhodotorula mucilaginosa, Exophiala dermatitidis and Staphylococcus aureus, and the same water extracts against S. aureus and Escherichia coli. To the best of our knowledge, this study represents the first report of the characterisation of pomegranate genetic resources from Istria at different levels, including the molecular, chemical, antimicrobial and nutritional properties. PMID:28867945

  13. Genetic engineering: a promising tool to engender physiological, biochemical and molecular stress resilience in green microalgae

    Directory of Open Access Journals (Sweden)

    Freddy eGuiheneuf

    2016-03-01

    Full Text Available As we march into the 21st century, the prevailing scenario of depleting energy resources, global warming and ever increasing issues of human health and food security will quadruple. In this context, genetic and metabolic engineering of green microalgae complete the quest towards a continuum of environmentally clean fuel and food production. Evolutionarily related, but unlike land plants, microalgae need nominal land or water, and are best described as unicellular autotrophs using light energy to fix atmospheric CO2 into algal biomass, mitigating fossil CO2 pollution in the process. Remarkably, a feature innate to most microalgae is synthesis and accumulation of lipids (60–65% of dry weight, carbohydrates and secondary metabolites like pigments and vitamins, especially when grown under abiotic stress conditions. Particularly fruitful, such an application of abiotic stress factors like nitrogen starvation , salinity, heat shock etc. can be used in a biorefinery concept for production of multiple valuable products. The focus of this mini-review underlies metabolic reorientation practices and tolerance mechanisms as applied to green microalgae under specific stress stimuli for a sustainable pollution-free future. Moreover, we entail current progress on genetic engineering as a promising tool to grasp adaptive processes for improving strains with potential biotechnological interests.

  14. Genetic, Biochemical, Nutritional and Antimicrobial Characteristics of Pomegranate (Punica granatum L.)
Grown in Istria.

    Science.gov (United States)

    Višnjevec, Ana Miklavčič; Ota, Ajda; Skrt, Mihaela; Butinar, Bojan; Možina, Sonja Smole; Cimerman, Nina Gunde; Nečemer, Marijan; Arbeiter, Alenka Baruca; Hladnik, Matjaž; Krapac, Marin; Ban, Dean; Bučar-Miklavčič, Milena; Ulrih, Nataša Poklar; Bandelj, Dunja

    2017-06-01

    This study characterises the genetic variability of local pomegranate ( Punica granatum L.) germplasm from the Slovenian and Croatian areas of Istria. The bioactive components and antioxidant and antimicrobial properties of ethanol and water extracts of different parts of pomegranate fruit were also determined, along with their preliminary nutritional characterisation. Twenty-six different genotypes identified with microsatellite analysis indicate the great diversity of pomegranate in Istria. The pomegranate fruit ethanol extracts represent rich sources of phenolic compounds (mean value of the mass fraction in exocarp and mesocarp expressed as gallic acid is 23 and 16 mg/g, respectively). The ethanol extracts of pomegranate exocarp and mesocarp showed the greatest antimicrobial activity against Candida albicans , Candida parapsilosis , Rhodotorula mucilaginosa , Exophiala dermatitidis and Staphylococcus aureus , and the same water extracts against S. aureus and Escherichia coli. To the best of our knowledge, this study represents the first report of the characterisation of pomegranate genetic resources from Istria at different levels, including the molecular, chemical, antimicrobial and nutritional properties.

  15. Enzymatic Reductive Dehalogenation Controls the Biosynthesis of Marine Bacterial Pyrroles.

    Science.gov (United States)

    El Gamal, Abrahim; Agarwal, Vinayak; Rahman, Imran; Moore, Bradley S

    2016-10-12

    Enzymes capable of performing dehalogenating reactions have attracted tremendous contemporary attention due to their potential application in the bioremediation of anthropogenic polyhalogenated persistent organic pollutants. Nature, in particular the marine environment, is also a prolific source of polyhalogenated organic natural products. The study of the biosynthesis of these natural products has furnished a diverse array of halogenation biocatalysts, but thus far no examples of dehalogenating enzymes have been reported from a secondary metabolic pathway. Here we show that the penultimate step in the biosynthesis of the highly brominated marine bacterial product pentabromopseudilin is catalyzed by an unusual debrominase Bmp8 that utilizes a redox thiol mechanism to remove the C-2 bromine atom of 2,3,4,5-tetrabromopyrrole to facilitate oxidative coupling to 2,4-dibromophenol. To the best of our knowledge, Bmp8 is first example of a dehalogenating enzyme from the established genetic and biochemical context of a natural product biosynthetic pathway.

  16. Duchenne/Becker muscular dystrophy: A report on clinical, biochemical, and genetic study in Gujarat population, India

    Directory of Open Access Journals (Sweden)

    Mandava V Rao

    2014-01-01

    Full Text Available Objective: In India, various groups have studied different regions to find out deletion pattern of dystrophin gene. We have investigated its deletion pattern among Duchenne/Becker muscular dystrophy (D/BMD patients across Gujarat. Moreover, in this study we also correlate the same with reading frame rule. However, we too consider various clinicopathological features to establish as adjunct indices when deletion detection fails. Materials and Methods: In this pilot study, a total of 88 D/BMD patients consulting at our centers in Gujarat, India were included. All patients were reviewed on basis of their clinical characteristics, tested by three primer sets of 10-plex, 9-plex, and 7-plex polymerase chain reaction (PCR for genetic analysis; whereas, biochemical indices were measured using automated biochemical analyzers. Results: The diagnosis of D/BMD was confirmed by multiplex-PCR (M-PCR in D/BMD patients. A number of 65 (73.86% out of 88 patients showed deletion in dystrophin gene. The exon 50 (58.46% was the most frequent deletion found in our study. The mean age of onset of DMD and BMD was 4.09 ΁ 0.15 and 7.14 ΁ 0.55 years, respectively. In patients, mean creatine phosphokinase (CPK, lactate dehydrogenase (LDH, and myoglobin levels were elevated significantly (P < 0.05 in comparison to controls. Addition to CPK, LDH and myoglobin are good adjunct when deletion detection failed. These data are further in accordance with world literature when correlated with frame rule. Conclusion: The analysis has been carried out for the first time for a total of 88 D/BMD patients particularly from Gujarat, India. More research is essential to elucidate specific mutation pattern in association with management and therapies of proband.

  17. Genetic and biochemical investigations of the role of MamP in redox control of iron biomineralization in Magnetospirillum magneticum.

    Science.gov (United States)

    Jones, Stephanie R; Wilson, Tiffany D; Brown, Margaret E; Rahn-Lee, Lilah; Yu, Yi; Fredriksen, Laura L; Ozyamak, Ertan; Komeili, Arash; Chang, Michelle C Y

    2015-03-31

    Magnetotactic bacteria have evolved complex subcellular machinery to construct linear chains of magnetite nanocrystals that allow the host cell to sense direction. Each mixed-valent iron nanoparticle is mineralized from soluble iron within a membrane-encapsulated vesicle termed the magnetosome, which serves as a specialized compartment that regulates the iron, redox, and pH environment of the growing mineral. To dissect the biological components that control this process, we have carried out a genetic and biochemical study of proteins proposed to function in iron mineralization. In this study, we show that the redox sites of c-type cytochromes of the Magnetospirillum magneticum AMB-1 magnetosome island, MamP and MamT, are essential to their physiological function and that ablation of one or both heme motifs leads to loss of function, suggesting that their ability to carry out redox chemistry in vivo is important. We also develop a method to heterologously express fully heme-loaded MamP from AMB-1 for in vitro biochemical studies, which show that its Fe(III)-Fe(II) redox couple is set at an unusual potential (-89 ± 11 mV) compared with other related cytochromes involved in iron reduction or oxidation. Despite its low reduction potential, it remains competent to oxidize Fe(II) to Fe(III) and mineralize iron to produce mixed-valent iron oxides. Finally, in vitro mineralization experiments suggest that Mms mineral-templating peptides from AMB-1 can modulate the iron redox chemistry of MamP.

  18. Genetic Characterization of Coenzyme A Biosynthesis Reveals Essential Distinctive Functions during Malaria Parasite Development in Blood and Mosquito

    Directory of Open Access Journals (Sweden)

    Robert J. Hart

    2017-06-01

    Full Text Available Coenzyme A (CoA is an essential universal cofactor for all prokaryotic and eukaryotic cells. In nearly all non-photosynthetic cells, CoA biosynthesis depends on the uptake and phosphorylation of vitamin B5 (pantothenic acid or pantothenate. Recently, putative pantothenate transporter (PAT and pantothenate kinases (PanKs were functionally characterized in P. yoelii. PAT and PanKs were shown to be dispensable for blood stage development, but they were essential for mosquito stages development. Yet, little is known about the cellular functions of the other enzymes of the CoA biosynthesis pathway in malaria parasite life cycle stages. All enzymes of this pathway were targeted for deletion or deletion/complementation analyses by knockout/knock-in plasmid constructs to reveal their essential roles in P. yoelii life cycle stages. The intermediate enzymes PPCS (Phosphopantothenylcysteine Synthase, PPCDC (Phosphopantothenylcysteine Decarboxylase were shown to be dispensable for asexual and sexual blood stage development, but they were essential for oocyst development and the production of sporozoites. However, the last two enzymes of this pathway, PPAT (Phosphopantetheine Adenylyltransferase and DPCK (Dephospho-CoA Kinase, were essential for blood stage development. These results indicate alternative first substrate requirement for the malaria parasite, other than the canonical pantothenate, for the synthesis of CoA in the blood but not inside the mosquito midgut. Collectively, our data shows that CoA de novo biosynthesis is essential for both blood and mosquito stages, and thus validates the enzymes of this pathway as potential antimalarial targets.

  19. Genetic Characterization of Coenzyme A Biosynthesis Reveals Essential Distinctive Functions during Malaria Parasite Development in Blood and Mosquito.

    Science.gov (United States)

    Hart, Robert J; Abraham, Amanah; Aly, Ahmed S I

    2017-01-01

    Coenzyme A (CoA) is an essential universal cofactor for all prokaryotic and eukaryotic cells. In nearly all non-photosynthetic cells, CoA biosynthesis depends on the uptake and phosphorylation of vitamin B5 (pantothenic acid or pantothenate). Recently, putative pantothenate transporter (PAT) and pantothenate kinases (PanKs) were functionally characterized in P. yoelii . PAT and PanKs were shown to be dispensable for blood stage development, but they were essential for mosquito stages development. Yet, little is known about the cellular functions of the other enzymes of the CoA biosynthesis pathway in malaria parasite life cycle stages. All enzymes of this pathway were targeted for deletion or deletion/complementation analyses by knockout/knock-in plasmid constructs to reveal their essential roles in P. yoelii life cycle stages. The intermediate enzymes PPCS (Phosphopantothenylcysteine Synthase), PPCDC (Phosphopantothenylcysteine Decarboxylase) were shown to be dispensable for asexual and sexual blood stage development, but they were essential for oocyst development and the production of sporozoites. However, the last two enzymes of this pathway, PPAT (Phosphopantetheine Adenylyltransferase) and DPCK (Dephospho-CoA Kinase), were essential for blood stage development. These results indicate alternative first substrate requirement for the malaria parasite, other than the canonical pantothenate, for the synthesis of CoA in the blood but not inside the mosquito midgut. Collectively, our data shows that CoA de novo biosynthesis is essential for both blood and mosquito stages, and thus validates the enzymes of this pathway as potential antimalarial targets.

  20. Biochemical and genetic role of apelin in essential hypertension and acute coronary syndrome.

    Science.gov (United States)

    Gupta, Mohit D; Girish, M P; Shah, Dhaval; Rain, Manjari; Mehta, Vimal; Tyagi, Sanjay; Trehan, Vijay; Pasha, Qadar

    2016-11-15

    Apelin-APJ pathway has emerged as a potent regulator of blood pressure (BP) and blood flow in vasculature and heart. Variants in apelin gene may affect the vascular tone in peripheral circulation or heart, thereby predisposing to cardiovascular diseases. The aim of our study was to investigate the association of two apelin gene polymorphisms rs3761581 and rs2235312, and apelin levels in patients with essential hypertension (EH) and acute coronary syndrome (ACS). The study comprised of three groups namely, (1) 118 healthy control subjects, (2) 92 EH patients, and (3) 60 ACS patients. DNA was extracted from peripheral blood leukocytes and genotyping was performed by SNaPshot method. Plasma apelin 13 levels were estimated using ELISA. EH and ACS patients had a significantly lower level of apelin 13, regardless of gender (p=0.003, p=0.017, respectively). Interestingly, the female EH and ACS patients had lower levels of apelin 13 than their male counterparts. The G allele of rs3761581 was more apparent in patients especially in ACS than the controls. Reduced apelin levels may enhance vasoconstriction to influence high BP and heart's workload in EH and ACS. Genetic involvement of apelin needs to be established in well-defined larger sample size. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  1. Biochemical and genetic variation of some Syrian wheat varieties using NIR, RAPD and AFLPs techniques

    International Nuclear Information System (INIS)

    Saleh, B.

    2012-01-01

    This study was performed to assess chemical components and genetic variability of five Syrian wheat varieties using NIR, RAPD and AFLP techniques. NIR technique showed that Cham6 was the best variety in term of wheat grain quality due to their lowest protein (%), hardness, water uptake and baking volume and the highest starch (%) compared to the other tested varieties. PCR amplifications with 21 RAPD primers and 13 AFLP PCs primer combinations gave 104 and 466 discernible loci of which 24 (18.823%) and 199 (45.527%) were polymorphic for the both techniques respectively. Our data indicated that the three techniques gave similar results regarding the degree of relatedness among the tested varieties. In the present investigation, AFLP fingerprinting was more efficient than the RAPD assay. Where the letter exhibited lower Marker Index (MI) average (0.219) compared to AFLP one (3.203). The pattern generated by RAPD, AFLPs markers or by NIR separated the five wheat varieties into two groups. The first group consists of two subclusters. The first subcluster involved Cham8 and Bohous6, while the second one includes Cham6 that is very closed to precedent varieties. The second group consists of Bohous9 and Cham7 that were also closely related. Based on this study, the use of NIR, RAPD and AFLP techniques could be a powerful tool to detect the effectiveness relationships of these technologies. (author)

  2. Phosphatidylserine biosynthesis in cultured Chinese hamster ovary cells. III. Genetic evidence for utilization of phosphatidylcholine and phosphatidylethanolamine as precursors

    International Nuclear Information System (INIS)

    Kuge, O.; Nishijima, M.; Akamatsu, Y.

    1986-01-01

    We reported that Chinese hamster ovary (CHO) cells contain two different serine-exchange enzymes (I and II) which catalyze the base-exchange reaction of phospholipid(s) with serine and that a phosphatidylserine-requiring mutant (strain PSA-3) of CHO cells is defective in serine-exchange enzyme I and lacks the ability to synthesize phosphatidylserine. In this study, we examined precursor phospholipids for phosphatidylserine biosynthesis in CHO cells. When mutant PSA-3 and parent (CHO-K1) cells were cultured with [ 32 P]phosphatidylcholine, phosphatidylserine in the parent accumulated radioactivity while that in the mutant was not labeled significantly. On the contrary, when cultured with [ 32 P]phosphatidylethanolamine, the mutant incorporated the label into phosphatidylserine more efficiently than the parent. Furthermore, we found that mutant PSA-3 grew normally in growth medium supplemented with 30 microM phosphatidylethanolamine as well as phosphatidylserine and that the biosynthesis of phosphatidylserine in the mutant was normal when cells were cultured in the presence of exogenous phosphatidylethanolamine. The simplest interpretation of these findings is that phosphatidylserine in CHO cells is biosynthesized through the following sequential reactions: phosphatidylcholine----phosphatidylserine----phosphatidylethanolamine--- - phosphatidylserine. The three reactions are catalyzed by serine-exchange enzyme I, phosphatidylserine decarboxylase, and serine-exchange enzyme II, respectively

  3. New insights into bacterial type II polyketide biosynthesis [version 1; referees: 2 approved

    Directory of Open Access Journals (Sweden)

    Zhuan Zhang

    2017-02-01

    Full Text Available Bacterial aromatic polyketides, exemplified by anthracyclines, angucyclines, tetracyclines, and pentangular polyphenols, are a large family of natural products with diverse structures and biological activities and are usually biosynthesized by type II polyketide synthases (PKSs. Since the starting point of biosynthesis and combinatorial biosynthesis in 1984–1985, there has been a continuous effort to investigate the biosynthetic logic of aromatic polyketides owing to the urgent need of developing promising therapeutic candidates from these compounds. Recently, significant advances in the structural and mechanistic identification of enzymes involved in aromatic polyketide biosynthesis have been made on the basis of novel genetic, biochemical, and chemical technologies. This review highlights the progress in bacterial type II PKSs in the past three years (2013–2016. Moreover, novel compounds discovered or created by genome mining and biosynthetic engineering are also included.

  4. Pathogenicity of novel ABCD1 variants: The need for biochemical testing in the era of advanced genetics.

    Science.gov (United States)

    Schackmann, Martin J A; Ofman, Rob; van Geel, Björn M; Dijkstra, Inge M E; van Engelen, Klaartje; Wanders, Ronald J A; Engelen, Marc; Kemp, Stephan

    2016-06-01

    X-linked adrenoleukodystrophy (ALD), a progressive neurodegenerative disease, is caused by mutations in ABCD1 and characterized by very-long-chain fatty acids (VLCFA) accumulation. In male patients, an increased plasma VLCFA levels in combination with a pathogenic mutation in ABCD1 confirms the diagnosis. Recent studies have shown that many women with ALD also develop myelopathy. Correct diagnosis is important for management including genetic counseling. Diagnosis in women can only be confirmed when VLCFA levels are elevated or when a known pathogenic ABCD1 mutation is identified. However, in 15-20% of women with ALD VLCFA plasma levels are not elevated. Demonstration that a novel sequence variant is pathogenic can be a challenge when VLCFA levels are in the normal range. Here we report two women with a clinical presentation compatible with ALD, an ABCD1 variation (p.Arg17His and p.Ser358Pro) of unknown significance, but with normal VLCFA levels. We developed a diagnostic test that is based on generating clonal cell lines that express only one of the two alleles. Subsequent biochemical studies enabled us to show that the two sequence variants were not pathogenic, thereby excluding the diagnosis ALD in these women. We conclude that the clonal approach is an important addition to the existing diagnostic array. Copyright © 2016 Elsevier Inc. All rights reserved.

  5. Evaluation of biochemical parameters and genetic markers for association with meat tenderness in South African feedlot cattle.

    Science.gov (United States)

    Frylinck, L; van Wyk, G L; Smith, T P L; Strydom, P E; van Marle-Köster, E; Webb, E C; Koohmaraie, M; Smith, M F

    2009-12-01

    A large proportion of South African feedlot cattle are crossbreds of Brahman (BrX, Bos indicus), and Simmental (SiX, Bos taurus). A sample of 20 grain fed bulls from each of these crossbreeds was used to compare meat quality with that of the small frame indigenous Nguni (NgX, Sanga) by evaluating a variety of biochemical and genetic parameters previously shown to be associated with meat tenderness. Shear force values were generally high (5.6kg average at 14days post mortem), with SiX animals higher than BrX or NgX (P=0.051) despite higher calpastatin:calpain ratio in BrX (P<0.05). Calpain activity and cold shortening were both correlated with tenderness for all classes. The sample size was too small to accurately estimate genotypic effects of previously published markers in the CAST and CAPN1 genes, but the allele frequencies suggest that only modest progress would be possible in these South African crossbreds using these markers.

  6. Assessment of a land-locked Atlantic salmon (Salmo salar L.) population as a potential genetic resource with a focus on long-chain polyunsaturated fatty acid biosynthesis.

    Science.gov (United States)

    Betancor, M B; Olsen, R E; Solstorm, D; Skulstad, O F; Tocher, D R

    2016-03-01

    The natural food for Atlantic salmon (Salmo salar) in freshwater has relatively lower levels of omega-3 (n-3) long-chain polyunsaturated fatty acids (LC-PUFA) than found in prey for post-smolt salmon in seawater. Land-locked salmon such as the Gullspång population feed exclusively on freshwater type lipids during its entire life cycle, a successful adaptation derived from divergent evolution. Studying land-locked populations may provide insights into the molecular and genetic control mechanisms that determine and regulate n-3 LC-PUFA biosynthesis and retention in Atlantic salmon. A two factorial study was performed comparing land-locked and farmed salmon parr fed diets formulated with fish or rapeseed oil for 8 weeks. The land-locked parr had higher capacity to synthesise n-3 LC-PUFA as indicated by higher expression and activity of desaturase and elongase enzymes. The data suggested that the land-locked salmon had reduced sensitivity to dietary fatty acid composition and that dietary docosahexaenoic acid (DHA) did not appear to suppress expression of LC-PUFA biosynthetic genes or activity of the biosynthesis pathway, probably an evolutionary adaptation to a natural diet lower in DHA. Increased biosynthetic activity did not translate to enhanced n-3 LC-PUFA contents in the flesh and diet was the only factor affecting this parameter. Additionally, high lipogenic and glycolytic potentials were found in land-locked salmon, together with decreased lipolysis which in turn could indicate increased use of carbohydrates as an energy source and a sparing of lipid. Copyright © 2015 Elsevier B.V. All rights reserved.

  7. Serine biosynthesis and transport defects.

    Science.gov (United States)

    El-Hattab, Ayman W

    2016-07-01

    l-serine is a non-essential amino acid that is biosynthesized via the enzymes phosphoglycerate dehydrogenase (PGDH), phosphoserine aminotransferase (PSAT), and phosphoserine phosphatase (PSP). Besides its role in protein synthesis, l-serine is a potent neurotrophic factor and a precursor of a number of essential compounds including phosphatidylserine, sphingomyelin, glycine, and d-serine. Serine biosynthesis defects result from impairments of PGDH, PSAT, or PSP leading to systemic serine deficiency. Serine biosynthesis defects present in a broad phenotypic spectrum that includes, at the severe end, Neu-Laxova syndrome, a lethal multiple congenital anomaly disease, intermediately, infantile serine biosynthesis defects with severe neurological manifestations and growth deficiency, and at the mild end, the childhood disease with intellectual disability. A serine transport defect resulting from deficiency of the ASCT1, the main transporter for serine in the central nervous system, has been recently described in children with neurological manifestations that overlap with those observed in serine biosynthesis defects. l-serine therapy may be beneficial in preventing or ameliorating symptoms in serine biosynthesis and transport defects, if started before neurological damage occurs. Herein, we review serine metabolism and transport, the clinical, biochemical, and molecular aspects of serine biosynthesis and transport defects, the mechanisms of these diseases, and the potential role of serine therapy. Copyright © 2016 Elsevier Inc. All rights reserved.

  8. Biochemical and genetic toxicity of the ionic liquid 1-octyl-3-methylimidazolium chloride on earthworms (Eisenia fetida).

    Science.gov (United States)

    Guo, Yingying; Liu, Tong; Zhang, Jun; Wang, Jinhua; Wang, Jun; Zhu, Lusheng; Yang, Jinhui

    2016-02-01

    Ionic liquids also known as "green solvents," are used in many fields. However, the dispersion of ionic liquids in soil systems is likely to cause damage to soil organisms. The objective of the present study was to investigate the toxicity of 1-octyl-3-methylimidazolium chloride ([C8 mim]Cl) on earthworms (Eisenia fetida). For this purpose, earthworms were exposed to different concentrations of [C8 mim]Cl (0 mg/kg, 5 mg/kg, 10 mg/kg, 20 mg/kg, and 40 mg/kg artificial soil) and sampled at 7 d, 14 d, 21 d, and 28 d. The results indicated that [C8 mim]Cl could cause an accumulation of reactive oxygen species (ROS) in earthworms, even at the lowest concentration (5 mg/kg). Compared with the controls, during the [C8 mim]Cl exposure period, the activities of superoxide dismutase (SOD) and catalase (CAT) decreased and then increased, whereas the activities of peroxidase (POD) and glutathione S-transferase (GST) increased. These changes in the activities of antioxidant enzymes and GST indicated that [C8 mim]Cl could induce oxidative damage in earthworms. The malondialdehyde content was increased by high levels of [C8 mim]Cl at 14 d and 28 d, indicating that [C8 mim]Cl could lead to lipid peroxidation in earthworms. In addition, the degree of DNA damage significantly increased with increasing [C8 mim]Cl concentrations and exposure time. The present study shows that [C8 mim]Cl caused biochemical and genetic toxicity in earthworms. © 2015 SETAC.

  9. Genetic and Biochemical Identification of a Novel Single-Stranded DNA-Binding Complex in Haloferax volcanii.

    Science.gov (United States)

    Stroud, Amy; Liddell, Susan; Allers, Thorsten

    2012-01-01

    Single-stranded DNA (ssDNA)-binding proteins play an essential role in DNA replication and repair. They use oligonucleotide/oligosaccharide-binding (OB)-folds, a five-stranded β-sheet coiled into a closed barrel, to bind to ssDNA thereby protecting and stabilizing the DNA. In eukaryotes the ssDNA-binding protein (SSB) is known as replication protein A (RPA) and consists of three distinct subunits that function as a heterotrimer. The bacterial homolog is termed SSB and functions as a homotetramer. In the archaeon Haloferax volcanii there are three genes encoding homologs of RPA. Two of the rpa genes (rpa1 and rpa3) exist in operons with a novel gene specific to Euryarchaeota; this gene encodes a protein that we have termed RPA-associated protein (rpap). The rpap genes encode proteins belonging to COG3390 group and feature OB-folds, suggesting that they might cooperate with RPA in binding to ssDNA. Our genetic analysis showed that rpa1 and rpa3 deletion mutants have differing phenotypes; only Δrpa3 strains are hypersensitive to DNA damaging agents. Deletion of the rpa3-associated gene rpap3 led to similar levels of DNA damage sensitivity, as did deletion of the rpa3 operon, suggesting that RPA3 and RPAP3 function in the same pathway. Protein pull-downs involving recombinant hexahistidine-tagged RPAs showed that RPA3 co-purifies with RPAP3, and RPA1 co-purifies with RPAP1. This indicates that the RPAs interact only with their respective associated proteins; this was corroborated by the inability to construct rpa1 rpap3 and rpa3 rpap1 double mutants. This is the first report investigating the individual function of the archaeal COG3390 RPA-associated proteins (RPAPs). We have shown genetically and biochemically that the RPAPs interact with their respective RPAs, and have uncovered a novel single-stranded DNA-binding complex that is unique to Euryarchaeota.

  10. Genetic Variation in Plant CYP51s Confers Resistance against Voriconazole, a Novel Inhibitor of Brassinosteroid-Dependent Sterol Biosynthesis

    Science.gov (United States)

    Rozhon, Wilfried; Husar, Sigrid; Kalaivanan, Florian; Khan, Mamoona; Idlhammer, Markus; Shumilina, Daria; Lange, Theo; Hoffmann, Thomas; Schwab, Wilfried; Fujioka, Shozo; Poppenberger, Brigitte

    2013-01-01

    Brassinosteroids (BRs) are plant steroid hormones with structural similarity to mammalian sex steroids and ecdysteroids from insects. The BRs are synthesized from sterols and are essential regulators of cell division, cell elongation and cell differentiation. In this work we show that voriconazole, an antifungal therapeutic drug used in human and veterinary medicine, severely impairs plant growth by inhibiting sterol-14α-demethylation and thereby interfering with BR production. The plant growth regulatory properties of voriconazole and related triazoles were identified in a screen for compounds with the ability to alter BR homeostasis. Voriconazole suppressed growth of the model plant Arabidopsis thaliana and of a wide range of both monocotyledonous and dicotyledonous plants. We uncover that voriconazole toxicity in plants is a result of a deficiency in BRs that stems from an inhibition of the cytochrome P450 CYP51, which catalyzes a step of BR-dependent sterol biosynthesis. Interestingly, we found that the woodland strawberry Fragaria vesca, a member of the Rosaceae, is naturally voriconazole resistant and that this resistance is conferred by the specific CYP51 variant of F. vesca. The potential of voriconazole as a novel tool for plant research is discussed. PMID:23335967

  11. Genetic heterogeneity and clinical variability in musculocontractural Ehlers-Danlos syndrome caused by impaired dermatan sulfate biosynthesis.

    Science.gov (United States)

    Syx, Delfien; Van Damme, Tim; Symoens, Sofie; Maiburg, Merel C; van de Laar, Ingrid; Morton, Jenny; Suri, Mohnish; Del Campo, Miguel; Hausser, Ingrid; Hermanns-Lê, Trinh; De Paepe, Anne; Malfait, Fransiska

    2015-05-01

    Bi-allelic variants in CHST14, encoding dermatan 4-O-sulfotransferase-1 (D4ST1), cause musculocontractural Ehlers-Danlos syndrome (MC-EDS), a recessive disorder characterized by connective tissue fragility, craniofacial abnormalities, congenital contractures, and developmental anomalies. Recently, the identification of bi-allelic variants in DSE, encoding dermatan sulfate epimerase-1 (DS-epi1), in a child with MC-EDS features, suggested locus heterogeneity for this condition. DS-epi1 and D4ST1 are crucial for biosynthesis of dermatan sulfate (DS) moieties in the hybrid chondroitin sulfate (CS)/DS glycosaminoglycans (GAGs). Here, we report four novel families with severe MC-EDS caused by unique homozygous CHST14 variants and the second family with a homozygous DSE missense variant, presenting a somewhat milder MC-EDS phenotype. The glycanation of the dermal DS proteoglycan decorin is impaired in fibroblasts from D4ST1- as well as DS-epi1-deficient patients. However, in D4ST1-deficiency, the decorin GAG is completely replaced by CS, whereas in DS-epi1-deficiency, still some DS moieties are present. The multisystemic abnormalities observed in our patients support a tight spatiotemporal control of the balance between CS and DS, which is crucial for multiple processes including cell differentiation, organ development, cell migration, coagulation, and connective tissue integrity. © 2015 WILEY PERIODICALS, INC.

  12. Acetylglutamate synthase in Neurospora crassa: characterization, localization, and genetic behavior of a regulatory enzyme of arginine biosynthesis

    International Nuclear Information System (INIS)

    Jacobson, J.A.

    1988-01-01

    This study describes the characterization and localization of the first enzyme of arginine biosynthesis in Neurospora crassa. A radioactive assay was developed to detect this enzyme whereby radioactive substrate and product molecules could be separated by ion-exchange chromatography. The enzyme was found to have a pH optimum of 9.0 and K/sub m/ values for glutamate and acetyl-CoA of approximately 4.7 and 0.45 mM, respectively. The enzyme was shown to be feedback inhibited by arginine. Half-maximal inhibition was observed at 0.13 mM arginine, a concentration which is similar to be in vivo cytosolic concentration of 0.2 mM. Arginine was found to act as a competitive inhibitor with respect to acetyl-CoA. Acetylglutamate synthase was localized to the mitochondrion. However, in contrast to the mitochondrial matrix location of the other ornithine biosynthetic enzymes, this enzyme was found to reside on the mitochondrial inner membrane

  13. Genetic variation in plant CYP51s confers resistance against voriconazole, a novel inhibitor of brassinosteroid-dependent sterol biosynthesis.

    Science.gov (United States)

    Rozhon, Wilfried; Husar, Sigrid; Kalaivanan, Florian; Khan, Mamoona; Idlhammer, Markus; Shumilina, Daria; Lange, Theo; Hoffmann, Thomas; Schwab, Wilfried; Fujioka, Shozo; Poppenberger, Brigitte

    2013-01-01

    Brassinosteroids (BRs) are plant steroid hormones with structural similarity to mammalian sex steroids and ecdysteroids from insects. The BRs are synthesized from sterols and are essential regulators of cell division, cell elongation and cell differentiation. In this work we show that voriconazole, an antifungal therapeutic drug used in human and veterinary medicine, severely impairs plant growth by inhibiting sterol-14α-demethylation and thereby interfering with BR production. The plant growth regulatory properties of voriconazole and related triazoles were identified in a screen for compounds with the ability to alter BR homeostasis. Voriconazole suppressed growth of the model plant Arabidopsis thaliana and of a wide range of both monocotyledonous and dicotyledonous plants. We uncover that voriconazole toxicity in plants is a result of a deficiency in BRs that stems from an inhibition of the cytochrome P450 CYP51, which catalyzes a step of BR-dependent sterol biosynthesis. Interestingly, we found that the woodland strawberry Fragaria vesca, a member of the Rosaceae, is naturally voriconazole resistant and that this resistance is conferred by the specific CYP51 variant of F. vesca. The potential of voriconazole as a novel tool for plant research is discussed.

  14. Genetic variation in plant CYP51s confers resistance against voriconazole, a novel inhibitor of brassinosteroid-dependent sterol biosynthesis.

    Directory of Open Access Journals (Sweden)

    Wilfried Rozhon

    Full Text Available Brassinosteroids (BRs are plant steroid hormones with structural similarity to mammalian sex steroids and ecdysteroids from insects. The BRs are synthesized from sterols and are essential regulators of cell division, cell elongation and cell differentiation. In this work we show that voriconazole, an antifungal therapeutic drug used in human and veterinary medicine, severely impairs plant growth by inhibiting sterol-14α-demethylation and thereby interfering with BR production. The plant growth regulatory properties of voriconazole and related triazoles were identified in a screen for compounds with the ability to alter BR homeostasis. Voriconazole suppressed growth of the model plant Arabidopsis thaliana and of a wide range of both monocotyledonous and dicotyledonous plants. We uncover that voriconazole toxicity in plants is a result of a deficiency in BRs that stems from an inhibition of the cytochrome P450 CYP51, which catalyzes a step of BR-dependent sterol biosynthesis. Interestingly, we found that the woodland strawberry Fragaria vesca, a member of the Rosaceae, is naturally voriconazole resistant and that this resistance is conferred by the specific CYP51 variant of F. vesca. The potential of voriconazole as a novel tool for plant research is discussed.

  15. Lotus tenuis x L. corniculatus interspecific hybridization as a means to breed bloat-safe pastures and gain insight into the genetic control of proanthocyanidin biosynthesis in legumes.

    Science.gov (United States)

    Escaray, Francisco J; Passeri, Valentina; Babuin, Florencia M; Marco, Francisco; Carrasco, Pedro; Damiani, Francesco; Pieckenstain, Fernando L; Paolocci, Francesco; Ruiz, Oscar A

    2014-02-03

    Proanthocyanidins (PAs) are secondary metabolites that strongly affect plant quality traits. The concentration and the structure of these metabolites influence the palatability and nutritional value of forage legumes. Hence, modulating PAs in the leaves of forage legumes is of paramount relevance for forage breeders worldwide. The lack of genetic variation in the leaf PA trait within the most important forage species and the difficulties in engineering this pathway via the ectopic expression of regulatory genes, prompted us to pursue alternative strategies to enhance this trait in forage legumes of agronomic interest. The Lotus genus includes forage species which accumulate PAs in edible organs and can thus be used as potential donor parents in breeding programs. We recovered a wild, diploid and PA-rich population of L. corniculatus and crossed with L. tenuis. The former grows in an alkaline-salty area in Spain while the latter is a diploid species, grown extensively in South American pastures, which does not accumulate PAs in the herbage. The resulting interspecific hybrids displayed several traits of outstanding agronomic relevance such as rhizome production, PA levels in edible tissues sufficient to prevent ruminal bloating (around 5 mg of PAs/g DW), biomass production similar to the cultivated parent and potential for adaptability to marginal lands. We show that PA levels correlate with expression levels of the R2R3MYB transcription factor TT2 and, in turn, with those of the key structural genes of the epicatechin and catechin biosynthetic pathways leading to PA biosynthesis. The L. tenuis x L. corniculatus hybrids, reported herein, represent the first example of the introgression of the PA trait in forage legumes to levels known to provide nutritional and health benefits to ruminants. Apart from PAs, the hybrids have additional traits which may prove useful to breed forage legumes with increased persistence and adaptability to marginal conditions. Finally, our

  16. A molecular genetic analysis of carotenoid biosynthesis and the effects of carotenoid mutations on other photosynthetic genes in Rhodobacter capsulatus

    Energy Technology Data Exchange (ETDEWEB)

    Armstrong, G.A.

    1989-04-01

    The nine known R. capsulatus carotenoid genes are contained within the 46 kilobase (kb) photosynthesis gene cluster. An 11 kb subcluster containing eight of these genes has been cloned and its nucleotide sequence determined. A new gene, crtK, has been located in the middle of the subcluster. The carotenoid gene cluster contains sequences homologous to Escherichia coli ..omega../sup 70/ promoters, rho-independent transcription terminators, and prokaryotic transcriptional factor binding sites. The phenotypes and genotypes of ten transposon Tn5.7 insertion mutations within the carotenoid gene cluster have been analyzed, by characterization of the carotenoids accumulated and high resolution mapping of the Tn5.7 insertions. The enzymatic blockages in previously uncharacterized early carotenoid mutants have been determined using a new in vitro synthesis system, suggesting specific roles for the CrtB and CrtE gene products. The expression of six of the eight carotenoid genes in the cluster is induced upon the shift from dark chemoheterotrophic to anaerobic photosynthetic growth. The magnitude of the induction is equivalent to that of genes encoding structural photosynthesis polypeptides, although the carotenoid genes are induced earlier after the growth shift. Different means of regulating photosynthesis genes in R. capsulatus are discussed, and a rationale for the temporal pattern of expression of the carotenoid genes during photosynthetic adaptation is presented. Comparison of the deduced amino acid sequences of the two dehydrogenases of the R. capsulatus carotenoid biosynthesis pathway reveals two regions of strong similarity. The effect of carotenoid mutations on the photosynthetic phenotype has been studied by examining growth rates, pigments, pigment-protein complexes and gene expression for a complete set of carotenoid mutants. 161 refs.

  17. Genetic and Functional Analysis of the Biosynthesis of a Non-Ribosomal Peptide Siderophore in Burkholderia xenovorans LB400.

    Directory of Open Access Journals (Sweden)

    María José Vargas-Straube

    Full Text Available B. xenovorans LB400 is a model bacterium for the study of the metabolism of aromatic compounds. The aim of this study was the genomic and functional characterization of a non-ribosomal peptide synthetase containing gene cluster that encodes a siderophore in B. xenovorans LB400. The mba gene cluster from strain LB400 encodes proteins involved in the biosynthesis and transport of a hydroxamate-type siderophore. Strain LB400 has a unique mba gene organization, although mba gene clusters have been observed in diverse Burkholderiales. Bioinformatic analysis revealed the presence of promoters in the mba gene cluster that strongly suggest regulation by the ferric uptake regulator protein (Fur and by the alternative RNA polymerase extracytoplasmic function sigma factor MbaF. Reverse transcriptase PCR analyses showed the expression of iron-regulated transcriptional units mbaFGHIJKL, mbaN, mbaABCE, mbaO, mbaP and mbaD genes under iron limitation. Chrome azurol S (CAS assay strongly suggests that strain LB400 synthesized a siderophore under iron limitation. Mass spectrometry ESI-MS and MALDI-TOF-MS analyses revealed that the siderophore is a non-ribosomal peptide, and forms an iron complex with a molecular mass of 676 Da. Based on bioinformatic prediction, CAS assay and MS analyses, we propose that the siderophore is L-Nδ-hydroxy-Nδ-formylOrn-D-β-hydroxyAsp-L-Ser-L-Nδ-hydroxy-Nδ-formylOrn-1,4-diaminobutane that is closely related to malleobactin-type siderophores reported in B. thailandensis.

  18. A molecular genetic analysis of carotenoid biosynthesis and the effects of carotenoid mutations on other photosynthetic genes in Rhodobacter capsulatus

    International Nuclear Information System (INIS)

    Armstrong, G.A.

    1989-04-01

    The nine known R. capsulatus carotenoid genes are contained within the 46 kilobase (kb) photosynthesis gene cluster. An 11 kb subcluster containing eight of these genes has been cloned and its nucleotide sequence determined. A new gene, crtK, has been located in the middle of the subcluster. The carotenoid gene cluster contains sequences homologous to Escherichia coli ω 70 promoters, rho-independent transcription terminators, and prokaryotic transcriptional factor binding sites. The phenotypes and genotypes of ten transposon Tn5.7 insertion mutations within the carotenoid gene cluster have been analyzed, by characterization of the carotenoids accumulated and high resolution mapping of the Tn5.7 insertions. The enzymatic blockages in previously uncharacterized early carotenoid mutants have been determined using a new in vitro synthesis system, suggesting specific roles for the CrtB and CrtE gene products. The expression of six of the eight carotenoid genes in the cluster is induced upon the shift from dark chemoheterotrophic to anaerobic photosynthetic growth. The magnitude of the induction is equivalent to that of genes encoding structural photosynthesis polypeptides, although the carotenoid genes are induced earlier after the growth shift. Different means of regulating photosynthesis genes in R. capsulatus are discussed, and a rationale for the temporal pattern of expression of the carotenoid genes during photosynthetic adaptation is presented. Comparison of the deduced amino acid sequences of the two dehydrogenases of the R. capsulatus carotenoid biosynthesis pathway reveals two regions of strong similarity. The effect of carotenoid mutations on the photosynthetic phenotype has been studied by examining growth rates, pigments, pigment-protein complexes and gene expression for a complete set of carotenoid mutants. 161 refs

  19. Simulation of quantitative characters by genes with biochemically definable action. III. The components of genetic effects in the inheritance of anthocyanins in Matthiola incana R. Br.

    Science.gov (United States)

    Jana, S; Seyffert, W

    1971-01-01

    In a self-pollinated plant species, Matthiola incana R. Br., six groups of isogenic lines were developed which were ideally suited for investigating the properties of individual genes controlling a quantitative character. Each group consisted of four homozygous parents for two alleles at each of the two loci in a common genetic background. A complete 4 × 4 diallel cross was obtained in each group. Because of the identical genetic background each diallel set could be considered as a genetic system of two loci. The biochemical functions of the alleles at each locus modifying the structure of the anthocyanin molecule were known. The phenotypes of the nine possible genotypes were qualitatively distinguishable by their flower colour differences. A quantitative measure of the phenotypic value associated with a genotype is the concentration of anthocyanins in flower tissues. In these simplified genetic systems, the nine phenotypic values could be expressed in terms of nine biometrical quantities, eight of which are attributable to the genetic effects of the alleles at the two loci under consideration. An unique solution of the set of nine equations in nine unknowns provided direct estimates of the parameters specifying additive, dominance and epistatic effects. Thus the effects of individual genes in a well-defined genetic background could be estimated by the use of a simple additive genetic model. An extension of the model provided estimates of the genetic parameters in different years and genetic backgrounds.Dominance was found to be the most important type of gene action in the inheritance of anthocyanin content in the flower tissues of M. incana. There was considerable epistasis, but the effect was very unstable over years and genetic backgrounds. The relative magnitude of additive effect was most stable. Heterosis was observed and was found to be largely due to dominance and additive × dominance interactions.

  20. Clinical, Biochemical, and Genetic Characterization of North American Patients With Erythropoietic Protoporphyria and X-linked Protoporphyria.

    Science.gov (United States)

    Balwani, Manisha; Naik, Hetanshi; Anderson, Karl E; Bissell, D Montgomery; Bloomer, Joseph; Bonkovsky, Herbert L; Phillips, John D; Overbey, Jessica R; Wang, Bruce; Singal, Ashwani K; Liu, Lawrence U; Desnick, Robert J

    2017-08-01

    Autosomal recessive erythropoietic protoporphyria (EPP) and X-linked protoporphyria (XLP) are rare photodermatoses presenting with variable degrees of painful phototoxicity that markedly affects quality of life. The clinical variability, determinants of severity, and genotype/phenotype correlations of these diseases are not well characterized. To describe the baseline clinical characteristics, genotypes, and determinants of disease severity in a large patient cohort with EPP or XLP. A prospective observational study was conducted among patients with confirmed diagnoses of EPP or XLP from November 1, 2010, to December 6, 2015, at 6 academic medical centers of the Porphyrias Consortium of the National Institutes of Health Rare Diseases Clinical Research Network. Detailed medical histories, including history of phototoxicity and treatment, were collected on standardized case report forms. Patients underwent baseline laboratory testing, total erythrocyte protoporphyrin (ePPIX) testing, and molecular genetic testing. Data were entered into a centralized database. Results of biochemical and genetic tests were explored for association with clinical phenotype in patients with EPP or XLP. Of the 226 patients in the study (113 female and 113 male patients; mean [SD] age, 36.7 [17.0] years), 186 (82.3%) had EPP with a FECH (OMIM 612386) mutation and the common low-expression FECH allele IVS3-48T>C, and only 1 patient had 2 FECH mutations. Twenty-two patients had XLP (9.7%; 10 male and 12 female patients), and 9 patients (4.0%) had elevated ePPIX levels and symptoms consistent with protoporphyria but no detectable mutation in the FECH or ALAS2 (OMIM 301300) gene. Samples of DNA could not be obtained from 8 patients. Patients' mean (SD) age at symptom onset was 4.4 (4.4) years. Anemia (107 [47.3%]), history of liver dysfunction (62 [27.4%]), and gallstones (53 [23.5%]) were commonly reported. Higher ePPIX levels were associated with earlier age of symptom onset (median e

  1. Convergent Transcription in the Butyrolactone Regulon in Streptomyces coelicolor Confers a Bistable Genetic Switch for Antibiotic Biosynthesis

    Science.gov (United States)

    Chatterjee, Anushree; Drews, Laurie; Mehra, Sarika; Takano, Eriko; Kaznessis, Yiannis N.; Hu, Wei-Shou

    2011-01-01

    cis-encoded antisense RNAs (cis asRNA) have been reported to participate in gene expression regulation in both eukaryotic and prokaryotic organisms. Its presence in Streptomyces coelicolor has also been reported recently; however, its role has yet to be fully investigated. Using mathematical modeling we explore the role of cis asRNA produced as a result of convergent transcription in scbA-scbR genetic switch. scbA and scbR gene pair, encoding repressor–amplifier proteins respectively, mediates the synthesis of a signaling molecule, the γ-butyrolactone SCB1 and controls the onset of antibiotic production. Our model considers that transcriptional interference caused by convergent transcription of two opposing RNA polymerases results in fatal collision and transcriptional termination, which suppresses transcription efficiency. Additionally, convergent transcription causes sense and antisense interactions between complementary sequences from opposing strands, rendering the full length transcript inaccessible for translation. We evaluated the role of transcriptional interference and the antisense effect conferred by convergent transcription on the behavior of scbA-scbR system. Stability analysis showed that while transcriptional interference affects the system, it is asRNA that confers scbA-scbR system the characteristics of a bistable switch in response to the signaling molecule SCB1. With its critical role of regulating the onset of antibiotic synthesis the bistable behavior offers this two gene system the needed robustness to be a genetic switch. The convergent two gene system with potential of transcriptional interference is a frequent feature in various genomes. The possibility of asRNA regulation in other such gene-pairs is yet to be examined. PMID:21765930

  2. Biochemical and genetical analysis reveal a new clade of biovar 3 Dickeya spp. strains isolated from potato in Europe

    NARCIS (Netherlands)

    Slawiak, M.; Beckhoven, van J.R.C.M.; Speksnijder, A.G.C.L.; Czajkowski, R.L.; Grabe, G.; Wolf, van der J.M.

    2009-01-01

    Sixty-five potato strains of the soft rot-causing plant pathogenic bacterium Dickeya spp., and two strains from hyacinth, were characterised using biochemical assays, REP-PCR genomic finger printing, 16S rDNA and dnaX sequence analysis. These methods were compared with nineteen strains representing

  3. Partial response to biotin therapy in a patient with holocarboxylase synthetase deficiency: clinical, biochemical, and molecular genetic aspects

    NARCIS (Netherlands)

    Santer, R.; Muhle, H.; Suormala, T.; Baumgartner, E. R.; Duran, M.; Yang, X.; Aoki, Y.; Suzuki, Y.; Stephani, U.

    2003-01-01

    We report the clinical course and biochemical findings of a 10-year-old, mentally retarded girl with late-onset holocarboxylase synthetase (HCS, gene symbol HLCS) deficiency and only partial response to biotin. On treatment, even with an unusually high dose of 200mg/day, activities of the

  4. Biosynthesis of the 22nd Genetically Encoded Amino Acid Pyrrolysine: Structure and Reaction Mechanism of PylC at 1.5Å Resolution

    KAUST Repository

    Quitterer, Felix

    2012-12-01

    The second step in the biosynthesis of the 22nd genetically encoded amino acid pyrrolysine (Pyl) is catalyzed by PylC that forms the pseudopeptide l-lysine-Nε-3R-methyl-d-ornithine. Here, we present six crystal structures of the monomeric active ligase in complex with substrates, reaction intermediates, and products including ATP, the non-hydrolyzable ATP analogue 5′-adenylyl-β-γ-imidodiphosphate, ADP, d-ornithine (d-Orn), l-lysine (Lys), phosphorylated d-Orn, l-lysine-Nε-d-ornithine, inorganic phosphate, carbonate, and Mg2 +. The overall structure of PylC reveals similarities to the superfamily of ATP-grasp enzymes; however, there exist unique structural and functional features for a topological control of successive substrate entry and product release. Furthermore, the presented high-resolution structures provide detailed insights into the reaction mechanism of isopeptide bond formation starting with phosphorylation of d-Orn by transfer of a phosphate moiety from activated ATP. The binding of Lys to the enzyme complex is then followed by an SN2 reaction resulting in l-lysine-Nε-d-ornithine and inorganic phosphate. Surprisingly, PylC harbors two adenine nucleotides bound at the active site, what has not been observed in any ATP-grasp protein analyzed to date. Whereas one ATP molecule is involved in catalysis, the second adenine nucleotide functions as a selective anchor for the C- and N-terminus of the Lys substrate and is responsible for protein stability as shown by mutagenesis. © 2012 Elsevier Ltd.

  5. Biochemical features of genetic Creutzfeldt-Jakob disease with valine-to-isoleucine substitution at codon 180 on the prion protein gene.

    Science.gov (United States)

    Ito, Yoko; Sanjo, Nobuo; Hizume, Masaki; Kobayashi, Atsushi; Ohgami, Tetsuya; Satoh, Katsuya; Hamaguchi, Tsuyoshi; Yamada, Masahito; Kitamoto, Tetsuyuki; Mizusawa, Hidehiro; Yokota, Takanori

    2018-02-19

    Valine-to-isoleucine substitution at codon 180 of the prion protein gene is only observed in patients with Creutzfeldt-Jakob disease and accounts for approximately half of all cases of genetic prion disease in Japan. In the present study, we investigated the biochemical characteristics of valine-to-isoleucine substitution at codon 180 in the prion protein gene, using samples obtained from the autopsied brains of seven patients with genetic Creutzfeldt-Jakob disease exhibiting this mutation (diagnoses confirmed via neuropathological examination). Among these patients, we observed an absence of diglycosylated and monoglycosylated forms of PrP res at codon 181. Our findings further indicated that the abnormal prion proteins were composed of at least three components, although smaller carboxyl-terminal fragments were predominant. Western blot analyses revealed large amounts of PrP res in the cerebral neocortices, where neuropathological examination revealed marked spongiosis. Relatively smaller amounts of PrP res were detected in the hippocampus, where milder spongiosis was observed, than in the cerebral neocortex. These findings indicate that abnormal prion proteins in the neocortex are associated with severe toxicity, resulting in severe spongiosis. Our findings further indicate that the valine-to-isoleucine substitution is not a polymorphism, but rather an authentic pathogenic mutation associated with specific biochemical characteristics that differ from those observed in sporadic Creutzfeldt-Jakob disease. Copyright © 2018 Elsevier Inc. All rights reserved.

  6. Genome-wide association mapping and biochemical markers reveal that seed ageing and longevity are intricately affected by genetic background and developmental and environmental conditions in barley.

    Science.gov (United States)

    Nagel, Manuela; Kranner, Ilse; Neumann, Kerstin; Rolletschek, Hardy; Seal, Charlotte E; Colville, Louise; Fernández-Marín, Beatriz; Börner, Andreas

    2015-06-01

    Globally, over 7.4 million accessions of crop seeds are stored in gene banks, and conservation of genotypic variation is pivotal for breeding. We combined genetic and biochemical approaches to obtain a broad overview of factors that influence seed storability and ageing in barley (Hordeum vulgare). Seeds from a germplasm collection of 175 genotypes from four continents grown in field plots with different nutrient supply were subjected to two artificial ageing regimes. Genome-wide association mapping revealed 107 marker trait associations, and hence, genotypic effects on seed ageing. Abiotic and biotic stresses were found to affect seed longevity. To address aspects of abiotic, including oxidative, stress, two major antioxidant groups were analysed. No correlation was found between seed deterioration and the lipid-soluble tocochromanols, nor with oil, starch and protein contents. Conversely, the water-soluble glutathione and related thiols were converted to disulphides, indicating a strong shift towards more oxidizing intracellular conditions, in seeds subjected to long-term dry storage at two temperatures or to two artificial ageing treatments. The data suggest that intracellular pH and (bio)chemical processes leading to seed deterioration were influenced by the type of ageing or storage. Moreover, seed response to ageing or storage treatment appears to be significantly influenced by both maternal environment and genetic background. © 2014 John Wiley & Sons Ltd.

  7. A Metabolic Gene Cluster in the Wheat W1 and the Barley Cer-cqu Loci Determines β-Diketone Biosynthesis and Glaucousness.

    Science.gov (United States)

    Hen-Avivi, Shelly; Savin, Orna; Racovita, Radu C; Lee, Wing-Sham; Adamski, Nikolai M; Malitsky, Sergey; Almekias-Siegl, Efrat; Levy, Matan; Vautrin, Sonia; Bergès, Hélène; Friedlander, Gilgi; Kartvelishvily, Elena; Ben-Zvi, Gil; Alkan, Noam; Uauy, Cristobal; Kanyuka, Kostya; Jetter, Reinhard; Distelfeld, Assaf; Aharoni, Asaph

    2016-06-01

    The glaucous appearance of wheat (Triticum aestivum) and barley (Hordeum vulgare) plants, that is the light bluish-gray look of flag leaf, stem, and spike surfaces, results from deposition of cuticular β-diketone wax on their surfaces; this phenotype is associated with high yield, especially under drought conditions. Despite extensive genetic and biochemical characterization, the molecular genetic basis underlying the biosynthesis of β-diketones remains unclear. Here, we discovered that the wheat W1 locus contains a metabolic gene cluster mediating β-diketone biosynthesis. The cluster comprises genes encoding proteins of several families including type-III polyketide synthases, hydrolases, and cytochrome P450s related to known fatty acid hydroxylases. The cluster region was identified in both genetic and physical maps of glaucous and glossy tetraploid wheat, demonstrating entirely different haplotypes in these accessions. Complementary evidence obtained through gene silencing in planta and heterologous expression in bacteria supports a model for a β-diketone biosynthesis pathway involving members of these three protein families. Mutations in homologous genes were identified in the barley eceriferum mutants defective in β-diketone biosynthesis, demonstrating a gene cluster also in the β-diketone biosynthesis Cer-cqu locus in barley. Hence, our findings open new opportunities to breed major cereal crops for surface features that impact yield and stress response. © 2016 American Society of Plant Biologists. All rights reserved.

  8. Arabinogalactan biosynthesis

    DEFF Research Database (Denmark)

    Poulsen, Christian Peter; Dilokpimol, Adiphol; Geshi, Naomi

    2015-01-01

    Arabinogalactan proteins are abundant cell surface proteoglycans in plants and are implicated to act as developmental markers during plant growth. We previously reported that AtGALT31A, AtGALT29A, and AtGLCAT14A-C, which are involved in the biosynthesis of arabinogalactan proteins, localize......GALT29A. Therefore, the electrostatic status of Y144, which is regulated by an unknown kinase/phosphatase system, may regulate AtGALT29A enzyme activity. Moreover, we have identified additional proteins, apyrase 3 (APY3; At1g14240) and UDPglucuronate epimerases 1 and 6 (GAE1, At4g30440; GAE6, At3g23820...

  9. Biochemical and genetical analysis reveal a new clade of biovar 3 Dickeya spp. strains isolated from potato in Europe

    OpenAIRE

    Slawiak, M.; Beckhoven, van, J.R.C.M.; Speksnijder, A.G.C.L.; Czajkowski, R.L.; Grabe, G.; Wolf, van der, J.M.

    2009-01-01

    Sixty-five potato strains of the soft rot-causing plant pathogenic bacterium Dickeya spp., and two strains from hyacinth, were characterised using biochemical assays, REP-PCR genomic finger printing, 16S rDNA and dnaX sequence analysis. These methods were compared with nineteen strains representing six Dickeya species which included the type strains. A group of twenty-two potato strains isolated between 2005-2007 in the Netherlands, Poland, Finland and Israel were characterised as belonging t...

  10. Genetic and biochemical diversity in the HCV NS5B RNA polymerase in the context of interferon α plus ribavirin therapy.

    Science.gov (United States)

    Cao, F; Donlin, M J; Turner, K; Cheng, X; Tavis, J E

    2011-05-01

    The hepatitis C virus (HCV) RNA polymerase (RdRp) may be a target of the drug ribavirin, and it is an object of drug development. Independent isolates of any HCV subtype differ genetically by approximately 10%, but the effects of this variation on enzymatic activity and drug sensitivity are poorly understood. We proposed that nucleotide use profiles (G/U ratio) among subtype 1b RdRps may reflect their use of ribavirin. Here, we characterized how subtype 1b genetic variation affects RNA polymerase activity and evaluated the G/U ratio as a surrogate for ribavirin use during pegylated interferon α and ribavirin therapy. Genetic and biochemical variation in the RdRp was compared between responders who would be largely sensitive to ribavirin and relapsers who would be mostly resistant. There were no consistent genetic differences between responder and relapser RdRps. RNA polymerization, RNA binding and primer usage varied widely among the RdRps, but these parameters did not differ significantly between the response groups. The G/U ratio among a set of subtype 1a RdRps increased rather than decreased following failed therapy, as would be expected if it reflected ribavirin use. Finally, RdRp activity was significantly associated with ALT levels. These data indicate that (i) current genetic approaches cannot predict RNA polymerase behaviour, (ii) the G/U ratio is not a surrogate for ribavirin use, (iii) RdRp activity may contribute to liver disease by modulating viral mRNA and antigen levels, and (iv) drug candidates should be tested against multiple patient-derived enzymes to ensure widespread efficacy even within a viral subtype. © 2010 Blackwell Publishing Ltd.

  11. Asparagus Spears as a Model to Study Heteroxylan Biosynthesis during Secondary Wall Development.

    Directory of Open Access Journals (Sweden)

    Lili Song

    Full Text Available Garden asparagus (Asparagus officinalis L. is a commercially important crop species utilized for its excellent source of vitamins, minerals and dietary fiber. However, after harvest the tissue hardens and its quality rapidly deteriorates because spear cell walls become rigidified due to lignification and substantial increases in heteroxylan content. This latter observation prompted us to investigate the in vitro xylan xylosyltransferase (XylT activity in asparagus. The current model system for studying heteroxylan biosynthesis, Arabidopsis, whilst a powerful genetic system, displays relatively low xylan XylT activity in in vitro microsomal preparations compared with garden asparagus therefore hampering our ability to study the molecular mechanism(s of heteroxylan assembly. Here, we analyzed physiological and biochemical changes of garden asparagus spears stored at 4 °C after harvest and detected a high level of xylan XylT activity that accounts for this increased heteroxylan. The xylan XylT catalytic activity is at least thirteen-fold higher than that reported for previously published species, including Arabidopsis and grasses. A biochemical assay was optimized and up to seven successive Xyl residues were incorporated to extend the xylotetraose (Xyl4 acceptor backbone. To further elucidate the xylan biosynthesis mechanism, we used RNA-seq to generate an Asparagus reference transcriptome and identified five putative xylan biosynthetic genes (AoIRX9, AoIRX9-L, AoIRX10, AoIRX14_A, AoIRX14_B with AoIRX9 having an expression profile that is distinct from the other genes. We propose that Asparagus provides an ideal biochemical system to investigate the biochemical aspects of heteroxylan biosynthesis and also offers the additional benefit of being able to study the lignification process during plant stem maturation.

  12. Asparagus Spears as a Model to Study Heteroxylan Biosynthesis during Secondary Wall Development.

    Science.gov (United States)

    Song, Lili; Zeng, Wei; Wu, Aimin; Picard, Kelsey; Lampugnani, Edwin R; Cheetamun, Roshan; Beahan, Cherie; Cassin, Andrew; Lonsdale, Andrew; Doblin, Monika S; Bacic, Antony

    2015-01-01

    Garden asparagus (Asparagus officinalis L.) is a commercially important crop species utilized for its excellent source of vitamins, minerals and dietary fiber. However, after harvest the tissue hardens and its quality rapidly deteriorates because spear cell walls become rigidified due to lignification and substantial increases in heteroxylan content. This latter observation prompted us to investigate the in vitro xylan xylosyltransferase (XylT) activity in asparagus. The current model system for studying heteroxylan biosynthesis, Arabidopsis, whilst a powerful genetic system, displays relatively low xylan XylT activity in in vitro microsomal preparations compared with garden asparagus therefore hampering our ability to study the molecular mechanism(s) of heteroxylan assembly. Here, we analyzed physiological and biochemical changes of garden asparagus spears stored at 4 °C after harvest and detected a high level of xylan XylT activity that accounts for this increased heteroxylan. The xylan XylT catalytic activity is at least thirteen-fold higher than that reported for previously published species, including Arabidopsis and grasses. A biochemical assay was optimized and up to seven successive Xyl residues were incorporated to extend the xylotetraose (Xyl4) acceptor backbone. To further elucidate the xylan biosynthesis mechanism, we used RNA-seq to generate an Asparagus reference transcriptome and identified five putative xylan biosynthetic genes (AoIRX9, AoIRX9-L, AoIRX10, AoIRX14_A, AoIRX14_B) with AoIRX9 having an expression profile that is distinct from the other genes. We propose that Asparagus provides an ideal biochemical system to investigate the biochemical aspects of heteroxylan biosynthesis and also offers the additional benefit of being able to study the lignification process during plant stem maturation.

  13. Genetic cytological and biochemical study of a tomato chlorophyll mutant of the xanthic type, obtained by irradiation of the seeds

    International Nuclear Information System (INIS)

    Lefort, M.; Duranton, J.; Galmiche, J.M.; Roux, E.

    1958-01-01

    Irradiation of Lycopersicum aesculantum seeds with increasing doses of X-rays and thermal neutrons leads to the appearance of chlorophyll mutations in the descendants of the irradiated seeds. A genetic study of one of these mutants of the xanthic type showed that it was a recessive mutant with typical mono-genetic separation, while the cytological study demonstrated that the differentiation of the plast stopped at the stage of elementary lamella. Finally it is shown that in the light, the mutation brings about a very large deviation of the carbon metabolism towards the synthesis of amino acids and proteins, at the expense of that of glucosides. (author) [fr

  14. Novel mutations in xanthine dehydrogenase/oxidase cause severe hypouricemia: biochemical and molecular genetic analysis in two Czech families with xanthinuria type I.

    Science.gov (United States)

    Stiburkova, Blanka; Krijt, Jakub; Vyletal, Petr; Bartl, Josef; Gerhatova, Eva; Korinek, Martin; Sebesta, Ivan

    2012-01-18

    The article describes the clinical, biochemical, enzymological and molecular genetics findings in two patients from two families with xanthinuria type I. Biochemical analysis using high performance liquid chromatography, allopurinol loading test and analysis of xanthine oxidase activity in plasma and of uromodulin excretion in urine were performed. Sequencing analysis of the xanthine dehydrogenase gene and the haplotype and statistical analyses of consanguinity were performed. Probands showed extremely low concentrations of uric acid, on seven occasions under the limit of detection. The concentration of uric acid in 38-year-old female was 15 μmol/L in serum and 0.04 mmol/L in urine. Excretion of xanthine in urine was 170 mmol/mol creatinine. The concentration of uric acid in 25-year-old male was 0.03 mmol/L in urine. Excretion of xanthine in urine was 141 mmol/mol creatinine. The allopurinol loading test confirmed xanthinuria type I. The xanthine oxidase activities in patients were 0 and 0.4 pmol/h/mL of plasma. We found three nonsense changes: p.P214QfsX4 and unpublished p.R825X and p.R881X. We found two nonconsanguineous compound heterozygotes with xanthinuria type I caused by three nonsense changes. The methods used did not confirm consanguinity in the probands, thus there might be an unconfirmed biological relationship or mutational hotspot. Copyright © 2011 Elsevier B.V. All rights reserved.

  15. Genetic Diversity and Biochemical Characteristics of Trichosporon asahii Isolated from Clinical Specimens, Houses of Patients with Summer-Type-Hypersensitivity Pneumonitis, and Environmental Materials

    Science.gov (United States)

    Sugita, Takashi; Ichikawa, Tomoe; Matsukura, Manami; Sueda, Mika; Takashima, Masako; Ikeda, Reiko; Nishikawa, Akemi; Shinoda, Takako

    2001-01-01

    Trichosporon asahii, which is distributed in the environment, is the major causative agent of the opportunistic infection trichosporonosis, and it also causes summer-type hypersensitivity pneumonitis (SHP). Random amplification of polymorphic DNA analysis was used to determine the intraspecies diversity of 39 T. asahii isolates from clinical specimens, SHP patients' houses, and environmental materials. The three primers used revealed 46 polymorphic bands. A phenogram was generated by the unweighted pair-group method with arithmetic mean. Clinical isolates formed a cluster, characterized by a 90% matching coefficient, but they did not cluster with strains isolated from SHP patients' houses or environmental sources. In addition, the biochemical characteristics of 86 strains from three sources were examined with 31 compounds using an ID32C kit, and a phenogram was constructed. The phenogram consisted of three major clusters. Cluster I included most of the clinical SHP isolates, and cluster II included most of the environmental isolates. Cluster III contained only one strain. A remarkable difference was found in the abilities of the strains belonging to clusters I and II to utilize six compounds. These results suggest that the genetic diversity and biochemical characteristics of T. asahii seem to be related to the source of the isolate. We also found a specific DNA fragment for the clinical isolates and strains isolated from SHP patients' houses. PMID:11427546

  16. Chronic effects of soft drink consumption on the health state of Wistar rats: A biochemical, genetic and histopathological study

    Science.gov (United States)

    ALKHEDAIDE, ADEL; SOLIMAN, MOHAMED MOHAMED; SALAH-ELDIN, ALAA-ELDIN; ISMAIL, TAMER AHMED; ALSHEHIRI, ZAFER SAAD; ATTIA, HOSSAM FOUAD

    2016-01-01

    The present study was performed to examine the effects of chronic soft drink consumption (SDC) on oxidative stress, biochemical alterations, gene biomarkers and histopathology of bone, liver and kidney. Free drinking water of adult male Wistar rats was substituted with three different soft drinks: Coca-Cola, Pepsi and 7-Up, for three consecutive months. The serum and organs were collected for examining the biochemical parameters associated with bone, liver and kidney functions. Semi-quantitative reverse transcription polymerase chain reaction was used to observe the changes in the expression of genes in the liver and kidney, which are associated with oxidative stress resistance. Histopathological investigations were performed to determine the changes in bone, liver and kidney tissues using hematoxylin and eosin stains. SDC affected liver, kidney and bone function biomarkers. Soft drinks increased oxidative stress, which is represented by an increase in malondialdehyde and a decrease in antioxidant levels. SDC affected serum mineral levels, particularly calcium and phosphorus. Soft drinks downregulated the expression levels of glutathione-S-transferase and super oxide dismutase in the liver compared with that of control rats. Rats administered Coca-Cola exhibited a hepatic decrease in the mRNA expression of α2-macroglobulin compared with rats administered Pepsi and 7-Up. On the other hand, SDC increased the mRNA expression of α1-acid glycoprotein. The present renal studies revealed that Coca-Cola increased the mRNA expression levels of desmin, angiotensinogen and angiotensinogen receptor compared with the other groups, together with mild congestion in renal histopathology. Deleterious histopathological changes were reported predominantly in the bone and liver of the Coca-Cola and Pepsi groups. In conclusion, a very strict caution must be considered with SDC due to the increase in oxidative stress biomarkers and disruption in the expression of certain genes

  17. Control of triacylglycerol biosynthesis in plants

    Energy Technology Data Exchange (ETDEWEB)

    1993-01-31

    Seeds of most species of the Umbelliferae (Apiaciae), Araliaceae, and Garryaceae families are characterized by their high content of the unusual C[sub 18] monounsaturated fatty acid petroselinic acid (18:l[Delta][sup 6cis]). Prior to a recent report of this lab, little was known of the biosynthetic origin of the cis[Delta][sup 6] double bond of petroselinic acid. Such knowledge may be of both biochemical and biotechnological significance. Because petroselinic acid is potentially the product of a novel desaturase, information regarding its synthesis may contribute to an understanding of fatty acid desaturation mechanisms in plants. Through chemical cleavage at its double bond, petroselinic acid can be used as a precursor of lauric acid (12:0), a component of detergents and surfactants, and adipic acid (6:0 dicarboxylic), the monomeric component of nylon 6,6. Therefore, the development of an agronomic source of an oil rich in petroselinic acid is of biotechnological interest. As such, studies of petroselinic acid biosynthesis may provide basic information required for any attempt to genetically engineer the production and accumulation of this fatty acid in an existing oilseed.

  18. Forward genetic screen for auxin-deficient mutants by cytokinin.

    Science.gov (United States)

    Wu, Lei; Luo, Pan; Di, Dong-Wei; Wang, Li; Wang, Ming; Lu, Cheng-Kai; Wei, Shao-Dong; Zhang, Li; Zhang, Tian-Zi; Amakorová, Petra; Strnad, Miroslav; Novák, Ondřej; Guo, Guang-Qin

    2015-07-06

    Identification of mutants with impairments in auxin biosynthesis and dynamics by forward genetic screening is hindered by the complexity, redundancy and necessity of the pathways involved. Furthermore, although a few auxin-deficient mutants have been recently identified by screening for altered responses to shade, ethylene, N-1-naphthylphthalamic acid (NPA) or cytokinin (CK), there is still a lack of robust markers for systematically isolating such mutants. We hypothesized that a potentially suitable phenotypic marker is root curling induced by CK, as observed in the auxin biosynthesis mutant CK-induced root curling 1 / tryptophan aminotransferase of Arabidopsis 1 (ckrc1/taa1). Phenotypic observations, genetic analyses and biochemical complementation tests of Arabidopsis seedlings displaying the trait in large-scale genetic screens showed that it can facilitate isolation of mutants with perturbations in auxin biosynthesis, transport and signaling. However, unlike transport/signaling mutants, the curled (or wavy) root phenotypes of auxin-deficient mutants were significantly induced by CKs and could be rescued by exogenous auxins. Mutants allelic to several known auxin biosynthesis mutants were re-isolated, but several new classes of auxin-deficient mutants were also isolated. The findings show that CK-induced root curling provides an effective marker for discovering genes involved in auxin biosynthesis or homeostasis.

  19. Genetic and biochemical characterization of ISP6, a small mitochondrial outer membrane protein associated with the protein translocation complex.

    OpenAIRE

    Kassenbrock, C K; Cao, W; Douglas, M G

    1993-01-01

    To search genetically for additional components of the protein translocation apparatus of mitochondria, we have used low fidelity PCR mutagenesis to generate temperature-sensitive mutants in the outer membrane translocation pore component ISP42. A high copy number suppressor of temperature-sensitive isp42 has been isolated and sequenced. This novel gene, denoted ISP6, encodes a 61 amino acid integral membrane protein of the mitochondrial outer membrane, which is oriented with its amino-termin...

  20. Genetic and Biochemical Studies on Mannose-Negative Mutants That Are Deficient in Phosphomannose Isomerase in Escherichia coli K-12

    Science.gov (United States)

    Markovitz, Alvin; Sydiskis, Robert J.; Lieberman, Michael M.

    1967-01-01

    Two mannose-negative mutants of Escherichia coli K-12 have been isolated. These mutants are deficient in the ability to synthesize phosphomannose isomerase and capsular polysaccharide when grown on glucose-containing media. Interrupted mating experiments to determine the kinetics of genetic transfer show that the two mannose-negative mutations map together between the histidine and tryptophan regions of the E. coli chromosome. PMID:4862193

  1. Genetic and biochemical characterization of rhizobacterial strains and their potential use in combination with chelants for assisted phytoremediation.

    Science.gov (United States)

    Cicatelli, Angela; Guarino, Francesco; Baldan, Enrico; Castiglione, Stefano

    2017-03-01

    Copper and zinc are essential micronutrients in plants but, at high concentrations, they are toxic. Assisted phytoremediation is an emerging "green" technology that aims to improve the efficiency of tolerant species to remove metals from soils through the use of chelants or microorganisms. Rhizobacteria can promote plant growth and tolerance and also affect the mobility, bioavailability, and complexation of metals. A pot experiment was conducted to evaluate the phytoremediation effectiveness of sunflowers cultivated in a Cu- and Zn-spiked soil, in the presence or absence of bacterial consortium and/or chelants. The consortium was constituted of two Stenotrophomonas maltophilia strains and one of Agrobacterium sp. These strains were previously isolated from the rhizosphere of maize plants cultivated on a metal-polluted soil and here molecularly and biochemically characterized. Results showed that the consortium improved sunflower growth and biomass production on the spiked soils. Sunflowers accumulated large amounts of metals in their roots and leaves; however, neither the bacterial consortium nor the chelants, singularly added to pots, influenced significantly Cu and Zn plant uptake. Furthermore, the consecutive soil amendment with the EDTA and bacterial consortium determined a consistent accumulation of metals in sunflowers, and it might be an alternative strategy to limit the use of EDTA and its associated environmental risks in phytoremediation.

  2. The Impact of Physical Activity and Dietary Measures on the Biochemical and Anthropometric Parameters in Obese Children. Is There Any Genetic Predisposition?

    Science.gov (United States)

    Zlatohlávek, Lukáš; Hubáček, Jaroslav Alois; Vrablík, Michal; Pejšová, Hana; Lánská, Věra; Češka, Richard

    2015-11-01

    The aim of the study was to monitor the importance of laboratory, anthropometric and genetic determination of the presence of risk factors for atherosclerosis, obesity, dyslipidemia and components of the metabolic syndrome in obese children and the response to dietary and regimen interventions in obese children. As a part of the study, 353 paediatric patients (46% boys, 54% girls) with obesity and dyslipidemia, aged 8-16 years, participated in a one-month lifestyle intervention programme. The programme involved a reduction of energy intake and supervised exercise programme consisting of 5 exercise units per day, each 50 minutes long. Standard biochemical methods were applied, including Lp-PLA2, as were anthropometric measurements and genetic analyses. During the reduction programme for the children there was a statistically significant decrease in all anthropometric indicators of bodyweight (pgenetic predisposition for obesity, as well as individuals with a better response to regimen interventions which could, among other things, be determined by the FTO and MC4R genotypes. Copyright© by the National Institute of Public Health, Prague 2015.

  3. Biosynthesis of antibiotic chuangxinmycin from Actinoplanes tsinanensis

    Directory of Open Access Journals (Sweden)

    Yuanyuan Shi

    2018-03-01

    Full Text Available Chuangxinmycin is an antibiotic isolated from Actinoplanes tsinanensis CPCC 200056 in the 1970s with a novel indole-dihydrothiopyran heterocyclic skeleton. Chuangxinmycin showed in vitro antibacterial activity and in vivo efficacy in mouse infection models as well as preliminary clinical trials. But the biosynthetic pathway of chuangxinmycin has been obscure since its discovery. Herein, we report the identification of a stretch of DNA from the genome of A. tsinanensis CPCC 200056 that encodes genes for biosynthesis of chuangxinmycin by bioinformatics analysis. The designated cxn cluster was then confirmed to be responsible for chuangxinmycin biosynthesis by direct cloning and heterologous expressing in Streptomyces coelicolor M1146. The cytochrome P450 CxnD was verified to be involved in the dihydrothiopyran ring closure reaction by the identification of seco-chuangxinmycin in S. coelicolor M1146 harboring the cxn gene cluster with an inactivated cxnD. Based on these results, a plausible biosynthetic pathway for chuangxinmycin biosynthesis was proposed, by hijacking the primary sulfur transfer system for sulfur incorporation. The identification of the biosynthetic gene cluster of chuangxinmycin paves the way for elucidating the detail biochemical machinery for chuangxinmycin biosynthesis, and provides the basis for the generation of novel chuangxinmycin derivatives by means of combinatorial biosynthesis and synthetic biology. KEY WORDS: Chuangxinmycin, Actinoplanes tsinanensis, Biosynthesis gene cluster, Heterologous expression, Cytochrome P450, Seco-chuangxinmycin, C–S bond formation, Sulfur incorporation

  4. Two enzymes involved in biosynthesis of the host-selective phytotoxin HC-toxin

    International Nuclear Information System (INIS)

    Walton, J.D.

    1987-01-01

    Cochliobolus carbonum race 1 produces a cyclic tetrapeptide HC-toxin, which is necessary for its exceptional virulence on certain varieties of maize. Previous genetic analysis of HC-toxin production by the fungus has indicated that a single genetic locus controls HC-toxin production. Enzymes involved in the biosynthesis of HC-toxin have been sought by following the precedents established for the biosynthetic enzymes of cyclic peptide antibiotics. Two enzymatic activities from C. carbonum race 1 were found, a D-alanine- and an L-proline-dependent ATP/PP/sub i/ exchange, which by biochemical and genetic criteria were shown to be involved in the biosynthesis of HC-toxin. These two activities were present in all tested race 1 isolates of C. carbonum, which produce HC-toxin, and in none of the tested race 2 and race 3 isolates, which do not produce the toxin. In a genetic cross between two isolates of C. carbonum differing at the tox locus, all tox + progeny had both activities, and all tox - progeny lacked both activities

  5. Fatty acid biosynthesis in actinomycetes

    Science.gov (United States)

    Gago, Gabriela; Diacovich, Lautaro; Arabolaza, Ana; Tsai, Shiou-Chuan; Gramajo, Hugo

    2011-01-01

    All organisms that produce fatty acids do so via a repeated cycle of reactions. In mammals and other animals, these reactions are catalyzed by a type I fatty acid synthase (FAS), a large multifunctional protein to which the growing chain is covalently attached. In contrast, most bacteria (and plants) contain a type II system in which each reaction is catalyzed by a discrete protein. The pathway of fatty acid biosynthesis in Escherichia coli is well established and has provided a foundation for elucidating the type II FAS pathways in other bacteria (White et al., 2005). However, fatty acid biosynthesis is more diverse in the phylum Actinobacteria: Mycobacterium, possess both FAS systems while Streptomyces species have only the multi-enzyme FAS II system and Corynebacterium species exclusively FAS I. In this review we present an overview of the genome organization, biochemical properties and physiological relevance of the two FAS systems in the three genera of actinomycetes mentioned above. We also address in detail the biochemical and structural properties of the acyl-CoA carboxylases (ACCases) that catalyzes the first committed step of fatty acid synthesis in actinomycetes, and discuss the molecular bases of their substrate specificity and the structure-based identification of new ACCase inhibitors with anti-mycobacterial properties. PMID:21204864

  6. Higher physiopathogenicity by Fasciola gigantica than by the genetically close F. hepatica: experimental long-term follow-up of biochemical markers.

    Science.gov (United States)

    Valero, M Adela; Bargues, M Dolores; Khoubbane, Messaoud; Artigas, Patricio; Quesada, Carla; Berinde, Lavinia; Ubeira, Florencio M; Mezo, Mercedes; Hernandez, Jose L; Agramunt, Veronica H; Mas-Coma, Santiago

    2016-01-01

    Fascioliasis is caused by Fasciola hepatica and F. gigantica. The latter, always considered secondary in human infection, nowadays appears increasingly involved in Africa and Asia. Unfortunately, little is known about its pathogenicity, mainly due to difficulties in assessing the moment a patient first becomes infected and the differential diagnosis with F. hepatica. A long-term, 24-week, experimental study comparing F. hepatica and F. gigantica was made for the first time in the same animal model host, Guirra sheep. Serum biochemical parameters of liver damage, serum electrolytes, protein metabolism, plasma proteins, carbohydrate metabolism, hepatic lipid metabolism and inflammation were analysed on a biweekly basis as morbidity indicators. Serum anti-Fasciola IgG, coproantigen and egg shedding were simultaneously followed up. rDNA and mtDNA sequencing and the morphometric study by computer image analysis system (CIAS) showed that fasciolids used fitted standard species characteristics. Results demonstrated that F. gigantica is more pathogenic, given its bigger size and biomass but not due to genetic differences which are few. Fasciola gigantica shows a delayed development of 1-2 weeks regarding both the biliary phase and the beginning of egg shedding, with respective consequences for biochemical modifications in the acute and chronic periods. The higher F. gigantica pathogenicity contrasts with previous studies which only reflected the faster development of F. hepatica observed in short-term experiments. © The Author 2015. Published by Oxford University Press on behalf of Royal Society of Tropical Medicine and Hygiene. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  7. The first joint congress of the South African Biochemical Society, South African Genetics Society and the South African Society for Microbiology at the University of the Witwatersrand, 29 June-4 July 1986

    International Nuclear Information System (INIS)

    1986-01-01

    The South African Biochemical Society, South African Genetics Society and the South African Society for Microbiology held a joint congress at the University of the Witwatersrand from 29 June - 4 July 1986. The papers delivered cover subjects such as Molecular biology, Genetics, Biochemistry, Medical biochemistry, Physiology, Zoology and Isotope and radiation sciences. Different isotopes are used in labelling studies of enzymes, nutrition, metabolism, viruses, bacteria and other biological assays done in the fields of Biochenmistry, Genetics and Microbiology. This work contains only the abstracts of these papers

  8. A Combined Genetic, Biochemical, and Biophysical Analysis of the A1 Phylloquinone Binding Site of Photosystem I from Green Algae

    Energy Technology Data Exchange (ETDEWEB)

    Kevin E. Redding

    2011-12-17

    This project has resulted in the increase in our understanding of how proteins interact with and influence the properties of bound cofactors. This information is important for several reasons, including providing essential information for the re-engineering of biological molecules, such as proteins, for either improved function or entirely new ones. In particular, we have found that a molecule, such as the phylloquinone used in Photosystem I (PS1), can be made a stronger electron donor by placing it in a hydrophobic environment surrounded by negative charges. In addition, the protein is constrained in its interactions with the phylloqinone, in that it must bind the cofactor tightly, but not in such a way that would stabilize the reduced (negatively-charged) version of the molecule. We have used a combination of molecular genetics, in order to make specific mutations in the region of the phylloquinone, and an advanced form of spectroscopy capable of monitoring the transfer of electrons within PS1 using living cells as the material. This approach turned out to produce a significant savings in time and supplies, as it allowed us to focus quickly on the mutants that produced interesting effects, without having to go through laborious purification of the affected proteins. We followed up selected mutants using other spectroscopic techniques in order to gain more specialized information. In addition to the main project funded by this work, this grant supported several related side-projects that also increased our understanding about related issues.

  9. A Combined Genetic, Biochemical, and Biophysical Analysis of the A1 Phylloquinone Binding Site of Photosystem I from Green Algae

    Energy Technology Data Exchange (ETDEWEB)

    Kevin E. Redding

    2008-05-31

    This project has resulted in the increase in our understanding of how proteins interact with and influence the properties of bound cofactors. This information is important for several reasons, including providing essential information for the re-engineering of biological molecules, such as proteins, for either improved function or entirely new ones. In particular, we have found that a molecule, such as the phylloquinone used in Photosystem I (PS1), can be made a stronger electron donor by placing it in a hydrophobic (greasy) environment surrounded by negative charges. In addition, the protein is constrained in its interactions with the phylloqinone, in that it must bind the cofactor tightly, but not in such a way that would stabilize the reduced (natively-charged) version of the molecule. We have used a combination of molecular genetics, in order to make specific mutations in the region of the phylloquinone, and an advanced form of spectroscopy capable of monitoring the transfer of electrons within PS1 using living cells as the material. This approach turned out to produce a significant savings in time and supplies, as it allowed us to focus quickly on the mutants that produced interesting effects, without having to go through laborious purification of the affected proteins. We followed up selected mutants using other spectroscopic techniques in order to gain more specialized information.

  10. Analysis of Arabidopsis mutants deficient in flavonoid biosynthesis

    International Nuclear Information System (INIS)

    Shirley, B.W.; Kubasek, W.L.; Storz, G.; Bruggemann, E.; Koornneef, M.; Ausubel, F.M.; Goodman, H.M.

    1995-01-01

    Eleven loci that play a role in the synthesis of flavonoids in Arabidopsis are described. Mutations at these loci, collectively named transparent testa (tt), disrupt the synthesis of brown pigments in the seed coat (testa). Several of these loci (tt3, tt4, tt5 and ttg) are also required for the accumulation of purple anthocyanins in leaves and stems and one locus (ttg) plays additional roles in trichome and root hair development. Specific functions were previously assigned to tt1-7 and ttg. Here, the results of additional genetic, biochemical and molecular analyses of these mutants are described. Genetic map positions were determined for tt8, tt9 and tt10. Thin-layer chromatography identified tissue- and locus-specific differences in the flavonols and anthocyanidins synthesized by mutant and wild-type plants. It was found that UV light reveals distinct differences in the floral tissues of tt3, tt4, tt5, tt6 and ttg, even though these tissues are indistinguishable under visible light. Evidence was also uncovered that tt8 and ttg specifically affect dihydroflavonol reductase gene expression. A summary of these and previously published results are incorporated into an overview of the genetics of flavonoid biosynthesis in Arabidopsis

  11. Genetic, serological and biochemical characterization of Leishmania tropica from foci in northern Palestine and discovery of zymodeme MON-307

    Directory of Open Access Journals (Sweden)

    Azmi Kifaya

    2012-06-01

    came. Conclusions The Palestinian strains that were assigned to different genetic groups differed in their MLEE profiles and their EF types. A new zymodeme, zymodeme MON-307 was discovered that seems to be unique to the northern part of the Palestinian West Bank. What seemed to be a straight forward classical situation of L. tropica causing anthroponotic CL in the Jenin District might be a more complex situation, owing to the presence of two separate sub-types of L. tropica that, possibly, indicates two separate transmission cycles involving two separate types of phlebotomine sand fly vector.

  12. Genetic polymorphisms of antioxidant enzymes CAT and SOD affect the outcome of clinical, biochemical, and anthropometric variables in people with obesity under a dietary intervention.

    Science.gov (United States)

    Hernández-Guerrero, César; Parra-Carriedo, Alicia; Ruiz-de-Santiago, Diana; Galicia-Castillo, Oscar; Buenrostro-Jáuregui, Mario; Díaz-Gutiérrez, Carmen

    2018-01-01

    Genetic polymorphisms of antioxidant enzymes CAT, GPX, and SOD are involved in the etiology of obesity and its principal comorbidities. The aim of the present study was to analyze the effect of aforementioned SNPs over the output of several variables in people with obesity after a nutritional intervention. The study included 92 Mexican women, which received a dietary intervention by 3 months. Participants were genotyped and stratified into two groups: (1) carriers; mutated homozygous plus heterozygous (CR) and (2) homozygous wild type (WT). A comparison between CR and WT was done in clinical (CV), biochemical (BV), and anthropometric variables (AV), at the beginning and at the end of the intervention. Participants ( n  = 92) showed statistically significant differences ( p  T GPX1 (rs1050450), - 251A>G SOD1 (rs2070424), and - 262C>T CAT (rs1001179). (B) Only CR showed statistically changes ( p  T CAT (rs7943316) and 47C>T SOD2 (rs4880). The dietary intervention effect was statistically significantly between the polymorphisms of 47C>T SOD2 and BMI, SBP, TBARS, total cholesterol, and C-LCL ( p  T CAT (rs7943316) and SBP, DBP, total cholesterol, and atherogenic index ( p  CAT enzymes.

  13. A Reverse-Genetics Mutational Analysis of the Barley HvDWARF Gene Results in Identification of a Series of Alleles and Mutants with Short Stature of Various Degree and Disturbance in BR Biosynthesis Allowing a New Insight into the Process

    Czech Academy of Sciences Publication Activity Database

    Gruszka, D.; Gorniak, M.; Glodowska, E.; Wierus, E.; Oklešťková, Jana; Janeczko, A.; Maluszynski, M.; Szarejko, I.

    2016-01-01

    Roč. 17, č. 4 (2016), s. 600 E-ISSN 1422-0067 R&D Projects: GA MŠk(CZ) LO1204 Institutional support: RVO:61389030 Keywords : barley * biosynthesis * brassinosteroids Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.226, year: 2016

  14. TarO-specific inhibitors of wall teichoic acid biosynthesis restore β-lactam efficacy against methicillin-resistant staphylococci.

    Science.gov (United States)

    Lee, Sang Ho; Wang, Hao; Labroli, Marc; Koseoglu, Sandra; Zuck, Paul; Mayhood, Todd; Gill, Charles; Mann, Paul; Sher, Xinwei; Ha, Sookhee; Yang, Shu-Wei; Mandal, Mihir; Yang, Christine; Liang, Lianzhu; Tan, Zheng; Tawa, Paul; Hou, Yan; Kuvelkar, Reshma; DeVito, Kristine; Wen, Xiujuan; Xiao, Jing; Batchlett, Michelle; Balibar, Carl J; Liu, Jenny; Xiao, Jianying; Murgolo, Nicholas; Garlisi, Charles G; Sheth, Payal R; Flattery, Amy; Su, Jing; Tan, Christopher; Roemer, Terry

    2016-03-09

    The widespread emergence of methicillin-resistant Staphylococcus aureus (MRSA) has dramatically eroded the efficacy of current β-lactam antibiotics and created an urgent need for new treatment options. We report an S. aureus phenotypic screening strategy involving chemical suppression of the growth inhibitory consequences of depleting late-stage wall teichoic acid biosynthesis. This enabled us to identify early-stage pathway-specific inhibitors of wall teichoic acid biosynthesis predicted to be chemically synergistic with β-lactams. We demonstrated by genetic and biochemical means that each of the new chemical series discovered, herein named tarocin A and tarocin B, inhibited the first step in wall teichoic acid biosynthesis (TarO). Tarocins do not have intrinsic bioactivity but rather demonstrated potent bactericidal synergy in combination with broad-spectrum β-lactam antibiotics against diverse clinical isolates of methicillin-resistant staphylococci as well as robust efficacy in a murine infection model of MRSA. Tarocins and other inhibitors of wall teichoic acid biosynthesis may provide a rational strategy to develop Gram-positive bactericidal β-lactam combination agents active against methicillin-resistant staphylococci. Copyright © 2016, American Association for the Advancement of Science.

  15. A directed-overflow and damage-control N-glycosidase in riboflavin biosynthesis

    Science.gov (United States)

    Frelin, Océane; Huang, Lili; Hasnain, Ghulam; Jeffryes, James G.; Ziemak, Michael J.; Rocca, James R.; Wang, Bing; Rice, Jennifer; Roje, Sanja; Yurgel, Svetlana N.; Gregory, Jesse F.; Edison, Arthur S.; Henry, Christopher S.; deCrécy-Lagard, Valérie; Hanson, Andrew D.

    2015-01-01

    Plants and bacteria synthesize the essential human micronutrient riboflavin (vitamin B2) via the same multistep pathway. The early intermediates of this pathway are notoriously reactive, and may be overproduced in vivo because riboflavin biosynthesis enzymes lack feedback controls. Here we demonstrate disposal of riboflavin intermediates by COG3236 (DUF1768), a protein of previously unknown function that is fused to two different riboflavin pathway enzymes in plants and bacteria (RIBR and RibA, respectively). We present cheminformatic, biochemical, genetic, and genomic evidence to show that: (i) plant and bacterial COG3236 proteins cleave the N-glycosidic bond of the first two intermediates of riboflavin biosynthesis, yielding relatively innocuous products; (ii) certain COG3236 proteins are in a multienzyme riboflavin biosynthesis complex that gives them privileged access to riboflavin intermediates; and (iii) COG3236 action in Arabidopsis thaliana and Escherichia coli helps maintain flavin levels. COG3236 proteins thus illustrate two emerging principles in chemical biology: directed overflow metabolism, in which excess flux is diverted out of a pathway, and the pre-emption of damage from reactive metabolites. PMID:25431972

  16. Evidence for synergistic control of glutamate biosynthesis by glutamate dehydrogenases and glutamate in Bacillus subtilis.

    Science.gov (United States)

    Stannek, Lorena; Thiele, Martin J; Ischebeck, Till; Gunka, Katrin; Hammer, Elke; Völker, Uwe; Commichau, Fabian M

    2015-09-01

    In the Gram-positive bacterium, Bacillus subtilis glutamate is synthesized by the glutamine synthetase and the glutamate synthase (GOGAT). During growth with carbon sources that exert carbon catabolite repression, the rocG glutamate dehydrogenase (GDH) gene is repressed and the transcription factor GltC activates the expression of the GOGAT encoding gltAB genes. In the presence of amino acids of the glutamate family, the GDH RocG is synthesized and the enzyme prevents GltC from binding to DNA. The dual control of glutamate biosynthesis allows the efficient utilization of the available nutrients. Here we provide genetic and biochemical evidence that, like RocG, also the paralogous GDH GudB can inhibit the transcription factor GltC, thereby controlling glutamate biosynthesis. Contradictory previous observations show that high level of GDH activity does not result in permanent inhibition of GltC. By controlling the intracellular levels of glutamate through feeding with exogenous arginine, we observed that the GDH-dependent control of GltC and thus expression of the gltAB genes inversely correlates with the glutamate pool. These results suggest that the B. subtilis GDHs RocG and GudB in fact act as glutamate sensors. In conclusion, the GDH-mediated control of glutamate biosynthesis seems to depend on the intracellular glutamate concentration. © 2015 Society for Applied Microbiology and John Wiley & Sons Ltd.

  17. How Embryophytic is the Biosynthesis of Phenylpropanoids and their Derivatives in Streptophyte Algae?

    Science.gov (United States)

    de Vries, Jan; de Vries, Sophie; Slamovits, Claudio H; Rose, Laura E; Archibald, John M

    2017-05-01

    The origin of land plants from algae is a long-standing question in evolutionary biology. It is becoming increasingly clear that many characters that were once assumed to be 'embryophyte specific' can in fact be found in their closest algal relatives, the streptophyte algae. One such case is the phenylpropanoid pathway. While biochemical data indicate that streptophyte algae harbor lignin-like components, the phenylpropanoid core pathway, which serves as the backbone of lignin biosynthesis, has been proposed to have arisen at the base of the land plants. Here we revisit this hypothesis using a wealth of new sequence data from streptophyte algae. Tracing the biochemical pathway towards lignin biogenesis, we show that most of the genes required for phenylpropanoid synthesis and the precursors for lignin production were already present in streptophyte algae. Nevertheless, phylogenetic analyses and protein structure predictions of one of the key enzyme classes in lignin production, cinnamyl alcohol dehydrogenase (CAD), suggest that CADs of streptophyte algae are more similar to sinapyl alcohol dehydrogenases (SADs). This suggests that the end-products of the pathway leading to lignin biosynthesis in streptophyte algae may facilitate the production of lignin-like compounds and defense molecules. We hypothesize that streptophyte algae already possessed the genetic toolkit from which the capacity to produce lignin later evolved in vascular plants. © The Author 2017. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

  18. Biochemical and Structural Characterization of WlbA from Bordetella pertussis and Chromobacterium violaceum: Enzymes Required for the Biosynthesis of 2,3-Diacetamido-2,3-dideoxy-d-mannuronic Acid

    Energy Technology Data Exchange (ETDEWEB)

    Thoden, James B.; Holden, Hazel M. (UW)

    2011-12-22

    The unusual sugar 2,3-diacetamido-2,3-dideoxy-d-mannuronic acid, or ManNAc3NAcA, has been observed in the lipopolysaccharides of both pathogenic and nonpathogenic Gram-negative bacteria. It is added to the lipopolysaccharides of these organisms by glycosyltransferases that use as substrates UDP-ManNAc3NAcA. Five enzymes are ultimately required for the biosynthesis of UDP-ManNAc3NAcA starting from UDP-N-acetylglucosamine. The second enzyme in the pathway, encoded by the wlba gene and referred to as WlbA, catalyzes the NAD-dependent oxidation of the C-3' hydroxyl group of the UDP-linked sugar. Here we describe a combined structural and functional investigation of the WlbA enzymes from Bordetella pertussis and Chromobacterium violaceum. For this investigation, ternary structures were determined in the presence of NAD(H) and substrate to 2.13 and 1.5 {angstrom} resolution, respectively. Both of the enzymes display octameric quaternary structures with their active sites positioned far apart. The octamers can be envisioned as tetramers of dimers. Kinetic studies demonstrate that the reaction mechanisms for these enzymes are sequential and that they do not require {alpha}-ketoglutarate for activity. These results are in sharp contrast to those recently reported for the WlbA enzymes from Pseudomonas aeruginosa and Thermus thermophilus, which function via ping-pong mechanisms that involve {alpha}-ketoglutarate. Taken together, the results reported here demonstrate that there are two distinct families of WlbA enzymes, which differ with respect to amino acid sequences, quaternary structures, active site architectures, and kinetic mechanisms.

  19. Combinatorial biosynthesis of medicinal plant secondary metabolites

    NARCIS (Netherlands)

    Julsing, Mattijs K.; Koulman, Albert; Woerdenbag, Herman J.; Quax, Wim J.; Kayser, Oliver

    2006-01-01

    Combinatorial biosynthesis is a new tool in the generation of novel natural products and for the production of rare and expensive natural products. The basic concept is combining metabolic pathways in different organisms on a genetic level. As a consequence heterologous organisms provide precursors

  20. Biosynthesis of polyhydroxyalkanotes in wildtype yeasts | Desuoky ...

    African Journals Online (AJOL)

    Biosynthesis of the biodegradable polymers polyhydroxyalkanotes (PHAs) are studied extensively in wild type and genetically modified prokaryotic cells, however the content and structure of PHA in wild type yeasts are not well documented. The purpose of this study was to screen forty yeast isolates collected from different ...

  1. Dissecting complex polyketide biosynthesis

    Directory of Open Access Journals (Sweden)

    Patrick Caffrey

    2012-10-01

    Full Text Available Numerous bioactive natural products are synthesised by modular polyketide synthases. These compounds can be made in high yield by native multienzyme assembly lines. In contrast, formation of analogues by genetically engineered systems is often considerably less efficient. Biochemical studies on intact polyketide synthase proteins have amassed a body of knowledge that is substantial but still incomplete. Recently, the constituent enzymes have been structurally characterised as discrete domains or didomains. These recombinant proteins have been used to reconstitute single extension cycles in vitro. This has given further insights into how the final stereochemistry of chiral centres in polyketides is determined. In addition, this approach has revealed how domains co-operate to ensure efficient transfer of growing intermediates along the assembly line. This work is leading towards more effective re-programming of these enzymes for use in synthesis of new medicinal compounds.

  2. DISSECTING COMPLEX POLYKETIDE BIOSYNTHESIS

    Directory of Open Access Journals (Sweden)

    Patrick Caffrey

    2012-10-01

    Full Text Available Numerous bioactive natural products are synthesised by modular polyketide synthases. These compounds can be made in high yield by native multienzyme assembly lines. However, formation of analogues by genetically engineered systems is often considerably less efficient. Biochemical studies on intact polyketide synthase proteins have amassed a body of knowledge that is substantial but still incomplete. Recently, the constituent enzymes have been structurally characterised as discrete domains or didomains. These recombinant proteins have been used to reconstitute single extension cycles in vitro. This has given further insights into how the final stereochemistry of chiral centres in polyketides is determined. In addition, this approach has revealed how domains co-operate to ensure efficient transfer of growing intermediates along the assembly line. This work is leading towards more effective re-programming of these enzymes for use in synthesis of new medicinal compounds.

  3. Variants in estrogen-biosynthesis genes CYP17 and CYP19 and breast cancer risk: a family-based genetic association study

    International Nuclear Information System (INIS)

    Ahsan, Habibul; Whittemore, Alice S; Chen, Yu; Senie, Ruby T; Hamilton, Steven P; Wang, Qiao; Gurvich, Irina; Santella, Regina M

    2005-01-01

    Case-control studies have reported inconsistent results concerning breast cancer risk and polymorphisms in genes that control endogenous estrogen biosynthesis. We report findings from the first family-based association study examining associations between female breast cancer risk and polymorphisms in two key estrogen-biosynthesis genes CYP17 (T→C promoter polymorphism) and CYP19 (TTTA repeat polymorphism). We conducted the study among 278 nuclear families containing one or more daughters with breast cancer, with a total of 1123 family members (702 with available constitutional DNA and questionnaire data and 421 without them). These nuclear families were selected from breast cancer families participating in the Metropolitan New York Registry, one of the six centers of the National Cancer Institute's Breast Cancer Family Registry. We used likelihood-based statistical methods to examine allelic associations. We found the CYP19 allele with 11 TTTA repeats to be associated with breast cancer risk in these families. We also found that maternal (but not paternal) carrier status of CYP19 alleles with 11 repeats tended to be associated with breast cancer risk in daughters (independently of the daughters' own genotype), suggesting a possible in utero effect of CYP19. We found no association of a woman's breast cancer risk either with her own or with her mother's CYP17 genotype. This family-based study indicates that a woman's personal and maternal carrier status of CYP19 11 TTTA repeat allele might be related to increased breast cancer risk. However, because this is the first study to report an association between CYP19 11 TTTA repeat allele and breast cancer, and because multiple comparisons have been made, the associations should be interpreted with caution and need confirmation in future family-based studies

  4. Inverse problem studies of biochemical systems with structure identification of S-systems by embedding training functions in a genetic algorithm.

    Science.gov (United States)

    Sarode, Ketan Dinkar; Kumar, V Ravi; Kulkarni, B D

    2016-05-01

    An efficient inverse problem approach for parameter estimation, state and structure identification from dynamic data by embedding training functions in a genetic algorithm methodology (ETFGA) is proposed for nonlinear dynamical biosystems using S-system canonical models. Use of multiple shooting and decomposition approach as training functions has been shown for handling of noisy datasets and computational efficiency in studying the inverse problem. The advantages of the methodology are brought out systematically by studying it for three biochemical model systems of interest. By studying a small-scale gene regulatory system described by a S-system model, the first example demonstrates the use of ETFGA for the multifold aims of the inverse problem. The estimation of a large number of parameters with simultaneous state and network identification is shown by training a generalized S-system canonical model with noisy datasets. The results of this study bring out the superior performance of ETFGA on comparison with other metaheuristic approaches. The second example studies the regulation of cAMP oscillations in Dictyostelium cells now assuming limited availability of noisy data. Here, flexibility of the approach to incorporate partial system information in the identification process is shown and its effect on accuracy and predictive ability of the estimated model are studied. The third example studies the phenomenological toy model of the regulation of circadian oscillations in Drosophila that follows rate laws different from S-system power-law. For the limited noisy data, using a priori information about properties of the system, we could estimate an alternate S-system model that showed robust oscillatory behavior with predictive abilities. Copyright © 2016 Elsevier Inc. All rights reserved.

  5. Dithiolopyrrolone Natural Products: Isolation, Synthesis and Biosynthesis

    Science.gov (United States)

    Qin, Zhiwei; Huang, Sheng; Yu, Yi; Deng, Hai

    2013-01-01

    Dithiolopyrrolones are a class of antibiotics that possess the unique pyrrolinonodithiole (4H-[1,2] dithiolo [4,3-b] pyrrol-5-one) skeleton linked to two variable acyl groups. To date, there are approximately 30 naturally occurring dithiolopyrrolone compounds, including holomycin, thiolutin, and aureothricin, and more recently thiomarinols, a unique class of hybrid marine bacterial natural products containing a dithiolopyrrolone framework linked by an amide bridge with an 8-hydroxyoctanoyl chain linked to a monic acid. Generally, dithiolopyrrolone antibiotics have broad-spectrum antibacterial activity against various microorganisms, including Gram-positive and Gram-negative bacteria, and even parasites. Holomycin appeared to be active against rifamycin-resistant bacteria and also inhibit the growth of the clinical pathogen methicillin-resistant Staphylococcus aureus N315. Its mode of action is believed to inhibit RNA synthesis although the exact mechanism has yet to be established in vitro. A recent work demonstrated that the fish pathogen Yersinia ruckeri employs an RNA methyltransferase for self-resistance during the holomycin production. Moreover, some dithiolopyrrolone derivatives have demonstrated promising antitumor activities. The biosynthetic gene clusters of holomycin have recently been identified in S. clavuligerus and characterized biochemically and genetically. The biosynthetic gene cluster of thiomarinol was also identified from the marine bacterium Pseudoalteromonas sp. SANK 73390, which was uniquely encoded by two independent pathways for pseudomonic acid and pyrrothine in a novel plasmid. The aim of this review is to give an overview about the isolations, characterizations, synthesis, biosynthesis, bioactivities and mode of action of this unique family of dithiolopyrrolone natural products, focusing on the period from 1940s until now. PMID:24141227

  6. Growth stage-based modulation in physiological and biochemical attributes of two genetically diverse wheat (Triticum aestivum L.) cultivars grown in salinized hydroponic culture.

    Science.gov (United States)

    Ashraf, Muhammad Arslan; Ashraf, Muhammad

    2016-04-01

    Hydroponic experiment was conducted to appraise variation in the salt tolerance potential of two wheat cultivars (salt tolerant, S-24, and moderately salt sensitive, MH-97) at different growth stages. These two wheat cultivars are not genetically related as evident from randomized polymorphic DNA analysis (random amplified polymorphic DNA (RAPD)) which revealed 28% genetic diversity. Salinity stress caused a marked reduction in grain yield of both wheat cultivars. However, cv. S-24 was superior to cv. MH-97 in maintaining grain yield under saline stress. Furthermore, salinity caused a significant variation in different physiological attributes measured at different growth stages. Salt stress caused considerable reduction in different water relation attributes of wheat plants. A significant reduction in leaf water, osmotic, and turgor potentials was recorded in both wheat cultivars at different growth stages. Maximal reduction in leaf water potential was recorded at the reproductive stage in both wheat cultivars. In contrast, maximal turgor potential was observed at the boot stage. Salt-induced adverse effects of salinity on different water relation attributes were more prominent in cv. MH-97 as compared to those in cv. S-24. Salt stress caused a substantial decrease in glycine betaine and alpha tocopherols. These biochemical attributes exhibited significant salt-induced variation at different growth stages in both wheat cultivars. For example, maximal accumulation of glycine betaine was evident at the early growth stages (vegetative and boot). However, cv. S-24 showed higher accumulation of this organic osmolyte, and this could be the reason for maintenance of higher turgor than that of cv. MH-97 under stress conditions. Salt stress significantly increased the endogenous levels of toxic ions (Na(+) and Cl(-)) and decreased essential cations (K(+) and Ca(2+)) in both wheat cultivars at different growth stages. Furthermore, K(+)/Na(+) and Ca(2+)/Na(+) ratios

  7. Structural Diversity and Biochemical and Microbiological Characteristics of Aflatoxins

    Directory of Open Access Journals (Sweden)

    Ketney Otto

    2014-12-01

    Full Text Available Among all mycotoxins, Aflatoxin B1 (AFB1 is considered to be the most carcinogenic, and it has been classified by the International Agency for Research on Cancer in Group 1 of human carcinogen. It signifies a high hazard because it contaminates a diversity of agricultural products such as nuts and derivatives, peanuts/hazelnuts, grains, seeds, cottonseed, milk, dairy food. In milk AFB1 is metabolized to aflatoxin M (AFM1 which is 4-hydroxy derivative of AFB1, it is formed in the liver and excreted in the milk into the mammary glands of both human and lactating animals which have been fed with AFB1 contaminated diet. After the food contamination, one part of the aflatoxin B1 which was present in the food is eliminated through the milk. At the molecular level aflatoxin biosynthesis involves several levels of transcriptional and post-transcriptional control, so the main stages subsequent biochemical and genetic constituents of aflatoxin biosynthesis have been demonstrated recently. Recent studies over the last few decades have shown that the metabolism of AFB is an essential component of hepatocarcinogenic, however it was shown that AFB1 is metabolized by cytochrome P450 oxidised to intermediates and other metabolites Therefore, the biotransformation process may also lead to the formation of carcinogenic metabolites.

  8. Genetics of trehalose biosynthesis in desert-derived Aureobasidium melanogenum and role of trehalose in the adaptation of the yeast to extreme environments.

    Science.gov (United States)

    Jiang, Hong; Liu, Guang-Lei; Chi, Zhe; Hu, Zhong; Chi, Zhen-Ming

    2018-04-01

    Melanin plays an important role in the stress adaptation of Aureobasidium melanogenum XJ5-1 isolated from the Taklimakan desert. A trehalose-6-phosphate synthase gene (TPS1 gene) was cloned from K5, characterized, and then deleted to determine the role of trehalose in the stress adaptation of the albino mutant K5. No stress response element and heat shock element were found in the promoter of the TPS1 gene. Deletion of the TPS1 gene in the albino mutant rendered a strain DT43 unable to synthesize any trehalose, but DT43 still could grow in glucose, suggesting that its hexokinase was insensitive to inhibition by trehalose-6-phosphate. Overexpression of the TPS1 gene enhanced trehalose biosynthesis in strain ET6. DT43 could not grow at 33 °C, whereas K5, ET6, and XJ5-1 could grow well at this temperature. Compared with K5 and ET6, DT43 was highly sensitive to heat shock treatment, high oxidation, and high desiccation, but all the three strains demonstrated the same sensitivity to UV light and high NaCl concentration. Therefore, trehalose played an important role in the adaptation of K5 to heat shock treatment, high oxidation, and high desiccation.

  9. Genetic analysis of the biosynthesis of non-ribosomal peptide- and polyketide-like antibiotics, iron uptake and biofilm formation by Bacillus subtilis A1/3.

    Science.gov (United States)

    Hofemeister, J; Conrad, B; Adler, B; Hofemeister, B; Feesche, J; Kucheryava, N; Steinborn, G; Franke, P; Grammel, N; Zwintscher, A; Leenders, F; Hitzeroth, G; Vater, J

    2004-11-01

    The Bacillus subtilis strain A1/3 shows exceptionally diverse antibiotic capacities compared to other B. subtilis strains. To analyze this phenomenon, mutants for the putative pantotheinyltransferase gene (pptS), and for several genes involved in non-ribosomal peptide synthesis and polyketide synthesis were constructed and characterized, using bioassays with blood cells, bacterial and fungal cells, and mass spectrometry. Among at least nine distinct bioactive compounds, five antibiotics and one siderophore activity were identified. The anti-fungal and hemolytic activities of strain A1/3 could be eliminated by mutation of the fen and srf genes essential for the synthesis of fengycins and surfactins. Both pptS- and dhb -type mutants were defective in iron uptake, indicating an inability to produce a 2,3-dihydroxybenzoate-type iron siderophore. Transposon mutants in the malonyl CoA transacylase gene resulted in the loss of hemolytic and anti-fungal activities due to the inhibition of bacillomycin L synthesis, and this led to the discovery of bmyLD-LA-LB* genes. In mutants bearing disruption mutations in polyketide (pksM- and/or pksR -like) genes, the biosynthesis of bacillaene and difficidins, respectively, was inactivated and was accompanied by the loss of discrete antibacterial activities. The formation of biofilms (pellicles) was shown to require the production of surfactins, but no other lipopeptides, indicating that surfactins serve specific developmental functions.

  10. Dissecting the Genetic Basis for Seed Coat Mucilage Heteroxylan Biosynthesis inPlantago ovataUsing Gamma Irradiation and Infrared Spectroscopy.

    Science.gov (United States)

    Tucker, Matthew R; Ma, Chao; Phan, Jana; Neumann, Kylie; Shirley, Neil J; Hahn, Michael G; Cozzolino, Daniel; Burton, Rachel A

    2017-01-01

    Seeds from the myxospermous species Plantago ovata release a polysaccharide-rich mucilage upon contact with water. This seed coat derived mucilage is composed predominantly of heteroxylan (HX) and is utilized as a gluten-free dietary fiber supplement to promote human colorectal health. In this study, a gamma-irradiated P. ovata population was generated and screened using histological stains and Fourier Transform Mid Infrared (FTMIR) spectroscopy to identify putative mutants showing defects in seed coat mucilage HX composition and/or structure. FTMIR analysis of dry seed revealed variation in regions of the IR spectra previously linked to xylan structure in Secale cereale (rye). Subsequent absorbance ratio and PCA multivariate analysis identified 22 putative mutant families with differences in the HX IR fingerprint region. Many of these showed distinct changes in the amount and subtle changes in structure of HX after mucilage extrusion, while 20% of the putative HX mutants identified by FTMIR showed no difference in staining patterns of extruded mucilage compared to wild-type. Transcriptional screening analysis of two putative reduced xylan in mucilage ( rxm ) mutants, rxm1 and rxm3 , revealed that changes in HX levels in rxm1 correlate with reduced transcription of known and novel genes associated with xylan synthesis, possibly indicative of specific co-regulatory units within the xylan biosynthetic pathway. These results confirm that FTMIR is a suitable method for identifying putative mutants with altered mucilage HX composition in P. ovata , and therefore forms a resource to identify novel genes involved in xylan biosynthesis.

  11. Biosynthesis of lipophilic compounds in tomato fruit | Angaman ...

    African Journals Online (AJOL)

    A study performed with chromoplasts to know the origin of the precursors for carotenoids biosynthesis using a variety of 14C-labelled precursors showed that the most important incorporation was found in lipids. This study aims to understand the biochemical and metabolic processes operating during tomato fruit ripening.

  12. Study of the genetic diversity of the aflatoxin biosynthesis cluster in Aspergillus section Flavi using insertion/deletion markers in peanut seeds from Georgia, USA.

    Science.gov (United States)

    Faustinelli, Paola C; Palencia, Edwin R; Sobolev, Victor S; Horn, Bruce W; Sheppard, Hank T; Lamb, Marshall C; Wang, Xinye M; Scheffler, Brian E; Martinez Castillo, Jaime; Arias, Renée S

    2017-01-01

    Aflatoxins are among the most powerful carcinogens in nature. The major aflatoxin-producing fungi are Aspergillus flavus and A. parasiticus. Numerous crops, including peanut, are susceptible to aflatoxin contamination by these fungi. There has been an increased use of RNA interference (RNAi) technology to control phytopathogenic fungi in recent years. In order to develop molecular tools targeting specific genes of these fungi for the control of aflatoxins, it is necessary to obtain their genome sequences. Although high-throughput sequencing is readily available, it is still impractical to sequence the genome of every isolate. Thus, in this work, the authors proposed a workflow that allowed prescreening of 238 Aspergillus section Flavi isolates from peanut seeds from Georgia, USA. The aflatoxin biosynthesis cluster (ABC) of the isolates was fingerprinted at 25 InDel (insertion/deletion) loci using capillary electrophoresis. All isolates were tested for aflatoxins using ultra-high-performance liquid chromatography. The neighbor-joining, three-dimension (3D) principal coordinate, and Structure analyses revealed that the Aspergillus isolates sampled consisted of three main groups determined by their capability to produce aflatoxins. Group I comprised 10 non-aflatoxigenic A. flavus; Group II included A. parasiticus; and Group III included mostly aflatoxigenic A. flavus and the three non-aflatoxigenic A. caelatus. Whole genomes of 10 representative isolates from different groups were sequenced. Although InDels in Aspergillus have been used by other research groups, this is the first time that the cluster analysis resulting from fingerprinting was followed by whole-genome sequencing of representative isolates. In our study, cluster analysis of ABC sequences validated the results obtained with fingerprinting. This shows that InDels used here can predict similarities at the genome level. Our results also revealed a relationship between groups and their capability to produce

  13. Lotus tenuis x L. corniculatus interspecific hybridization as a means to breed bloat-safe pastures and gain insight into the genetic control of proanthocyanidin biosynthesis in legumes

    OpenAIRE

    Escaray, Francisco J; Passeri, Valentina; Babuin, Florencia M; Marco, Francisco; Carrasco, Pedro; Damiani, Francesco; Pieckenstain, Fernando L; Paolocci, Francesco; Ruiz, Oscar A

    2014-01-01

    Background Proanthocyanidins (PAs) are secondary metabolites that strongly affect plant quality traits. The concentration and the structure of these metabolites influence the palatability and nutritional value of forage legumes. Hence, modulating PAs in the leaves of forage legumes is of paramount relevance for forage breeders worldwide. The lack of genetic variation in the leaf PA trait within the most important forage species and the difficulties in engineering this pathway via the ectopic ...

  14. Exploring chemical diversity of α-pyrone antibiotics: molecular basis of myxopyronin biosynthesis.

    Science.gov (United States)

    Sucipto, Hilda; Wenzel, Silke C; Müller, Rolf

    2013-09-02

    Myxopyronins and corallopyronins are structurally related α-pyrone antibiotics from myxobacteria. They are thought to represent a highly promising compound class for the development of broad-spectrum antibacterial therapeutic agents, because of their ability to inhibit RNA polymerase through interaction with the "switch region", a recently identified novel drug target. Here we describe the identification and characterization of the myxopyronin biosynthetic pathway from Myxococcus fulvus Mx f50. A detailed comparison with the recently identified corallopyronin biosynthetic pathway revealed the genetic and biochemical basis, thus explaining the observed structural differences between the two natural product families. Directed mutagenesis procedures for M. fulvus Mx f50 were developed to enable functional studies and pathway modifications. Our work provided new insights into myxopyronin biosynthesis and led to the production of a novel and unexpected myxopyronin derivative. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  15. Genes associated with 2-methylisoborneol biosynthesis in cyanobacteria: isolation, characterization, and expression in response to light.

    Directory of Open Access Journals (Sweden)

    Zhongjie Wang

    Full Text Available The volatile microbial metabolite 2-methylisoborneol (2-MIB is a root cause of taste and odor issues in freshwater. Although current evidence suggests that 2-MIB is not toxic, this compound degrades water quality and presents problems for water treatment. To address these issues, cyanobacteria and actinomycetes, the major producers of 2-MIB, have been investigated extensively. In this study, two 2-MIB producing strains, coded as Pseudanabaena sp. and Planktothricoids raciborskii, were used in order to elucidate the genetic background, light regulation, and biochemical mechanisms of 2-MIB biosynthesis in cyanobacteria. Genome walking and PCR methods revealed that two adjacent genes, SAM-dependent methyltransferanse gene and monoterpene cyclase gene, are responsible for GPP methylation and subsequent cyclization to 2-MIB in cyanobacteria. These two genes are located in between two homologous cyclic nucleotide-binding protein genes that may be members of the Crp-Fnr regulator family. Together, this sequence of genes forms a putative operon. The synthesis of 2-MIB is similar in cyanobacteria and actinomycetes. Comparison of the gene arrangement and functional sites between cyanobacteria and other organisms revealed that gene recombination and gene transfer probably occurred during the evolution of 2-MIB-associated genes. All the microorganisms examined have a common origin of 2-MIB biosynthesis capacity, but cyanobacteria represent a unique evolutionary lineage. Gene expression analysis suggested that light is a crucial, but not the only, active regulatory factor for the transcription of 2-MIB synthesis genes. This light-regulated process is immediate and transient. This study is the first to identify the genetic background and evolution of 2-MIB biosynthesis in cyanobacteria, thus enhancing current knowledge on 2-MIB contamination of freshwater.

  16. Genetic and biochemical basis of Gall Midge resistance in some cultivars of Indica Rice. Final report for the period 1 October 1980 - 30 November 1986

    International Nuclear Information System (INIS)

    Reddy, G.M.

    1986-01-01

    The stability of high productivity of modern rice varieties is greatly affected by insect pests. Rice gall midge is a serious insect pest of rice that is prevalent in several south eastern asian countries. Gall midge resistance has been mainly attributed to antibiosis. No progress has so far been made in identifying the exact biochemical nature of resistance. In Indica subspecies the understanding of chemical nature of disease would be helpful in the control of the disease and also in breeding programme aimed at developing resistance varieties. Studies were undertaken to establish the biochemical basis of resistance. Biochemical characterization of resistant and susceptible varieties were carried out. The parameters considered were: total sugar and reducing sugar content, total phenol content, amino acid profile, post infectional changes in sugar and phenol, isozyme studies. 2 figs, 6 tabs

  17. Medicago truncatula CYP716A12 is a multifunctional oxidase involved in the biosynthesis of hemolytic saponins.

    Science.gov (United States)

    Carelli, Maria; Biazzi, Elisa; Panara, Francesco; Tava, Aldo; Scaramelli, Laura; Porceddu, Andrea; Graham, Neil; Odoardi, Miriam; Piano, Efisio; Arcioni, Sergio; May, Sean; Scotti, Carla; Calderini, Ornella

    2011-08-01

    Saponins, a group of glycosidic compounds present in several plant species, have aglycone moieties that are formed using triterpenoid or steroidal skeletons. In spite of their importance as antimicrobial compounds and their possible benefits for human health, knowledge of the genetic control of saponin biosynthesis is still poorly understood. In the Medicago genus, the hemolytic activity of saponins is related to the nature of their aglycone moieties. We have identified a cytochrome P450 gene (CYP716A12) involved in saponin synthesis in Medicago truncatula using a combined genetic and biochemical approach. Genetic loss-of-function analysis and complementation studies showed that CYP716A12 is responsible for an early step in the saponin biosynthetic pathway. Mutants in CYP716A12 were unable to produce hemolytic saponins and only synthetized soyasaponins, and were thus named lacking hemolytic activity (lha). In vitro enzymatic activity assays indicate that CYP716A12 catalyzes the oxidation of β-amyrin and erythrodiol at the C-28 position, yielding oleanolic acid. Transcriptome changes in the lha mutant showed a modulation in the main steps of triterpenic saponin biosynthetic pathway: squalene cyclization, β-amyrin oxidation, and glycosylation. The analysis of CYP716A12 expression in planta is reported together with the sapogenin content in different tissues and stages. This article provides evidence for CYP716A12 being a key gene in hemolytic saponin biosynthesis.

  18. Medicago truncatula CYP716A12 Is a Multifunctional Oxidase Involved in the Biosynthesis of Hemolytic Saponins[W

    Science.gov (United States)

    Carelli, Maria; Biazzi, Elisa; Panara, Francesco; Tava, Aldo; Scaramelli, Laura; Porceddu, Andrea; Graham, Neil; Odoardi, Miriam; Piano, Efisio; Arcioni, Sergio; May, Sean; Scotti, Carla; Calderini, Ornella

    2011-01-01

    Saponins, a group of glycosidic compounds present in several plant species, have aglycone moieties that are formed using triterpenoid or steroidal skeletons. In spite of their importance as antimicrobial compounds and their possible benefits for human health, knowledge of the genetic control of saponin biosynthesis is still poorly understood. In the Medicago genus, the hemolytic activity of saponins is related to the nature of their aglycone moieties. We have identified a cytochrome P450 gene (CYP716A12) involved in saponin synthesis in Medicago truncatula using a combined genetic and biochemical approach. Genetic loss-of-function analysis and complementation studies showed that CYP716A12 is responsible for an early step in the saponin biosynthetic pathway. Mutants in CYP716A12 were unable to produce hemolytic saponins and only synthetized soyasaponins, and were thus named lacking hemolytic activity (lha). In vitro enzymatic activity assays indicate that CYP716A12 catalyzes the oxidation of β-amyrin and erythrodiol at the C-28 position, yielding oleanolic acid. Transcriptome changes in the lha mutant showed a modulation in the main steps of triterpenic saponin biosynthetic pathway: squalene cyclization, β-amyrin oxidation, and glycosylation. The analysis of CYP716A12 expression in planta is reported together with the sapogenin content in different tissues and stages. This article provides evidence for CYP716A12 being a key gene in hemolytic saponin biosynthesis. PMID:21821776

  19. Effects of Azospirillum brasilense with genetically modified auxin biosynthesis gene ipdC upon the diversity of the indigenous microbiota of the wheat rhizosphere.

    Science.gov (United States)

    Baudoin, Ezékiel; Lerner, Anat; Mirza, M Sajjad; El Zemrany, Hamdy; Prigent-Combaret, Claire; Jurkevich, Edouard; Spaepen, Stijn; Vanderleyden, Jos; Nazaret, Sylvie; Okon, Yaacov; Moënne-Loccoz, Yvan

    2010-04-01

    The phytostimulatory properties of Azospirillum inoculants, which entail production of the phytohormone indole-3-acetic acid (IAA), can be enhanced by genetic means. However, it is not known whether this could affect their interactions with indigenous soil microbes. Here, wheat seeds were inoculated with the wild-type strain Azospirillum brasilense Sp245 or one of three genetically modified (GM) derivatives and grown for one month. The GM derivatives contained a plasmid vector harboring the indole-3-pyruvate/phenylpyruvate decarboxylase gene ipdC (IAA production) controlled either by the constitutive promoter PnptII or the root exudate-responsive promoter PsbpA, or by an empty vector (GM control). All inoculants displayed equal rhizosphere population densities. Only inoculation with either ipdC construct increased shoot biomass compared with the non-inoculated control. At one month after inoculation, automated ribosomal intergenic spacer analysis (ARISA) revealed that the effect of the PsbpA construct on bacterial community structure differed from that of the GM control, which was confirmed by 16S rDNA-based denaturing gradient gel electrophoresis (DGGE). The fungal community was sensitive to inoculation with the PsbpA construct and especially the GM control, based on ARISA data. Overall, fungal and bacterial communities displayed distinct responses to inoculation of GM A. brasilense phytostimulators, whose effects could differ from those of the wild-type.

  20. Control of triacylglycerol biosynthesis in plants. Technical progress report

    Energy Technology Data Exchange (ETDEWEB)

    1993-01-31

    Seeds of most species of the Umbelliferae (Apiaciae), Araliaceae, and Garryaceae families are characterized by their high content of the unusual C{sub 18} monounsaturated fatty acid petroselinic acid (18:l{Delta}{sup 6cis}). Prior to a recent report of this lab, little was known of the biosynthetic origin of the cis{Delta}{sup 6} double bond of petroselinic acid. Such knowledge may be of both biochemical and biotechnological significance. Because petroselinic acid is potentially the product of a novel desaturase, information regarding its synthesis may contribute to an understanding of fatty acid desaturation mechanisms in plants. Through chemical cleavage at its double bond, petroselinic acid can be used as a precursor of lauric acid (12:0), a component of detergents and surfactants, and adipic acid (6:0 dicarboxylic), the monomeric component of nylon 6,6. Therefore, the development of an agronomic source of an oil rich in petroselinic acid is of biotechnological interest. As such, studies of petroselinic acid biosynthesis may provide basic information required for any attempt to genetically engineer the production and accumulation of this fatty acid in an existing oilseed.

  1. Characterization of major enzymes and genes involved in flavonoid and proanthocyanidin biosynthesis during fruit development in strawberry (Fragaria x ananassa).

    NARCIS (Netherlands)

    Almeida, J.R.; Amico, d' E.; Preuss, A.; Carbone, F.; Vos, de C.H.

    2007-01-01

    The biosynthesis of flavonoids and proanthocyanidins was studied in cultivated strawberry (Fragaria x ananassa) by combining biochemical and molecular approaches. Chemical analyses showed that ripe strawberries accumulate high amounts of pelargonidin-derived anthocyanins, and a larger pool of

  2. Biosynthesis of tylophora alkaloids

    International Nuclear Information System (INIS)

    Mulchandani, N.B.; Iyer, S.S.; Badheka, L.P.

    1974-01-01

    Using labelled precursors, biosynthesis of the tylophora alkaloids, tylophorine, tylophorinidine and tylophorinide has been investigated in Tylophora asthmatica plants. The radioactive precursors, phenylalanine-2- 14 C, benzoic acid-1- 14 C, benzoic acid-ring 14 C, acetate-2- 14 C, ornithine-5- 14 C, acetate-2- 14 C, ornithine-5- 14 C and cinnamic acid-2- 14 C were administered to the plants individually by wick technique. Tylophorine was isolated in each case and assayed for its radioactivity to find out the incorporation of the label into it. The results indicate that: (1) phenylalanine via cinnamic acid is an important precursor in the biosynthesis of tylophorine (2) orinithine participates in tylophorine biosynthesis via pyrroline and (3) tylophorinidine may be a direct precursor of tylophorine. (M.G.B.)

  3. Aflatoxin biosynthesis: current frontiers.

    Science.gov (United States)

    Roze, Ludmila V; Hong, Sung-Yong; Linz, John E

    2013-01-01

    Aflatoxins are among the principal mycotoxins that contaminate economically important food and feed crops. Aflatoxin B1 is the most potent naturally occurring carcinogen known and is also an immunosuppressant. Occurrence of aflatoxins in crops has vast economic and human health impacts worldwide. Thus, the study of aflatoxin biosynthesis has become a focal point in attempts to reduce human exposure to aflatoxins. This review highlights recent advances in the field of aflatoxin biosynthesis and explores the functional connection between aflatoxin biosynthesis, endomembrane trafficking, and response to oxidative stress. Dissection of the regulatory mechanisms involves a complete comprehension of the aflatoxin biosynthetic process and the dynamic network of transcription factors that orchestrates coordinated expression of the target genes. Despite advancements in the field, development of a safe and effective multifaceted approach to solve the aflatoxin food contamination problem is still required.

  4. Estimating P-coverage of biosynthetic pathways in DNA libraries and screening by genetic selection: biotin biosynthesis in the marine microorganism Chromohalobacter.

    Science.gov (United States)

    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.

  5. Discovery, Biosynthesis and Stress-Related Accumulation of Dolabradiene-Derived Defenses in Maize.

    Science.gov (United States)

    Mafu, Sibongile; Ding, Yezhang; Murphy, Katherine M; Yaacoobi, Omar; Addison, J Bennett; Wang, Qiang; Shen, Zhouxin; Briggs, Steven P; Bohlmann, Jörg; Castro-Falcon, Gabriel; Hughes, Chambers C; Betsiashvili, Mariam; Huffaker, Alisa; Schmelz, Eric A; Zerbe, Philipp

    2018-04-01

    Terpenoids are a major component of maize ( Zea mays ) chemical defenses that mediate responses to herbivores, pathogens, and other environmental challenges. Here, we describe the biosynthesis and elicited production of a class of maize diterpenoids, named dolabralexins. Dolabralexin biosynthesis involves the sequential activity of two diterpene synthases, ENT -COPALYL DIPHOSPHATE SYNTHASE (ZmAN2) and KAURENE SYNTHASE-LIKE4 (ZmKSL4). Together, ZmAN2 and ZmKSL4 form the diterpene hydrocarbon dolabradiene. In addition, we biochemically characterized a cytochrome P450 monooxygenase, ZmCYP71Z16, which catalyzes the oxygenation of dolabradiene to yield the epoxides 15,16-epoxydolabrene (epoxydolabrene) and 3β-hydroxy-15,16-epoxydolabrene (epoxydolabranol). The absence of dolabradiene and epoxydolabranol in Zman2 mutants under elicited conditions confirmed the in vivo biosynthetic requirement of ZmAN2. Combined mass spectrometry and NMR experiments demonstrated that much of the epoxydolabranol is further converted into 3β,15,16-trihydroxydolabrene (trihydroxydolabrene). Metabolite profiling of field-grown maize root tissues indicated that dolabralexin biosynthesis is widespread across common maize cultivars, with trihydroxydolabrene as the predominant diterpenoid. Oxidative stress induced dolabralexin accumulation and transcript expression of ZmAN2 and ZmKSL4 in root tissues, and metabolite and transcript accumulation were up-regulated in response to elicitation with the fungal pathogens Fusarium verticillioides and Fusarium graminearum Consistently, epoxydolabranol significantly inhibited the growth of both pathogens in vitro at 10 µg mL -1 , while trihydroxydolabrene-mediated inhibition was specific to F verticillioides These findings suggest that dolabralexins have defense-related roles in maize stress interactions and expand the known chemical space of diterpenoid defenses as genetic targets for understanding and ultimately improving maize resilience. © 2018

  6. Biosynthesis of secondary metabolites in sugarcane

    Directory of Open Access Journals (Sweden)

    S.C. França

    2001-12-01

    Full Text Available A set of genes related to secondary metabolism was extracted from the sugarcane expressed sequence tag (SUCEST database and was used to investigate both the gene expression pattern of key enzymes regulating the main biosynthetic secondary metabolism pathways and the major classes of metabolites involved in the response of sugarcane to environmental and developmental cues. The SUCEST database was constructed with tissues in different physiological conditions which had been collected under varied situation of environmental stress. This database allows researchers to identify and characterize the expressed genes of a wide range of putative enzymes able to catalyze steps in the phenylpropanoid, isoprenoid and other pathways of the special metabolic mechanisms involved in the response of sugarcane to environmental changes. Our results show that sugarcane cDNAs encoded putative ultra-violet induced sesquiterpene cyclases (SC; chalcone synthase (CHS, the first enzyme in the pathway branch for flavonoid biosynthesis; isoflavone synthase (IFS, involved in plant defense and root nodulation; isoflavone reductase (IFR, a key enzyme in phenylpropanoid phytoalexin biosynthesis; and caffeic acid-O-methyltransferase, a key enzyme in the biosynthesis of lignin cell wall precursors. High levels of CHS transcripts from plantlets infected with Herbaspirillum rubri or Gluconacetobacter diazotroficans suggests that agents of biotic stress can elicit flavonoid biosynthesis in sugarcane. From this data we have predicted the profile of isoprenoid and phenylpropanoid metabolism in sugarcane and pointed the branches of secondary metabolism activated during tissue-specific stages of development and the adaptive response of sugarcane to agents of biotic and abiotic stress, although our assignment of enzyme function should be confirmed by careful biochemical and genetic supporting evidence.Este trabalho foi realizado com os objetivos de gerar uma coleção de genes

  7. Genetics

    Science.gov (United States)

    ... Likelihood of getting certain diseases Mental abilities Natural talents An abnormal trait (anomaly) that is passed down ... one of them has a genetic disorder. Information Human beings have cells with 46 chromosomes . These consist ...

  8. Combinatorial Biosynthesis – Potential and Problems

    Science.gov (United States)

    Floss, Heinz G.

    2007-01-01

    Because of their ecological functions, natural products have been optimized in evolution for interaction with biological systems and receptors. However, they have not necessarily been optimized for other desirable drug properties and thus can often be improved by structural modification. Using examples from the literature, this paper reviews the opportunities for increasing structural diversity among natural products by combinatorial biosynthesis, i.e., the genetic manipulation of biosynthetic pathways. It distinguishes between combinatorial biosynthesis in a narrower sense to generate libraries of modified structures, and metabolic engineering for the targeted formation of specific structural analogs. Some of the problems and limitations encountered with these approaches are also discussed. Work from the author’s laboratory on ansamycin antibiotics is presented which illustrates some of the opportunities and limitations. PMID:16414140

  9. Mathematical model of fructan biosynthesis and polymer length distribution in plants

    DEFF Research Database (Denmark)

    Rasmussen, Gitte Susanne; Thornley, John H. M.; Parsons, Anthony J.

    2013-01-01

    Background and Aims There are many unresolved issues concerning the biochemistry of fructan biosynthesis. The aim of this paper is to address some of these by means of modelling mathematically the biochemical processes.Methods A model has been constructed for the step-by-step synthesis of fructan...... adjustments.Conclusions Mathematical modelling of fructan biosynthesis can provide a useful tool for helping elucidate the details of the biosynthetic processes....

  10. Three Cytochromes P450 are Sufficient to Reconstitute the Biosynthesis of Camalexin, a Major Arabidopsis Antibiotic**

    Science.gov (United States)

    Klein, Andrew P.; Anarat-Cappillino, Gülbenk

    2013-01-01

    Bringing it all together The discovery of the key missing step in the biosynthesis of camalexin, a model antibiotic from Arabidopsis, was uncovered through in vitro biochemical characterization. The coupling of Trp- and Cys-derived fragments via C–S bond formation is promoted by an unusual cytochrome P450 CYP71A13. The first in vitro reconstitution of camalexin biosynthesis from Trp and Cys was achieved using just three P450s. PMID:24151049

  11. Sex differences in prostaglandin biosynthesis in neutrophils during acute inflammation

    OpenAIRE

    Pace, Simona; Rossi, Antonietta; Krauth, Verena; Dehm, Friederike; Troisi, Fabiana; Bilancia, Rossella; Weinigel, Christina; Rummler, Silke; Werz, Oliver; Sautebin, Lidia

    2017-01-01

    The severity and course of inflammatory processes differ between women and men, but the biochemical mechanisms underlying these sex differences are elusive. Prostaglandins (PG) and leukotrienes (LT) are lipid mediators linked to inflammation. We demonstrated superior LT biosynthesis in human neutrophils and monocytes, and in mouse macrophages from females, and we confirmed these sex differences in vivo where female mice produced more LTs during zymosan-induced peritonitis versus males. Here, ...

  12. Genetics

    International Nuclear Information System (INIS)

    Hubitschek, H.E.

    1975-01-01

    Progress is reported on the following research projects: genetic effects of high LET radiations; genetic regulation, alteration, and repair; chromosome replication and the division cycle of Escherichia coli; effects of radioisotope decay in the DNA of microorganisms; initiation and termination of DNA replication in Bacillus subtilis; mutagenesis in mouse myeloma cells; lethal and mutagenic effects of near-uv radiation; effect of 8-methoxypsoralen on photodynamic lethality and mutagenicity in Escherichia coli; DNA repair of the lethal effects of far-uv; and near uv irradiation of bacterial cells

  13. Genetic background of Porphyromonas gingivalis capsule biosynthesis

    NARCIS (Netherlands)

    Brunner, J.

    2011-01-01

    Paradontitis is een inflammatoire aandoening van het weefsel rond de tanden. Het ontstaat door een bacteriële infectie van het tandvlees waarna door de daaropvolgende ontstekingsreactie uiteindelijk het bot rond de tanden wordt aangetast. Dit kan zelfs leiden tot tanduitval. Paradontitis is een

  14. Genetics

    DEFF Research Database (Denmark)

    Christensen, Kaare; McGue, Matt

    2016-01-01

    The sequenced genomes of individuals aged ≥80 years, who were highly educated, self-referred volunteers and with no self-reported chronic diseases were compared to young controls. In these data, healthy ageing is a distinct phenotype from exceptional longevity and genetic factors that protect...

  15. A Novel TetR Family Transcriptional Regulator, CalR3, Negatively Controls Calcimycin Biosynthesis in Streptomyces chartreusis NRRL 3882

    Directory of Open Access Journals (Sweden)

    Lixia Gou

    2017-11-01

    Full Text Available Calcimycin is a unique ionophoric antibiotic that is widely used in biochemical and pharmaceutical applications, but the genetic basis underlying the regulatory mechanisms of calcimycin biosynthesis are unclear. Here, we identified the calR3 gene, which encodes a novel TetR family transcriptional regulator and exerts a negative effect on calcimycin biosynthesis. Disruption of calR3 in Streptomyces chartreusis NRRL 3882 led to significantly increased calcimycin and its intermediate cezomycin. Gene expression analysis showed that the transcription of calR3 and its adjacent calT gene were dramatically enhanced (30- and 171-fold, respectively in GLX26 (ΔcalR3 mutants compared with the wild-type strains. Two CalR3-binding sites within the bidirectional calR3-calT promoter region were identified using a DNase I footprinting assay, indicating that CalR3 directly repressed the transcription of its own gene and the calT gene. In vitro electrophoretic mobility shift assays suggested that both calcimycin and cezomycin can act as CalR3 ligands to induce CalR3 to dissociate from its binding sites. These findings indicate negative feedback for the regulation of CalR3 in calcimycin biosynthesis and suggest that calcimycin production can be improved by manipulating its biosynthetic machinery.

  16. A Novel TetR Family Transcriptional Regulator, CalR3, Negatively Controls Calcimycin Biosynthesis inStreptomyces chartreusisNRRL 3882.

    Science.gov (United States)

    Gou, Lixia; Han, Tiesheng; Wang, Xiaoxia; Ge, Jingxuan; Liu, Wenxiu; Hu, Fen; Wang, Zhijun

    2017-01-01

    Calcimycin is a unique ionophoric antibiotic that is widely used in biochemical and pharmaceutical applications, but the genetic basis underlying the regulatory mechanisms of calcimycin biosynthesis are unclear. Here, we identified the calR3 gene, which encodes a novel TetR family transcriptional regulator and exerts a negative effect on calcimycin biosynthesis. Disruption of calR3 in Streptomyces chartreusis NRRL 3882 led to significantly increased calcimycin and its intermediate cezomycin. Gene expression analysis showed that the transcription of calR3 and its adjacent calT gene were dramatically enhanced (30- and 171-fold, respectively) in GLX26 (Δ calR3 ) mutants compared with the wild-type strains. Two CalR3-binding sites within the bidirectional calR3-calT promoter region were identified using a DNase I footprinting assay, indicating that CalR3 directly repressed the transcription of its own gene and the calT gene. In vitro electrophoretic mobility shift assays suggested that both calcimycin and cezomycin can act as CalR3 ligands to induce CalR3 to dissociate from its binding sites. These findings indicate negative feedback for the regulation of CalR3 in calcimycin biosynthesis and suggest that calcimycin production can be improved by manipulating its biosynthetic machinery.

  17. Discovery of new regulatory genes of lipopeptide biosynthesis in Pseudomonas fluorescens

    NARCIS (Netherlands)

    Song, C.; Aundy, K.; Mortel, van de J.E.; Raaijmakers, J.M.

    2014-01-01

    Pseudomonas fluorescens SS101 produces the cyclic lipopeptide massetolide with diverse functions in antimicrobial activity, motility, and biofilm formation. To understand how massetolide biosynthesis is genetically regulated in SS101, c. 8000 random plasposon mutants were screened for reduced or

  18. Effect of low temperature on highly unsaturated fatty acid biosynthesis in activated sludge.

    Science.gov (United States)

    He, Su; Ding, Li-Li; Xu, Ke; Geng, Jin-Ju; Ren, Hong-Qiang

    2016-07-01

    Low temperature is a limiting factor for the microbial activity of activated sludge for sewage treatment plant in winter. Highly unsaturated fatty acid (UFA) biosynthesis, phospholipid fatty acid (PLFA) constituents and microbial structure in activated sludge at low temperature were investigated. Over 12 gigabases of metagenomic sequence data were generated with the Illumina HiSeq 2000 platform. The result showed 43.11% of phospholipid fatty acid (PLFA) in the activated sludge participated in UFA biosynthesis, and γ-Linolenic could be converted to Arachidonic acid at low temperature. The highly UFA biosynthesis in activated sludge was n-6 highly UFA biosynthesis, rather than n-3 highly UFA biosynthesis. The microbial community structures of activated sludge were analyzed by PLFA and high-throughput sequencing (HiSeq) simultaneously. Acidovorax, Pseudomonas, Flavobacterium and Polaromonas occupied higher percentage at 5°C, and genetic changes of highly UFA biosynthesis derived from microbial community structures change. Copyright © 2016. Published by Elsevier Ltd.

  19. Biosynthesis of the red antibiotic, prodigiosin, in Serratia

    DEFF Research Database (Denmark)

    Williamson, Neil R; Simonsen, Henrik Toft; Ahmed, Raef A A

    2005-01-01

    from Serratia sp. ATCC 39006. The biosynthetic intermediates accumulating in each mutant have been analysed by LC-MS, cross-feeding and genetic complementation studies. Based on these results we assign specific roles in the biosynthesis of MBC to the following Pig proteins: PigI, PigG, PigA, PigJ, Pig...

  20. Expression profiles of genes involved in tanshinone biosynthesis of ...

    Indian Academy of Sciences (India)

    Expression profiles of genes involved in tanshinone biosynthesis of two. Salvia miltiorrhiza genotypes with different tanshinone contents. Zhenqiao Song, Jianhua Wang and Xingfeng Li. J. Genet. 95, 433–439. Table 1. S. miltiorrhiza genes and primer pairs used for qRT-PCR. Gene. GenBank accession. Primer name.

  1. Predictive hypotheses are ineffectual in resolving complex biochemical systems.

    Science.gov (United States)

    Fry, Michael

    2018-03-20

    Scientific hypotheses may either predict particular unknown facts or accommodate previously-known data. Although affirmed predictions are intuitively more rewarding than accommodations of established facts, opinions divide whether predictive hypotheses are also epistemically superior to accommodation hypotheses. This paper examines the contribution of predictive hypotheses to discoveries of several bio-molecular systems. Having all the necessary elements of the system known beforehand, an abstract predictive hypothesis of semiconservative mode of DNA replication was successfully affirmed. However, in defining the genetic code whose biochemical basis was unclear, hypotheses were only partially effective and supplementary experimentation was required for its conclusive definition. Markedly, hypotheses were entirely inept in predicting workings of complex systems that included unknown elements. Thus, hypotheses did not predict the existence and function of mRNA, the multiple unidentified components of the protein biosynthesis machinery, or the manifold unknown constituents of the ubiquitin-proteasome system of protein breakdown. Consequently, because of their inability to envision unknown entities, predictive hypotheses did not contribute to the elucidation of cation theories remained the sole instrument to explain complex bio-molecular systems, the philosophical question of alleged advantage of predictive over accommodative hypotheses became inconsequential.

  2. Developing New Antibiotics with Combinatorial Biosynthesis

    Science.gov (United States)

    Pohl, Nicola L.

    2000-11-01

    Polyketide synthases (PKSs), a class of enzymes found in soil bacteria that produce antibiotics such as erythromycin, string together acetate units using basic organic reactions. The manipulation of the sequence of these reactions at the genetic level has resulted in an alteration of the corresponding chemical structure of the antibiotic produced by the bacteria. This process, called combinatorial biosynthesis, allows the generation of many presently unknown complex structures that can be tested for antibacterial activity, thereby contributing to the race against antibiotic-resistant infectious bacteria.

  3. Final Report on Regulation of Guaiacyl and Syringyl Monolignol Biosynthesis

    Energy Technology Data Exchange (ETDEWEB)

    Vincent L. Chiang

    2006-03-09

    The focus of this research is to understand syringyl monolignol biosynthesis that leads to the formation of syringyl lignin, a type of lignin that can be easily removed during biomass conversion. We have achieved the three originally proposed goals for this project. (1) SAD and CAD genes (enzyme catalytic and kinetic properties) and their functional relevance to CAld5H/AldOMT pathway, (2) spatiotemporal expression patterns of Cald5H, AldOMT, SAD and CAD genes, and (3) functions of CAld5H, AldOMT, and SAD genes in vivo using transgenic aspen. Furthermore, we also found that microRNA might be involved in the upstream regulatory network of lignin biosynthesis and wood formation. The achievements are as below. (1) Based on biochemical and molecular studies, we discovered a novel syringyl-specific alcohol dehydrogenase (SAD) involved in monolignol biosynthesis in angiosperm trees. Through CAld5H/OMT/SAD mediation, syringyl monolignol biosynthesis branches out from guaiacyl pathway at coniferaldehyde; (2) The function of CAld5H gene in this syringyl monolignol biosynthesis pathway also was confirmed in vivo in transgenic Populus; (3) The proposed major monolignol biosynthesis pathways were further supported by the involving biochemical functions of CCR based on a detailed kinetic study; (4) Gene promoter activity analysis also supported the cell-type specific expression of SAD and CAD genes in xylem tissue, consistent with the cell-specific locations of SAD and CAD proteins and with the proposed pathways; (5) We have developed a novel small interfering RNA (siRNA)-mediated stable gene-silencing system in transgenic plants; (6) Using the siRNA and P. trichocarpa transformation/regeneration systems we are currently producing transgenic P. trichocarpa to investigate the interactive functions of CAD and SAD in regulating guaiacyl and syringyl lignin biosynthesis; (7) We have cloned for the first time from a tree species, P. trichocarpa, small regulatory RNAs termed micro

  4. [Optimization of oxytetracycline biosynthesis].

    Science.gov (United States)

    Maksimova, E A; Falkov, N N; Izmaĭlov, N N; Romanchuk, N N

    1988-06-01

    It was shown that rising of temperature up to 30 degrees C at the stage of the oxytetracycline-producing organism growth promoted acceleration of the culture growth rate and increasing of the antibiotic concentration by the 114th hour of the biosynthetic process. For the apparatus used in the study optimal aeration and agitation conditions were developed. To provide optimal parameters during biosynthesis of oxytetracycline, it was recommended to use the aeration rate of 1 v/v.min and the specific mechanical power for mixing of not less than 1 kW/m3.

  5. Biosynthesis of Rishirilide B

    Directory of Open Access Journals (Sweden)

    Philipp Schwarzer

    2018-03-01

    Full Text Available Rishirilide B was isolated from Streptomyces rishiriensis and Streptomyces bottropensis on the basis of its inhibitory activity towards alpha-2-macroglobulin. The biosynthesis of rishirilide B was investigated by feeding experiments with different 13C labelled precursors using the heterologous host Streptomyces albus J1074::cos4 containing a cosmid encoding of the gene cluster responsible for rishirilide B production. NMR spectroscopic analysis of labelled compounds demonstrate that the tricyclic backbone of rishirilide B is a polyketide synthesized from nine acetate units. One of the acetate units is decarboxylated to give a methyl group. The origin of the starter unit was determined to be isobutyrate.

  6. The effect of Deficient Muscarinic Signaling on Commonly Reported Biochemical Effects in Schizophrenia and Convergence with Genetic Susceptibility Loci in explaining symptom dimensions of psychosis.

    Directory of Open Access Journals (Sweden)

    Costa eVakalopoulos

    2014-12-01

    Full Text Available With the advent of DSM 5 criticism has generally centred on a lack of biological validity of the diagnostic criteria. Part of the problem in describing a nosology of psychosis is the tacit assumption of multiple genetic causes each with an incremental loading on the clinical picture that fails to differentiate a clear underlying pathophysiology of high impact. The aim of this paper is to consolidate a primary theory of deficient muscarinic signalling underlying key clinical features of schizophrenia and its regulation by several important genetic associations including neuregulin, DISC and dysbindin. Secondary reductions in markers for GABAergic function and changes in the levels of interneuron calcium binding proteins parvalbumin and calbindin can be attributed to dysfunctional muscarinic transduction. A parallel association exists for cytokine production. The convergent pathway hypothesis is likewise used to model dopaminergic and glutamatergic theories of schizophrenia. The negative symptom dimension is correlated with dysfunction of Akt and ERK transduction, a major point of convergence. The present paradigm predicts the importance of a recent finding of a deletion in a copy number variant of PLCB1 and its potential use if replicated, as one of the first testable biological markers differentiating schizophrenia from bipolar disorder and further subtyping of schizophrenia into deficit and non-deficit. Potential limitations of PLCB1 as a prospective marker are also discussed.

  7. Biosynthesis of nanoparticles using microbes- a review.

    Science.gov (United States)

    Hulkoti, Nasreen I; Taranath, T C

    2014-09-01

    The biosynthesis of nanoparticles by microorganism is a green and eco-friendly technology. This review focuses on the use of consortium of diverse microorganisms belonging to both prokaryotes and eukaryotes for the synthesis of metallic nanoparticles viz. silver, gold, platinum, zirconium, palladium, iron, cadmium and metal oxides such as titanium oxide, zinc oxide, etc. These microorganisms include bacteria, actinomycetes, fungi and algae. The synthesis of nanoparticles may be intracellular or extracellular. The several workers have reported that NADH dependent nitrate reductase enzyme plays a vital role in the conversion of metallic ions to nanoparticles. The FTIR study reveals that diverse biomolecules viz. carboxyl group, primary and secondary amines, amide I, II, and III bands etc serve as a tool for bioreduction and capping agents there by offering stability to particles by preventing agglomeration and growth. The size and shape of the nanoparticles vary with the organism employed and conditions employed during the synthesis which included pH, temperature and substrate concentration. The microorganisms provide diverse environment for biosynthesis of nanoparticles. These particles are safe and eco-friendly with a lot of applications in medicine, agriculture, cosmetic industry, drug delivery and biochemical sensors. The challenges for redressal include optimal production and minimal time to obtain desired size and shape, to enhance the stability of nanoparticles and optimization of specific microorganisms for specific application. Copyright © 2014 Elsevier B.V. All rights reserved.

  8. (+)-Germacrene A Biosynthesis

    Science.gov (United States)

    de Kraker, Jan-Willem; Franssen, Maurice C.R.; de Groot, Aede; König, Wilfried A.; Bouwmeester, Harro J.

    1998-01-01

    The leaves and especially the roots of chicory (Cichorium intybus L.) contain high concentrations of bitter sesquiterpene lactones such as the guianolides lactupicrin, lactucin, and 8-deoxylactucin. Eudesmanolides and germacranolides are present in smaller amounts. Their postulated biosynthesis through the mevalonate-farnesyl diphosphate-germacradiene pathway has now been confirmed by the isolation of a (+)-germacrene A synthase from chicory roots. This sesquiterpene cyclase was purified 200-fold using a combination of anion-exchange and dye-ligand chromatography. It has a Km value of 6.6 μm, an estimated molecular mass of 54 kD, and a (broad) pH optimum around 6.7. Germacrene A, the enzymatic product, proved to be much more stable than reported in literature. Its heat-induced Cope rearrangement into (−)-β-elemene was utilized to determine its absolute configuration on an enantioselective gas chromatography column. To our knowledge, until now in sesquiterpene biosynthesis, germacrene A has only been reported as an (postulated) enzyme-bound intermediate, which, instead of being released, is subjected to additional cyclization(s) by the same enzyme that generated it from farnesyl diphosphate. However, in chicory germacrene A is released from the sesquiterpene cyclase. Apparently, subsequent oxidations and/or glucosylation of the germacrane skeleton, together with a germacrene cyclase, determine whether guaiane- or eudesmane-type sesquiterpene lactones are produced. PMID:9701594

  9. Identification and functional analysis of gene cluster involvement in biosynthesis of the cyclic lipopeptide antibiotic pelgipeptin produced by Paenibacillus elgii

    Directory of Open Access Journals (Sweden)

    Qian Chao-Dong

    2012-09-01

    Full Text Available Abstract Background Pelgipeptin, a potent antibacterial and antifungal agent, is a non-ribosomally synthesised lipopeptide antibiotic. This compound consists of a β-hydroxy fatty acid and nine amino acids. To date, there is no information about its biosynthetic pathway. Results A potential pelgipeptin synthetase gene cluster (plp was identified from Paenibacillus elgii B69 through genome analysis. The gene cluster spans 40.8 kb with eight open reading frames. Among the genes in this cluster, three large genes, plpD, plpE, and plpF, were shown to encode non-ribosomal peptide synthetases (NRPSs, with one, seven, and one module(s, respectively. Bioinformatic analysis of the substrate specificity of all nine adenylation domains indicated that the sequence of the NRPS modules is well collinear with the order of amino acids in pelgipeptin. Additional biochemical analysis of four recombinant adenylation domains (PlpD A1, PlpE A1, PlpE A3, and PlpF A1 provided further evidence that the plp gene cluster involved in pelgipeptin biosynthesis. Conclusions In this study, a gene cluster (plp responsible for the biosynthesis of pelgipeptin was identified from the genome sequence of Paenibacillus elgii B69. The identification of the plp gene cluster provides an opportunity to develop novel lipopeptide antibiotics by genetic engineering.

  10. Improved phytoaccumulation of cadmium by genetically modified tobacco plants (Nicotiana tabacum L.). Physiological and biochemical response of the transformants to cadmium toxicity

    International Nuclear Information System (INIS)

    Gorinova, N.; Nedkovska, M.; Todorovska, E.; Simova-Stoilova, L.; Stoyanova, Z.; Georgieva, K.; Demirevska-Kepova, K.; Atanassov, A.; Herzig, R.

    2007-01-01

    The response of tobacco plants (Nicotiana tabacum L.)-non-transformed and transformed with a metallothionein gene MThis from Silene vulgaris L. - to increase cadmium supply in the nutrient solution was compared. The transgenic plants accumulated significantly more Cd both in the roots and the leaves. Visual toxicity symptoms and disturbance in water balance were correlated with Cd tissue content. Treatment with 300 μM CdCl 2 resulted in inhibition of photosynthesis and mobilization of the ascorbate-glutathione cycle. Treatment with 500 μM CdCl 2 led to irreversible damage of photosynthesis and oxidative stress. An appearance of a new peroxidase isoform and changes in the leaf polypeptide pattern were observed at the highest Cd concentration. The level of non-protein thiols gradually increased following the Cd treatment both in transgenic and non-transformed plants. - Genetic transformation of Nicotiana tabacum L. by metallothionein gene improved phytoaccumulation of cadmium

  11. Increased Insulin following an Oral Glucose Load, Genetic Variation near the Melatonin Receptor MTNR1B, but No Biochemical Evidence of Endothelial Dysfunction in Young Asian Men and Women.

    Directory of Open Access Journals (Sweden)

    Maria A Matuszek

    Full Text Available To identify biochemical and genetic variation relating to increased risk of developing type 2 diabetes mellitus and cardiovascular disease in young, lean male and female adults of different ethnicities.Fasting blood and urine and non-fasting blood following oral glucose intake were analysed in 90 Caucasians, South Asians and South East/East Asians.There were no differences in age, birthweight, blood pressure, body mass index, percent body fat, total energy, percentage of macronutrient intake, microalbumin, leptin, cortisol, adrenocorticotropic hormone, nitric oxide metabolites, C-reactive protein, homocysteine, tumor necrosis factor-α, interleukin-6, von Willebrand factor, vascular cell adhesion molecule-1, plasminogen activator inhibitor-1, and tissue plasminogen activator. Fasting total cholesterol (P = .000, triglycerides (P = .050, low density lipoprotein (P = .009 and non-fasting blood glucose (15 min (P = .024 were elevated in South Asians compared with Caucasians, but there was no significant difference in glucose area under curve (AUC. Non-fasting insulin in South Asians (15-120 min, in South East/East Asians (60-120 min, and insulin AUC in South Asians and South East/East Asians, were elevated compared with Caucasians (P≤0.006. The molar ratio of C-peptide AUC/Insulin AUC (P = .045 and adiponectin (P = .037 were lower in South Asians compared with Caucasians. A significant difference in allele frequency distributions in Caucasians and South Asians was found for rs2166706 (P = 0.022 and rs10830963 (P = 0.009, which are both near the melatonin receptor MTNR1B.Elevated non-fasting insulin exists in young South Asians of normal fasting glucose and insulin. Hepatic clearance of insulin may be reduced in South Asians. No current biochemical evidence exists of endothelial dysfunction at this stage of development. MTNR1B signalling may be a useful therapeutic target in Asian populations in the prevention of type 2 diabetes mellitus.

  12. Genetic and biochemical characterisation of CTX-M-37 extended-spectrum β-lactamase from an Enterobacter cloacae clinical isolate from Mongolia.

    Science.gov (United States)

    Lee, Kyungwon; Yong, Dongeun; Jeong, Seok Hoon; Tulgaa, Khosbayar; Docquier, Jean-Denis; Rossolini, Gian Maria; Chong, Yunsop

    2017-09-01

    The aims of this study were to determine the resistance level of a bla CTX-M-37 -carrying Enterobacter cloacae isolate from Mongolia, to analyse kinetic parameters of the purified enzyme and to compare the genetic environment of the gene. Minimum inhibitory concentrations (MICs) were determined using the Clinical and Laboratory Standards Institute (CLSI) agar dilution method. Purified CTX-M-37 enzyme was used to determined kinetic parameters. The genetic environment of the blaCTX-M-37 gene in E. cloacae was compared with a Kluyvera cryocrescens isolate. The E. cloacae isolate showed relatively low-level resistance to cefotaxime (MIC=16mg/L) compared with a CTX-M-3-producing strain (MIC=256mg/L), and CTX-M-37 had a lower k cat /K m value for cefotaxime (2.0μM -1 s -1 ) compared with CTX-M-3 (3.5μM -1 s -1 ), possibly due to Asn114Asp substitution. The bla CTX-M-37 gene in the E. cloacae isolate was carried on a conjugative plasmid and was associated with an ISEcp1 element containing the -35 and -10 putative promoter sequences TTGAAA and TACAAT, respectively, unlike in the K. cryocrescens isolate. The CTX-M-37-producing E. cloacae isolate showed relatively low-level resistance to cefotaxime and the purified enzyme had lower kinetic parameters as the result of Asn114Asp substitution. Presence of an ISEcp1 element and putative promoters upstream of the bla CTX-M-37 gene in E. cloacae, but not in the K. cryocrescens isolate, indicated their roles in mobilisation and expression of the gene. Copyright © 2017 International Society for Chemotherapy of Infection and Cancer. Published by Elsevier Ltd. All rights reserved.

  13. Phosphoribosyl Diphosphate (PRPP): Biosynthesis, Enzymology, Utilization, and Metabolic Significance

    DEFF Research Database (Denmark)

    Hove-Jensen, Bjarne; Andersen, Kasper R; Kilstrup, Mogens

    2017-01-01

    . PRPP is utilized in the biosynthesis of purine and pyrimidine nucleotides, the amino acids histidine and tryptophan, the cofactors NAD and tetrahydromethanopterin, arabinosyl monophosphodecaprenol, and certain aminoglycoside antibiotics. The participation of PRPP in each of these metabolic pathways...... analysis. PRPP, furthermore, is an effector molecule of purine and pyrimidine nucleotide biosynthesis, either by binding to PurR or PyrR regulatory proteins or as an allosteric activator of carbamoylphosphate synthetase. Genetic analyses have disclosed a number of mutants altered in the PRPP synthase...

  14. Serum Biochemical Phenotypes in the Domestic Dog.

    Directory of Open Access Journals (Sweden)

    Yu-Mei Chang

    Full Text Available The serum or plasma biochemical profile is essential in the diagnosis and monitoring of systemic disease in veterinary medicine, but current reference intervals typically take no account of breed-specific differences. Breed-specific hematological phenotypes have been documented in the domestic dog, but little has been published on serum biochemical phenotypes in this species. Serum biochemical profiles of dogs in which all measurements fell within the existing reference intervals were retrieved from a large veterinary database. Serum biochemical profiles from 3045 dogs were retrieved, of which 1495 had an accompanying normal glucose concentration. Sixty pure breeds plus a mixed breed control group were represented by at least 10 individuals. All analytes, except for sodium, chloride and glucose, showed variation with age. Total protein, globulin, potassium, chloride, creatinine, cholesterol, total bilirubin, ALT, CK, amylase, and lipase varied between sexes. Neutering status significantly impacted all analytes except albumin, sodium, calcium, urea, and glucose. Principal component analysis of serum biochemical data revealed 36 pure breeds with distinctive phenotypes. Furthermore, comparative analysis identified 23 breeds with significant differences from the mixed breed group in all biochemical analytes except urea and glucose. Eighteen breeds were identified by both principal component and comparative analysis. Tentative reference intervals were generated for breeds with a distinctive phenotype identified by comparative analysis and represented by at least 120 individuals. This is the first large-scale analysis of breed-specific serum biochemical phenotypes in the domestic dog and highlights potential genetic components of biochemical traits in this species.

  15. Biochemical and Genetic Characterization of Coagulin, a New Antilisterial Bacteriocin in the Pediocin Family of Bacteriocins, Produced by Bacillus coagulans I4

    Science.gov (United States)

    Le Marrec, Claire; Hyronimus, Bertrand; Bressollier, Philippe; Verneuil, Bernard; Urdaci, Maria C.

    2000-01-01

    A plasmid-linked antimicrobial peptide, named coagulin, produced by Bacillus coagulans I4 has recently been reported (B. Hyronimus, C. Le Marrec and M. C. Urdaci, J. Appl. Microbiol. 85:42–50, 1998). In the present study, the complete, unambiguous primary amino acid sequence of the peptide was obtained by a combination of both N-terminal sequencing of purified peptide and the complete sequence deduced from the structural gene harbored by plasmid I4. Data revealed that this peptide of 44 residues has an amino acid sequence similar to that described for pediocins AcH and PA-1, produced by different Pediococcus acidilactici strains and 100% identical. Coagulin and pediocin differed only by a single amino acid at their C terminus. Analysis of the genetic determinants revealed the presence, on the pI4 DNA, of the entire 3.5-kb operon of four genes described for pediocin AcH and PA-1 production. No extended homology was observed between pSMB74 from P. acidilactici and pI4 when analyzing the regions upstream and downstream of the operon. An oppositely oriented gene immediately dowstream of the bacteriocin operon specifies a 474-amino-acid protein which shows homology to Mob-Pre (plasmid recombination enzyme) proteins encoded by several small plasmids extracted from gram-positive bacteria. This is the first report of a pediocin-like peptide appearing naturally in a non-lactic acid bacterium genus. PMID:11097892

  16. Improvement of Neutral Lipid and Polyunsaturated Fatty Acid Biosynthesis by Overexpressing a Type 2 Diacylglycerol Acyltransferase in Marine Diatom Phaeodactylum tricornutum

    Directory of Open Access Journals (Sweden)

    Ying-Fang Niu

    2013-11-01

    Full Text Available Microalgae have been emerging as an important source for the production of bioactive compounds. Marine diatoms can store high amounts of lipid and grow quite quickly. However, the genetic and biochemical characteristics of fatty acid biosynthesis in diatoms remain unclear. Glycerophospholipids are integral as structural and functional components of cellular membranes, as well as precursors of various lipid mediators. In addition, diacylglycerol acyltransferase (DGAT is a key enzyme that catalyzes the last step of triacylglyceride (TAG biosynthesis. However, a comprehensive sequence-structure and functional analysis of DGAT in diatoms is lacking. In this study, an isoform of diacylglycerol acyltransferase type 2 of the marine diatom Phaeodactylum tricornutum was characterized. Surprisingly, DGAT2 overexpression in P. tricornutum stimulated more oil bodies, and the neutral lipid content increased by 35%. The fatty acid composition showed a significant increase in the proportion of polyunsaturated fatty acids; in particular, EPA was increased by 76.2%. Moreover, the growth rate of transgenic microalgae remained similar, thereby maintaining a high biomass. Our results suggest that increased DGAT2 expression could alter fatty acid profile in the diatom, and the results thus represent a valuable strategy for polyunsaturated fatty acid production by genetic manipulation.

  17. Chloroplast genes in Chlamydomonas affecting organelle ribosomes. Genetic and biochemical analysis of analysis of antibiotic-resistant mutants at several gene loci.

    Science.gov (United States)

    Conde, M F; Boynton, J E; Gillham, N W; Harris, E H; Tingle, C L; Wang, W L

    1975-10-03

    Six chloroplast gene mutants of Chlamydomonas reinhardtii resistant to spectinomycin, erythromycin, or streptomycin have been assessed for antibiotic resistance of their chloroplast ribosomes. Four of these mutations clearly confer high levels of antibiotic resistance on the chloroplast ribosomes both in vivo. Although one mutant resistant to streptomycin and one resistant to spectinomycin have chloroplast ribosomes as sensitive to antibiotics as those of wild type in vivo, these mutations can be shown to alter the wildtype sensitivity of chloroplast ribosomes in polynucleotide-directed amino acid incorporation in vitro. Genetic analysis of these six chloroplast mutants and three similar mutants (Sager, 1972), two of which have been shown to affect chloroplast ribosomes (Mets and Bogorad, 1972; Schlanger and Sager, 1974), indicates that in Chlamydomonas at least three chloroplast gene loci can affect streptomycin resistance of chloroplast ribosomes and that two can affect erythromycin resistance. The three spectinomycin-resistant mutants examined appear to be alleles at a single chloroplast gene locus, but may represent mutations at two different sites within the same gene. Unlike wild type, the streptomycin and spectinomycin resistant mutants which have chloroplast ribosomes sensitive to antibiotics in vivo, grow well in the presence of antibiotic by respiring exogenously supplied acetate as a carbon source, and have normal levels of cytochrome oxidase activity and cyanide-sensitive respiration. We conclude that mitochondrial protein synthesis in these mutants is resistant to these antibiotics, whereas in wild type it is sensitive. To explain the behavior of these two chloroplast gene mutants as well as other one-step mutants which are resistant both photosynthetically and when respiring acetate in the dark, we have postulated that a mutation in a single chloroplast gene may result in alteration of both chloroplast and mitochondrial ribosomes. Mitochondrial

  18. Capsular glucan and intracellular glycogen of Mycobacterium tuberculosis: biosynthesis and impact on the persistence in mice

    DEFF Research Database (Denmark)

    Sambou, Tounkang; Dinadayala, Premkumar; Stadthagen, Gustavo

    2008-01-01

    of these bacteria, orthologues of the glg genes involved in the biosynthesis of glycogen in Escherichia coli were identified in M. tuberculosis H37Rv and inactivated by allelic replacement. Biochemical analyses of the mutants and complemented strains indicated that the synthesis of glucan and glycogen involves...

  19. Genetic control of cuticular wax compounds in Eucalyptus globulus.

    Science.gov (United States)

    Gosney, Benjamin J; Potts, Brad M; O'Reilly-Wapstra, Julianne M; Vaillancourt, René E; Fitzgerald, Hugh; Davies, Noel W; Freeman, Jules S

    2016-01-01

    Plant cuticular wax compounds perform functions that are essential for the survival of terrestrial plants. Despite their importance, the genetic control of these compounds is poorly understood outside of model taxa. Here we investigate the genetic basis of variation in cuticular compounds in Eucalyptus globulus using quantitative genetic and quantitative trait loci (QTL) analyses. Quantitative genetic analysis was conducted using 246 open-pollinated progeny from 13 native sub-races throughout the geographic range. QTL analysis was conducted using 112 clonally replicated progeny from an outcross F2 population. Nine compounds exhibited significant genetic variation among sub-races with three exhibiting signals of diversifying selection. Fifty-two QTL were found with co-location of QTL for related compounds commonly observed. Notable among these was the QTL for five wax esters, which co-located with a gene from the KCS family, previously implicated in the biosynthesis of cuticular waxes in Arabidopsis. In combination, the QTL and quantitative genetic analyses suggest the variation and differentiation in cuticular wax compounds within E. globulus has a complex genetic origin. Sub-races exhibited independent latitudinal and longitudinal differentiation in cuticular wax compounds, likely reflecting processes such as historic gene flow and diversifying selection acting upon genes that have diverse functions in distinct biochemical pathways. © 2015 University of Tasmania New Phytologist © 2015 New Phytologist Trust.

  20. Zincophorin – biosynthesis in Streptomyces griseus and antibiotic properties

    Directory of Open Access Journals (Sweden)

    Walther, Elisabeth

    2016-11-01

    Full Text Available Zincophorin is a polyketide antibiotic that possesses potent activity against Gram-positive bacteria, including human pathogens. While a number of total syntheses of this highly functionalized natural product were reported since its initial discovery, the genetic basis for the biosynthesis of zincophorin has remained unclear. In this study, the co-linearity inherent to polyketide pathways was used to identify the zincophorin biosynthesis gene cluster in the genome of the natural producer HKI 0741. Interestingly, the same locus is fully conserved in the streptomycin-producing actinomycete IFO 13350, suggesting that the latter bacterium is also capable of zincophorin biosynthesis. Biological profiling of zincophorin revealed a dose-dependent inhibition of the Gram-positive bacterium . The antibacterial effect, however, is accompanied by cytotoxicity. Antibiotic and cytotoxic activities were completely abolished upon esterification of the carboxylic acid group in zincophorin.

  1. RESULTS OF THE SELECTION OF BREEDING SAMPLES OF CARROT BASED ON BIOCHEMICAL COMPOSITION

    OpenAIRE

    V. K. Cherkasova; O. N. Shabetya

    2014-01-01

    12 samples of carrot were analyzed for biochemical components in roots. 5 genotypes with high content of vitamin C, β-carotene, and total sugar were selected as genetic sources of high biochemical components.

  2. Molecular analysis of "de novo" purine biosynthesis in solanaceous species and in Arabidopsis thaliana

    DEFF Research Database (Denmark)

    van der Graaff, Eric; Hooykaas, Paul; Lein, Wolfgang

    2004-01-01

    Purine nucleotides are essential components to sustain plant growth and development. In plants they are either synthesized "de novo" during the process of purine biosynthesis or are recycled from purine bases and purine nucleosides throughout the salvage pathway. Comparison between animals......, microorganisms and Arabidopsis, the first plant species with a completely sequenced genome, shows that plants principally use the same biochemical steps to synthesize purine nucleotides and possess all the essential genes and enzymes. Here we report on the cloning and molecular analysis of the complete purine...... biosynthesis pathway in plants, and the in planta functional analysis of PRPP (5-phosphoribosyl-1-pyrophoshate) amidotransferase (ATase), catalyzing the first committed step of the "de novo" purine biosynthesis. The cloning of the genes involved in the purine biosynthesis pathway was attained by a screening...

  3. Biosynthesis and chemical synthesis of presilphiperfolanol natural products.

    Science.gov (United States)

    Hong, Allen Y; Stoltz, Brian M

    2014-05-19

    Presilphiperfolanols constitute a family of biosynthetically important sesquiterpenes which can rearrange to diverse sesquiterpenoid skeletons. While the origin of these natural products can be traced to simple linear terpene precursors, the details of the enzymatic cyclization mechanism that forms the stereochemically dense tricyclic skeleton has required extensive biochemical, computational, and synthetic investigation. Parallel efforts to prepare the unique and intriguing structures of these compounds by total synthesis have also inspired novel strategies, thus resulting in four synthetic approaches and two completed syntheses. While the biosynthesis and chemical synthesis studies performed to date have provided much insight into the role and properties of these molecules, emerging questions regarding the biosynthesis of newer members of the family and subtle details of rearrangement mechanisms have yet to be explored. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  4. Raman spectroscopic biochemical mapping of tissues

    Science.gov (United States)

    Stone, Nicholas; Hart Prieto, Maria C.; Kendall, Catherine A.; Shetty, Geeta; Barr, Hugh

    2006-02-01

    Advances in technologies have brought us closer to routine spectroscopic diagnosis of early malignant disease. However, there is still a poor understanding of the carcinogenesis process. For example it is not known whether many cancers follow a logical sequence from dysplasia, to carcinoma in situ, to invasion. Biochemical tissue changes, triggered by genetic mutations, precede morphological and structural changes. These can be probed using Raman or FTIR microspectroscopy and the spectra analysed for biochemical constituents. Local microscopic distribution of various constituents can then be visualised. Raman mapping has been performed on a number of tissues including oesophagus, breast, bladder and prostate. The biochemical constituents have been calculated at each point using basis spectra and least squares analysis. The residual of the least squares fit indicates any unfit spectral components. The biochemical distribution will be compared with the defined histopathological boundaries. The distribution of nucleic acids, glycogen, actin, collagen I, III, IV, lipids and others appear to follow expected patterns.

  5. Jasmonate mediates salt-induced nicotine biosynthesis in tobacco (Nicotiana tabacum L.

    Directory of Open Access Journals (Sweden)

    Xiaodong Chen

    2016-04-01

    Full Text Available Jasmonate (JA, as an important signal, plays a key role in multiple processes of plant growth, development and stress response. Nicotine and related pyridine alkaloids in tobacco (Nicotiana tabacum L. are essential secondary metabolites. Whether environmental factors control nicotine biosynthesis and the underlying mechanism remains previously unreported. Here, we applied physiological and biochemical approaches to investigate how salt stress affects nicotine biosynthesis in tobacco. We found that salt stress induced the biosynthesis of JA, which subsequently triggered the activation of JA-responsive gene expression and, ultimately, nicotine synthesis. Bioinformatics analysis revealed the existence of many NtMYC2a-recognized G-box motifs in the promoter regions of NtLOX, NtAOS, NtAOC and NtOPR genes. Applying exogenous JA increased nicotine content, while suppressing JA biosynthesis reduced nicotine biosynthesis. Salt treatment could not efficiently induce nicotine biosynthesis in transgenic anti-COI1 tobacco plants. These results demonstrate that JA acts as the essential signal which triggers nicotine biosynthesis in tobacco after salt stress.

  6. [Advances and prospects of taxol biosynthesis by endophytic fungi].

    Science.gov (United States)

    Zhao, Kai; Yu, Lu; Jin, Yuyan; Ma, Xueling; Liu, Dan; Wang, Xiaohua; Wang, Xin

    2016-08-25

    Taxol is one of the most important chemotherapeutic drugs against cancer. Taxol has been mainly extracted from the bark of yews for a long time. However, methods for the extraction of taxol from the bark of Taxus species were inefficient and environmentally costly. As a result of the high ecological toll exacted on trees with the potential for Pacific yew extinction, investigators began to look for other methods of taxol production. Recently, increasing efforts have been made to develop alternative means of taxol production, such as using complete chemical synthesis, semi-synthesis, Taxus spp. plant cell culture and microbe fermentation. Using microbe fermentation in the production of taxol would be a very prospective method for obtaining a large amount of taxol. Therefore, it is necessary to understand the molecular basis and genetic regulation mechanisms of taxol biosynthesis by endophytic fungi, which may be helpful to construct the genetic engineering strain with high taxol output. In this paper, the taxol biosynthesis pathway from Taxus cells and the advantages of taxol biosynthesis by endophytic fungi were discussed. The study on the isolation and biodiversity of taxol-producing endophytic fungi and the taxol biosynthesis related genes are also discussed.

  7. Carotenoid Biosynthesis in Fusarium

    Directory of Open Access Journals (Sweden)

    Javier Avalos

    2017-07-01

    Full Text Available Many fungi of the genus Fusarium stand out for the complexity of their secondary metabolism. Individual species may differ in their metabolic capacities, but they usually share the ability to synthesize carotenoids, a family of hydrophobic terpenoid pigments widely distributed in nature. Early studies on carotenoid biosynthesis in Fusarium aquaeductuum have been recently extended in Fusarium fujikuroi and Fusarium oxysporum, well-known biotechnological and phytopathogenic models, respectively. The major Fusarium carotenoid is neurosporaxanthin, a carboxylic xanthophyll synthesized from geranylgeranyl pyrophosphate through the activity of four enzymes, encoded by the genes carRA, carB, carT and carD. These fungi produce also minor amounts of β-carotene, which may be cleaved by the CarX oxygenase to produce retinal, the rhodopsin’s chromophore. The genes needed to produce retinal are organized in a gene cluster with a rhodopsin gene, while other carotenoid genes are not linked. In the investigated Fusarium species, the synthesis of carotenoids is induced by light through the transcriptional induction of the structural genes. In some species, deep-pigmented mutants with up-regulated expression of these genes are affected in the regulatory gene carS. The molecular mechanisms underlying the control by light and by the CarS protein are currently under investigation.

  8. Biosynthesis of Tetrahydroisoquinoline Antibiotics.

    Science.gov (United States)

    Tang, Gong-Li; Tang, Man-Cheng; Song, Li-Qiang; Zhang, Yue

    2016-01-01

    The tetrahydroisoquinoline (THIQ) alkaloids are naturally occurring antibiotics isolated from a variety of microorganisms and marine invertebrates. This family of natural products exhibit broad spectrum antimicrobial and strong antitumor activities, and the potency of clinical application has been validated by the marketing of ecteinascidin 743 (ET-743) as anticancer drug. In the past 20 years, the biosynthetic gene cluster of six THIQ antibiotics has been characterized including saframycin Mx1 from Myxococcus xanthus, safracin-B from Pseudomonas fluorescens, saframycin A, naphthyridinomycin, and quinocarcin from Streptomyces, as well as ET-743 from Ecteinascidia turbinata. This review gives a brief summary of the current status in understanding the molecular logic for the biosynthesis of these natural products, which provides new insights on the biosynthetic machinery involved in the nonribosomal peptide synthetase system. The proposal of the THIQ biosynthetic pathway not only shows nature's route to generate such complex molecules, but also set the stage to develop a different process for production of ET-743 by synthetic biology.

  9. Stereoselectivity in Polyphenol Biosynthesis

    Science.gov (United States)

    Lewis, Norman G.; Davin, Laurence B.

    1992-01-01

    Stereoselectivity plays an important role in the late stages of phenyl-propanoid metabolism, affording lignins, lignans, and neolignans. Stereoselectivity is manifested during monolignol (glucoside) synthesis, e.g., where the geometry (E or Z) of the pendant double bond affects the specificity of UDPG:coniferyl alcohol glucosyltransferases in different species. Such findings are viewed to have important ramifications in monolignol transport and storage processes, with roles for both E- and Z-monolignols and their glucosides in lignin/lignan biosynthesis being envisaged. Stereoselectivity is also of great importance in enantiose-lective enzymatic processes affording optically active lignans. Thus, cell-free extracts from Forsythia species were demonstrated to synthesize the enantiomerically pure lignans, (-)-secoisolariciresinol, and (-)-pinoresinol, when NAD(P)H, H2O2 and E-coniferyl alcohol were added. Progress toward elucidating the enzymatic steps involved in such highly stereoselective processes is discussed. Also described are preliminary studies aimed at developing methodologies to determine the subcellular location of late-stage phenylpropanoid metabolites (e.g., coniferyl alcohol) and key enzymes thereof, in intact tissue or cells. This knowledge is essential if questions regarding lignin and lignan tissue specificity and regulation of these processes are to be deciphered.

  10. Glycolipid biosynthesis in cyanobacteria

    International Nuclear Information System (INIS)

    Van Dusen, W.J.; Jaworski, J.G.

    1987-01-01

    The biosynthesis of monogalactosyldiacyl-glycerol (MGDG) was studied in five different cyanobacteria. Previous work has shown Anabaena variabilis to synthesize both MGDG and monoglucosyl-diacylglycerol (MG1cDG) with MG1cDG being the precursor of MGDG. They have examined four other cyanobacteria to determine if a similar relationship exists. The cyanobacteria studied were Anabaena variabilis, Chlorogloeopsis sp., Schizothrix calcicola, Anacystis nidulans, and Anacystis marina. Each were grown in liquid culture and lipids were labeled with 14 C]CO 2 for 20 min., 1.0 hr, 1.0 hr + 10 hr chase. Glycolipids were analyzed by initial separation of MGDG and MG1cDG by TLC followed by further analysis by HPLC. Complete separation of molecular species was obtained isocratically on an ODS column. All of the cyanobacteria labeled 16-C and 18-C fatty acids except for A. marina which labeled only 14-C and 16-C fatty acids. Desaturation of the fatty acids could be observed in the 1.0 hr and chase experiments. All were capable of labeling both MG1cDG and MGDG with the precursor-product relationship being observed. There does not appear to be a direct relationship between the epimerization of the sugar moiety and fatty acid desaturation

  11. Biosynthesis of silver nanoparticles.

    Science.gov (United States)

    Poulose, Subin; Panda, Tapobrata; Nair, Praseetha P; Théodore, Thomas

    2014-02-01

    Metal nanoparticles have unique optical, electronic, and catalytic properties. There exist well-defined physical and chemical processes for their preparation. Those processes often yield small quantities of nanoparticles having undesired morphology, and involve high temperatures for the reaction and the use of hazardous chemicals. Relatively, the older technique of bioremediation of metals uses either microorganisms or their components for the production of nanoparticles. The nanoparticles obtained from bacteria, fungi, algae, plants and their components, etc. appear environment-friendly, as toxic chemicals are not used in the processes. In addition to this, the formation of nanoparticles takes place at almost normal temperature and pressure. Control of the shape and size of the nanoparticles is possible by appropriate selection of the pH and temperature. Three important steps are the bioconversion of Ag+ ions, conversion of desired crystals to nanoparticles, and nanoparticle stability. Generally, nanoparticles are characterized by the UV-visible spectroscopy and use of the electron microscope. Silver nanoparticles are used as antimicrobial agents and they possess antifungal, anti-inflammatory, and anti-angiogenic properties. This review highlights the biosynthesis of silver nanoparticles by various organisms, possible mechanisms of their synthesis, their characterization, and applications of silver nanoparticles.

  12. The Biosynthesis of Capuramycin-type Antibiotics

    Science.gov (United States)

    Cai, Wenlong; Goswami, Anwesha; Yang, Zhaoyong; Liu, Xiaodong; Green, Keith D.; Barnard-Britson, Sandra; Baba, Satoshi; Funabashi, Masanori; Nonaka, Koichi; Sunkara, Manjula; Morris, Andrew J.; Spork, Anatol P.; Ducho, Christian; Garneau-Tsodikova, Sylvie; Thorson, Jon S.; Van Lanen, Steven G.

    2015-01-01

    A-500359s, A-503083s, and A-102395 are capuramycin-type nucleoside antibiotics that were discovered using a screen to identify inhibitors of bacterial translocase I, an essential enzyme in peptidoglycan cell wall biosynthesis. Like the parent capuramycin, A-500359s and A-503083s consist of three structural components: a uridine-5′-carboxamide (CarU), a rare unsaturated hexuronic acid, and an aminocaprolactam, the last of which is substituted by an unusual arylamine-containing polyamide in A-102395. The biosynthetic gene clusters for A-500359s and A-503083s have been reported, and two genes encoding a putative non-heme Fe(II)-dependent α-ketoglutarate:UMP dioxygenase and an l-Thr:uridine-5′-aldehyde transaldolase were uncovered, suggesting that C–C bond formation during assembly of the high carbon (C6) sugar backbone of CarU proceeds from the precursors UMP and l-Thr to form 5′-C-glycyluridine (C7) as a biosynthetic intermediate. Here, isotopic enrichment studies with the producer of A-503083s were used to indeed establish l-Thr as the direct source of the carboxamide of CarU. With this knowledge, the A-102395 gene cluster was subsequently cloned and characterized. A genetic system in the A-102395-producing strain was developed, permitting the inactivation of several genes, including those encoding the dioxygenase (cpr19) and transaldolase (cpr25), which abolished the production of A-102395, thus confirming their role in biosynthesis. Heterologous production of recombinant Cpr19 and CapK, the transaldolase homolog involved in A-503083 biosynthesis, confirmed their expected function. Finally, a phosphotransferase (Cpr17) conferring self-resistance was functionally characterized. The results provide the opportunity to use comparative genomics along with in vivo and in vitro approaches to probe the biosynthetic mechanism of these intriguing structures. PMID:25855790

  13. Ectopic expression of a basic helix-loop-helix gene transactivates parallel pathways of proanthocyanidin biosynthesis. structure, expression analysis, and genetic control of leucoanthocyanidin 4-reductase and anthocyanidin reductase genes in Lotus corniculatus.

    Science.gov (United States)

    Paolocci, Francesco; Robbins, Mark P; Madeo, Laura; Arcioni, Sergio; Martens, Stefan; Damiani, Francesco

    2007-01-01

    Proanthocyanidins (PAs) are plant secondary metabolites and are composed primarily of catechin and epicatechin units in higher plant species. Due to the ability of PAs to bind reversibly with plant proteins to improve digestion and reduce bloat, engineering this pathway in leaves is a major goal for forage breeders. Here, we report the cloning and expression analysis of anthocyanidin reductase (ANR) and leucoanthocyanidin 4-reductase (LAR), two genes encoding enzymes committed to epicatechin and catechin biosynthesis, respectively, in Lotus corniculatus. We show the presence of two LAR gene families (LAR1 and LAR2) and that the steady-state levels of ANR and LAR1 genes correlate with the levels of PAs in leaves of wild-type and transgenic plants. Interestingly, ANR and LAR1, but not LAR2, genes produced active proteins following heterologous expression in Escherichia coli and are affected by the same basic helix-loop-helix transcription factor that promotes PA accumulation in cells of palisade and spongy mesophyll. This study provides direct evidence that the same subclass of transcription factors can mediate the expression of the structural genes of both branches of PA biosynthesis.

  14. A Simple Approach to Study Designs in Complex Biochemical ...

    Indian Academy of Sciences (India)

    Somdatta Sinha

    Protein sequences. • Biochemical & Genetic information. REVERSE ENGINEERING. LARGE NETWORKS. FORWARD ENGINEERING. All designs that are not physically forbidden are realizable, but not all realizable designs are functionally effective. (in relation to context and constraints of the system and environment).

  15. Abnormal Ergosterol Biosynthesis Activates Transcriptional Responses to Antifungal Azoles.

    Science.gov (United States)

    Hu, Chengcheng; Zhou, Mi; Wang, Wenzhao; Sun, Xianyun; Yarden, Oded; Li, Shaojie

    2018-01-01

    (s). This study demonstrates, by detailed genetic and chemical analysis, that transcriptional responses by a major efflux pump and genes of the ergosterol biosynthesis pathway to ergosterol biosynthesis inhibitors can be independent of the presence of the drugs and are linked with the accumulation of ergosterol intermediate(s).

  16. Lignin biosynthesis in wheat (Triticum aestivum L.): its response to waterlogging and association with hormonal levels.

    Science.gov (United States)

    Nguyen, Tran-Nguyen; Son, SeungHyun; Jordan, Mark C; Levin, David B; Ayele, Belay T

    2016-01-25

    Lignin is an important structural component of plant cell wall that confers mechanical strength and tolerance against biotic and abiotic stressors; however it affects the use of biomass such as wheat straw for some industrial applications such as biofuel production. Genetic alteration of lignin quantity and quality has been considered as a viable option to overcome this problem. However, the molecular mechanisms underlying lignin formation in wheat biomass has not been studied. Combining molecular and biochemical approaches, the present study investigated the transcriptional regulation of lignin biosynthesis in two wheat cultivars with varying lodging characteristics and also in response to waterlogging. It also examined the association of lignin level in tissues with that of plant hormones implicated in the control of lignin biosynthesis. Analysis of lignin biosynthesis in the two wheat cultivars revealed a close association of lodging resistance with internode lignin content and expression of 4-coumarate:CoA ligase1 (4CL1), p-coumarate 3-hydroxylase1 (C3H1), cinnamoyl-CoA reductase2 (CCR2), ferulate 5-hydroxylase2 (F5H2) and caffeic acid O-methyltransferase2 (COMT2), which are among the genes highly expressed in wheat tissues, implying the importance of these genes in mediating lignin deposition in wheat stem. Waterlogging of wheat plants reduced internode lignin content, and this effect is accompanied by transcriptional repression of three of the genes characterized as highly expressed in wheat internode including phenylalanine ammonia-lyase6 (PAL6), CCR2 and F5H2, and decreased activity of PAL. Expression of the other genes was, however, induced by waterlogging, suggesting their role in the synthesis of other phenylpropanoid-derived molecules with roles in stress responses. Moreover, difference in internode lignin content between cultivars or change in its level due to waterlogging is associated with the level of cytokinin. Lodging resistance, tolerance against

  17. Cross-Neutralizing Antibodies in HIV-1 Individuals Infected by Subtypes B, F1, C or the B/Bbr Variant in Relation to the Genetics and Biochemical Characteristics of the env Gene.

    Directory of Open Access Journals (Sweden)

    Dalziza Victalina de Almeida

    Full Text Available Various HIV-1 env genetic and biochemical features impact the elicitation of cross-reactive neutralizing antibodies in natural infections. Thus, we aimed to investigate cross-neutralizing antibodies in individuals infected with HIV-1 env subtypes B, F1, C or the B/Bbr variant as well as env characteristics. Therefore, plasma samples from Brazilian chronically HIV-1 infected individuals were submitted to the TZM-bl neutralization assay. We also analyzed putative N-glycosylation sites (PNGLs and the size of gp120 variable domains in the context of HIV-1 subtypes prevalent in Brazil. We observed a greater breadth and potency of the anti-Env neutralizing response in individuals infected with the F1 or B HIV-1 subtypes compared with the C subtype and the variant B/Bbr. We observed greater V1 B/Bbr and smaller V4 F1 than those of other subtypes (p<0.005, however neither was there a correlation verified between the variable region length and neutralization potency, nor between PNLG and HIV-1 subtypes. The enrichment of W at top of V3 loop in weak neutralizing response viruses and the P in viruses with higher neutralization susceptibility was statistically significant (p = 0.013. Some other signatures sites were associated to HIV-1 subtype-specific F1 and B/Bbr samples might influence in the distinct neutralizing response. These results indicate that a single amino acid substitution may lead to a distinct conformational exposure or load in the association domain of the trimer of gp120 and interfere with the induction power of the neutralizing response, which affects the sensitivity of the neutralizing antibody and has significant implications for vaccine design.

  18. Cross-Neutralizing Antibodies in HIV-1 Individuals Infected by Subtypes B, F1, C or the B/Bbr Variant in Relation to the Genetics and Biochemical Characteristics of the env Gene.

    Science.gov (United States)

    de Almeida, Dalziza Victalina; Macieira, Karine Venegas; Grinsztejn, Beatriz Gilda Jegerhorn; Veloso Dos Santos, Valdiléa Gonçalves; Guimarães, Monick Lindenmeyer

    2016-01-01

    Various HIV-1 env genetic and biochemical features impact the elicitation of cross-reactive neutralizing antibodies in natural infections. Thus, we aimed to investigate cross-neutralizing antibodies in individuals infected with HIV-1 env subtypes B, F1, C or the B/Bbr variant as well as env characteristics. Therefore, plasma samples from Brazilian chronically HIV-1 infected individuals were submitted to the TZM-bl neutralization assay. We also analyzed putative N-glycosylation sites (PNGLs) and the size of gp120 variable domains in the context of HIV-1 subtypes prevalent in Brazil. We observed a greater breadth and potency of the anti-Env neutralizing response in individuals infected with the F1 or B HIV-1 subtypes compared with the C subtype and the variant B/Bbr. We observed greater V1 B/Bbr and smaller V4 F1 than those of other subtypes (pHIV-1 subtypes. The enrichment of W at top of V3 loop in weak neutralizing response viruses and the P in viruses with higher neutralization susceptibility was statistically significant (p = 0.013). Some other signatures sites were associated to HIV-1 subtype-specific F1 and B/Bbr samples might influence in the distinct neutralizing response. These results indicate that a single amino acid substitution may lead to a distinct conformational exposure or load in the association domain of the trimer of gp120 and interfere with the induction power of the neutralizing response, which affects the sensitivity of the neutralizing antibody and has significant implications for vaccine design.

  19. Biochemical and genetic diversity of carbohydrate-fermenting and obligate amino acid-fermenting hyper-ammonia-producing bacteria from Nellore steers fed tropical forages and supplemented with casein.

    Science.gov (United States)

    Bento, Cláudia Braga Pereira; de Azevedo, Analice Cláudia; Detmann, Edenio; Mantovani, Hilário Cuquetto

    2015-02-14

    Dietary protein plays a major role in ruminant nutrition, and protein supplementation is a widespread practice among farmers in the tropics. Ruminal bacteria are the main agents of dietary protein and amino acid degradation, yet few studies have focused on the isolation and characterization of hyper-ammonia-producing bacteria in animals fed tropical diets or supplemented with rumen-degradable proteins. This work investigated the bacterial community diversity of the rumen of Nellore steers fed tropical forages, with or without casein supplementation. We also isolated and characterized ruminal bacteria showing high levels of ammonia production. Polymerase chain reaction-denaturing gradient gel electrophoresis analysis indicated no differences in the ruminal bacterial community composition between the control and supplemented animals. Amino acid-fermenting bacteria (n = 250) were isolated from crossbred Nellore steers fed Tifton 85 (Cynodon sp.) using trypticase as the sole carbon and organic nitrogen source in the enrichment and isolation media. The deamination rates in isolates obtained from steers supplemented with casein showed a higher incidence of deamination rates >350 nmol NH3 mg protein(-1) min(-1) (P < 0.05), whereas isolates obtained from steers without supplementation showed deamination rates <200 nmol NH3 mg protein(-1) min(-1). Although most isolates (84%) could ferment carbohydrates, none could hydrolyze proteins or use urea to sustain growth. All isolates were sensitive to lasalocid and monensin (1 μmol l(-1)), and similarity analysis of the 16S rRNA sequences indicated a predominance of bacteria from the order Clostridiales, with variable homology (73-99%) to known bacterial species. These results expand what is known about the biochemical and genetic diversity of hyper-ammonia-producing bacteria, and emphasize the role of carbohydrate-fermenting bacteria in ammonia production in the rumen.

  20. Oxylipin Pathway in the Biosynthesis of Fresh Tomato Volatiles

    OpenAIRE

    YILMAZ, Emin

    2001-01-01

    Fresh tomato volatiles are formed in intact fruit during ripening and upon tissue disruption. There are different pathways involved in the biosynthesis of these volatiles. The oxylipin pathway uses free unsaturated fatty acids with the sequential action of lipoxygenase, hydroperoxide lyase and alcohol dehydrogenase to produce volatile aldehyde and alcohol compounds. Oxylipin volatiles are the most important components in fresh tomato aroma. In order to genetically improve the quality of tomat...

  1. Engineered polyketide biosynthesis and biocatalysis in Escherichia coli

    OpenAIRE

    Gao, Xue; Wang, Peng; Tang, Yi

    2010-01-01

    Polyketides are important bioactive natural products biosynthesized by bacteria, fungi, and plants. The enzymes that synthesize polyketides are collectively referred to as polyketide synthases (PKSs). Because many of the natural hosts that produce polyketides are difficult to culture or manipulate, establishing a universal heterologous host that is genetically tractable has become an important goal toward the engineered biosynthesis of polyketides and analogues. Here, we summarize the recent ...

  2. Coupled biochemical genetic and karyomorphological analyses for ...

    African Journals Online (AJOL)

    S. esocinus showed five bands, S. curvifrons five, S. niger seven, S. labiatus and S. plagiostomus each showed six bands; they also showed species characteristic bands. Karyotypic study of these was carried out. The diploid chromosome numbers recorded were 98 in S. niger (24 m + 32 sm + 22 st + 20 t), 98 in S. esocinus ...

  3. RxnFinder: biochemical reaction search engines using molecular structures, molecular fragments and reaction similarity.

    Science.gov (United States)

    Hu, Qian-Nan; Deng, Zhe; Hu, Huanan; Cao, Dong-Sheng; Liang, Yi-Zeng

    2011-09-01

    Biochemical reactions play a key role to help sustain life and allow cells to grow. RxnFinder was developed to search biochemical reactions from KEGG reaction database using three search criteria: molecular structures, molecular fragments and reaction similarity. RxnFinder is helpful to get reference reactions for biosynthesis and xenobiotics metabolism. RxnFinder is freely available via: http://sdd.whu.edu.cn/rxnfinder. qnhu@whu.edu.cn.

  4. Recent advances in combinatorial biosynthesis for drug discovery

    Directory of Open Access Journals (Sweden)

    Sun H

    2015-02-01

    Full Text Available Huihua Sun,1,* Zihe Liu,1,* Huimin Zhao,1,2 Ee Lui Ang1 1Metabolic Engineering Research Laboratory, Institute of Chemical and Engineering Sciences, Agency for Science, Technology and Research, Singapore; 2Department of Chemical and Biomolecular Engineering, University of Illinois at Urbana-Champaign, Urbana, IL, USA *These authors contributed equally to this work Abstract: Because of extraordinary structural diversity and broad biological activities, natural products have played a significant role in drug discovery. These therapeutically important secondary metabolites are assembled and modified by dedicated biosynthetic pathways in their host living organisms. Traditionally, chemists have attempted to synthesize natural product analogs that are important sources of new drugs. However, the extraordinary structural complexity of natural products sometimes makes it challenging for traditional chemical synthesis, which usually involves multiple steps, harsh conditions, toxic organic solvents, and byproduct wastes. In contrast, combinatorial biosynthesis exploits substrate promiscuity and employs engineered enzymes and pathways to produce novel “unnatural” natural products, substantially expanding the structural diversity of natural products with potential pharmaceutical value. Thus, combinatorial biosynthesis provides an environmentally friendly way to produce natural product analogs. Efficient expression of the combinatorial biosynthetic pathway in genetically tractable heterologous hosts can increase the titer of the compound, eventually resulting in less expensive drugs. In this review, we will discuss three major strategies for combinatorial biosynthesis: 1 precursor-directed biosynthesis; 2 enzyme-level modification, which includes swapping of the entire domains, modules and subunits, site-specific mutagenesis, and directed evolution; 3 pathway-level recombination. Recent examples of combinatorial biosynthesis employing these

  5. Comparison of 454-ESTs from Huperzia serrata and Phlegmariurus carinatus reveals putative genes involved in lycopodium alkaloid biosynthesis and developmental regulation

    Directory of Open Access Journals (Sweden)

    Steinmetz André

    2010-09-01

    Full Text Available Abstract Background Plants of the Huperziaceae family, which comprise the two genera Huperzia and Phlegmariurus, produce various types of lycopodium alkaloids that are used to treat a number of human ailments, such as contusions, swellings and strains. Huperzine A, which belongs to the lycodine type of lycopodium alkaloids, has been used as an anti-Alzheimer's disease drug candidate. Despite their medical importance, little genomic or transcriptomic data are available for the members of this family. We used massive parallel pyrosequencing on the Roche 454-GS FLX Titanium platform to generate a substantial EST dataset for Huperzia serrata (H. serrata and Phlegmariurus carinatus (P. carinatus as representative members of the Huperzia and Phlegmariurus genera, respectively. H. serrata and P. carinatus are important plants for research on the biosynthesis of lycopodium alkaloids. We focused on gene discovery in the areas of bioactive compound biosynthesis and transcriptional regulation as well as genetic marker detection in these species. Results For H. serrata, 36,763 unique putative transcripts were generated from 140,930 reads totaling over 57,028,559 base pairs; for P. carinatus, 31,812 unique putative transcripts were generated from 79,920 reads totaling over 30,498,684 base pairs. Using BLASTX searches of public databases, 16,274 (44.3% unique putative transcripts from H. serrata and 14,070 (44.2% from P. carinatus were assigned to at least one protein. Gene Ontology (GO and Kyoto Encyclopedia of Genes and Genomes (KEGG orthology annotations revealed that the functions of the unique putative transcripts from these two species cover a similarly broad set of molecular functions, biological processes and biochemical pathways. In particular, a total of 20 H. serrata candidate cytochrome P450 genes, which are more abundant in leaves than in roots and might be involved in lycopodium alkaloid biosynthesis, were found based on the comparison of H

  6. On the Adaptive Design Rules of Biochemical Networks in Evolution

    Directory of Open Access Journals (Sweden)

    Bor-Sen Chen

    2007-01-01

    Full Text Available Biochemical networks are the backbones of physiological systems of organisms. Therefore, a biochemical network should be sufficiently robust (not sensitive to tolerate genetic mutations and environmental changes in the evolutionary process. In this study, based on the robustness and sensitivity criteria of biochemical networks, the adaptive design rules are developed for natural selection in the evolutionary process. This will provide insights into the robust adaptive mechanism of biochemical networks in the evolutionary process. We find that if a mutated biochemical network satisfies the robustness and sensitivity criteria of natural selection, there is a high probability for the biochemical network to prevail during natural selection in the evolutionary process. Since there are various mutated biochemical networks that can satisfy these criteria but have some differences in phenotype, the biochemical networks increase their diversities in the evolutionary process. The robustness of a biochemical network enables co-option so that new phenotypes can be generated in evolution. The proposed robust adaptive design rules of natural selection gain much insight into the evolutionary mechanism and provide a systematic robust biochemical circuit design method of biochemical networks for biotechnological and therapeutic purposes in the future.

  7. Minimum set of cytochromes P450 for reconstituting the biosynthesis of camalexin, a major Arabidopsis antibiotic.

    Science.gov (United States)

    Klein, Andrew P; Anarat-Cappillino, Gülbenk; Sattely, Elizabeth S

    2013-12-16

    Bringing it all together: The missing key step in the biosynthesis of camalexin was uncovered by in vitro biochemical characterization. The coupling of Trp- and Cys-derived fragments through CS bond formation is promoted by an unusual cytochrome P450 CYP71A13. The in vitro reconstitution of the camalexin biosynthesis (left) from Trp and Cys was achieved using just three cytochromes P450. IAN=indole-3-acetonitrile. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  8. Auxin biosynthesis and storage forms

    Science.gov (United States)

    Strader, Lucia C.

    2013-01-01

    The plant hormone auxin drives plant growth and morphogenesis. The levels and distribution of the active auxin indole-3-acetic acid (IAA) are tightly controlled through synthesis, inactivation, and transport. Many auxin precursors and modified auxin forms, used to regulate auxin homeostasis, have been identified; however, very little is known about the integration of multiple auxin biosynthesis and inactivation pathways. This review discusses the many ways auxin levels are regulated through biosynthesis, storage forms, and inactivation, and the potential roles modified auxins play in regulating the bioactive pool of auxin to affect plant growth and development. PMID:23580748

  9. Hypericin: chemical synthesis and biosynthesis.

    Science.gov (United States)

    Huang, Lin-Fang; Wang, Zeng-Hui; Chen, Shi-Lin

    2014-02-01

    Hypericin is one of the most important phenanthoperylene quinones extracted mainly from plants of the genus Hypericum belonging to the sections Euhypericum and Campylosporus of Keller's classification. Widespread attention to the antiviral and anti-tumor properties of hypericin has spurred investigations of the chemical synthesis and biosynthesis of this unique compound. However, the synthetic strategies are challenging for organic and biological chemists. In this review, specific significant advances in total synthesis, semi-synthesis, and biosynthesis in the past decades are summarized. Copyright © 2014 China Pharmaceutical University. Published by Elsevier B.V. All rights reserved.

  10. Biochemical adaptation to ocean acidification.

    Science.gov (United States)

    Stillman, Jonathon H; Paganini, Adam W

    2015-06-01

    The change in oceanic carbonate chemistry due to increased atmospheric PCO2  has caused pH to decline in marine surface waters, a phenomenon known as ocean acidification (OA). The effects of OA on organisms have been shown to be widespread among diverse taxa from a wide range of habitats. The majority of studies of organismal response to OA are in short-term exposures to future levels of PCO2 . From such studies, much information has been gathered on plastic responses organisms may make in the future that are beneficial or harmful to fitness. Relatively few studies have examined whether organisms can adapt to negative-fitness consequences of plastic responses to OA. We outline major approaches that have been used to study the adaptive potential for organisms to OA, which include comparative studies and experimental evolution. Organisms that inhabit a range of pH environments (e.g. pH gradients at volcanic CO2 seeps or in upwelling zones) have great potential for studies that identify adaptive shifts that have occurred through evolution. Comparative studies have advanced our understanding of adaptation to OA by linking whole-organism responses with cellular mechanisms. Such optimization of function provides a link between genetic variation and adaptive evolution in tuning optimal function of rate-limiting cellular processes in different pH conditions. For example, in experimental evolution studies of organisms with short generation times (e.g. phytoplankton), hundreds of generations of growth under future conditions has resulted in fixed differences in gene expression related to acid-base regulation. However, biochemical mechanisms for adaptive responses to OA have yet to be fully characterized, and are likely to be more complex than simply changes in gene expression or protein modification. Finally, we present a hypothesis regarding an unexplored area for biochemical adaptation to ocean acidification. In this hypothesis, proteins and membranes exposed to the

  11. Arogenate Dehydratase Isoforms Differentially Regulate Anthocyanin Biosynthesis in Arabidopsis thaliana.

    Science.gov (United States)

    Chen, Qingbo; Man, Cong; Li, Danning; Tan, Huijuan; Xie, Ye; Huang, Jirong

    2016-12-05

    Anthocyanins, a group of L-phenylalanine (Phe)-derived flavonoids, have been demonstrated to play important roles in plant stress resistance and interactions between plants and insects. Although the anthocyanin biosynthetic pathway and its regulatory mechanisms have been extensively studied, it remains unclear whether the level of Phe supply affects anthocyanin biosynthesis. Here, we investigated the roles of arogenate dehydratases (ADTs), the key enzymes that catalyze the conversion of arogenate into Phe, in sucrose-induced anthocyanin biosynthesis in Arabidopsis. Genetic analysis showed that all six ADT isoforms function redundantly in anthocyanin biosynthesis but have differential contributions. ADT2 contributes the most to anthocyanin accumulation, followed by ADT1 and ADT3, and ADT4-ADT6. We found that anthocyanin content is positively correlated with the levels of Phe and sucrose-induced ADT transcripts in seedlings. Consistently, addition of Phe to the medium could dramatically increase anthocyanin content in the wild-type plants and rescue the phenotype of the adt1 adt3 double mutant regarding the anthocyanin accumulation. Moreover, transgenic plants overexpressing ADT4, which appears to be less sensitive to Phe than overexpression of ADT2, hyperaccumulate Phe and produce elevated level of anthocyanins. Taken together, our results suggest that the level of Phe is an important regulatory factor for sustaining anthocyanin biosynthesis. Copyright © 2016 The Author. Published by Elsevier Inc. All rights reserved.

  12. Defining Determinants and Dynamics and Cellulose Microfibril Biosynthesis, Assembly and Degredation OSP Number: 63079/A001

    Energy Technology Data Exchange (ETDEWEB)

    None, None

    2013-12-01

    been based on the idea that the most effective way to address this long standing and highly complex question is to adopt a broad ‘systems approach’. Accordingly, we assembled a multi-disciplinary collaborative team with collective expertise in plant biology and molecular genetics, polymer structure and chemistry, enzyme biochemistry and biochemical engineering. We used a spectrum of cutting edge technologies, including plant functional genomics, chemical genetics, live cell imaging, advanced microscopy, high energy X-ray spectroscopy and nanotechnology, to study the molecular determinants of cellulose microfibril structure. Importantly, this research effort was closely coupled with an analytical pipeline to characterize the effects of altering microfibril architecture on bioconversion potential, with the goal of generating predictive models to help guide the identification, development and implementation of new feedstocks. This project therefore spanned core basic science and applied research, in line with the goals of the program. Over the course of the project, accomplishments included: - Establishing platforms through reverse and forward genetics to identify and manipulate candidate genes that influence cellulose microfibril synthesis and structure in a model C3 grass, Brachypodium distachyon and a model C4 grass Setaria viridis; Identifying and characterizing the effects of a number of cellulose biosynthesis inhibitors (CBIs), and particularly those that target monocots with the aim of generating resistance loci; Developing protocols for the use of high energy X-ray diffraction (XRD) to study the structure and organization of cellulose microfibrils in plant walls, notably those in Arabidopsis and Brachypodium; Using the chemical and genetic based inhibition strategies to develop new mechanistic models of cellulose microfibril crystallization, and of how altering microfibril architecture influences digestibility.

  13. (vitamin B1) biosynthesis genes

    African Journals Online (AJOL)

    In this study, the gene transcripts of first two enzymes in thiamine biosynthesis pathway, THIC and THI1/THI4 were identified and amplified from oil palm tissues. Primers were designed based on sequence comparison of the genes from Arabidopsis thaliana, Zea mays, Oryza sativa and Alnus glutinosa. Oil palm's responses ...

  14. Problems in Protein Biosynthesis

    Science.gov (United States)

    Lengyel, Peter

    1966-01-01

    Outline of the steps in protein synthesis. Nature of the genetic code. The use of synthetic oligo- and polynucleotides in deciphering the code. Structure of the code: relatedness of synonym codons. The wobble hypothesis. Chain initiation and N-formyl-methionine. Chain termination and nonsense codons. Mistakes in translation: ambiguity in vitro. Suppressor mutations resulting in ambiguity. Limitations in the universality of the code. Attempts to determine the particular codons used by a species. Mechanisms of suppression, caused by (a) abnormal aminoacyl-tRNA, (b) ribosomal malfunction. Effect of streptomycin. The problem of "reading" a nucleic acid template. Different ribosomal mutants and DNA polymerase mutants might cause different mistakes. The possibility of involvement of allosteric proteins in template reading. PMID:5338560

  15. The Arabidopsis Vacuolar Sorting Receptor1 Is Required for Osmotic Stress-Induced Abscisic Acid Biosynthesis

    KAUST Repository

    Wang, Zhen-Yu

    2014-11-21

    Osmotic stress activates the biosynthesis of the phytohormone abscisic acid (ABA) through a pathway that is rate limited by the carotenoid cleavage enzyme 9-cis-epoxycarotenoid dioxygenase (NCED). To understand the signal transduction mechanism underlying the activation of ABA biosynthesis, we performed a forward genetic screen to isolate mutants defective in osmotic stress regulation of the NCED3 gene. Here, we identified the Arabidopsis (Arabidopsis thaliana) Vacuolar Sorting Receptor1 (VSR1) as a unique regulator of ABA biosynthesis. The vsr1 mutant not only shows increased sensitivity to osmotic stress, but also is defective in the feedback regulation of ABA biosynthesis by ABA. Further analysis revealed that vacuolar trafficking mediated by VSR1 is required for osmotic stress-responsive ABA biosynthesis and osmotic stress tolerance. Moreover, under osmotic stress conditions, the membrane potential, calcium flux, and vacuolar pH changes in the vsr1 mutant differ from those in the wild type. Given that manipulation of the intracellular pH is sufficient to modulate the expression of ABA biosynthesis genes, including NCED3, and ABA accumulation, we propose that intracellular pH changes caused by osmotic stress may play a signaling role in regulating ABA biosynthesis and that this regulation is dependent on functional VSR1.

  16. Regulation of neurosteroid biosynthesis by neurotransmitters and neuropeptides

    Directory of Open Access Journals (Sweden)

    Jean-Luc eDo-Rego

    2012-01-01

    Full Text Available The enzymatic pathways leading to the synthesis of bioactive steroids in the brain are now almost completely elucidated in various groups of vertebrates and, during the last decade, the neuronal mechanisms involved in the regulation of neurosteroid production have received increasing attention. This report reviews the current knowledge concerning the effects of neurotransmitters, peptide hormones and neuropeptides on the biosynthesis of neurosteroids. Anatomical studies have been carried out to visualize the neurotransmitter- or neuropeptide-containing fibers contacting steroid-synthesizing neurons as well as the neurotransmitter, peptide hormones or neuropeptide receptors expressed in these neurons. Biochemical experiments have been conducted to investigate the effects of neurotransmitters, peptide hormones or neuropeptides on neurosteroid biosynthesis, and to characterize the type of receptors involved. Thus, it has been found that glutamate, acting through kainate and/or AMPA receptors, rapidly inactivates P450arom, and that melatonin produced by the pineal gland and eye inhibits the biosynthesis of 7-hydroxypregnenolone (7-OH-5P, while prolactin produced by the adenohypophysis enhances the formation of 7-OH-5P. It has also been demonstrated that the biosynthesis of neurosteroids is inhibited by GABA, acting through GABAA receptors, and neuropeptide Y, acting through Y1 receptors. In contrast, it has been shown that the octadecaneuropetide ODN, acting through central-type benzodiazepine receptors, the triakontatetraneuropeptide TTN, acting though peripheral-type benzodiazepine receptors, and vasotocine, acting through V1a-like receptors, stimulate the production of neurosteroids. Since neurosteroids are implicated in the control of various neurophysiological and behavioral processes, these data suggest that some of the neurophysiological effects exerted by neurotransmitters and neuropeptides may be mediated via the regulation

  17. Isoprenoid-derived plant signaling molecules: biosynthesis and biological importance

    Czech Academy of Sciences Publication Activity Database

    Tarkowská, Danuše; Strnad, Miroslav

    2018-01-01

    Roč. 247, č. 5 (2018), s. 1051-1066 ISSN 0032-0935 R&D Projects: GA MŠk(CZ) LO1204 Institutional support: RVO:61389030 Keywords : Dimethylallyl diphosphate * Isopentenyl diphosphate * Isoprenoids * Phytoecdysteroids * Plant hormones * Terpenoids Subject RIV: EB - Genetics ; Molecular Biology OBOR OECD: Biochemical research methods Impact factor: 3.361, year: 2016

  18. 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.

  19. Cysteine Biosynthesis Controls Serratia marcescens Phospholipase Activity.

    Science.gov (United States)

    Anderson, Mark T; Mitchell, Lindsay A; Mobley, Harry L T

    2017-08-15

    Serratia marcescens causes health care-associated opportunistic infections that can be difficult to treat due to a high incidence of antibiotic resistance. One of the many secreted proteins of S. marcescens is the PhlA phospholipase enzyme. Genes involved in the production and secretion of PhlA were identified by screening a transposon insertion library for phospholipase-deficient mutants on phosphatidylcholine-containing medium. Mutations were identified in four genes ( cyaA , crp , fliJ , and fliP ) that are involved in the flagellum-dependent PhlA secretion pathway. An additional phospholipase-deficient isolate harbored a transposon insertion in the cysE gene encoding a predicted serine O -acetyltransferase required for cysteine biosynthesis. The cysE requirement for extracellular phospholipase activity was confirmed using a fluorogenic phospholipase substrate. Phospholipase activity was restored to the cysE mutant by the addition of exogenous l-cysteine or O -acetylserine to the culture medium and by genetic complementation. Additionally, phlA transcript levels were decreased 6-fold in bacteria lacking cysE and were restored with added cysteine, indicating a role for cysteine-dependent transcriptional regulation of S. marcescens phospholipase activity. S. marcescens cysE mutants also exhibited a defect in swarming motility that was correlated with reduced levels of flhD and fliA flagellar regulator gene transcription. Together, these findings suggest a model in which cysteine is required for the regulation of both extracellular phospholipase activity and surface motility in S. marcescens IMPORTANCE Serratia marcescens is known to secrete multiple extracellular enzymes, but PhlA is unusual in that this protein is thought to be exported by the flagellar transport apparatus. In this study, we demonstrate that both extracellular phospholipase activity and flagellar function are dependent on the cysteine biosynthesis pathway. Furthermore, a disruption of cysteine

  20. Acylphloroglucinol Biosynthesis in Strawberry Fruit1

    Science.gov (United States)

    Song, Chuankui; Ring, Ludwig; Hoffmann, Thomas; Huang, Fong-Chin; Slovin, Janet; Schwab, Wilfried

    2015-01-01

    Phenolics have health-promoting properties and are a major group of metabolites in fruit crops. Through reverse genetic analysis of the functions of four ripening-related genes in the octoploid strawberry (Fragaria × ananassa), we discovered four acylphloroglucinol (APG)-glucosides as native Fragaria spp. fruit metabolites whose levels were differently regulated in the transgenic fruits. The biosynthesis of the APG aglycones was investigated by examination of the enzymatic properties of three recombinant Fragaria vesca chalcone synthase (FvCHS) proteins. CHS is involved in anthocyanin biosynthesis during ripening. The F. vesca enzymes readily catalyzed the condensation of two intermediates in branched-chain amino acid metabolism, isovaleryl-Coenzyme A (CoA) and isobutyryl-CoA, with three molecules of malonyl-CoA to form phlorisovalerophenone and phlorisobutyrophenone, respectively, and formed naringenin chalcone when 4-coumaroyl-CoA was used as starter molecule. Isovaleryl-CoA was the preferred starter substrate of FvCHS2-1. Suppression of CHS activity in both transient and stable CHS-silenced fruit resulted in a substantial decrease of APG glucosides and anthocyanins and enhanced levels of volatiles derived from branched-chain amino acids. The proposed APG pathway was confirmed by feeding isotopically labeled amino acids. Thus, Fragaria spp. plants have the capacity to synthesize pharmaceutically important APGs using dual functional CHS/(phloriso)valerophenone synthases that are expressed during fruit ripening. Duplication and adaptive evolution of CHS is the most probable scenario and might be generally applicable to other plants. The results highlight that important promiscuous gene function may be missed when annotation relies solely on in silico analysis. PMID:26169681

  1. Biosynthesis of Anthocyanins and Their Regulation in Colored Grapes

    Directory of Open Access Journals (Sweden)

    Guo-Liang Yan

    2010-12-01

    Full Text Available Anthocyanins, synthesized via the flavonoid pathway, are a class of crucial phenolic compounds which are fundamentally responsible for the red color of grapes and wines. As the most important natural colorants in grapes and their products, anthocyanins are also widely studied for their numerous beneficial effects on human health. In recent years, the biosynthetic pathway of anthocyanins in grapes has been thoroughly investigated. Their intracellular transportation and accumulation have also been further clarified. Additionally, the genetic mechanism regulating their biosynthesis and the phytohormone influences on them are better understood. Furthermore, due to their importance in the quality of wine grapes, the effects of the environmental factors and viticulture practices on anthocyanin accumulation are being investigated increasingly. The present paper summarizes both the basic information and the most recent advances in the study of the anthocyanin biosynthesis in red grapes, emphasizing their gene structure, the transcriptional factors and the diverse exterior regulation factors.

  2. [Biosynthesis of opioid peptides].

    Science.gov (United States)

    Rossier, J

    1988-01-01

    The endogenous opioid peptides all contain the enkephalin sequence Tyr-Gly-Gly-Phe-Met and Tyr-Gly-Gly-Phe-Leu at their aminoterminus. Three distinct families of these peptides (endorphins, enkephalins and dynorphins) are present in different neuronal pathways within the central nervous system. Molecular genetics have shown that these three families of opioid peptides are derived from three distinct precursors. Pro-opiomelanocortin gives rise to the endorphins, as well as adrenocorticotropic hormone (ACTH) and the melanotropic hormones (MSH's). [Met] enkephalin, [Leu] enkephalin and the related heptapeptide [Met] enkephalin-Arg6-Phe7 and octapeptide [Met] enkephalin-Arg6-Gly7-Leu8 are derived from proenkephalin. The third family is derived from prodynorphin and includes dynorphin A, dynorphin B (also known as rimorphin) and alpha- and beta-neo-endorphin. The structure of the genes coding for these precursors are similar, suggesting the possibility of one common ancestral gene. The most common scheme for enzymatic maturation of precursors proposes the action of a trypsin-like endopeptidase followed by a carboxypeptidase B-like exopeptidase. However, we have provided evidence that this combination of trypsin-like and carboxypeptidase B-like enzymes may not be the only mechanism for liberating enkephalin from low molecular weight enkephalin-containing peptides. Indeed, endo-oligopeptidase A, an enzyme, known to hydrolyze the Phe5-Ser6 bond of bradykinin and the Arg8-Arg9 bond of neurotensin, has been shown to produce, by a single cleavage, [Leu] enkephalin or [Met] enkephalin from small enkephalin-containing peptides, (Camargo et al., 1987, J. Neurochem. 48, 1258-1263).(ABSTRACT TRUNCATED AT 250 WORDS)

  3. Lysine biosynthesis in microbes: relevance as drug target and prospects for β-lactam antibiotics production.

    Science.gov (United States)

    Fazius, Felicitas; Zaehle, Christoph; Brock, Matthias

    2013-05-01

    Plants as well as pro- and eukaryotic microorganisms are able to synthesise lysine via de novo synthesis. While plants and bacteria, with some exceptions, rely on variations of the meso-diaminopimelate pathway for lysine biosynthesis, fungi exclusively use the α-aminoadipate pathway. Although bacteria and fungi are, in principle, both suitable as lysine producers, current industrial fermentations rely on the use of bacteria. In contrast, fungi are important producers of β-lactam antibiotics such as penicillins or cephalosporins. The synthesis of these antibiotics strictly depends on α-aminoadipate deriving from lysine biosynthesis. Interestingly, despite the resulting industrial importance of the fungal α-aminoadipate pathway, biochemical reactions leading to α-aminoadipate formation have only been studied on a limited number of fungal species. In this respect, just recently an essential isomerisation reaction required for the formation of α-aminoadipate has been elucidated in detail. This review summarises biochemical pathways leading to lysine production, discusses the suitability of interrupting lysine biosynthesis as target for new antibacterial and antifungal compounds and emphasises on biochemical reactions involved in the formation of α-aminoadipate in fungi as an essential intermediate for both, lysine and β-lactam antibiotics production.

  4. Measures of Biochemical Sociology

    Science.gov (United States)

    Snell, Joel; Marsh, Mitchell

    2008-01-01

    In a previous article, the authors introduced a new sub field in sociology that we labeled "biochemical sociology." We introduced the definition of a sociology that encompasses sociological measures, psychological measures, and biological indicators Snell & Marsh (2003). In this article, we want to demonstrate a research strategy that would assess…

  5. Biochemical Education in Brazil.

    Science.gov (United States)

    Vella, F.

    1988-01-01

    Described are discussions held concerning the problems of biochemical education in Brazil at a meeting of the Sociedade Brazileira de Bioquimica in April 1988. Also discussed are other visits that were made to universities in Brazil. Three major recommendations to improve the state of biochemistry education in Brazil are presented. (CW)

  6. A novel and widespread class of ketosynthase is responsible for the head-to-head condensation of two acyl moieties in bacterial pyrone biosynthesis

    Directory of Open Access Journals (Sweden)

    Darko Kresovic

    2015-08-01

    Full Text Available The biosynthesis of photopyrones, novel quorum sensing signals in Photorhabdus, has been studied by heterologous expression of the photopyrone synthase PpyS catalyzing the head-to-head condensation of two acyl moieties. The biochemical mechanism of pyrone formation has been investigated by amino acid exchange and bioinformatic analysis. Additionally, the evolutionary origin of PpyS has been studied by phylogenetic analyses also revealing homologous enzymes in Pseudomonas sp. GM30 responsible for the biosynthesis of pseudopyronines including a novel derivative. Moreover this novel class of ketosynthases is only distantly related to other pyrone-forming enzymes identified in the biosynthesis of the potent antibiotics myxopyronin and corallopyronin.

  7. Identification of aminotransferase genes for biosynthesis of aminoglycoside antibiotics from soil DNA.

    Science.gov (United States)

    Nagaya, Atsushi; Takeyama, Satoko; Tamegai, Hideyuki

    2005-07-01

    Aminoglycoside has been known as a clinically important antibiotic for a long time, but genetic information for the biosynthesis of aminoglycoside is still insufficient. In this study, we tried to clone aminoglycoside-biosynthetic genes from soil DNA for accumulation of genetic information. We chose the genes encoding L-glutamine:(2-deoxy-)scyllo-inosose aminotransferase as the target, because it is specific for all types of aminoglycoside biosynthesis. By degenerate PCR, we obtained 33 individual clones that were homologous with aminotransferase genes in aminoglycoside biosynthesis. Phylogenetic analysis and alignment of these genes showed that horizontal gene transfer has occurred in the soil. Among these, several quite interesting genes were obtained. Some genes probably originated from non-actinomycetes, and some were far from the known homologs. These genes can be useful markers for the isolation of entire gene clusters and originating organisms.

  8. Three Principles of Diversity-Generating Biosynthesis.

    Science.gov (United States)

    Gu, Wenjia; Schmidt, Eric W

    2017-10-17

    Natural products are significant therapeutic agents and valuable drug leads. This is likely owing to their three-dimensional structural complexity, which enables them to form complex interactions with biological targets. Enzymes from natural product biosynthetic pathways show great potential to generate natural product-like compounds and libraries. Many challenges still remain in biosynthesis, such as how to rationally synthesize small molecules with novel structures and how to generate maximum chemical diversity. In this Account, we describe recent advances from our laboratory in the synthesis of natural product-like libraries using natural biosynthetic machinery. Our work has focused on the pat and tru biosynthetic pathways to patellamides, trunkamide, and related compounds from cyanobacterial symbionts in marine tunicates. These belong to the cyanobactin class of natural products, which are part of the larger group of ribosomally synthesized and post-translationally modified peptides (RiPPs). These results have enabled the synthesis of rationally designed small molecules and libraries covering more than 1 million estimated derivatives. Because the RiPPs are translated on the ribosome and then enzymatically modified, they are highly compatible with recombinant technologies. This is important because it means that the resulting natural products, their derivatives, and wholly new compounds can be synthesized using the tools of genetic engineering. The RiPPs also represent possibly the most widespread group of bioactive natural products, although this is in part because of the broad definition of what constitutes a RiPP. In addition, the underlying ideas may form the basis for broad-substrate biosynthetic pathways beyond the RiPPs. For example, some of the ideas about kinetic ordering of broad substrate pathways may apply to polyketide or nonribosomal peptide biosynthesis as well. While making these products, we have sought to understand what makes biosynthetic

  9. In Vivo Roles of Fatty Acid Biosynthesis Enzymes in Biosynthesis of Biotin and α-Lipoic Acid in Corynebacterium glutamicum.

    Science.gov (United States)

    Ikeda, Masato; Nagashima, Takashi; Nakamura, Eri; Kato, Ryosuke; Ohshita, Masakazu; Hayashi, Mikiro; Takeno, Seiki

    2017-10-01

    For fatty acid biosynthesis, Corynebacterium glutamicum uses two type I fatty acid synthases (FAS-I), FasA and FasB, in addition to acetyl-coenzyme A (CoA) carboxylase (ACC) consisting of AccBC, AccD1, and AccE. The in vivo roles of the enzymes in supplying precursors for biotin and α-lipoic acid remain unclear. Here, we report genetic evidence demonstrating that the biosynthesis of these cofactors is linked to fatty acid biosynthesis through the FAS-I pathway. For this study, we used wild-type C. glutamicum and its derived biotin vitamer producer BFI-5, which was engineered to express Escherichia coli bioBF and Bacillus subtilis bioI Disruption of either fasA or fasB in strain BFI-5 led to decreased production of biotin vitamers, whereas its amplification contributed to increased production, with a larger impact of fasA in both cases. Double disruptions of fasA and fasB resulted in no biotin vitamer production. The acc genes showed a positive effect on production when amplified simultaneously. Augmented fatty acid biosynthesis was also reflected in pimelic acid production when carbon flow was blocked at the BioF reaction. These results indicate that carbon flow down the FAS-I pathway is destined for channeling into the biotin biosynthesis pathway, and that FasA in particular has a significant impact on precursor supply. In contrast, fasB disruption resulted in auxotrophy for lipoic acid or its precursor octanoic acid in both wild-type and BFI-5 strains. The phenotypes were fully complemented by plasmid-mediated expression of fasB but not fasA These results reveal that FasB plays a specific physiological role in lipoic acid biosynthesis in C. glutamicum IMPORTANCE For the de novo biosynthesis of fatty acids, C. glutamicum exceptionally uses a eukaryotic multifunctional type I fatty acid synthase (FAS-I) system comprising FasA and FasB, in contrast to most bacteria, such as E. coli and B. subtilis , which use an individual nonaggregating type II fatty acid synthase

  10. Arabidopsis acetyl-amido synthetase GH3.5 involvement in camalexin biosynthesis through conjugation of indole-3-carboxylic acid and cysteine and upregulation of camalexin biosynthesis genes.

    Science.gov (United States)

    Wang, Mu-Yang; Liu, Xue-Ting; Chen, Ying; Xu, Xiao-Jing; Yu, Biao; Zhang, Shu-Qun; Li, Qun; He, Zu-Hua

    2012-07-01

    Camalexin (3-thiazol-2'-yl-indole) is the major phytoalexin found in Arabidopsis thaliana. Several key intermediates and corresponding enzymes have been identified in camalexin biosynthesis through mutant screening and biochemical experiments. Camalexin is formed when indole-3-acetonitrile (IAN) is catalyzed by the cytochrome P450 monooxygenase CYP71A13. Here, we demonstrate that the Arabidopsis GH3.5 protein, a multifunctional acetyl-amido synthetase, is involved in camalexin biosynthesis via conjugating indole-3-carboxylic acid (ICA) and cysteine (Cys) and regulating camalexin biosynthesis genes. Camalexin levels were increased in the activation-tagged mutant gh3.5-1D in both Col-0 and cyp71A13-2 mutant backgrounds after pathogen infection. The recombinant GH3.5 protein catalyzed the conjugation of ICA and Cys to form a possible intermediate indole-3-acyl-cysteinate (ICA(Cys)) in vitro. In support of the in vitro reaction, feeding with ICA and Cys increased camalexin levels in Col-0 and gh3.5-1D. Dihydrocamalexic acid (DHCA), the precursor of camalexin and the substrate for PAD3, was accumulated in gh3.5-1D/pad3-1, suggesting that ICA(Cys) could be an additional precursor of DHCA for camalexin biosynthesis. Furthermore, expression of the major camalexin biosynthesis genes CYP79B2, CYP71A12, CYP71A13 and PAD3 was strongly induced in gh3.5-1D. Our study suggests that GH3.5 is involved in camalexin biosynthesis through direct catalyzation of the formation of ICA(Cys), and upregulation of the major biosynthetic pathway genes. © 2012 Institute of Botany, Chinese Academy of Sciences.

  11. ASSESMENT OF BIOCHEMICAL ATTRIBUTES OF PRAECITRULLUS FISTULOSUS TREATED WITH MUTAGENS

    OpenAIRE

    Mehreen Khan

    2016-01-01

    Plants are well known to have certain primary and secondary metabolites collectively are known as biochemicals that plays an important role for human health as their medicinal properties. The aim of present study was to enhance and evaluate biochemical profile of Praecitrullus fistulosus by induced mutagenesis to cause genetic variations, plant leaves were treated with different chemical and physical mutagens. Colchicine and Ethidium bromide were used as chemical mutagens. While Ultraviolet (...

  12. Comparison of Effect of Brassinosteroid and Gibberellin Biosynthesis Inhibitors on Growth of Rice Seedlings

    Directory of Open Access Journals (Sweden)

    Tadashi Matusmoto

    2016-01-01

    Full Text Available Brassinosteroid (BR and gibberellin (GA are two predominant plant hormones that regulate plant cell elongation. Mutants disrupt the biosynthesis of these hormones and display different degrees of dwarf phenotypes in rice. Although the role of each plant hormone in promoting the longitudinal growth of plants has been extensively studied using genetic methods, their relationship is still poorly understood. In this study, we used two specific inhibitors targeting BR and GA biosynthesis to investigate the roles of BR and GA in growth of rice seedlings. Yucaizol, a specific inhibitor of BR biosynthesis, and Trinexapac-ethyl, a commercially available inhibitor of GA biosynthesis, were used. The effect of Yucaizol on rice seedlings indicated that Yucaizol significantly retarded stem elongation. The IC50 value was found to be approximately 0.8 μmol/L. Yucaizol also induced small leaf angle phenocopy in rice seedlings, similarly to BR-deficient rice, while Trinexapac-ethyl did not. When Yucaizol combined with Trinexapac-ethyl was applied to the rice plants, the mixture of these two inhibitors retarded stem elongation of rice at lower doses. Our results suggest that the use of a BR biosynthesis inhibitor combined with a GA biosynthesis inhibitor may be useful in the development of new technologies for controlling rice plant height.

  13. Genome of wild olive and the evolution of oil biosynthesis.

    Science.gov (United States)

    Unver, Turgay; Wu, Zhangyan; Sterck, Lieven; Turktas, Mine; Lohaus, Rolf; Li, Zhen; Yang, Ming; He, Lijuan; Deng, Tianquan; Escalante, Francisco Javier; Llorens, Carlos; Roig, Francisco J; Parmaksiz, Iskender; Dundar, Ekrem; Xie, Fuliang; Zhang, Baohong; Ipek, Arif; Uranbey, Serkan; Erayman, Mustafa; Ilhan, Emre; Badad, Oussama; Ghazal, Hassan; Lightfoot, David A; Kasarla, Pavan; Colantonio, Vincent; Tombuloglu, Huseyin; Hernandez, Pilar; Mete, Nurengin; Cetin, Oznur; Van Montagu, Marc; Yang, Huanming; Gao, Qiang; Dorado, Gabriel; Van de Peer, Yves

    2017-10-31

    Here we present the genome sequence and annotation of the wild olive tree ( Olea europaea var. sylvestris ), called oleaster, which is considered an ancestor of cultivated olive trees. More than 50,000 protein-coding genes were predicted, a majority of which could be anchored to 23 pseudochromosomes obtained through a newly constructed genetic map. The oleaster genome contains signatures of two Oleaceae lineage-specific paleopolyploidy events, dated at ∼28 and ∼59 Mya. These events contributed to the expansion and neofunctionalization of genes and gene families that play important roles in oil biosynthesis. The functional divergence of oil biosynthesis pathway genes, such as FAD2 , SACPD, EAR , and ACPTE , following duplication, has been responsible for the differential accumulation of oleic and linoleic acids produced in olive compared with sesame, a closely related oil crop. Duplicated oleaster FAD2 genes are regulated by an siRNA derived from a transposable element-rich region, leading to suppressed levels of FAD2 gene expression. Additionally, neofunctionalization of members of the SACPD gene family has led to increased expression of SACPD2 , 3 , 5 , and 7 , consequently resulting in an increased desaturation of steric acid. Taken together, decreased FAD2 expression and increased SACPD expression likely explain the accumulation of exceptionally high levels of oleic acid in olive. The oleaster genome thus provides important insights into the evolution of oil biosynthesis and will be a valuable resource for oil crop genomics.

  14. Vitamin B biosynthesis in plants.

    Science.gov (United States)

    Roje, Sanja

    2007-07-01

    The vitamin B complex comprises water-soluble enzyme cofactors and their derivatives that are essential contributors to diverse metabolic processes in plants as well as in animals and microorganisms. Seven vitamins form this complex: B1 (thiamin (1)), B2 (riboflavin (2)), B3 (niacin (3)), B5 (pantothenic acid (4)), B6 (pyridoxine, pyridoxal (5), and pyridoxamine), B8 (biotin (6)), and B9 (folate (7)). All seven B vitamins are required in the human diet for proper nutrition because humans lack enzymes to synthesize these compounds de novo. This review aims to summarize the present knowledge of vitamin B biosynthesis in plants.

  15. Transcriptomic variation in proanthocyanidin biosynthesis pathway genes in soybean (Glycine spp.).

    Science.gov (United States)

    Ha, Jungmin; Kim, Myoyeon; Kim, Moon Young; Lee, Taeyoung; Yoon, Min Young; Lee, Jayern; Lee, Yeong-Ho; Kang, Young-Gyu; Park, Jun Seong; Lee, John Hwan; Lee, Suk-Ha

    2018-04-01

    Proanthocyanidins are oligomeric or polymeric end products of flavonoid metabolic pathways starting with the central phenylpropanoid pathway. Although soybean (Glycine spp.) seeds represent a major source of nutrients for the human diet, as well as components for the cosmetics industry as a result of their high levels of flavonoid metabolites, including isoflavonoids, anthocyanins and proanthocyanidins, the genetic regulatory mechanisms underlying proanthocyanidin biosynthesis in soybean remain unclear. We evaluated interspecific and intraspecific variability in flavonoid components in soybean using 43 cultivars, landraces and wild soybean accessions. We performed transcriptomic profiling of genes encoding enzymes involved in flavonoid biosynthesis using three soybean genotypes, Hwangkeum (elite cultivar), IT109098 (landrace) and IT182932 (wild accession), in seeds. We identified a Glycine max landrace, IT109098, with a proanthocyanidin content as high as that of wild soybean. Different homologous genes for anthocyanidin reductase, which is involved in proanthocyanidin biosynthesis, were detected as differentially expressed genes between IT109098 and IT182932 compared to Hwangkeum. We detected major differences in the transcriptional levels of genes involved in the biosynthesis of proanthocyanidin and anthocyanin among genotypes beginning at the early stage of seed development. The results of the present study provide insights into the underlying genetic variation in proanthocyanidin biosynthesis among soybean genotypes. © 2017 Society of Chemical Industry. © 2017 Society of Chemical Industry.

  16. Solving the puzzles of cutin and suberin polymer biosynthesis.

    Science.gov (United States)

    Beisson, Fred; Li-Beisson, Yonghua; Pollard, Mike

    2012-06-01

    Cutin and suberin are insoluble lipid polymers that provide critical barrier functions to the cell wall of certain plant tissues, including the epidermis, endodermis and periderm. Genes that are specific to the biosynthesis of cutins and/or aliphatic suberins have been identified, mainly in Arabidopsis thaliana. They notably encode acyltransferases, oxidases and transporters, which may have either well-defined or more debatable biochemical functions. However, despite these advances, important aspects of cutin and suberin synthesis remain obscure. Central questions include whether fatty acyl monomers or oligomers are exported, and the extent of extracellular assembly and attachment to the cell wall. These issues are reviewed. Greater emphasis on chemistry and biochemistry will be required to solve these unknowns and link structure with function. Copyright © 2012 Elsevier Ltd. All rights reserved.

  17. Engineered biosynthesis of bacterial aromatic polyketides in Escherichia coli.

    Science.gov (United States)

    Zhang, Wenjun; Li, Yanran; Tang, Yi

    2008-12-30

    Bacterial aromatic polyketides are important therapeutic compounds including front line antibiotics and anticancer drugs. It is one of the last remaining major classes of natural products of which the biosynthesis has not been reconstituted in the genetically superior host Escherichia coli. Here, we demonstrate the engineered biosynthesis of bacterial aromatic polyketides in E. coli by using a dissected and reassembled fungal polyketide synthase (PKS). The minimal PKS of the megasynthase PKS4 from Gibberella fujikuroi was extracted by using two approaches. The first approach yielded a stand-alone Ketosynthase (KS)_malonyl-CoA:ACP transferase (MAT) didomain and an acyl-carrier protein (ACP) domain, whereas the second approach yielded a compact PKS (PKS_WJ) that consists of KS, MAT, and ACP on a single polypeptide. Both minimal PKSs produced nonfungal polyketides cyclized via different regioselectivity, whereas the fungal-specific C2-C7 cyclization mode was not observed. The kinetic properties of the two minimal PKSs were characterized to confirm both PKSs can synthesize polyketides with similar efficiency as the parent PKS4 megasynthase. Both minimal PKSs interacted effectively with exogenous polyketide cyclases as demonstrated by the synthesis of predominantly PK8 3 or NonaSEK4 6 in the presence of a C9-C14 or a C7-C12 cyclase, respectively. When PKS_WJ and downstream tailoring enzymes were expressed in E. coli, the expected nonaketide anthraquinone SEK26 was recovered in good titer. High-cell density fermentation was performed to demonstrate the scale-up potential of the in vivo platform for the biosynthesis of bacterial polyketides. Using engineered fungal PKSs can therefore be a general approach toward the heterologous biosynthesis of bacterial aromatic polyketides in E. coli.

  18. Starch Biosynthesis in the Developing Endosperms of Grasses and Cereals

    Directory of Open Access Journals (Sweden)

    Ian J. Tetlow

    2017-12-01

    Full Text Available The starch-rich endosperms of the Poaceae, which includes wild grasses and their domesticated descendents the cereals, have provided humankind and their livestock with the bulk of their daily calories since the dawn of civilization up to the present day. There are currently unprecedented pressures on global food supplies, largely resulting from population growth, loss of agricultural land that is linked to increased urbanization, and climate change. Since cereal yields essentially underpin world food and feed supply, it is critical that we understand the biological factors contributing to crop yields. In particular, it is important to understand the biochemical pathway that is involved in starch biosynthesis, since this pathway is the major yield determinant in the seeds of six out of the top seven crops grown worldwide. This review outlines the critical stages of growth and development of the endosperm tissue in the Poaceae, including discussion of carbon provision to the growing sink tissue. The main body of the review presents a current view of our understanding of storage starch biosynthesis, which occurs inside the amyloplasts of developing endosperms.

  19. Plant amino acid-derived vitamins: biosynthesis and function.

    Science.gov (United States)

    Miret, Javier A; Munné-Bosch, Sergi

    2014-04-01

    Vitamins are essential organic compounds for humans, having lost the ability to de novo synthesize them. Hence, they represent dietary requirements, which are covered by plants as the main dietary source of most vitamins (through food or livestock's feed). Most vitamins synthesized by plants present amino acids as precursors (B1, B2, B3, B5, B7, B9 and E) and are therefore linked to plant nitrogen metabolism. Amino acids play different roles in their biosynthesis and metabolism, either incorporated into the backbone of the vitamin or as amino, sulfur or one-carbon group donors. There is a high natural variation in vitamin contents in crops and its exploitation through breeding, metabolic engineering and agronomic practices can enhance their nutritional quality. While the underlying biochemical roles of vitamins as cosubstrates or cofactors are usually common for most eukaryotes, the impact of vitamins B and E in metabolism and physiology can be quite different on plants and animals. Here, we first aim at giving an overview of the biosynthesis of amino acid-derived vitamins in plants, with a particular focus on how this knowledge can be exploited to increase vitamin contents in crops. Second, we will focus on the functions of these vitamins in both plants and animals (and humans in particular), to unravel common and specific roles for vitamins in evolutionary distant organisms, in which these amino acid-derived vitamins play, however, an essential role.

  20. Synthesis and assessment of date palm genetic diversity studies

    Science.gov (United States)

    A thorough assessment of genetic diversity and population differentiation of Phoenix dactylifera are critical for its dynamic conservation and sustainable utilization of its genetic diversity. Estimates of genetic diversity based on phenotypic, biochemical and molecular markers; and fruit quality tr...

  1. Biosynthesis of bacterial aromatic polyketides.

    Science.gov (United States)

    Zhan, Jixun

    2009-01-01

    Aromatic polyketides represent important members of the family of polyketides, which have displayed a wide assortment of bioactive properties, such as antibacterial, antitumor, and antiviral activities. Bacterial aromatic polyketides are mainly synthesized by type II polyketide synthases (PKSs). Whereas malonyl-CoA is exclusively used as the extender unit, starter units can vary in different aromatic polyketide biosynthetic pathways, leading to a variety of polyketide backbones. Once the polyketide chains are elongated by the minimal PKSs to the full length, the immediate tailoring enzymes including ketoreductases, oxygenases and cyclases will work on the nascent chains to form aromatic structures, which will be further decorated by those late tailoring enzymes such as methyltransferases and glycosyltransferases. The mechanistic studies on the biosynthetic pathways of aromatic polyketides such as oxytetracycline and pradimicin A have been extensively carried out in recent years. Engineered biosynthesis of novel "unnatural" polyketides has been achieved in heterologous hosts such as Streptomyces coelicolor and Escherichia coli. This review covers the most recent advances in aromatic polyketide biosynthesis, which provide new enzymes or methods for building novel polyketide biosynthetic machinery.

  2. Genetic and perinatal effects of abused substances

    Energy Technology Data Exchange (ETDEWEB)

    Brande, M.C.; Zimmerman, A.M.

    1987-01-01

    This book provides an overview of the effects of several abused drugs, including opiates, cannabinoids, alcohol, nicotine, and cocaine, with special emphasis on the actions of these substances at the molecular and cellular levels. The first half deals with genetic effects, including molecular genetics, biochemical genetics, pharmacogenetics, cytogenetics, and genetic toxicity. The second half focuses on perinatal effects and covers: drug abuse during pregnancy; biochemical aspects of marihuana on male reproduction; and long-term behavioral and neuroendocrine effects of perinatal alcohol exposure.

  3. Direct Ionic Regulation of the Activity of Myo-Inositol Biosynthesis Enzymes in Mozambique Tilapia.

    Directory of Open Access Journals (Sweden)

    Fernando D Villarreal

    Full Text Available Myo-inositol (Ins is a major compatible osmolyte in many cells, including those of Mozambique tilapia (Oreochromis mossambicus. Ins biosynthesis is highly up-regulated in tilapia and other euryhaline fish exposed to hyperosmotic stress. In this study, enzymatic regulation of two enzymes of Ins biosynthesis, Ins phosphate synthase (MIPS and inositol monophosphatase (IMPase, by direct ionic effects is analyzed. Specific MIPS and IMPase isoforms from Mozambique tilapia (MIPS-160 and IMPase 1 were selected based on experimental, phylogenetic, and structural evidence supporting their role for Ins biosynthesis during hyperosmotic stress. Recombinant tilapia IMPase 1 and MIPS-160 activity was assayed in vitro at ionic conditions that mimic changes in the intracellular milieu during hyperosmotic stress. The in vitro activities of MIPS-160 and IMPase 1 are highest at alkaline pH of 8.8. IMPase 1 catalytic efficiency is strongly increased during hyperosmolality (particularly for the substrate D-Ins-3-phosphate, Ins-3P, mainly as a result of [Na+] elevation. Furthermore, the substrate-specificity of IMPase 1 towards D-Ins-1-phosphate (Ins-1P is lower than towards Ins-3P. Because MIPS catalysis results in Ins-3P this results represents additional evidence for IMPase 1 being the isoform that mediates Ins biosynthesis in tilapia. Our data collectively demonstrate that the Ins biosynthesis enzymes are activated under ionic conditions that cells are exposed to during hypertonicity, resulting in Ins accumulation, which, in turn, results in restoration of intracellular ion homeostasis. We propose that the unique and direct ionic regulation of the activities of Ins biosynthesis enzymes represents an efficient biochemical feedback loop for regulation of intracellular physiological ion homeostasis during hyperosmotic stress.

  4. Direct Ionic Regulation of the Activity of Myo-Inositol Biosynthesis Enzymes in Mozambique Tilapia.

    Science.gov (United States)

    Villarreal, Fernando D; Kültz, Dietmar

    2015-01-01

    Myo-inositol (Ins) is a major compatible osmolyte in many cells, including those of Mozambique tilapia (Oreochromis mossambicus). Ins biosynthesis is highly up-regulated in tilapia and other euryhaline fish exposed to hyperosmotic stress. In this study, enzymatic regulation of two enzymes of Ins biosynthesis, Ins phosphate synthase (MIPS) and inositol monophosphatase (IMPase), by direct ionic effects is analyzed. Specific MIPS and IMPase isoforms from Mozambique tilapia (MIPS-160 and IMPase 1) were selected based on experimental, phylogenetic, and structural evidence supporting their role for Ins biosynthesis during hyperosmotic stress. Recombinant tilapia IMPase 1 and MIPS-160 activity was assayed in vitro at ionic conditions that mimic changes in the intracellular milieu during hyperosmotic stress. The in vitro activities of MIPS-160 and IMPase 1 are highest at alkaline pH of 8.8. IMPase 1 catalytic efficiency is strongly increased during hyperosmolality (particularly for the substrate D-Ins-3-phosphate, Ins-3P), mainly as a result of [Na+] elevation. Furthermore, the substrate-specificity of IMPase 1 towards D-Ins-1-phosphate (Ins-1P) is lower than towards Ins-3P. Because MIPS catalysis results in Ins-3P this results represents additional evidence for IMPase 1 being the isoform that mediates Ins biosynthesis in tilapia. Our data collectively demonstrate that the Ins biosynthesis enzymes are activated under ionic conditions that cells are exposed to during hypertonicity, resulting in Ins accumulation, which, in turn, results in restoration of intracellular ion homeostasis. We propose that the unique and direct ionic regulation of the activities of Ins biosynthesis enzymes represents an efficient biochemical feedback loop for regulation of intracellular physiological ion homeostasis during hyperosmotic stress.

  5. Biochemical Hypermedia: Galactose Metabolism.

    Directory of Open Access Journals (Sweden)

    J.K. Sugai

    2013-05-01

    Full Text Available Introduction: Animations of biochemical processes and virtual laboratory environments lead to true molecular simulations. The use of interactive software’s in education can improve cognitive capacity, better learning and, mainly, it makes information acquisition easier. Material and Methods: This work presents the development of a biochemical hypermedia to understanding of the galactose metabolism. It was developed with the help of concept maps, ISIS Draw, ADOBE Photoshop and FLASH MX Program. Results and Discussion: A step by step animation process shows the enzymatic reactions of galactose conversion to glucose-1-phosphate (to glycogen synthesis, glucose-6-phosphate (glycolysis intermediary, UDP-galactose (substrate to mucopolysaccharides synthesis and collagen’s glycosylation. There are navigation guide that allow scrolling the mouse over the names of the components of enzymatic reactions of via the metabolism of galactose. Thus, explanatory text box, chemical structures and animation of the actions of enzymes appear to navigator. Upon completion of the module, the user’s response to the proposed exercise can be checked immediately through text box with interactive content of the answer. Conclusion: This hypermedia was presented for undergraduate students (UFSC who revealed that it was extremely effective in promoting the understanding of the theme.

  6. Regulatory variability of camalexin biosynthesis.

    Science.gov (United States)

    Schuhegger, Regina; Rauhut, Thomas; Glawischnig, Erich

    2007-05-01

    The anthranilate synthase ASA1, CYP79B2 and CYP71B15 (PAD3) are biosynthetic genes of the Arabidopsis phytoalexin camalexin, which are induced after pathogen infection and abiotic treatments like silver nitrate spraying. The natural variation of camalexin biosynthesis in response to Pseudomonas syringae infection was determined in several ecotypes, and differential CYP71B15 regulation as a potential basis for this variation was investigated. The expression of camalexin biosynthetic genes was restricted to the tissue undergoing cell death. After droplet infection with Alternaria alternata, a potent camalexin inducer in the Col-0 ecotype, camalexin formation and the induction of ASA1, CYP79B2 and CYP71B15 were strictly co-localized with the infection site.

  7. Diphthamide biosynthesis requires an organic radical generated by an iron-sulphur enzyme

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Yang; Zhu, Xuling; Torelli, Andrew T; Lee, Michael; Dzikovski, Boris; Koralewski, Rachel M; Wang, Eileen; Freed, Jack; Krebs, Carsten; Ealick, Steve E; Lin, Hening [Cornell; (Penn)

    2010-08-30

    Archaeal and eukaryotic translation elongation factor 2 contain a unique post-translationally modified histidine residue called diphthamide, which is the target of diphtheria toxin. The biosynthesis of diphthamide was proposed to involve three steps, with the first being the formation of a C-C bond between the histidine residue and the 3-amino-3-carboxypropyl group of S-adenosyl-l-methionine (SAM). However, further details of the biosynthesis remain unknown. Here we present structural and biochemical evidence showing that the first step of diphthamide biosynthesis in the archaeon Pyrococcus horikoshii uses a novel iron-sulphur-cluster enzyme, Dph2. Dph2 is a homodimer and each of its monomers can bind a [4Fe-4S] cluster. Biochemical data suggest that unlike the enzymes in the radical SAM superfamily, Dph2 does not form the canonical 5'-deoxyadenosyl radical. Instead, it breaks the Cγ,Met-S bond of SAM and generates a 3-amino-3-carboxypropyl radical. Our results suggest that P. horikoshii Dph2 represents a previously unknown, SAM-dependent, [4Fe-4S]-containing enzyme that catalyses unprecedented chemistry.

  8. The biochemical womb of schizophrenia: A review.

    Science.gov (United States)

    Gaur, N; Gautam, S; Gaur, M; Sharma, P; Dadheech, G; Mishra, S

    2008-10-01

    The conclusive identification of specific etiological factors or pathogenic processes in the illness of schizophrenia has remained elusive despite great technological progress. The convergence of state-of-art scientific studies in molecular genetics, molecular neuropathophysiology, in vivo brain imaging and psychopharmacology, however, indicates that we may be coming much closer to understanding the genesis of schizophrenia. In near future, the diagnosis and assessment of schizophrenia using biochemical markers may become a "dream come true" for the medical community as well as for the general population. An understanding of the biochemistry/ visa vis pathophysiology of schizophrenia is essential to the discovery of preventive measures and therapeutic intervention.

  9. The Arabidopsis transcription factor ANAC032 represses anthocyanin biosynthesis in response to high sucrose and oxidative and abiotic stresses

    Directory of Open Access Journals (Sweden)

    Kashif Mahmood

    2016-10-01

    Full Text Available Production of anthocyanins is one of the adaptive responses employed by plants during stress conditions. During stress, anthocyanin biosynthesis is mainly regulated at the transcriptional level via a complex interplay between activators and repressors of anthocyanin biosynthesis genes. In this study, we investigated the role of a NAC transcription factor, ANAC032, in the regulation of anthocyanin biosynthesis during stress conditions. ANAC032 expression was found to be induced by exogenous sucrose as well as high light stress. Using biochemical, molecular and transgenic approaches, we show that ANAC032 represses anthocyanin biosynthesis in response to sucrose treatment, high light and oxidative stress. ANAC032 was found to negatively affect anthocyanin accumulation and the expression of anthocyanin biosynthesis (DFR, ANS/LDOX and positive regulatory (TT8 genes as demonstrated in overexpression line (35S:ANAC032 compared to wild-type under high light stress. The chimeric repressor line (35S:ANAC032-SRDX exhibited the opposite expression patterns for these genes. The negative impact of ANAC032 on the expression of DFR, ANS/LDOX and TT8 was found to be correlated with the altered expression of negative regulators of anthocyanin biosynthesis, AtMYBL2 and SPL9. In addition to this, ANAC032 also repressed the MeJA- and ABA-induced anthocyanin biosynthesis. As a result, transgenic lines overexpressing ANAC032 (35S:ANAC032 produced drastically reduced levels of anthocyanin pigment compared to wild-type when challenged with salinity stress. However, transgenic chimeric repressor lines (35S:ANAC032-SRDX exhibited the opposite phenotype. Our results suggest that ANAC032 functions as a negative regulator of anthocyanin biosynthesis in Arabidopsis thaliana during stress conditions.

  10. The Arabidopsis Transcription Factor ANAC032 Represses Anthocyanin Biosynthesis in Response to High Sucrose and Oxidative and Abiotic Stresses.

    Science.gov (United States)

    Mahmood, Kashif; Xu, Zhenhua; El-Kereamy, Ashraf; Casaretto, José A; Rothstein, Steven J

    2016-01-01

    Production of anthocyanins is one of the adaptive responses employed by plants during stress conditions. During stress, anthocyanin biosynthesis is mainly regulated at the transcriptional level via a complex interplay between activators and repressors of anthocyanin biosynthesis genes. In this study, we investigated the role of a NAC transcription factor, ANAC032, in the regulation of anthocyanin biosynthesis during stress conditions. ANAC032 expression was found to be induced by exogenous sucrose as well as high light (HL) stress. Using biochemical, molecular and transgenic approaches, we show that ANAC032 represses anthocyanin biosynthesis in response to sucrose treatment, HL and oxidative stress. ANAC032 was found to negatively affect anthocyanin accumulation and the expression of anthocyanin biosynthesis ( DFR, ANS/LDOX) and positive regulatory ( TT8) genes as demonstrated in overexpression line (35S:ANAC032) compared to wild-type under HL stress. The chimeric repressor line (35S:ANAC032-SRDX) exhibited the opposite expression patterns for these genes. The negative impact of ANAC032 on the expression of DFR, ANS/LDOX and TT8 was found to be correlated with the altered expression of negative regulators of anthocyanin biosynthesis, AtMYBL2 and SPL9 . In addition to this, ANAC032 also repressed the MeJA- and ABA-induced anthocyanin biosynthesis. As a result, transgenic lines overexpressing ANAC032 (35S:ANAC032) produced drastically reduced levels of anthocyanin pigment compared to wild-type when challenged with salinity stress. However, transgenic chimeric repressor lines (35S:ANAC032-SRDX) exhibited the opposite phenotype. Our results suggest that ANAC032 functions as a negative regulator of anthocyanin biosynthesis in Arabidopsis thaliana during stress conditions.

  11. An Unusual Protector-Protégé Strategy for the Biosynthesis of Purine Nucleoside Antibiotics.

    Science.gov (United States)

    Wu, Pan; Wan, Dan; Xu, Gudan; Wang, Gui; Ma, Hongmin; Wang, Tingting; Gao, Yaojie; Qi, Jianzhao; Chen, Xiaoxia; Zhu, Jian; Li, Yong-Quan; Deng, Zixin; Chen, Wenqing

    2017-02-16

    Pentostatin (PTN, deoxycoformycin) and arabinofuranosyladenine (Ara-A, vidarabine) are purine nucleoside antibiotics used clinically to treat hematological cancers and human DNA virus infections, respectively. PTN has a 1,3-diazepine ring, and Ara-A is an adenosine analog with an intriguing epimerization at the C-2' hydroxyl group. However, the logic underlying the biosynthesis of these interesting molecules has long remained elusive. Here, we report that the biosynthesis of PTN and Ara-A employs an unusual protector-protégé strategy. To our surprise, we determined that a single gene cluster governs PTN and Ara-A biosynthesis via two independent pathways. Moreover, we verified that PenB functions as a reversible oxidoreductase for the final step of PTN. Remarkably, we provided the first direct biochemical evidence that PTN can protect Ara-A from deamination by selective inhibition of the host adenosine deaminase. These findings expand our knowledge of natural product biosynthesis and open the way for target-directed genome mining of Ara-A/PTN-related antibiotics. Copyright © 2017 Elsevier Ltd. All rights reserved.

  12. Agrobacterium mediated transient gene silencing (AMTS in Stevia rebaudiana: insights into steviol glycoside biosynthesis pathway.

    Directory of Open Access Journals (Sweden)

    Praveen Guleria

    Full Text Available Steviol glycoside biosynthesis pathway has emerged as bifurcation from ent-kaurenoic acid, substrate of methyl erythritol phosphate pathway that also leads to gibberellin biosynthesis. However, the genetic regulation of steviol glycoside biosynthesis has not been studied. So, in present study RNA interference (RNAi based Agrobacterium mediated transient gene silencing (AMTS approach was followed. SrKA13H and three SrUGTs (SrUGT85C2, SrUGT74G1 and SrUGT76G1 genes encoding ent-kaurenoic acid-13 hydroxylase and three UDP glycosyltransferases of steviol glycoside biosynthesis pathway were silenced in Stevia rebaudiana to understand its molecular mechanism and association with gibberellins.RNAi mediated AMTS of SrKA13H and three SrUGTs has significantly reduced the expression of targeted endogenous genes as well as total steviol glycoside accumulation. While gibberellins (GA3 content was significantly enhanced on AMTS of SrUGT85C2 and SrKA13H. Silencing of SrKA13H and SrUGT85C2 was found to block the metabolite flux of steviol glycoside pathway and shifted it towards GA3 biosynthesis. Further, molecular docking of three SrUGT proteins has documented highest affinity of SrUGT76G1 for the substrates of alternate pathways synthesizing steviol glycosides. This could be a plausible reason for maximum reduction in steviol glycoside content on silencing of SrUGT76G1 than other genes.SrKA13H and SrUGT85C2 were identified as regulatory genes influencing carbon flux between steviol glycoside and gibberellin biosynthesis. This study has also documented the existence of alternate steviol glycoside biosynthesis route.

  13. Agrobacterium Mediated Transient Gene Silencing (AMTS) in Stevia rebaudiana: Insights into Steviol Glycoside Biosynthesis Pathway

    Science.gov (United States)

    Guleria, Praveen; Yadav, Sudesh Kumar

    2013-01-01

    Background Steviol glycoside biosynthesis pathway has emerged as bifurcation from ent-kaurenoic acid, substrate of methyl erythritol phosphate pathway that also leads to gibberellin biosynthesis. However, the genetic regulation of steviol glycoside biosynthesis has not been studied. So, in present study RNA interference (RNAi) based Agrobacterium mediated transient gene silencing (AMTS) approach was followed. SrKA13H and three SrUGTs (SrUGT85C2, SrUGT74G1 and SrUGT76G1) genes encoding ent-kaurenoic acid-13 hydroxylase and three UDP glycosyltransferases of steviol glycoside biosynthesis pathway were silenced in Stevia rebaudiana to understand its molecular mechanism and association with gibberellins. Methodology/Principal Findings RNAi mediated AMTS of SrKA13H and three SrUGTs has significantly reduced the expression of targeted endogenous genes as well as total steviol glycoside accumulation. While gibberellins (GA3) content was significantly enhanced on AMTS of SrUGT85C2 and SrKA13H. Silencing of SrKA13H and SrUGT85C2 was found to block the metabolite flux of steviol glycoside pathway and shifted it towards GA3 biosynthesis. Further, molecular docking of three SrUGT proteins has documented highest affinity of SrUGT76G1 for the substrates of alternate pathways synthesizing steviol glycosides. This could be a plausible reason for maximum reduction in steviol glycoside content on silencing of SrUGT76G1 than other genes. Conclusions SrKA13H and SrUGT85C2 were identified as regulatory genes influencing carbon flux between steviol glycoside and gibberellin biosynthesis. This study has also documented the existence of alternate steviol glycoside biosynthesis route. PMID:24023961

  14. Genetic structure and origin of a tetraploid toad species Bufo danatensis Pisanetz, 1978 (Amphibia, Bufonidae) from central Asia: Description of biochemical polymorphism and comparison of heterozygosity levels in diploid and tetraploid species

    Energy Technology Data Exchange (ETDEWEB)

    Mezhzherin, S.V.; Pisanets, E.M. [Schmalhausen Institute of Zoology, Kiev (Ukraine)

    1995-01-01

    Comparison of individual variation at 24 biochemical loci in members of the species complex of Palearctic green toads showed that the heterozygosity of the tetraploid species Bufo danatensis (H{sub obs} = 0.45) was significantly higher than that of the diploid species B. viridis, B. sp., and B. raddei (H{sub obs} = 0.009 - 0.103). Such difference can be explained only by a hybrid origin of the tetraploid species. Individual electrophoretic variability of the polyploid toad species is associated with an allelic variation that is manifested in constantly heterozygous spectra as the gene dosage effect. At the population level, this phenomenon found in Pamir toads is caused by irregular meiosis in founders of the population or by directional changes in gene regulation. Genotypic distributions in zones of contact of the diploid and tetraploid taxons demonstrate the possibility of restricted introgressive hybridization.

  15. Metabolite and light regulation of metabolism in plants: lessons from the study of a single biochemical pathway

    Directory of Open Access Journals (Sweden)

    I.C. Oliveira

    2001-05-01

    Full Text Available We are using molecular, biochemical, and genetic approaches to study the structural and regulatory genes controlling the assimilation of inorganic nitrogen into the amino acids glutamine, glutamate, aspartate and asparagine. These amino acids serve as the principal nitrogen-transport amino acids in most crop and higher plants including Arabidopsis thaliana. We have begun to investigate the regulatory mechanisms controlling nitrogen assimilation into these amino acids in plants using molecular and genetic approaches in Arabidopsis. The synthesis of the amide amino acids glutamine and asparagine is subject to tight regulation in response to environmental factors such as light and to metabolic factors such as sucrose and amino acids. For instance, light induces the expression of glutamine synthetase (GLN2 and represses expression of asparagine synthetase (ASN1 genes. This reciprocal regulation of GLN2 and ASN1 genes by light is reflected at the level of transcription and at the level of glutamine and asparagine biosynthesis. Moreover, we have shown that the regulation of these genes is also reciprocally controlled by both organic nitrogen and carbon metabolites. We have recently used a reverse genetic approach to study putative components of such metabolic sensing mechanisms in plants that may be conserved in evolution. These components include an Arabidopsis homolog for a glutamate receptor gene originally found in animal systems and a plant PII gene, which is a homolog of a component of the bacterial Ntr system. Based on our observations on the biology of both structural and regulatory genes of the nitrogen assimilatory pathway, we have developed a model for metabolic control of the genes involved in the nitrogen assimilatory pathway in plants.

  16. RESULTS OF THE SELECTION OF BREEDING SAMPLES OF CARROT BASED ON BIOCHEMICAL COMPOSITION

    Directory of Open Access Journals (Sweden)

    V. K. Cherkasova

    2014-01-01

    Full Text Available 12 samples of carrot were analyzed for biochemical components in roots. 5 genotypes with high content of vitamin C, β-carotene, and total sugar were selected as genetic sources of high biochemical components.

  17. Biotechnology for a renewable resources chemicals and fuels industry, biochemical engineering R and D

    Energy Technology Data Exchange (ETDEWEB)

    Villet, R.H.

    1980-04-01

    To establish an effective biotechnology of biomass processing for the production of fuels and chemicals, an integration of research in biochemical engineering, microbial genetics, and biochemistry is required. Reduction of the costs of producing chemicals and fuels from renewable resources will hinge on extensive research in biochemical engineering.

  18. The transcriptome of Populus in elevated CO2 reveals increased anthocyanin biosynthesis during delayed autumnal senescence

    Energy Technology Data Exchange (ETDEWEB)

    Tallis, M.J.; Rogers, A.; Lin, Y.; Zhang, J.; Street, N. R.; Miglietta, F.; Karnosky, D. F.; Angelis, P. D.; Calfapietra, C.; Taylor, G.

    2010-03-01

    The delay in autumnal senescence that has occurred in recent decades has been linked to rising temperatures. Here, we suggest that increasing atmospheric CO{sub 2} may partly account for delayed autumnal senescence and for the first time, through transcriptome analysis, identify gene expression changes associated with this delay. Using a plantation of Populus x euramericana grown in elevated [CO{sub 2}] (e[CO{sub 2}]) with free-air CO{sub 2} enrichment (FACE) technology, we investigated the molecular and biochemical basis of this response. A Populus cDNA microarray was used to identify genes representing multiple biochemical pathways influenced by e[CO{sub 2}] during senescence. Gene expression changes were confirmed through real-time quantitative PCR, and leaf biochemical assays. Pathways for secondary metabolism and glycolysis were significantly up-regulated by e[CO{sub 2}] during senescence, in particular, those related to anthocyanin biosynthesis. Expressed sequence tags (ESTs) representing the two most significantly up-regulated transcripts in e[CO{sub 2}], LDOX (leucoanthocyanidin dioxgenase) and DFR (dihydroflavonol reductase), gave (e[CO{sub 2}]/ambient CO{sub 2} (a[CO{sub 2}])) expression ratios of 39.6 and 19.3, respectively. We showed that in e[CO{sub 2}] there was increased autumnal leaf sugar accumulation and up-regulation of genes determining anthocyanin biosynthesis which, we propose, prolongs leaf longevity during natural autumnal senescence.

  19. The pomegranate (Punica granatum L.) genome and the genomics of punicalagin biosynthesis.

    Science.gov (United States)

    Qin, Gaihua; Xu, Chunyan; Ming, Ray; Tang, Haibao; Guyot, Romain; Kramer, Elena M; Hu, Yudong; Yi, Xingkai; Qi, Yongjie; Xu, Xiangyang; Gao, Zhenghui; Pan, Haifa; Jian, Jianbo; Tian, Yinping; Yue, Zhen; Xu, Yiliu

    2017-09-01

    Pomegranate (Punica granatum L.) is a perennial fruit crop grown since ancient times that has been planted worldwide and is known for its functional metabolites, particularly punicalagins. We have sequenced and assembled the pomegranate genome with 328 Mb anchored into nine pseudo-chromosomes and annotated 29 229 gene models. A Myrtales lineage-specific whole-genome duplication event was detected that occurred in the common ancestor before the divergence of pomegranate and Eucalyptus. Repetitive sequences accounted for 46.1% of the assembled genome. We found that the integument development gene INNER NO OUTER (INO) was under positive selection and potentially contributed to the development of the fleshy outer layer of the seed coat, an edible part of pomegranate fruit. The genes encoding the enzymes for synthesis and degradation of lignin, hemicelluloses and cellulose were also differentially expressed between soft- and hard-seeded varieties, reflecting differences in their accumulation in cultivars differing in seed hardness. Candidate genes for punicalagin biosynthesis were identified and their expression patterns indicated that gallic acid synthesis in tissues could follow different biochemical pathways. The genome sequence of pomegranate provides a valuable resource for the dissection of many biological and biochemical traits and also provides important insights for the acceleration of breeding. Elucidation of the biochemical pathway(s) involved in punicalagin biosynthesis could assist breeding efforts to increase production of this bioactive compound. © 2017 The Authors The Plant Journal © 2017 John Wiley & Sons Ltd.

  20. Monoterpene biosynthesis potential of plant subcellular compartments

    NARCIS (Netherlands)

    Dong, L.; Jongedijk, E.J.; Bouwmeester, H.J.; Krol, van der A.R.

    2016-01-01

    Subcellular monoterpene biosynthesis capacity based on local geranyl diphosphate (GDP) availability or locally boosted GDP production was determined for plastids, cytosol and mitochondria. A geraniol synthase (GES) was targeted to plastids, cytosol, or mitochondria. Transient expression in Nicotiana

  1. Antibacterial Targets in Fatty Acid Biosynthesis

    Science.gov (United States)

    Wright, H. Tonie; Reynolds, Kevin A.

    2008-01-01

    Summary The fatty acid biosynthesis pathway is an attractive but still largely unexploited target for development of new anti-bacterial agents. The extended use of the anti-tuberculosis drug isoniazid and the antiseptic triclosan, which are inhibitors of fatty acid biosynthesis, validates this pathway as a target for anti-bacterial development. Differences in subcellular organization of the bacterial and eukaryotic multi-enzyme fatty acid synthase systems offer the prospect of inhibitors with host vs. target specificity. Platensimycin, platencin, and phomallenic acids, newly discovered natural product inhibitors of the condensation steps in fatty acid biosynthesis, represent new classes of compounds with antibiotic potential. An almost complete catalogue of crystal structures for the enzymes of the type II fatty acid biosynthesis pathway can now be exploited in the rational design of new inhibitors, as well as the recently published crystal structures of type I FAS complexes. PMID:17707686

  2. Lincomycin, cultivation of producing strains and biosynthesis

    Czech Academy of Sciences Publication Activity Database

    Spížek, Jaroslav; Řezanka, Tomáš

    2004-01-01

    Roč. 63, - (2004), s. 510-519 ISSN 0175-7598 Institutional research plan: CEZ:AV0Z5020903 Keywords : lincomycin * cultivation * biosynthesis Subject RIV: EE - Microbiology, Virology Impact factor: 2.358, year: 2004

  3. Control of tylosin biosynthesis in Streptomyces fradiae.

    Science.gov (United States)

    Cundliffe, Eric

    2008-09-01

    Tylosin biosynthesis is controlled in cascade fashion by multiple transcriptional regulators, acting positively or negatively, in conjunction with a signalling ligand that acts as a classical inducer. The roles of regulatory gene products have been characterized by a combination of gene expression analysis and fermentation studies, using engineered strains of S. fradiae in which specific genes were inactivated or overexpressed. Among various novel features of the regulatory model, involvement of the signalling ligand is not essential for tylosin biosynthesis.

  4. Overcoming challenges in engineering the genetic code

    OpenAIRE

    Lajoie, MJ; Söll, D; Church, GM

    2015-01-01

    Withstanding 3.5 billion years of genetic drift, the canonical genetic code remains such a fundamental foundation for the complexity of life that it is highly conserved across all three phylogenetic domains. Genome engineering technologies are now making it possible to rationally change the genetic code, offering resistance to viruses, genetic isolation from horizontal gene transfer, and prevention of environmental escape by genetically modified organisms. We discuss the biochemical, genetic,...

  5. Fitness effects of fluctuations in biochemical networks

    Science.gov (United States)

    Tanase-Nicola, Sorin

    2009-03-01

    The concentration of many cellular components fluctuates not only as a response to external and internal inputs but also due to random birth and death events of individual molecules. This biochemical noise affects the capacity of every individual cell in a population to respond and adapt to the environment. While the sources and effects of biochemical fluctuations on individual cells have been intensively studied, the effects of noise on the growth rate of a population of cells are much less understood. We present a model of the cell cycle in which the growth and division of individual cells are coupled with the noisy dynamics of their internal components. The model allows us to compute the contribution of the biochemical noise to the average growth rate of a population of cells as a function of the noise strength and the correlation time of the fluctuations. We show that, due to fluctuations, the growth rate of a population of cells is always larger than the average growth rate of a individual cell and can be larger even than a corresponding deterministic model. In most relevant cases it is assumed that the average concentration of a cellular component is close to a value that maximizes the population growth as given by the external, environmental, conditions and the internal cellular regulation. In such cases we show that contribution of fluctuations to the growth rate is negative and increases with the sensitivity of the biochemical networks to the noise sources and the noise correlation time. We also discuss how the selection pressure due to fluctuations affects the structure and parameters of genetic regulatory networks.

  6. Revealing complexity and specificity in the activation of lipase-mediated oxylipin biosynthesis: a specific role of the Nicotiana attenuata GLA1 lipase in the activation of jasmonic acid biosynthesis in leaves and roots.

    Science.gov (United States)

    Bonaventure, Gustavo; Schuck, Stefan; Baldwin, Ian T

    2011-09-01

    The activation of enzymatic oxylipin biosynthesis upon wounding, herbivory and pathogen attack depends on the biochemical activation of lipases that make polyunsaturated fatty acids (PUFAs) available to lipoxygenases (LOXs). The identity and number of the lipases involved in this process remain controversial and they probably differ among plant species. Analysis of transgenic Nicotiana attenuata plants (ir-gla1) stably reduced in the expression of the NaGLA1 gene showed that this plastidial glycerolipase is a major supplier of trienoic fatty acids for jasmonic acid (JA) biosynthesis in leaves and roots after wounding and simulated herbivory, but not during infection with the oomycete Phytophthora parasitica (var. nicotianae). NaGLA1 was not essential for the developmental control of JA biosynthesis in flowers and for the biosynthesis of C(6) volatiles by the hydroperoxide lyase (HPL) pathway; however, it affected the metabolism of divinyl ethers (DVEs) early during infection with P. parasitica (var. nicotianae) and the accumulation of NaDES1 and NaLOX1 mRNAs. Profiling of lysolipids by LC-MS/MS was consistent with a rapid activation of NaGLA1 and indicated that this lipase utilizes different lipid classes as substrates. The results revealed the complexity and specificity of the regulation of lipase-mediated oxylipin biosynthesis, highlighting the existence of pathway- and stimulus-specific lipases. © 2011 Blackwell Publishing Ltd.

  7. Advances in the genetic dissection of plant cell walls: tools and resources available in Miscanthus

    Directory of Open Access Journals (Sweden)

    Gancho eSlavov

    2013-07-01

    Full Text Available Tropical C4 grasses from the genus Miscanthus are believed to have great potential as biomass crops. However, Miscanthus species are essentially undomesticated, and genetic, molecular and bioinformatics tools are in very early stages of development. Furthermore, similar to other crops targeted as lignocellulosic feedstocks, the efficient utilisation of biomass is hampered by our limited knowledge of the structural organisation of the plant cell wall and the underlying genetic components that control this organisation. The Institute of Biological, Environmental and Rural Sciences (IBERS has assembled an extensive collection of germplasm for several species of Miscanthus. In addition, an integrated, multidisciplinary research programme at IBERS aims to inform accelerated breeding for biomass productivity and composition, while also generating novel fundamental knowledge. Here we review recent advances with respect to the genetic characterisation of the cell wall in Miscanthus. First, we present a summary of recent and on-going biochemical studies, including prospects and limitations for the development of powerful phenotyping approaches. Second, we review current knowledge about genetic variation for cell wall characteristics of Miscanthus and illustrate how phenotypic data, combined with high-density arrays of single nucleotide polymorphisms, are being used in genome-wide association studies to generate testable hypotheses and guide biological discovery. Finally, we provide an overview of the current knowledge about the molecular biology of cell wall biosynthesis in Miscanthus and closely related grasses, discuss the key conceptual and technological bottlenecks, and outline the short-term prospects for progress in this field.

  8. Advances in the genetic dissection of plant cell walls: tools and resources available in Miscanthus.

    Science.gov (United States)

    Slavov, Gancho; Allison, Gordon; Bosch, Maurice

    2013-01-01

    Tropical C4 grasses from the genus Miscanthus are believed to have great potential as biomass crops. However, Miscanthus species are essentially undomesticated, and genetic, molecular and bioinformatics tools are in very early stages of development. Furthermore, similar to other crops targeted as lignocellulosic feedstocks, the efficient utilization of biomass is hampered by our limited knowledge of the structural organization of the plant cell wall and the underlying genetic components that control this organization. The Institute of Biological, Environmental and Rural Sciences (IBERS) has assembled an extensive collection of germplasm for several species of Miscanthus. In addition, an integrated, multidisciplinary research programme at IBERS aims to inform accelerated breeding for biomass productivity and composition, while also generating fundamental knowledge. Here we review recent advances with respect to the genetic characterization of the cell wall in Miscanthus. First, we present a summary of recent and on-going biochemical studies, including prospects and limitations for the development of powerful phenotyping approaches. Second, we review current knowledge about genetic variation for cell wall characteristics of Miscanthus and illustrate how phenotypic data, combined with high-density arrays of single-nucleotide polymorphisms, are being used in genome-wide association studies to generate testable hypotheses and guide biological discovery. Finally, we provide an overview of the current knowledge about the molecular biology of cell wall biosynthesis in Miscanthus and closely related grasses, discuss the key conceptual and technological bottlenecks, and outline the short-term prospects for progress in this field.

  9. Biosynthesis of promatrix metalloproteinase-9/chondroitin sulphate proteoglycan heteromer involves a Rottlerin-sensitive pathway.

    Directory of Open Access Journals (Sweden)

    Nabin Malla

    Full Text Available BACKGROUND: Previously we have shown that a fraction of the matrix metalloproteinase-9 (MMP-9 synthesized by the macrophage cell line THP-1 was bound to a chondroitin sulphate proteoglycan (CSPG core protein as a reduction sensitive heteromer. Several biochemical properties of the enzyme were changed when it was bound to the CSPG. METHODOLOGY/PRINCIPAL FINDINGS: By use of affinity chromatography, zymography, and radioactive labelling, various macrophage stimulators were tested for their effect on the synthesis of the proMMP-9/CSPG heteromer and its components by THP-1 cells. Of the stimulators, only PMA largely increased the biosynthesis of the heteromer. As PMA is an activator of PKC, we determined which PKC isoenzymes were expressed by performing RT-PCR and Western Blotting. Subsequently specific inhibitors were used to investigate their involvement in the biosynthesis of the heteromer. Of the inhibitors, only Rottlerin repressed the biosynthesis of proMMP-9/CSPG and its two components. Much lower concentrations of Rottlerin were needed to reduce the amount of CSPG than what was needed to repress the synthesis of the heteromer and MMP-9. Furthermore, Rottlerin caused a minor reduction in the activation of the PKC isoenzymes δ, ε, θ and υ (PKD3 in both control and PMA exposed cells. CONCLUSIONS/SIGNIFICANCE: The biosynthesis of the proMMP-9/CSPG heteromer and proMMP-9 in THP-1 cells involves a Rottlerin-sensitive pathway that is different from the Rottlerin sensitive pathway involved in the CSPG biosynthesis. MMP-9 and CSPGs are known to be involved in various physiological and pathological processes. Formation of complexes may influence both the specificity and localization of the enzyme. Therefore, knowledge about biosynthetic pathways and factors involved in the formation of the MMP-9/CSPG heteromer may contribute to insight in the heteromers biological function as well as pointing to future targets for therapeutic agents.

  10. The Catalytic Diversity of Multimodular Polyketide Synthases: Natural Product Biosynthesis Beyond Textbook Assembly Rules.

    Science.gov (United States)

    Gulder, Tobias A M; Freeman, Michael F; Piel, Jörn

    2011-03-01

    Bacterial multimodular polyketide synthases (PKSs) are responsible for the biosynthesis of a wide range of pharmacologically active natural products. These megaenzymes contain numerous catalytic and structural domains and act as biochemical templates to generate complex polyketides in an assembly line-like fashion. While the prototypical PKS is composed of only a few different domain types that are fused together in a combinatorial fashion, an increasing number of enzymes is being found that contain additional components. These domains can introduce remarkably diverse modifications into polyketides. This review discusses our current understanding of such noncanonical domains and their role in expanding the biosynthetic versatility of bacterial PKSs.

  11. Genetics and developmental biology

    International Nuclear Information System (INIS)

    Barnett, W.E.

    1975-01-01

    Progress is reported on research activities in the fields of mutagenesis in Haemophilus influenzae and Escherichia coli; radioinduced chromosomal aberrations in mammalian germ cells; effects of uv radiation on xeroderma pigmentosum skin cells; mutations in Chinese hamster ovary cells; radioinduced hemoglobin variants in the mouse; analysis of mutants in yeast; Drosophila genetics; biochemical genetics of Neurospora; DNA polymerase activity in Xenopus laevis oocytes; uv-induced damage in Bacillus subtilis; and others

  12. Anthocyanin Biosynthesis and Degradation Mechanisms in Solanaceous Vegetables: A Review

    Science.gov (United States)

    Liu, Ying; Tikunov, Yury; Schouten, Rob E.; Marcelis, Leo F. M.; Visser, Richard G. F.; Bovy, Arnaud

    2018-01-01

    Anthocyanins are a group of polyphenolic pigments that are ubiquitously found in the plant kingdom. In plants, anthocyanins play a role not only in reproduction, by attracting pollinators and seed dispersers, but also in protection against various abiotic and biotic stresses. There is accumulating evidence that anthocyanins have health-promoting properties, which makes anthocyanin metabolism an interesting target for breeders and researchers. In this review, the state of the art knowledge concerning anthocyanins in the Solanaceous vegetables, i.e., pepper, tomato, eggplant, and potato, is discussed, including biochemistry and biological function of anthocyanins, as well as their genetic and environmental regulation. Anthocyanin accumulation is determined by the balance between biosynthesis and degradation. Although the anthocyanin biosynthetic pathway has been well-studied in Solanaceous vegetables, more research is needed on the inhibition of biosynthesis and, in particular, the anthocyanin degradation mechanisms if we want to control anthocyanin content of Solanaceous vegetables. In addition, anthocyanin metabolism is distinctly affected by environmental conditions, but the molecular regulation of these effects is poorly understood. Existing knowledge is summarized and current gaps in our understanding are highlighted and discussed, to create opportunities for the development of anthocyanin-rich crops through breeding and environmental management. PMID:29594099

  13. Anthocyanin Biosynthesis and Degradation Mechanisms in Solanaceous Vegetables: A Review

    Directory of Open Access Journals (Sweden)

    Ying Liu

    2018-03-01

    Full Text Available Anthocyanins are a group of polyphenolic pigments that are ubiquitously found in the plant kingdom. In plants, anthocyanins play a role not only in reproduction, by attracting pollinators and seed dispersers, but also in protection against various abiotic and biotic stresses. There is accumulating evidence that anthocyanins have health-promoting properties, which makes anthocyanin metabolism an interesting target for breeders and researchers. In this review, the state of the art knowledge concerning anthocyanins in the Solanaceous vegetables, i.e., pepper, tomato, eggplant, and potato, is discussed, including biochemistry and biological function of anthocyanins, as well as their genetic and environmental regulation. Anthocyanin accumulation is determined by the balance between biosynthesis and degradation. Although the anthocyanin biosynthetic pathway has been well-studied in Solanaceous vegetables, more research is needed on the inhibition of biosynthesis and, in particular, the anthocyanin degradation mechanisms if we want to control anthocyanin content of Solanaceous vegetables. In addition, anthocyanin metabolism is distinctly affected by environmental conditions, but the molecular regulation of these effects is poorly understood. Existing knowledge is summarized and current gaps in our understanding are highlighted and discussed, to create opportunities for the development of anthocyanin-rich crops through breeding and environmental management.

  14. Cloning and characterization of a norbelladine 4'-O-methyltransferase involved in the biosynthesis of the Alzheimer's drug galanthamine in Narcissus sp. aff. pseudonarcissus.

    Science.gov (United States)

    Kilgore, Matthew B; Augustin, Megan M; Starks, Courtney M; O'Neil-Johnson, Mark; May, Gregory D; Crow, John A; Kutchan, Toni M

    2014-01-01

    Galanthamine is an Amaryllidaceae alkaloid used to treat the symptoms of Alzheimer's disease. This compound is primarily isolated from daffodil (Narcissus spp.), snowdrop (Galanthus spp.), and summer snowflake (Leucojum aestivum). Despite its importance as a medicine, no genes involved in the biosynthetic pathway of galanthamine have been identified. This absence of genetic information on biosynthetic pathways is a limiting factor in the development of synthetic biology platforms for many important botanical medicines. The paucity of information is largely due to the limitations of traditional methods for finding biochemical pathway enzymes and genes in non-model organisms. A new bioinformatic approach using several recent technological improvements was applied to search for genes in the proposed galanthamine biosynthetic pathway, first targeting methyltransferases due to strong signature amino acid sequences in the proteins. Using Illumina sequencing, a de novo transcriptome assembly was constructed for daffodil. BLAST was used to identify sequences that contain signatures for plant O-methyltransferases in this transcriptome. The program HAYSTACK was then used to identify methyltransferases that fit a model for galanthamine biosynthesis in leaf, bulb and inflorescence tissues. One candidate gene for the methylation of norbelladine to 4'-O-methylnorbelladine in the proposed galanthamine biosynthetic pathway was identified. This methyltransferase cDNA was expressed in E. coli and the protein purified by affinity chromatography. The resulting protein was found to be a norbelladine 4'-O-methyltransferase (NpN4OMT) of the proposed galanthamine biosynthetic pathway.

  15. Cloning and characterization of a norbelladine 4'-O-methyltransferase involved in the biosynthesis of the Alzheimer's drug galanthamine in Narcissus sp. aff. pseudonarcissus.

    Directory of Open Access Journals (Sweden)

    Matthew B Kilgore

    Full Text Available Galanthamine is an Amaryllidaceae alkaloid used to treat the symptoms of Alzheimer's disease. This compound is primarily isolated from daffodil (Narcissus spp., snowdrop (Galanthus spp., and summer snowflake (Leucojum aestivum. Despite its importance as a medicine, no genes involved in the biosynthetic pathway of galanthamine have been identified. This absence of genetic information on biosynthetic pathways is a limiting factor in the development of synthetic biology platforms for many important botanical medicines. The paucity of information is largely due to the limitations of traditional methods for finding biochemical pathway enzymes and genes in non-model organisms. A new bioinformatic approach using several recent technological improvements was applied to search for genes in the proposed galanthamine biosynthetic pathway, first targeting methyltransferases due to strong signature amino acid sequences in the proteins. Using Illumina sequencing, a de novo transcriptome assembly was constructed for daffodil. BLAST was used to identify sequences that contain signatures for plant O-methyltransferases in this transcriptome. The program HAYSTACK was then used to identify methyltransferases that fit a model for galanthamine biosynthesis in leaf, bulb and inflorescence tissues. One candidate gene for the methylation of norbelladine to 4'-O-methylnorbelladine in the proposed galanthamine biosynthetic pathway was identified. This methyltransferase cDNA was expressed in E. coli and the protein purified by affinity chromatography. The resulting protein was found to be a norbelladine 4'-O-methyltransferase (NpN4OMT of the proposed galanthamine biosynthetic pathway.

  16. Genetic disorders of collagen.

    OpenAIRE

    Tsipouras, P; Ramirez, F

    1987-01-01

    Osteogenesis imperfecta, Ehlers-Danlos syndrome, and Marfan syndrome form a group of genetic disorders of connective tissue. These disorders exhibit remarkable clinical heterogeneity which reflects their underlying biochemical and molecular differences. Defects in collagen types I and III have been found in all three syndromes.

  17. Medical Genetics In Clinical Practice

    African Journals Online (AJOL)

    1974-08-24

    Aug 24, 1974 ... Laboratory facilities for cytogenetic and biochemical investigation are an essential feature of such a genetic department. The purpose of this article is to review the clinical activities during 1973 of the Department of Human. Genetics, University of Cape Town, to present an analysis of the medical problems ...

  18. Biochemical effect of chocolate colouring and flavouring like substances on thyroid function and protein biosynthesis.

    Science.gov (United States)

    el-Saadany, S S

    1991-01-01

    Synthetic chocolate colourant, flavourant and the mixture of both were administered to healthy adult male albino rats to evaluate their effect on the nucleic acids metabolism, i.e. deoxyribonucleic and ribonucleic acids (DNA and RNA), total serum protein, thyroid hormones (T4 and T3) and nuclease enzymes, i.e. cytoplasmic- and mitochondrial deoxyribonuclease and ribonuclease (DNase and RNase) in brain, liver, and kidneys. Also, the activity of the fundamental enzymes of the oxidative pentose phosphate pathway, i.e. cytoplasmic and mitochondrial glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase (G-6-PD and 6-PGD), as well as total lipids and cholesterol contents in the same organs were studied. Ingestion of the studied food additives significantly increased serum protein, RNA and T4 hormone, while, DNA and T3 hormone were insignificantly elevated. In connection with this, the hydrolytic enzymes of nucleic acids (DNase and RNase activities) were stimulated by all studied food additives and in all mentioned organs. The activity of G-6-PD and 6-PGD in both cytoplasmic and mitochondrial fractions of all studied organs were increased. The highest increase was noticed in rats fed on diets supplemented with the mixture of both colourant and flavourant followed by colourant then flavourant, respectively.

  19. Current Understanding on Aflatoxin Biosynthesis and Future Perspective in Reducing Aflatoxin Contamination

    OpenAIRE

    Jiujiang Yu

    2012-01-01

    Traditional molecular techniques have been used in research in discovering the genes and enzymes that are involved in aflatoxin formation and genetic regulation. We cloned most, if not all, of the aflatoxin pathway genes. A consensus gene cluster for aflatoxin biosynthesis was discovered in 2005. The factors that affect aflatoxin formation have been studied. In this report, the author summarized the current status of research progress and future possibilities that may be used for solving afla...

  20. Current Understanding on Aflatoxin Biosynthesis and Future Perspective in Reducing Aflatoxin Contamination

    Science.gov (United States)

    Yu, Jiujiang

    2012-01-01

    Traditional molecular techniques have been used in research in discovering the genes and enzymes that are involved in aflatoxin formation and genetic regulation. We cloned most, if not all, of the aflatoxin pathway genes. A consensus gene cluster for aflatoxin biosynthesis was discovered in 2005. The factors that affect aflatoxin formation have been studied. In this report, the author summarized the current status of research progress and future possibilities that may be used for solving aflatoxin contamination. PMID:23202305

  1. Evolution of the Phosphatidylcholine Biosynthesis Pathways in Green Algae: Combinatorial Diversity of Methyltransferases.

    Science.gov (United States)

    Hirashima, Takashi; Toyoshima, Masakazu; Moriyama, Takashi; Sato, Naoki

    2018-01-01

    Phosphatidylcholine (PC) is one of the most common phospholipids in eukaryotes, although some green algae such as Chlamydomonas reinhardtii are known to lack PC. Recently, we detected PC in four species in the genus Chlamydomonas: C. applanata NIES-2202, C. asymmetrica NIES-2207, C. debaryana NIES-2212, and C. sphaeroides NIES-2242. To reveal the PC biosynthesis pathways in green algae and the evolutionary scenario involved in their diversity, we analyzed the PC biosynthesis genes in these four algae using draft genome sequences. Homology searches suggested that PC in these species is synthesized by phosphoethanolamine-N-methyltransferase (PEAMT) and/or phosphatidylethanolamine-N-methyltransferase (PEMT), both of which are absent in C. reinhardtii. Recombinant PEAMTs from these algae showed methyltransferase activity for phosphoethanolamine but not for monomethyl phosphoethanolamine in vitro, in contrast to land plant PEAMT, which catalyzes the three methylations from phosphoethanolamine to phosphocholine. This suggested an involvement of other methyltransferases in PC biosynthesis. Here, we characterized the putative phospholipid-N-methyltransferase (PLMT) genes of these species by genetic and phylogenetic analysis. Complementation assays using a PC biosynthesis-deficient yeast suggested that the PLMTs of these algae can synthesize PC from phosphatidylethanolamine. These results indicated that the PC biosynthesis pathways in green algae differ from those of land plants, although the enzymes involved are homologous. Phylogenetic analysis suggested that the PEAMTs and PLMTs in these algae were inherited from the common ancestor of green algae. The absence of PC biosynthesis in many Chlamydomonas species is likely a result of parallel losses of PEAMT and PLMT in this genus.

  2. Dual Roles of Glutathione in Ecdysone Biosynthesis and Antioxidant Function During Larval Development in Drosophila.

    Science.gov (United States)

    Enya, Sora; Yamamoto, Chikana; Mizuno, Hajime; Esaki, Tsuyoshi; Lin, Hsin-Kuang; Iga, Masatoshi; Morohashi, Kana; Hirano, Yota; Kataoka, Hiroshi; Masujima, Tsutomu; Shimada-Niwa, Yuko; Niwa, Ryusuke

    2017-12-01

    hypothesis that a primary role of GSH in early D. melanogaster larval development is ecdysteroid biosynthesis, independent from the antioxidant role of GSH. Copyright © 2017 by the Genetics Society of America.

  3. Microbial biosynthesis of secondary metabolites involved in biocontrol

    DEFF Research Database (Denmark)

    Hennessy, Rosanna Catherine; Michelsen, Charlotte Frydenlund; Olsson, Stefan

    secondary metabolite biosynthesis gene clusters. A combination of random and targeted mutagenesis, together with MALDI-TOF imaging mass spectrometry, linked two non-ribosomal peptides (NRPs) designated nunapeptin and nunamycin respectively, to antifungal activity against Rhizoctonia solani, Pythium...... aphanidermatum and Fusarium graminearum1, 2. In order to unravel the complex genetic regulation of these large NRP synthetase gene clusters, antisense RNAs (asRNAs) and CRISPR/Cas9 based systems are being tested and developed as tools to target transcripts of interest and elucidate gene function3, 4....... To investigate the effect of purified nunamycin and nunapeptin at the omics level against pathogenic fungi, an NRP production platform is being developed which, could additionally provide a source of antifungal compounds for industrial applications (e.g. food production, pharmaceutical, personal care). Methods...

  4. Biosynthesis of therapeutic natural products using synthetic biology.

    Science.gov (United States)

    Awan, Ali R; Shaw, William M; Ellis, Tom

    2016-10-01

    Natural products are a group of bioactive structurally diverse chemicals produced by microorganisms and plants. These molecules and their derivatives have contributed to over a third of the therapeutic drugs produced in the last century. However, over the last few decades traditional drug discovery pipelines from natural products have become far less productive and far more expensive. One recent development with promise to combat this trend is the application of synthetic biology to therapeutic natural product biosynthesis. Synthetic biology is a young discipline with roots in systems biology, genetic engineering, and metabolic engineering. In this review, we discuss the use of synthetic biology to engineer improved yields of existing therapeutic natural products. We further describe the use of synthetic biology to combine and express natural product biosynthetic genes in unprecedented ways, and how this holds promise for opening up completely new avenues for drug discovery and production. Copyright © 2016 Elsevier B.V. All rights reserved.

  5. Ouroboros - Playing A Biochemical

    Directory of Open Access Journals (Sweden)

    D. T. Rodrigues

    2014-08-01

    Full Text Available Ouroboros: Playing A Biochemical RODRIGUES,D.T.1,2;GAYER, M.C.1,2; ESCOTO, D.F.1; DENARDIN, E.L.G.2, ROEHRS, R.1,2 1Interdisciplinary Research Group on Teaching Practice, Graduate Program in Biochemistry, Unipampa, RS, Brazil 2Laboratory of Physicochemical Studies and Natural Products, Post Graduate Program in Biochemistry, Unipampa, RS, Brazil Introduction: Currently, teachers seek different alternatives to enhance the teaching-learning process. Innovative teaching methodologies are increasingly common tools in educational routine. The use of games, electronic or conventional, is an effective tool to assist in learning and also to raise the social interaction between students. Objective: In this sense our work aims to evaluate the card game and "Ouroboros" board as a teaching and learning tool in biochemistry for a graduating class in Natural Sciences. Materials and methods: The class gathered 22 students of BSc in Natural Sciences. Each letter contained a question across the board that was drawn to a group to answer within the allotted time. The questions related concepts of metabolism, organic and inorganic chemical reactions, bioenergetics, etc.. Before the game application, students underwent a pre-test with four issues involving the content that was being developed. Soon after, the game was applied. Then again questions were asked. Data analysis was performed from the ratio of the number of correct pre-test and post-test answers. Results and discussion: In the pre-test 18.1% of the students knew all issues, 18.1% got 3 correct answers, 40.9% answered only 2 questions correctly and 22.7% did not hit any. In post-test 45.4% answered all the questions right, 31.8% got 3 questions and 22.7% got 2 correct answers. The results show a significant improvement of the students about the field of content taught through the game. Conclusion: Generally, traditional approaches of chemistry and biochemistry are abstract and complex. Thus, through games

  6. Sphingolipid biosynthesis upregulation by TOR complex 2-Ypk1 signaling during yeast adaptive response to acetic acid stress.

    Science.gov (United States)

    Guerreiro, Joana F; Muir, Alexander; Ramachandran, Subramaniam; Thorner, Jeremy; Sá-Correia, Isabel

    2016-12-01

    Acetic acid-induced inhibition of yeast growth and metabolism limits the productivity of industrial fermentation processes, especially when lignocellulosic hydrolysates are used as feedstock in industrial biotechnology. Tolerance to acetic acid of food spoilage yeasts is also a problem in the preservation of acidic foods and beverages. Thus understanding the molecular mechanisms underlying adaptation and tolerance to acetic acid stress is increasingly important in industrial biotechnology and the food industry. Prior genetic screens for Saccharomyces cerevisiae mutants with increased sensitivity to acetic acid identified loss-of-function mutations in the YPK1 gene, which encodes a protein kinase activated by the target of rapamycin (TOR) complex 2 (TORC2). We show in the present study by several independent criteria that TORC2-Ypk1 signaling is stimulated in response to acetic acid stress. Moreover, we demonstrate that TORC2-mediated Ypk1 phosphorylation and activation is necessary for acetic acid tolerance, and occurs independently of Hrk1, a protein kinase previously implicated in the cellular response to acetic acid. In addition, we show that TORC2-Ypk1-mediated activation of l-serine:palmitoyl-CoA acyltransferase, the enzyme complex that catalyzes the first committed step of sphingolipid biosynthesis, is required for acetic acid tolerance. Furthermore, analysis of the sphingolipid pathway using inhibitors and mutants indicates that it is production of certain complex sphingolipids that contributes to conferring acetic acid tolerance. Consistent with that conclusion, promoting sphingolipid synthesis by adding exogenous long-chain base precursor phytosphingosine to the growth medium enhanced acetic acid tolerance. Thus appropriate modulation of the TORC2-Ypk1-sphingolipid axis in industrial yeast strains may have utility in improving fermentations of acetic acid-containing feedstocks. © 2016 The Author(s); published by Portland Press Limited on behalf of the

  7. The Spatial Organization of Glucosinolate Biosynthesis

    DEFF Research Database (Denmark)

    Nintemann, Sebastian

    between the individual classes of glucosinolates under constitutive and induced conditions and identified the source tissues of these defense compounds. Protein-protein interaction studies were carried out to investigate the subcellular organization of glucosinolate biosynthesis. We identified a family...... resistance and nutritional value and many plant specialized metabolites are of high value due to their health promoting characteristics. Glucosinolates are defense compounds found in many crops from the Brassicaceae family and are of high interest because of their nutritional and antinutritional properties...... cells is an open question. Likewise, it is not known how glucosinolate biosynthesis is orchestrated at the subcellular level. These open questions were addressed with several approaches in this project, with the aim of shedding light on the spatial organization of glucosinolate biosynthesis from...

  8. Functional and Evolutionary Relationship between Arginine Biosynthesis and Prokaryotic Lysine Biosynthesis through α-Aminoadipate

    Science.gov (United States)

    Miyazaki, Junichi; Kobashi, Nobuyuki; Nishiyama, Makoto; Yamane, Hisakazu

    2001-01-01

    Our previous studies revealed that lysine is synthesized through α-aminoadipate in an extremely thermophilic bacterium, Thermus thermophilus HB27. Sequence analysis of a gene cluster involved in the lysine biosynthesis of this microorganism suggested that the conversion from α-aminoadipate to lysine proceeds in a way similar to that of arginine biosynthesis. In the present study, we cloned an argD homolog of T. thermophilus HB27 which was not included in the previously cloned lysine biosynthetic gene cluster and determined the nucleotide sequence. A knockout of the argD-like gene, now termed lysJ, in T. thermophilus HB27 showed that this gene is essential for lysine biosynthesis in this bacterium. The lysJ gene was cloned into a plasmid and overexpressed in Escherichia coli, and the LysJ protein was purified to homogeneity. When the catalytic activity of LysJ was analyzed in a reverse reaction in the putative pathway, LysJ was found to transfer the ɛ-amino group of N2-acetyllysine, a putative intermediate in lysine biosynthesis, to 2-oxoglutarate. When N2-acetylornithine, a substrate for arginine biosynthesis, was used as the substrate for the reaction, LysJ transferred the δ-amino group of N2-acetylornithine to 2-oxoglutarate 16 times more efficiently than when N2-acetyllysine was the amino donor. All these results suggest that lysine biosynthesis in T. thermophilus HB27 is functionally and evolutionarily related to arginine biosynthesis. PMID:11489859

  9. Triterpenoid biosynthesis in Euphorbia lathyris latex

    International Nuclear Information System (INIS)

    Hawkins, D.R.

    1987-11-01

    The structures of triterpenols, not previously been known, from Euphorbia lathyris latex are reported. A method for quantifying very small amounts of these compounds was developed. Concerning the biochemistry of the latex, no exogenous cofactors were required for the biosynthesis and the addition of compounds such as NADPAH and ATP do not stimulate the biosynthesis. The addition of DTE or a similar anti-oxidant was found to help reduce the oxidation of the latex, thus increasing the length of time that the latex remains active. The requirement of a divalent cation and the preference for Mn in the pellet was observed. The effect of several inhibitors on the biosynthesis of the triterpenoids was examined. Mevinolin was found to inhibit the biosynthesis of the triterpenoids from acetate, but not mevalonate. A dixon plot of the inhibition of acetate incorporation showed an I 50 concentration of 3.2 μM. Fenpropimorph was found to have little or no effect on the biosynthesis. Tridemorph was found to inhibit the biosynthesis of all of the triterpenoids with an I 50 of 4 μM. It was also observed that the cyclopropyl containing triterpenols, cycloartenol and 24-methylenecycloartenol were inhibited much more strongly than those containing an 8-9 double bond, lanosterol and 24-methylenelanosterol. The evidence indicates, but does not definetely prove, that lanosterol and 24-methylenelanosterol are not made from cycloartenol and 24-methylenecycloartenol via a ring-opening enzyme such as cycloeucalenol-obtusifoliol isomerase. The possibilty that cycloartenol is made via lanosterol was investigated by synthesizing 4-R-4- 3 H-mevalonic acid and incubating latex with a mixture of this and 14 C-mevalonic acid. From the 3 H/ 14 C ratio it was shown that cycloartenol and 24-methylenecycloartenol are not made via an intermediate containing as 8-9 double bond. 88 refs., 15 figs., 30 tabs

  10. Combinatorial Biosynthesis of Polyketides – A Perspective

    Science.gov (United States)

    Wong, Fong T.; Khosla, Chaitan

    2012-01-01

    Since their discovery, polyketide synthases have been attractive targets of biosynthetic engineering to make “unnatural” natural products. Although combinatorial biosynthesis has made encouraging advances over the past two decades, the field remains in its infancy. In this enzyme-centric perspective, we discuss the scientific and technological challenges that could accelerate the adoption of combinatorial biosynthesis as a method of choice for the preparation of encoded libraries of bioactive small molecules. Borrowing a page from the protein structure prediction community, we propose a periodic challenge program to vet the most promising methods in the field, and to foster the collective development of useful tools and algorithms. PMID:22342766

  11. Method for determining heterologous biosynthesis pathways

    KAUST Repository

    Gao, Xin

    2017-08-10

    The present invention relates to a method and system for dynamically analyzing, determining, predicting and displaying ranked suitable heterologous biosynthesis pathways for a specified host. The present invention addresses the problem of finding suitable pathways for the endogenous metabolism of a host organism because the efficacy of heterologous biosynthesis is affected by competing endogenous pathways. The present invention is called MRE (Metabolic Route Explorer), and it was conceived and developed to systematically and dynamically search for, determine, analyze, and display promising heterologous pathways while considering competing endogenous reactions in a given host organism.

  12. Convergent Evolution of Ergothioneine Biosynthesis in Cyanobacteria.

    Science.gov (United States)

    Liao, Cangsong; Seebeck, Florian P

    2017-11-02

    Biosynthesis of N-α-trimethyl-2-thiohistidine (ergothioneine) is a frequent trait in cyanobacteria. This sulfur compound may provide essential relief from oxidative stress related to oxygenic photosynthesis. The central steps in ergothioneine biosynthesis are catalyzed by a histidine methyltransferase and an iron-dependent sulfoxide synthase. In this report, we present evidence that some cyanobacteria recruited and adapted a sulfoxide synthase from a different biosynthetic pathway to make ergothioneine. The discovery of a second origin of ergothioneine production underscores the physiological importance of this metabolite and highlights the evolutionary malleability of the thiohistidine biosynthetic machinery. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  13. Biosynthesis of silver nanoparticles by Aspergillus niger , Fusarium ...

    African Journals Online (AJOL)

    ... scanning electron microscope (SEM). Results indicate the synthesis of silver nanoparticles in the reaction mixture. The synthesis of nanoparticles would be suitable for developing a microbial nanotechnology biosynthesis process for mass scale production. Keywords: Silver nanoparticles, biosynthesis, fungi, Aspergillus.

  14. Nutritional and biochemical therapies for neurodegenerative diseases

    Directory of Open Access Journals (Sweden)

    George E. Barreto

    2015-04-01

    Full Text Available Neurodegenerative diseases such as Parkinson’s disease, Huntington and Alzheimer’s disease are characterized by neuronal death and loss in different areas of the brain. Downstream signaling mechanisms associated to cellular death/survival are altered, where mitochondrial damage and inflammation, dysfunctional autophagy process, and accumulation of toxins proteins play a central role in the pathogenesis of these diseases. The disabling effects of these diseases on health system are high and greatly affect the health and daily lifestyle of patients. In this context, pharmacological and non-pharmacological therapies, which are used in palliative and preventive treatments, have been widely assessed in human patients, as well as animal and cellular models in the last decades. However, the genetics and epigenetics factors of any disease can cause different paths in its progression. Nutritional and biochemical therapy approaches by activation or manipulation of different transcription factors such as Nrf2, PPARα, CREB and TEFB in animal and cellular models have shown protective effects against neurodegeneration. Some of these therapies include caloric restriction diet, use of glutathione precursors and Mediterranean diet. This work highlights the evidences of different nutritional and biochemical approaches for the treatment of neurodegenerative diseases and how novel research approaches, such as the use of systems biology, will allow a better comprehension of key processes and biological responses involved in these diseases.

  15. Hemoglobin Variants: Biochemical Properties and Clinical Correlates

    Science.gov (United States)

    Thom, Christopher S.; Dickson, Claire F.; Gell, David A.; Weiss, Mitchell J.

    2013-01-01

    Diseases affecting hemoglobin synthesis and function are extremely common worldwide. More than 1000 naturally occurring human hemoglobin variants with single amino acid substitutions throughout the molecule have been discovered, mainly through their clinical and/or laboratory manifestations. These variants alter hemoglobin structure and biochemical properties with physiological effects ranging from insignificant to severe. Studies of these mutations in patients and in the laboratory have produced a wealth of information on hemoglobin biochemistry and biology with significant implications for hematology practice. More generally, landmark studies of hemoglobin performed over the past 60 years have established important paradigms for the disciplines of structural biology, genetics, biochemistry, and medicine. Here we review the major classes of hemoglobin variants, emphasizing general concepts and illustrative examples. PMID:23388674

  16. Biosynthesis of Polyunsaturated Fatty Acids in Marine Invertebrates: Recent Advances in Molecular Mechanisms

    Science.gov (United States)

    Monroig, Óscar; Tocher, Douglas R.; Navarro, Juan C.

    2013-01-01

    Virtually all polyunsaturated fatty acids (PUFA) originate from primary producers but can be modified by bioconversions as they pass up the food chain in a process termed trophic upgrading. Therefore, although the main primary producers of PUFA in the marine environment are microalgae, higher trophic levels have metabolic pathways that can produce novel and unique PUFA. However, little is known about the pathways of PUFA biosynthesis and metabolism in the levels between primary producers and fish that are largely filled by invertebrates. It has become increasingly apparent that, in addition to trophic upgrading, de novo synthesis of PUFA is possible in some lower animals. The unequivocal identification of PUFA biosynthetic pathways in many invertebrates is complicated by the presence of other organisms within them. These organisms include bacteria and algae with PUFA biosynthesis pathways, and range from intestinal flora to symbiotic relationships that can involve PUFA translocation to host organisms. This emphasizes the importance of studying biosynthetic pathways at a molecular level, and the continual expansion of genomic resources and advances in molecular analysis is facilitating this. The present paper highlights recent research into the molecular and biochemical mechanisms of PUFA biosynthesis in marine invertebrates, particularly focusing on cephalopod molluscs. PMID:24152561

  17. Biochemistry and Molecular Biology of Carotenoid Biosynthesis in Chili Peppers (Capsicum spp.

    Directory of Open Access Journals (Sweden)

    María del Rocío Gómez-García

    2013-09-01

    Full Text Available Capsicum species produce fruits that synthesize and accumulate carotenoid pigments, which are responsible for the fruits’ yellow, orange and red colors. Chili peppers have been used as an experimental model for studying the biochemical and molecular aspects of carotenoid biosynthesis. Most reports refer to the characterization of carotenoids and content determination in chili pepper fruits from different species, cultivars, varieties or genotypes. The types and levels of carotenoids differ between different chili pepper fruits, and they are also influenced by environmental conditions. Yellow-orange colors of chili pepper fruits are mainly due to the accumulation of α- and β-carotene, zeaxanthin, lutein and β-cryptoxanthin. Carotenoids such as capsanthin, capsorubin and capsanthin-5,6-epoxide confer the red colors. Chromoplasts are the sites of carotenoid pigment synthesis and storage. According to the most accepted theory, the synthesis of carotenoids in chili peppers is controlled by three loci: c1, c2 and y. Several enzymes participating in carotenoid biosynthesis in chili pepper fruits have been isolated and characterized, and the corresponding gene sequences have been reported. However, there is currently limited information on the molecular mechanisms that regulate this biosynthetic pathway. Approaches to gain more knowledge of the regulation of carotenoid biosynthesis are discussed.

  18. Competition between ethanol clearance and retinoic acid biosynthesis in the induction of fetal alcohol syndrome.

    Science.gov (United States)

    Shabtai, Yehuda; Fainsod, Abraham

    2018-04-01

    Several models have been proposed to explain the neurodevelopmental syndrome induced by exposure of human embryos to alcohol, which is known as fetal alcohol spectrum disorder (FASD). One of the proposed models suggests a competition for the enzymes required for the biosynthesis of retinoic acid. The outcome of such competition is development under conditions of reduced retinoic acid signaling. Retinoic acid is one of the biologically active metabolites of vitamin A (retinol), and regulates numerous embryonic and differentiation processes. The developmental malformations characteristic of FASD resemble those observed in vitamin A deficiency syndrome as well as from inhibition of retinoic acid biosynthesis or signaling in experimental models. There is extensive biochemical and enzymatic overlap between ethanol clearance and retinoic acid biosynthesis. Several lines of evidence suggest that in the embryo, the competition takes place between acetaldehyde and retinaldehyde for the aldehyde dehydrogenase activity available. In adults, this competition also extends to the alcohol dehydrogenase activity. Ethanol-induced developmental defects can be ameliorated by increasing the levels of retinol, retinaldehyde, or retinaldehyde dehydrogenase. Acetaldehyde inhibits the production of retinoic acid by retinaldehyde dehydrogenase, further supporting the competition model. All of the evidence supports the reduction of retinoic acid signaling as the etiological trigger in the induction of FASD.

  19. Control of biotin biosynthesis in mycobacteria by a pyruvate carboxylase dependent metabolic signal.

    Science.gov (United States)

    Lazar, Nathaniel; Fay, Allison; Nandakumar, Madhumitha; Boyle, Kerry E; Xavier, Joao; Rhee, Kyu; Glickman, Michael S

    2017-12-01

    Biotin is an essential cofactor utilized by all domains of life, but only synthesized by bacteria, fungi and plants, making biotin biosynthesis a target for antimicrobial development. To understand biotin biosynthesis in mycobacteria, we executed a genetic screen in Mycobacterium smegmatis for biotin auxotrophs and identified pyruvate carboxylase (Pyc) as required for biotin biosynthesis. The biotin auxotrophy of the pyc::tn strain is due to failure to transcriptionally induce late stage biotin biosynthetic genes in low biotin conditions. Loss of bioQ, the repressor of biotin biosynthesis, in the pyc::tn strain reverted biotin auxotrophy, as did reconstituting the last step of the pathway through heterologous expression of BioB and provision of its substrate DTB. The role of Pyc in biotin regulation required its catalytic activities and could be supported by M. tuberculosis Pyc. Quantitation of the kinetics of depletion of biotinylated proteins after biotin withdrawal revealed that Pyc is the most rapidly depleted biotinylated protein and metabolomics revealed a broad metabolic shift in wild type cells upon biotin withdrawal which was blunted in cell lacking Pyc. Our data indicate that mycobacterial cells monitor biotin sufficiency through a metabolic signal generated by dysfunction of a biotinylated protein of central metabolism. © 2017 John Wiley & Sons Ltd.

  20. Biosynthesis and metabolic pathways of pivalic acid

    Czech Academy of Sciences Publication Activity Database

    Řezanka, Tomáš; Kolouchová, I.; Čejková, A.; Sigler, Karel

    2012-01-01

    Roč. 95, č. 6 (2012), s. 1371-1376 ISSN 0175-7598 R&D Projects: GA ČR(CZ) GAP503/11/0215 Institutional support: RVO:61388971 Keywords : Pivalic acid * Isooctane * Biosynthesis Subject RIV: EE - Microbiology, Virology Impact factor: 3.689, year: 2012

  1. Biosynthesis of silver nanoparticles synthesized by Aspergillus

    Indian Academy of Sciences (India)

    In the present study, biosynthesis of silver nanoparticles and its antioxidant, antimicrobial and cytotoxic activities were investigated. Silver nanoparticles were extracellularly synthesized using Aspergillus flavus and the formation of nanoparticles was observed after 72 h of incubation. The results recorded from colour ...

  2. Bile acid biosynthesis and its regulation

    Directory of Open Access Journals (Sweden)

    Areta Hebanowska

    2010-10-01

    Full Text Available Bile acid biosynthesis is the main pathway of cholesterol catabolism. Bile acids are more soluble than cholesterol so are easier to excrete. As amphipathic molecules they participate in lipid digestion and absorption in the intestine and they help to excrete free cholesterol with bile. They are also ligands for nuclear receptors regulating the expression of genes involved in cholesterol metabolism. Interconversion of cholesterol into bile acids is an important point of its homeostasis. Seventeen enzymes are engaged in this process and many of them are cytochromes P450. Bile acid synthesis initiation may proceed with the “classical” pathway (starting with cholesterol hydroxylation at the C7α position or the “alternative” pathway (starting with cholesterol hydroxylation at the C27 position. Two additional pathways are possible, though their quantitative significance is small (initiated with cholesterol hydroxylations of C24 and C25 positions. Oxysterols produced are not only intermediates of bile acid biosynthesis but also important regulators of metabolism. Bile acid biosynthesis takes place in the liver, but some enzymes are also present in other organs, where they participate in regulation of cholesterol metabolism. Those enzymes are potential targets for new drugs against cholesterol metabolism disturbances. This article is a brief description of the bile acid biosynthesis pathway and participating enzymes.

  3. Unedoside derivatives in Nuxia and their biosynthesis

    DEFF Research Database (Denmark)

    Jensen, Søren Rosendal; Ravnkilde, Lene; Schripsema, Jan

    1998-01-01

    isolated, while from N. oppositifolia 2 "-acetyl-3 "-benzoyl-nuxioside was obtained. Both plants contained verbascoside. The biosynthesis of unedoside in N. floribunda was investigated and deoxyloganic acid was found to be a precursor, similar to wh;lt was found for the eight-carbon iridoids in Thunbergia...

  4. Biosynthesis of furanochromones in Pimpinella monoica

    Indian Academy of Sciences (India)

    polyketide origin of their aromatic and pyrone rings while the furan ring originates via an acetate-mevalonate pathway. The plant also utilises glycine and leucine as substrate via acetate. Biotransformation of 3-H-visnagin to (6) but not to (2) was also observed. Keywords. Biosynthesis; furochromones; polyketide origin; ...

  5. Biosynthesis of silver nanoparticles synthesized by Aspergillus ...

    Indian Academy of Sciences (India)

    In the present study, biosynthesis of silver nanoparticles and its antioxidant, antimicrobial and cytotoxic activities were investigated. Silver nanoparticles were extracellularly synthesized using Aspergillus flavus and the formation of nanoparticles was observed after 72 h of incubation. The results recorded from colour ...

  6. Viral serine palmitoyltransferase induces metabolic switch in sphingolipid biosynthesis and is required for infection of a marine alga.

    Science.gov (United States)

    Ziv, Carmit; Malitsky, Sergey; Othman, Alaa; Ben-Dor, Shifra; Wei, Yu; Zheng, Shuning; Aharoni, Asaph; Hornemann, Thorsten; Vardi, Assaf

    2016-03-29

    Marine viruses are the most abundant biological entities in the oceans shaping community structure and nutrient cycling. The interaction between the bloom-forming alga Emiliania huxleyi and its specific large dsDNA virus (EhV) is a major factor determining the fate of carbon in the ocean, thus serving as a key host-pathogen model system. The EhV genome encodes for a set of genes involved in the de novo sphingolipid biosynthesis, not reported in any viral genome to date. We combined detailed lipidomic and biochemical analyses to characterize the functional role of this virus-encoded pathway during lytic viral infection. We identified a major metabolic shift, mediated by differential substrate specificity of virus-encoded serine palmitoyltransferase, a key enzyme of sphingolipid biosynthesis. Consequently, unique viral glycosphingolipids, composed of unusual hydroxylated C17 sphingoid bases (t17:0) were highly enriched in the infected cells, and their synthesis was found to be essential for viral assembly. These findings uncover the biochemical bases of the virus-induced metabolic rewiring of the host sphingolipid biosynthesis during the chemical "arms race" in the ocean.

  7. Biochemical characterization of xylan xylosyltransferases involved in wood formation in poplar.

    Science.gov (United States)

    Lee, Chanhui; Zhong, Ruiqin; Ye, Zheng-Hua

    2012-03-01

    The major polysaccharides in dicot wood biomass are cellulose and xylan. Although wood-associated cellulose synthase genes responsible for cellulose biosynthesis have been characterized, wood-associated xylan synthase genes have not been biochemically identified. A recent report by Lee et al. (2012) provides the first biochemical evidence that two functionally non-redundant Arabidopsis GT43 members are xylosyltransferases (XylTs) that function cooperatively in the elongation of the xylan backbone. We further extend this finding in the current report demonstrating that two poplar (Populus trichocarpa) GT43 glycosyltransferases, PtrGT43B and PtrGT43C, are xylan XylTs involved in wood formation. We show that microsomes from transgenic tobacco BY2 cells coexpressing PtrGT43B and PtrGT43C exhibited a high XylT activity capable of generating β-(1,4)-linked xylooligosaccharides, whereas little XylT activity was detected in microsomes with expression of PtrGT43B or PtrGT43C alone. These findings indicate that poplar GT43 members are XylTs that act cooperatively in catalyzing the successive transfer of xylosyl residues during xylan backbone biosynthesis, which provides further support of the hypothesis that the biochemical functions of GT43 members in vascular plants are evolutionarily conserved.

  8. Comprehensive Characterization for Ginsenosides Biosynthesis in Ginseng Root by Integration Analysis of Chemical and Transcriptome

    Directory of Open Access Journals (Sweden)

    Jing-Jing Zhang

    2017-05-01

    Full Text Available Herbgenomics provides a global platform to explore the genetics and biology of herbs on the genome level. Panax ginseng C.A. Meyer is an important medicinal plant with numerous pharmaceutical effects. Previous reports mainly discussed the transcriptome of ginseng at the organ level. However, based on mass spectrometry imaging analyses, the ginsenosides varied among different tissues. In this work, ginseng root was separated into three tissues—periderm, cortex and stele—each for five duplicates. The chemical analysis and transcriptome analysis were conducted simultaneously. Gene-encoding enzymes involved in ginsenosides biosynthesis and modification were studied based on gene and molecule data. Eight widely-used ginsenosides were distributed unevenly in ginseng roots. A total of 182,881 unigenes were assembled with an N50 contig size of 1374 bp. About 21,000 of these unigenes were positively correlated with the content of ginsenosides. Additionally, we identified 192 transcripts encoding enzymes involved in two triterpenoid biosynthesis pathways and 290 transcripts encoding UDP-glycosyltransferases (UGTs. Of these UGTs, 195 UGTs (67.2% were more highly expressed in the periderm, and that seven UGTs and one UGT were specifically expressed in the periderm and stele, respectively. This genetic resource will help to improve the interpretation on complex mechanisms of ginsenosides biosynthesis, accumulation, and transportation.

  9. The complete coenzyme B12 biosynthesis gene cluster of Lactobacillus reuteri CRL1098

    NARCIS (Netherlands)

    Santos, F.; Vera, J.L.; van der Heijden, R.; Valdez, G.; de Vos, W.M.; Sesma, F.; Hugenholtz, J.

    2008-01-01

    The coenzyme B12 production pathway in Lactobacillus reuteri has been deduced using a combination of genetic, biochemical and bioinformatics approaches. The coenzyme B12 gene cluster of Lb. reuteri CRL1098 has the unique feature of clustering together the cbi, cob and hem genes. It consists of 29

  10. The complete coenzyme B12 biosynthesis gene cluster of Lactobacillus reuteri CRL 1098

    NARCIS (Netherlands)

    Santos, dos F.; Vera, J.L.; Heijden, van der R.; Valdez, G.F.; Vos, de W.M.; Sesma, F.; Hugenholtz, J.

    2008-01-01

    The coenzyme B12 production pathway in Lactobacillus reuteri has been deduced using a combination of genetic, biochemical and bioinformatics approaches. The coenzyme B12 gene cluster of Lb. reuteri CRL1098 has the unique feature of clustering together the cbi, cob and hem genes. It consists of 29

  11. Transcriptome sequencing and expression analysis of terpenoid biosynthesis genes in Litsea cubeba.

    Directory of Open Access Journals (Sweden)

    Xiao-Jiao Han

    Full Text Available BACKGROUND: Aromatic essential oils extracted from fresh fruits of Litsea cubeba (Lour. Pers., have diverse medical and economic values. The dominant components in these essential oils are monoterpenes and sesquiterpenes. Understanding the molecular mechanisms of terpenoid biosynthesis is essential for improving the yield and quality of terpenes. However, the 40 available L. cubeba nucleotide sequences in the public databases are insufficient for studying the molecular mechanisms. Thus, high-throughput transcriptome sequencing of L. cubeba is necessary to generate large quantities of transcript sequences for the purpose of gene discovery, especially terpenoid biosynthesis related genes. RESULTS: Using Illumina paired-end sequencing, approximately 23.5 million high-quality reads were generated. De novo assembly yielded 68,648 unigenes with an average length of 834 bp. A total of 38,439 (56% unigenes were annotated for their functions, and 35,732 and 25,806 unigenes could be aligned to the GO and COG database, respectively. By searching against the Kyoto Encyclopedia of Genes and Genomes Pathway database (KEGG, 16,130 unigenes were assigned to 297 KEGG pathways, and 61 unigenes, which contained the mevalonate and 2-C-methyl-D-erythritol 4-phosphate pathways, could be related to terpenoid backbone biosynthesis. Of the 12,963 unigenes, 285 were annotated to the terpenoid pathways using the PlantCyc database. Additionally, 14 terpene synthase genes were identified from the transcriptome. The expression patterns of the 16 genes related to terpenoid biosynthesis were analyzed by RT-qPCR to explore their putative functions. CONCLUSION: RNA sequencing was effective in identifying a large quantity of sequence information. To our knowledge, this study is the first exploration of the L. cubeba transcriptome, and the substantial amount of transcripts obtained will accelerate the understanding of the molecular mechanisms of essential oils biosynthesis. The

  12. Genes involved in long-chain alkene biosynthesis in Micrococcus luteus

    Energy Technology Data Exchange (ETDEWEB)

    Beller, Harry R.; Goh, Ee-Been; Keasling, Jay D.

    2010-01-07

    Aliphatic hydrocarbons are highly appealing targets for advanced cellulosic biofuels, as they are already predominant components of petroleum-based gasoline and diesel fuels. We have studied alkene biosynthesis in Micrococcus luteus ATCC 4698, a close relative of Sarcina lutea (now Kocuria rhizophila), which four decades ago was reported to biosynthesize iso- and anteiso branched, long-chain alkenes. The underlying biochemistry and genetics of alkene biosynthesis were not elucidated in those studies. We show here that heterologous expression of a three-gene cluster from M. luteus (Mlut_13230-13250) in a fatty-acid overproducing E. coli strain resulted in production of long-chain alkenes, predominantly 27:3 and 29:3 (no. carbon atoms: no. C=C bonds). Heterologous expression of Mlut_13230 (oleA) alone produced no long-chain alkenes but unsaturated aliphatic monoketones, predominantly 27:2, and in vitro studies with the purified Mlut_13230 protein and tetradecanoyl-CoA produced the same C27 monoketone. Gas chromatography-time of flight mass spectrometry confirmed the elemental composition of all detected long-chain alkenes and monoketones (putative intermediates of alkene biosynthesis). Negative controls demonstrated that the M. luteus genes were responsible for production of these metabolites. Studies with wild-type M. luteus showed that the transcript copy number of Mlut_13230-13250 and the concentrations of 29:1 alkene isomers (the dominant alkenes produced by this strain) generally corresponded with bacterial population over time. We propose a metabolic pathway for alkene biosynthesis starting with acyl-CoA (or -ACP) thioesters and involving decarboxylative Claisen condensation as a key step, which we believe is catalyzed by OleA. Such activity is consistent with our data and with the homology (including the conserved Cys-His-Asn catalytic triad) of Mlut_13230 (OleA) to FabH (?-ketoacyl-ACP synthase III), which catalyzes decarboxylative Claisen condensation during

  13. Autotaxin : biochemical and functional studies

    NARCIS (Netherlands)

    Houben, Anna Jacoba Sara

    2012-01-01

    This thesis focuses on autotaxin (ATX), the main enzyme responsible for the production of lysophosphatidic acid (LPA). The ATX-LPA receptor axis has a wide implication in health and disease. The studies described in this thesis aim at characterizing the biochemical and functional properties of ATX,

  14. Serum biochemical changes accompanying prolonged ...

    African Journals Online (AJOL)

    Jane

    2010-10-18

    Oct 18, 2010 ... Toxicological evaluation of the whole fruit of Lagenaria breviflora was carried out using the serum biochemical changes accompanying prolonged administration of the ethanolic extract of the fruit in ... glucose in rats administered with the extract of 8000 mg/kg body weight increased two and half-fold.

  15. Journal of Genetics | Indian Academy of Sciences

    Indian Academy of Sciences (India)

    Home; Journals; Journal of Genetics; Volume 95; Issue 2. Expression profiles of genes involved in tanshinone biosynthesis of two Salvia miltiorrhiza genotypes with different tanshinone contents. ZHENQIAO SONG JIANHUA WANG XINGFENG LI. RESEARCH NOTE Volume 95 Issue 2 June 2016 pp 433-439 ...

  16. Synergy and Target Promiscuity Drive Structural Divergence in Bacterial Alkylquinolone Biosynthesis.

    Science.gov (United States)

    Wu, Yihan; Seyedsayamdost, Mohammad R

    2017-12-21

    Microbial natural products are genetically encoded by dedicated biosynthetic gene clusters (BGCs). A given BGC usually produces a family of related compounds that share a core but contain variable substituents. Though common, the reasons underlying this divergent biosynthesis are in general unknown. Herein, we have addressed this issue using the hydroxyalkylquinoline (HAQ) family of natural products synthesized by Burkholderia thailandensis. Investigations into the detailed functions of two analogs show that they act synergistically in inhibiting bacterial growth. One analog is a nanomolar inhibitor of pyrimidine biosynthesis and at the same time disrupts the proton motive force. A second analog inhibits the cytochrome bc 1 complex as well as pyrimidine biogenesis. These results provide a functional rationale for the divergent nature of HAQs. They imply that synergy and target promiscuity are driving forces for the evolution of tailoring enzymes that diversify the products of the HAQ biosynthetic pathway. Copyright © 2017 Elsevier Ltd. All rights reserved.

  17. Overcoming Challenges in Engineering the Genetic Code.

    Science.gov (United States)

    Lajoie, M J; Söll, D; Church, G M

    2016-02-27

    Withstanding 3.5 billion years of genetic drift, the canonical genetic code remains such a fundamental foundation for the complexity of life that it is highly conserved across all three phylogenetic domains. Genome engineering technologies are now making it possible to rationally change the genetic code, offering resistance to viruses, genetic isolation from horizontal gene transfer, and prevention of environmental escape by genetically modified organisms. We discuss the biochemical, genetic, and technological challenges that must be overcome in order to engineer the genetic code. Copyright © 2015 Elsevier Ltd. All rights reserved.

  18. Combinatorial biosynthesis of polyketides--a perspective.

    Science.gov (United States)

    Wong, Fong T; Khosla, Chaitan

    2012-04-01

    Since their discovery, polyketide synthases have been attractive targets of biosynthetic engineering to make 'unnatural' natural products. Although combinatorial biosynthesis has made encouraging advances over the past two decades, the field remains in its infancy. In this enzyme-centric perspective, we discuss the scientific and technological challenges that could accelerate the adoption of combinatorial biosynthesis as a method of choice for the preparation of encoded libraries of bioactive small molecules. Borrowing a page from the protein structure prediction community, we propose a periodic challenge program to vet the most promising methods in the field, and to foster the collective development of useful tools and algorithms. Copyright © 2012 Elsevier Ltd. All rights reserved.

  19. Structural basis for phosphatidylinositol-phosphate biosynthesis

    Science.gov (United States)

    Clarke, Oliver B.; Tomasek, David; Jorge, Carla D.; Dufrisne, Meagan Belcher; Kim, Minah; Banerjee, Surajit; Rajashankar, Kanagalaghatta R.; Shapiro, Lawrence; Hendrickson, Wayne A.; Santos, Helena; Mancia, Filippo

    2015-10-01

    Phosphatidylinositol is critical for intracellular signalling and anchoring of carbohydrates and proteins to outer cellular membranes. The defining step in phosphatidylinositol biosynthesis is catalysed by CDP-alcohol phosphotransferases, transmembrane enzymes that use CDP-diacylglycerol as donor substrate for this reaction, and either inositol in eukaryotes or inositol phosphate in prokaryotes as the acceptor alcohol. Here we report the structures of a related enzyme, the phosphatidylinositol-phosphate synthase from Renibacterium salmoninarum, with and without bound CDP-diacylglycerol to 3.6 and 2.5 Å resolution, respectively. These structures reveal the location of the acceptor site, and the molecular determinants of substrate specificity and catalysis. Functional characterization of the 40%-identical ortholog from Mycobacterium tuberculosis, a potential target for the development of novel anti-tuberculosis drugs, supports the proposed mechanism of substrate binding and catalysis. This work therefore provides a structural and functional framework to understand the mechanism of phosphatidylinositol-phosphate biosynthesis.

  20. Occurrence and biosynthesis of carotenoids in phytoplankton.

    Science.gov (United States)

    Huang, Jim Junhui; Lin, Shaoling; Xu, Wenwen; Cheung, Peter Chi Keung

    2017-09-01

    Naturally occurring carotenoids are important sources of antioxidants, anti-cancer compounds and anti-inflammatory agents and there is thus considerable market demand for their pharmaceutical applications. Carotenoids are widely distributed in marine and freshwater organisms including microalgae, phytoplankton, crustaceans and fish, as well as in terrestrial plants and birds. Recently, phytoplankton-derived carotenoids have received much attention due to their abundance, rapid rate of biosynthesis and unique composition. The carotenoids that accumulate in particular phytoplankton phyla are synthesized by specific enzymes and play unique physiological roles. This review focuses on studies related to the occurrence of carotenoids in different phytoplankton phyla and the molecular aspects of their biosynthesis. Recent biotechnological advances in the isolation and characterization of some representative carotenoid synthases in phytoplankton are also discussed. Copyright © 2017 Elsevier Inc. All rights reserved.

  1. Origin of Life: Pathways of the 20 Standard Amino Acids of the Genetic Code

    Science.gov (United States)

    Hall, A.; Acker-Moorehead, M.; Onyilagha, J.

    2017-11-01

    How nature used four nucleotides to build its proteins and form genetic code is intriguing. Stereochemical, Coevolution, and Adaptive theories have been propounded. We updated biosynthesis pathways and give insight into ancient evolutionary events.

  2. Microbial biosynthesis of nontoxic gold nanoparticles

    Energy Technology Data Exchange (ETDEWEB)

    Roy, Swarup, E-mail: swaruproy@klyuniv.ac.in [Department of Biochemistry and Biophysics, University of Kalyani, Kalyani 741235, West Bengal (India); Das, Tapan Kumar [Department of Biochemistry and Biophysics, University of Kalyani, Kalyani 741235, West Bengal (India); Maiti, Guru Prasad [Department of Molecular Biology and Biotechnology, University of Kalyani, Kalyani 741235, West Bengal (India); Department of Anesthesiology, Texas Tech University Health science Center, 3601 4th Street, Lubbock, TX 79430 (United States); Basu, Utpal [Department of Molecular Biology and Biotechnology, University of Kalyani, Kalyani 741235, West Bengal (India)

    2016-01-15

    Graphical abstract: The manuscript deals with the fungus mediated optimized biologically synthesized GNPs using Aspergillus foetidus and characterization of biosynthesized GNPs using various physico-chemical methods. The fairly stable synthesized nanoparticles have size in the range of 10–40 nm. Cytotoxicity study of biosynthesized GNPs on Human lung cancer cell line A549 showed no significant toxicity of GNPs. - Highlights: • A novel biosynthesis process of GNPs using Aspergillus foetidus. • Biosynthesized GNPs are in the range of 10–40 nm as observed from TEM. • This process of synthesis is an optimized biosynthesis process of GNPs. • Biosynthesized GNPs are noncytotoxic against A549 cell line. - Abstract: We study the extracellular biosynthesis of gold nanoparticles (GNPs) using the fungal species Aspergillus foetidus. The formation of GNPs were initially monitored by visual observation and then characterized with the help of various characterization techniques. X-ray diffraction (XRD) results revealed distinctive formation of face centered cubic crystalline GNPs. From field emission scanning electron microscopy (FESEM) the morphology of the nanoparticles were found to be roughly spherical and within the size range of 30–50 nm. The spherical and polydispersed GNPs in the range of 10–40 nm were observed by transmission electron microscopy (TEM) analysis. It was established that alkaline pH, 1 mM gold salt concentration and 75 °C temperature were the respective optimum parameter for biosynthesis of GNPs. Cell cytotoxicity of GNP was compared with that of normal gold salt solution on A549 cell. The A549 cell growth in presence of GNPs was found to be comparatively less toxic than the gold ion.

  3. Microbial biosynthesis of nontoxic gold nanoparticles

    International Nuclear Information System (INIS)

    Roy, Swarup; Das, Tapan Kumar; Maiti, Guru Prasad; Basu, Utpal

    2016-01-01

    Graphical abstract: The manuscript deals with the fungus mediated optimized biologically synthesized GNPs using Aspergillus foetidus and characterization of biosynthesized GNPs using various physico-chemical methods. The fairly stable synthesized nanoparticles have size in the range of 10–40 nm. Cytotoxicity study of biosynthesized GNPs on Human lung cancer cell line A549 showed no significant toxicity of GNPs. - Highlights: • A novel biosynthesis process of GNPs using Aspergillus foetidus. • Biosynthesized GNPs are in the range of 10–40 nm as observed from TEM. • This process of synthesis is an optimized biosynthesis process of GNPs. • Biosynthesized GNPs are noncytotoxic against A549 cell line. - Abstract: We study the extracellular biosynthesis of gold nanoparticles (GNPs) using the fungal species Aspergillus foetidus. The formation of GNPs were initially monitored by visual observation and then characterized with the help of various characterization techniques. X-ray diffraction (XRD) results revealed distinctive formation of face centered cubic crystalline GNPs. From field emission scanning electron microscopy (FESEM) the morphology of the nanoparticles were found to be roughly spherical and within the size range of 30–50 nm. The spherical and polydispersed GNPs in the range of 10–40 nm were observed by transmission electron microscopy (TEM) analysis. It was established that alkaline pH, 1 mM gold salt concentration and 75 °C temperature were the respective optimum parameter for biosynthesis of GNPs. Cell cytotoxicity of GNP was compared with that of normal gold salt solution on A549 cell. The A549 cell growth in presence of GNPs was found to be comparatively less toxic than the gold ion.

  4. Tetrahydrobiopterin biosynthesis, utilization and pharmacological effects.

    Science.gov (United States)

    Werner-Felmayer, G; Golderer, G; Werner, E R

    2002-04-01

    Tetrahydrobiopterin (H4-biopterin) is an essential cofactor of a set of enzymes that are of central metabolic importance, i.e. the hydroxylases of the three aromatic amino acids phenylalanine, tyrosine, and tryptophan, of ether lipid oxidase, and of the three nitric oxide synthase (NOS) isoenzymes. As a consequence, H4-biopterin plays a key role in a vast number of biological processes and pathological states associated with neurotransmitter formation, vasorelaxation, and immune response. In mammals, its biosynthesis is controlled by hormones, cytokines and certain immune stimuli. This review aims to summarize recent developments concerning regulation of H4-biopterin biosynthetic and regulatory enzymes and pharmacological effects of H4-biopterin in various conditions, e.g. endothelial dysfunction or apoptosis of neuronal cells. Also, approaches towards gene therapy of diseases like the different forms of phenylketonuria or of Parkinson's disease are reviewed. Additional emphasis is given to H4-biopterin biosynthesis and function in non-mammalian species such as fruit fly, zebra fish, fungi, slime molds, the bacterium Nocardia as well as to the parasitic protozoan genus of Leishmania that is not capable of pteridine biosynthesis but has evolved a sophisticated salvage network for scavenging various pteridine compounds, notably folate and biopterin.

  5. Exopolysaccharide biosynthesis by Lactobacillus helveticus ATCC 15807.

    Science.gov (United States)

    Torino, M I; Mozzi, F; Font de Valdez, G

    2005-08-01

    Exopolysaccharide (EPS) production and the activities of the enzymes involved in sugar nucleotide biosynthesis in Lactobacillus helveticus ATCC 15807 under controlled pH conditions were investigated. Batch fermentations using lactose as energy source showed higher EPS synthesis by L. helveticus ATCC 15807 at pH 4.5 with respect to pH 6.2, the enzyme alpha-phosphoglucomutase (alpha-PGM) being correlated with both total and specific EPS production. When glucose was used as carbon source instead of lactose, the lower EPS synthesis obtained was linked to a decrease in alpha-PGM and galactose 1-phosphate-uridyltransferase (GalT) activities, the reduction of the latter being more pronounced. Higher EPS production by L. helveticus ATCC 15807 at the acidic constant pH of 4.5 requires that both alpha-PGM and GalT activities are high. These enzymes are needed to synthesize UDP-glucose and UDP-galactose for supplying the corresponding monomers for EPS biosynthesis. Although differences are observed in EPS production by this strain regarding the energy source (lactose or glucose), the monomeric composition of the polymers produced is independent of the carbohydrate used. The obtained results contribute to a better understanding of the physiological factors that affect EPS biosynthesis by lactobacilli, which could help in the correct handling of the fermentation parameters within the fermented dairy industry.

  6. Lipopolysaccharide Structure and Biosynthesis in Helicobacter pylori.

    Science.gov (United States)

    Li, Hong; Liao, Tingting; Debowski, Aleksandra W; Tang, Hong; Nilsson, Hans-Olof; Stubbs, Keith A; Marshall, Barry J; Benghezal, Mohammed

    2016-12-01

    This review covers the current knowledge and gaps in Helicobacter pylori lipopolysaccharide (LPS) structure and biosynthesis. H. pylori is a Gram-negative bacterium which colonizes the luminal surface of the human gastric epithelium. Both a constitutive alteration of the lipid A preventing TLR4 elicitation and host mimicry of the Lewis antigen decorated O-antigen of H. pylori LPS promote immune escape and chronic infection. To date, the complete structure of H. pylori LPS is not available, and the proposed model is a linear arrangement composed of the inner core defined as the hexa-saccharide (Kdo-LD-Hep-LD-Hep-DD-Hep-Gal-Glc), the outer core composed of a conserved trisaccharide (-GlcNAc-Fuc-DD-Hep-) linked to the third heptose of the inner core, the glucan, the heptan and a variable O-antigen, generally consisting of a poly-LacNAc decorated with Lewis antigens. Although the glycosyltransferases (GTs) responsible for the biosynthesis of the H. pylori O-antigen chains have been identified and characterized, there are many gaps in regard to the biosynthesis of the core LPS. These limitations warrant additional mutagenesis and structural studies to obtain the complete LPS structure and corresponding biosynthetic pathway of this important gastric bacterium. © 2016 John Wiley & Sons Ltd.

  7. Pattern Selection by Dynamical Biochemical Signals

    Science.gov (United States)

    Palau-Ortin, David; Formosa-Jordan, Pau; Sancho, José M.; Ibañes, Marta

    2015-01-01

    The development of multicellular organisms involves cells to decide their fate upon the action of biochemical signals. This decision is often spatiotemporally coordinated such that a spatial pattern arises. The dynamics that drive pattern formation usually involve genetic nonlinear interactions and positive feedback loops. These complex dynamics may enable multiple stable patterns for the same conditions. Under these circumstances, pattern formation in a developing tissue involves a selection process: why is a certain pattern formed and not another stable one? Herein we computationally address this issue in the context of the Notch signaling pathway. We characterize a dynamical mechanism for developmental selection of a specific pattern through spatiotemporal changes of the control parameters of the dynamics, in contrast to commonly studied situations in which initial conditions and noise determine which pattern is selected among multiple stable ones. This mechanism can be understood as a path along the parameter space driven by a sequence of biochemical signals. We characterize the selection process for three different scenarios of this dynamical mechanism that can take place during development: the signal either 1) acts in all the cells at the same time, 2) acts only within a cluster of cells, or 3) propagates along the tissue. We found that key elements for pattern selection are the destabilization of the initial pattern, the subsequent exploration of other patterns determined by the spatiotemporal symmetry of the parameter changes, and the speeds of the path compared to the timescales of the pattern formation process itself. Each scenario enables the selection of different types of patterns and creates these elements in distinct ways, resulting in different features. Our approach extends the concept of selection involved in cellular decision-making, usually applied to cell-autonomous decisions, to systems that collectively make decisions through cell

  8. Biosynthesis and biotransformation of bile acids

    Directory of Open Access Journals (Sweden)

    Šarenac Tanja M.

    2017-01-01

    Full Text Available Bile acids are steroidal compounds, which contain 24 carbon atoms. They can be classified into two major groups: primary and secondary. The most abundant bile acids: The primary bile acids include cholic acid and chenodeoxycholic acid, while the major secondary bile acids are deoxycholic acid and litocholic acid. Bile acids are important physiological agents for intestinal absorption of nutrients and are used for biliary lipid secretion, toxic metabolites and xenobiotics. The aim of this paper is to analyze biosynthesis and biotransformation of bile acids, as preparation for practical usage in laboratory and clinical conditions. Topic: Biosynthesis and biotransformation of bile acids: The biosynthesis of bile acids is the dominant metabolic pathway for catabolism of cholesterol in humans. The classical route of biosynthesis of bile acids is embarking on the conversion of cholesterol into 7α-hydroxycholesterol using enzyme 7α-cholesterol hydroxylase (CYP7A1. This enzyme is one of the microsomal cytochrome P450 enzyme is localized exclusively in the liver. Classical road is the main road in the biosynthesis of bile acids, and its total contribution amounts to 90% for people, and 75% in mice. CYP 7A1 enzyme is considered to be sensitive to the inhibition of carbon monoxide, and the condition for the effect of NADPH, the oxygen, lecithin, and the NADPH-cytochrome P450 reductase. Bile acids are important signaling molecules and metabolic controls which activate the nuclear receptor and the G protein-coupled receptors (GPCR, a signaling lipid regulation of the liver, glucose and energy homeostasis. Also, bile acids maintain metabolic homeostasis. Biotransformation of bile acids: The conversion of cholesterol into bile acids just important for maintenance of cholesterol homeostasis, but also to prevent the accumulation of cholesterol, triglycerides and toxic metabolites as well as violations of the liver and other organs. Enterohepatic circulation of

  9. Physiological and biochemical characteristics of tobacco transgenic plants expressing bacterial dioxygenase

    Czech Academy of Sciences Publication Activity Database

    Piruzian, S.; Goldenkova, V.; Lenets, A.; Cvikrová, Milena; Macháčková, Ivana; Kobets, N.; Mett, V.

    2002-01-01

    Roč. 49, č. 6 (2002), s. 817-822 ISSN 1021-4437 R&D Projects: GA MŠk LN00A081; GA ČR GA206/00/1354 Institutional research plan: CEZ:AV0Z5038910 Keywords : phenylalanine ammonia lyase * polyphenol oxidases * biosynthesis Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 0.102, year: 2002

  10. Biochemical markers of bone turnover

    International Nuclear Information System (INIS)

    Kim, Deog Yoon

    1999-01-01

    Biochemical markers of bone turnover has received increasing attention over the past few years, because of the need for sensitivity and specific tool in the clinical investigation of osteoporosis. Bone markers should be unique to bone, reflect changes of bone less, and should be correlated with radiocalcium kinetics, histomorphometry, or changes in bone mass. The markers also should be useful in monitoring treatment efficacy. Although no bone marker has been established to meet all these criteria, currently osteocalcin and pyridinium crosslinks are the most efficient markers to assess the level of bone turnover in the menopausal and senile osteoporosis. Recently, N-terminal telopeptide (NTX), C-terminal telopeptide (CTX) and bone specific alkaline phosphatase are considered as new valid markers of bone turnover. Recent data suggest that CTX and free deoxypyridinoline could predict the subsequent risk of hip fracture of elderly women. Treatment of postmenopausal women with estrogen, calcitonin and bisphosphonates demonstrated rapid decrease of the levels of bone markers that correlated with the long-term increase of bone mass. Factors such as circadian rhythms, diet, age, sex, bone mass and renal function affect the results of biochemical markers and should be appropriately adjusted whenever possible. Each biochemical markers of bone turnover may have its own specific advantages and limitations. Recent advances in research will provide more sensitive and specific assays

  11. Biochemical markers of bone turnover

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Deog Yoon [College of Medicine, Kyunghee Univ., Seoul (Korea, Republic of)

    1999-08-01

    Biochemical markers of bone turnover has received increasing attention over the past few years, because of the need for sensitivity and specific tool in the clinical investigation of osteoporosis. Bone markers should be unique to bone, reflect changes of bone less, and should be correlated with radiocalcium kinetics, histomorphometry, or changes in bone mass. The markers also should be useful in monitoring treatment efficacy. Although no bone marker has been established to meet all these criteria, currently osteocalcin and pyridinium crosslinks are the most efficient markers to assess the level of bone turnover in the menopausal and senile osteoporosis. Recently, N-terminal telopeptide (NTX), C-terminal telopeptide (CTX) and bone specific alkaline phosphatase are considered as new valid markers of bone turnover. Recent data suggest that CTX and free deoxypyridinoline could predict the subsequent risk of hip fracture of elderly women. Treatment of postmenopausal women with estrogen, calcitonin and bisphosphonates demonstrated rapid decrease of the levels of bone markers that correlated with the long-term increase of bone mass. Factors such as circadian rhythms, diet, age, sex, bone mass and renal function affect the results of biochemical markers and should be appropriately adjusted whenever possible. Each biochemical markers of bone turnover may have its own specific advantages and limitations. Recent advances in research will provide more sensitive and specific assays.

  12. Physiological and biochemical responses of halophyte Kalidium ...

    African Journals Online (AJOL)

    In this study, the physiological and biochemical responses of a halophyte Kalidium foliatum to salinity were studied. In order to reflect salt-tolerance in K. foliatum and to analyze the physiological and biochemical mechanism for its salt tolerance, salinity threshold and biochemical parameters were studied. A halophyte ...

  13. Study of some biochemical and genetic risk factors for ...

    African Journals Online (AJOL)

    The aim of this study was to evaluate the relation between asymmetric dimethylarginine (ADMA ), high sensitive C-reactive protein (hs-CRP), monocyte chemoattractant protein-1 (MCP-1) ( both serum levels and the genotypes of the MCP-1 A-2518G polymorphism) with the development of carotid atherosclerosis in systemic ...

  14. Biochemical and genetic diagnosis of Smith-Lemli- Opitz syndrome ...

    African Journals Online (AJOL)

    brain development, when cholesterol binds covalently to sonic hedgehog protein involved in signalling to responding cells.[6] Engel king[7] investigated the pathogenesis in a .... QIAamp DNA Mini Kit from Qiagen (USA) and screened for mutations in the DHCR7 gene using quantitative realtime polymerase chain reaction ...

  15. Genetic and biochemical evidences reveal novel insights into the ...

    Indian Academy of Sciences (India)

    2016-10-26

    Oct 26, 2016 ... repair. Here, we reveal a crucial role for Sae2 in DNA replication stress. We show that different mutant alleles of SAE2 cause hypersensitivity to genotoxic agents, and when ... way, known as the DNA damage response is a collective cell ...... replication fork reversal catalyzed by Mycobacterium tubercu-.

  16. Assessment of genetic and biochemical diversity of ecologically ...

    African Journals Online (AJOL)

    USER

    2010-03-22

    Mar 22, 2010 ... g sucrose; 1 ml triton-X-100 and pH 8.0 was adjusted with conc. HCl) (Skovgaard and Rosendahl, ... The enzymes were separated by SDS polyacrylamide gel electro- phoresis in a discontinuous buffer ... Triton-x-100, 1 ml) and 2nd washing buffer (50 mM Tris buffer) in shaking condition, and then it was ...

  17. Genetic and biochemical evidences reveal novel insights into the ...

    Indian Academy of Sciences (India)

    2016-10-26

    mediated abrogation of replication stress- related defects in ... Branched DNA intermediates; DNA replication stress; genome integrity; MRX complex; Sae2 nuclease ...... stranded DNA structures into faster migrating cleavage prod-.

  18. Assessment of genetic and biochemical diversity of ecologically ...

    African Journals Online (AJOL)

    USER

    2010-03-22

    Mar 22, 2010 ... Protease, cellulase, glutamate dehydrogenase, pectinase and acid phosphatase designated band P220.16, C472.18, GLD130.21, Pe569.12 and AP472.12, respectively, were common in all the isolates and four monomorphic RAPD bands viz; 818, 512, 298 and 201 bp were also diversified in the isolates.

  19. Biochemical population genetics of the black mussel Choromytilus ...

    African Journals Online (AJOL)

    Gene products of nine loci were examined by horizontal starch-gel electrophoresis in five samples of black mussels, Choromytilus meridionalis, from the south-western Cape coast. Allelic frequency variation for four polymorphic proteins suggests no racial differences between west and south coast populations.

  20. Erythropoietic protoporphyria in Denmark: Demographic, biochemical and genetic characteristics

    DEFF Research Database (Denmark)

    Brusgaard, Klaus

    .7-408.6, and 0.8-21.5 µmol/L, respectively, and 4.7-110.0 (mean 41.6) and 0.8-7.2 (mean 2.8) µmol/L, respectively, when excluding 33 samples from the liver transplanted patient. All 608 samples except 10 had increased Zn-PPIX concentration in relation to the reference interval. DNA analysis was performed...

  1. Biochemical and genetic diagnosis of Smith-Lemli- Opitz syndrome ...

    African Journals Online (AJOL)

    -ups in four South African families of European ancestry with suspected SLOS in a range of presentations, from early fatality, congenital ... All the patients were of European ancestry, and the mutations reflected those in European studies.

  2. Biochemical population genetics of the black mussel Choromytilus ...

    African Journals Online (AJOL)

    Gene products of nine loci were examined by horizontal starch-gel electrophoresis in five samples of black ..... clinally from Hennanus to Marcus Island but the trend was not statistically significant. Discussion. Our results ... gene flow may proceed in a 'stepping-stone' fashion along the coast by sporadic recruitment of locally ...

  3. Assessment of genetic and biochemical diversity of ecologically ...

    African Journals Online (AJOL)

    Protease, cellulase, glutamate dehydrogenase, pectinase and acid phosphatase designated band P220.16, C472.18, GLD130.21, Pe569.12 and AP472.12, respectively, were common in all the isolates and four monomorphic RAPD bands viz; 818, 512, 298 and 201 bp were also diversified in the isolates. This common ...

  4. The p450 monooxygenase BcABA1 is essential for abscisic acid biosynthesis in Botrytis cinerea

    DEFF Research Database (Denmark)

    Siewers, V.; Smedsgaard, Jørn; Tudzynski, P.

    2004-01-01

    The phytopathogenic ascomycete Botrytis cinerea is known to produce abscisic acid (ABA), which is thought to be involved in host-pathogen interaction. Biochemical analyses had previously shown that, in contrast to higher plants, the fungal ABA biosynthesis probably does not proceed via carotenoids...... but involves direct cyclization of farnesyl diphosphate and subsequent oxidation steps. We present here evidence that this "direct" pathway is indeed the only one used by an ABA-overproducing strain of B. cinerea. Targeted inactivation of the gene bccpr1 encoding a cytochrome P450 oxidoreductase reduced...

  5. Complete Biosynthesis of Anthocyanins Using E. coli Polycultures.

    Science.gov (United States)

    Jones, J Andrew; Vernacchio, Victoria R; Collins, Shannon M; Shirke, Abhijit N; Xiu, Yu; Englaender, Jacob A; Cress, Brady F; McCutcheon, Catherine C; Linhardt, Robert J; Gross, Richard A; Koffas, Mattheos A G

    2017-06-06

    Fermentation-based chemical production strategies provide a feasible route for the rapid, safe, and sustainable production of a wide variety of important chemical products, ranging from fuels to pharmaceuticals. These strategies have yet to find wide industrial utilization due to their inability to economically compete with traditional extraction and chemical production methods. Here, we engineer for the first time the complex microbial biosynthesis of an anthocyanin plant natural product, starting from sugar. This was accomplished through the development of a synthetic, 4-strain Escherichia coli polyculture collectively expressing 15 exogenous or modified pathway enzymes from diverse plants and other microbes. This synthetic consortium-based approach enables the functional expression and connection of lengthy pathways while effectively managing the accompanying metabolic burden. The de novo production of specific anthocyanin molecules, such as calistephin, has been an elusive metabolic engineering target for over a decade. The utilization of our polyculture strategy affords milligram-per-liter production titers. This study also lays the groundwork for significant advances in strain and process design toward the development of cost-competitive biochemical production hosts through nontraditional methodologies. IMPORTANCE To efficiently express active extensive recombinant pathways with high flux in microbial hosts requires careful balance and allocation of metabolic resources such as ATP, reducing equivalents, and malonyl coenzyme A (malonyl-CoA), as well as various other pathway-dependent cofactors and precursors. To address this issue, we report the design, characterization, and implementation of the first synthetic 4-strain polyculture. Division of the overexpression of 15 enzymes and transcription factors over 4 independent strain modules allowed for the division of metabolic burden and for independent strain optimization for module-specific metabolite needs

  6. Rapid Identification of Airborne Biological Particles by Flow Cytometry, Gas Chromatography, and Genetic Probes

    National Research Council Canada - National Science Library

    Wick, Charles

    1997-01-01

    .... Physical characteristics of the particles, it was proposed, could be detected by flow cytometry, while their biochemical profiles could be determined by gas chromatography, and their genetic identity...

  7. Over-expression of the transcription factor HlMYB3 in transgenic hop (Humulus lupulus L. cv. Tettnanger) modulates the expression of genes involved in the biosynthesis of flavonoids and phloroglucinols

    Czech Academy of Sciences Publication Activity Database

    Gatica-Arias, A.; Stanke, M.; Häntzschel, K.R.; Matoušek, Jaroslav; Weber, G.

    2013-01-01

    Roč. 113, č. 2 (2013), s. 279-289 ISSN 0167-6857 R&D Projects: GA ČR GA521/08/0740 Institutional support: RVO:60077344 Keywords : Hop * R2R3 MYB transcription factors * Genetic transformation * Flavonoid biosynthesis Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.612, year: 2013

  8. Salt Stress Represses Soybean Seed Germination by Negatively Regulating GA Biosynthesis While Positively Mediating ABA Biosynthesis

    Directory of Open Access Journals (Sweden)

    Kai Shu

    2017-08-01

    Full Text Available Soybean is an important and staple oilseed crop worldwide. Salinity stress has adverse effects on soybean development periods, especially on seed germination and post-germinative growth. Improving seed germination and emergence will have positive effects under salt stress conditions on agricultural production. Here we report that NaCl delays soybean seed germination by negatively regulating gibberellin (GA while positively mediating abscisic acid (ABA biogenesis, which leads to a decrease in the GA/ABA ratio. This study suggests that fluridone (FLUN, an ABA biogenesis inhibitor, might be a potential plant growth regulator that can promote soybean seed germination under saline stress. Different soybean cultivars, which possessed distinct genetic backgrounds, showed a similar repressed phenotype during seed germination under exogenous NaCl application. Biochemical analysis revealed that NaCl treatment led to high MDA (malondialdehyde level during germination and the post-germinative growth stages. Furthermore, catalase, superoxide dismutase, and peroxidase activities also changed after NaCl treatment. Subsequent quantitative Real-Time Polymerase Chain Reaction analysis showed that the transcription levels of ABA and GA biogenesis and signaling genes were altered after NaCl treatment. In line with this, phytohormone measurement also revealed that NaCl considerably down-regulated active GA1, GA3, and GA4 levels, whereas the ABA content was up-regulated; and therefore ratios, such as GA1/ABA, GA3/ABA, and GA4/ABA, are decreased. Consistent with the hormonal quantification, FLUN partially rescued the delayed-germination phenotype caused by NaCl-treatment. Altogether, these results demonstrate that NaCl stress inhibits soybean seed germination by decreasing the GA/ABA ratio, and that FLUN might be a potential plant growth regulator that could promote soybean seed germination under salinity stress.

  9. The transcription factor AtMYB75/PAP1 regulates the expression of flavonoid biosynthesis genes in transgenic hop (Humulus lupulus L.)

    Czech Academy of Sciences Publication Activity Database

    Gatica-Arias, A.; Farag, M.A.; Häntzschel, K.R.; Matoušek, Jaroslav; Weber, G.

    2012-01-01

    Roč. 65, 7-8 (2012), s. 103-111 ISSN 1866-5195 R&D Projects: GA ČR GA521/08/0740 Institutional research plan: CEZ:AV0Z50510513 Institutional support: RVO:60077344 Keywords : metabolic engineering * Humulus lupulus L. * transcription factors * flavonoid biosynthesis Subject RIV: EB - Genetic s ; Molecular Biology

  10. GouR, a TetR Family Transcriptional Regulator, Coordinates the Biosynthesis and Export of Gougerotin in Streptomyces graminearus

    OpenAIRE

    Wei, Junhong; Tian, Yuqing; Niu, Guoqing; Tan, Huarong

    2014-01-01

    Gougerotin is a peptidyl nucleoside antibiotic. It functions as a specific inhibitor of protein synthesis by binding ribosomal peptidyl transferase and exhibits a broad spectrum of biological activities. gouR, situated in the gougerotin biosynthetic gene cluster, encodes a TetR family transcriptional regulatory protein. Gene disruption and genetic complementation revealed that gouR plays an important role in the biosynthesis of gougerotin. Transcriptional analysis suggested that GouR represse...

  11. Transformation of Aspergillus parasiticus with a homologous gene (pyrG) involved in pyrimidine biosynthesis

    International Nuclear Information System (INIS)

    Skory, C.D.; Horng, J.S.; Pestka, J.J.; Linz, J.E.

    1990-01-01

    The lack of efficient transformation methods for aflatoxigenic Aspergillus parasiticus has been a major constraint for the study of aflatoxin biosynthesis at the genetic level. A transformation system with efficiencies of 30 to 50 stable transformants per μg of DNA was developed for A. parasiticus by using homologous pyrG gene. The pyrG gene from A. parasiticus was isolated by in situ plaque hybridization of a lambda genomic DNA library. Uridine auxotrophs of A. parasiticus ATCC 36537, a mutant blocked in aflatoxin biosynthesis, were isolated by selection on 5-fluoroorotic acid following nitrosoguanidine mutagenesis. Isolates with mutations in the pyrG gene resulting in elimination of orotidine monophosphate (OMP) decarboxylase activity were detected by assaying cell extracts for their ability to convert [ 14 C]OMP to [ 14 C]UMP. Transformation of A. parasiticus pyrG protoplasts with the homologous pyrG gene restored the fungal cells to prototrophy. Enzymatic analysis of cell extracts of transformant clones demonstrated that these extracts had the ability to convert [ 14 C]OMP to [ 14 C]UMP. Southern analysis of DNA purified from transformant clones indicated that both pUC19 vector sequences and pyrG sequences were integrated into the genome. The development of this pyrG transformation system should allow cloning of the aflatoxin-biosynthetic genes, which will be useful in studying the regulation of aflatoxin biosynthesis and may ultimately provide a means for controlling aflatoxin production in the field

  12. The mbo operon is specific and essential for biosynthesis of mangotoxin in Pseudomonas syringae.

    Science.gov (United States)

    Carrión, Víctor J; Arrebola, Eva; Cazorla, Francisco M; Murillo, Jesús; de Vicente, Antonio

    2012-01-01

    Mangotoxin is an antimetabolite toxin produced by certain Pseudomonas syringae pv. syringae strains. This toxin is an oligopeptide that inhibits ornithine N-acetyl transferase, a key enzyme in the biosynthesis of ornithine and arginine. Previous studies have reported the involvement of the putative nonribosomal peptide synthetase MgoA in virulence and mangotoxin production. In this study, we analyse a new chromosomal region of P. syringae pv. syringae UMAF0158, which contains six coding sequences arranged as an operon (mbo operon). The mbo operon was detected in only mangotoxin-producing strains, and it was shown to be essential for the biosynthesis of this toxin. Mutants in each of the six ORFs of the mbo operon were partially or completely impaired in the production of the toxin. In addition, Pseudomonas spp. mangotoxin non-producer strains transformed with the mbo operon gained the ability to produce mangotoxin, indicating that this operon contains all the genetic information necessary for mangotoxin biosynthesis. The generation of a single transcript for the mbo operon was confirmed and supported by the allocation of a unique promoter and Rho-independent terminator. The phylogenetic analysis of the P. syringae strains harbouring the mbo operon revealed that these strains clustered together.

  13. The MIEL1 E3 Ubiquitin Ligase Negatively Regulates Cuticular Wax Biosynthesis in Arabidopsis Stems.

    Science.gov (United States)

    Lee, Hong Gil; Kim, Juyoung; Suh, Mi Chung; Seo, Pil Joon

    2017-07-01

    Cuticular wax is an important hydrophobic layer that covers the plant aerial surface. Cuticular wax biosynthesis is shaped by multiple layers of regulation. In particular, a pair of R2R3-type MYB transcription factors, MYB96 and MYB30, are known to be the main participants in cuticular wax accumulation. Here, we report that the MYB30-INTERACTING E3 LIGASE 1 (MIEL1) E3 ubiquitin ligase controls the protein stability of the two MYB transcription factors and thereby wax biosynthesis in Arabidopsis. MIEL1-deficient miel1 mutants exhibit increased wax accumulation in stems, with up-regulation of wax biosynthetic genes targeted by MYB96 and MYB30. Genetic analysis reveals that wax accumulation of the miel1 mutant is compromised by myb96 or myb30 mutation, but MYB96 is mainly epistatic to MIEL1, playing a predominant role in cuticular wax deposition. These observations indicate that the MIEL1-MYB96 module is important for balanced cuticular wax biosynthesis in developing inflorescence stems. © The Author 2017. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

  14. De Novo Assembly and Comparative Transcriptome Analysis Provide Insight into Lysine Biosynthesis in Toona sinensis Roem

    Directory of Open Access Journals (Sweden)

    Xia Zhang

    2016-01-01

    Full Text Available Toona sinensis Roem is a popular leafy vegetable in Chinese cuisine and is also used as a traditional Chinese medicine. In this study, leaf samples were collected from the same plant on two development stages and then used for high-throughput Illumina RNA-sequencing (RNA-Seq. 125,884 transcripts and 54,628 unigenes were obtained through de novo assembly. A total of 25,570 could be annotated with known biological functions, which indicated that the T. sinensis leaves and shoots were undergoing multiple developmental processes especially for active metabolic processes. Analysis of differentially expressed unigenes between the two libraries showed that the lysine biosynthesis was an enriched KEGG pathway, and candidate genes involved in the lysine biosynthesis pathway in T. sinensis leaves and shoots were identified. Our results provide a primary analysis of the gene expression files of T. sinensis leaf and shoot on different development stages and afford a valuable resource for genetic and genomic research on plant lysine biosynthesis.

  15. New Insights into the Biosynthesis Pathway of Polyketide Alkaloid Argimycins P in Streptomyces argillaceus

    Directory of Open Access Journals (Sweden)

    Suhui Ye

    2018-02-01

    Full Text Available Argimycins P are a recently identified family of polyketide alkaloids encoded by the cryptic gene cluster arp of Streptomyces argillaceus. These compounds contain either a piperideine ring, or a piperidine ring which may be fused to a five membered ring, and a polyene side chain, which is bound in some cases to an N-acetylcysteine moiety. The arp cluster consists of 11 genes coding for structural proteins, two for regulatory proteins and one for a hypothetical protein. Herein, we have characterized the post-piperideine ring biosynthesis steps of argimycins P through the generation of mutants in arp genes, the identification and characterization of compounds accumulated by those mutants, and cross-feeding experiments between mutants. Based in these results, a biosynthesis pathway is proposed assigning roles to every arp gene product. The regulation of the arp cluster is also addressed by inactivating/overexpressing the positive SARP-like arpRI and the negative TetR-like arpRII transcriptional regulators and determining the effect on argimycins P production, and through gene expression analyses (reverse transcription PCR and quantitative real-time PCR of arp genes in regulatory mutants in comparison to the wild type strain. These findings will contribute to deepen the knowledge on the biosynthesis of piperidine-containing polyketides and provide tools that can be used to generate new analogs by genetic engineering and/or biocatalysis.

  16. New Insights into the Biosynthesis Pathway of Polyketide Alkaloid Argimycins P in Streptomyces argillaceus

    Science.gov (United States)

    Ye, Suhui; Braña, Alfredo F.; González-Sabín, Javier; Morís, Francisco; Olano, Carlos; Salas, José A.; Méndez, Carmen

    2018-01-01

    Argimycins P are a recently identified family of polyketide alkaloids encoded by the cryptic gene cluster arp of Streptomyces argillaceus. These compounds contain either a piperideine ring, or a piperidine ring which may be fused to a five membered ring, and a polyene side chain, which is bound in some cases to an N-acetylcysteine moiety. The arp cluster consists of 11 genes coding for structural proteins, two for regulatory proteins and one for a hypothetical protein. Herein, we have characterized the post-piperideine ring biosynthesis steps of argimycins P through the generation of mutants in arp genes, the identification and characterization of compounds accumulated by those mutants, and cross-feeding experiments between mutants. Based in these results, a biosynthesis pathway is proposed assigning roles to every arp gene product. The regulation of the arp cluster is also addressed by inactivating/overexpressing the positive SARP-like arpRI and the negative TetR-like arpRII transcriptional regulators and determining the effect on argimycins P production, and through gene expression analyses (reverse transcription PCR and quantitative real-time PCR) of arp genes in regulatory mutants in comparison to the wild type strain. These findings will contribute to deepen the knowledge on the biosynthesis of piperidine-containing polyketides and provide tools that can be used to generate new analogs by genetic engineering and/or biocatalysis. PMID:29503641

  17. Integration of Transcriptome, Proteome and Metabolism Data Reveals the Alkaloids Biosynthesis in Macleaya cordata and Macleaya microcarpa

    Science.gov (United States)

    Liu, Fuqing; Huang, Peng; Zhu, Pengcheng; Chen, Jinjun; Shi, Mingming; Guo, Fang; Cheng, Pi; Zeng, Jing; Liao, Yifang; Gong, Jing; Zhang, Hong-Mei; Wang, Depeng; Guo, An-Yuan; Xiong, Xingyao

    2013-01-01

    Background The Macleaya spp., including Macleaya cordata and Macleaya microcarpa, are traditional anti-virus, inflammation eliminating, and insecticide herb medicines for their isoquinoline alkaloids. They are also known as the basis of the popular natural animal food addictive in Europe. However, few studies especially at genomics level were conducted on them. Hence, we performed the Macleaya spp. transcriptome and integrated it with iTRAQ proteome analysis in order to identify potential genes involved in alkaloids biosynthesis. Methodology and Principal Findings We elaborately designed the transcriptome, proteome and metabolism profiling for 10 samples of both species to explore their alkaloids biosynthesis. From the transcriptome data, we obtained 69367 and 78255 unigenes for M. cordata and M. microcarpa, in which about two thirds of them were similar to sequences in public databases. By metabolism profiling, reverse patterns for alkaloids sanguinarine, chelerythrine, protopine, and allocryptopine were observed in different organs of two species. We characterized the expressions of enzymes in alkaloid biosynthesis pathways. We also identified more than 1000 proteins from iTRAQ proteome data. Our results strongly suggest that the root maybe the organ for major alkaloids biosynthesis of Macleaya spp. Except for biosynthesis, the alkaloids storage and transport were also important for their accumulation. The ultrastructure of laticifers by SEM helps us to prove the alkaloids maybe accumulated in the mature roots. Conclusions/Significance To our knowledge this is the first study to elucidate the genetic makeup of Macleaya spp. This work provides clues to the identification of the potential modulate genes involved in alkaloids biosynthesis in Macleaya spp., and sheds light on researches for non-model medicinal plants by integrating different high-throughput technologies. PMID:23326424

  18. Heme biosynthesis and its regulation : Toward understanding and improvement of heme biosynthesis in filamentous fungi.

    NARCIS (Netherlands)

    S. de Weert; P.J. Punt; Christien Lokman; C.A. van den Hondel; A.C. Franken; A.F. Ram

    2011-01-01

    Heme biosynthesis in fungal host strains has acquired considerable interest in relation to the production of secreted heme-containing peroxidases. Class II peroxidase enzymes have been suggested as eco-friendly replacements of polluting chemical processes in industry. These peroxidases are naturally

  19. Heme biosynthesis and its regulation: Towards understanding and improvement of heme biosynthesis in filamentous fungi

    NARCIS (Netherlands)

    Franken, A.C.W.; Lokman, B.C.; Ram, A.F.J.; Punt, P.J.; Hondel, C.A.M.J.J. van den; Weert, S. de

    2011-01-01

    Heme biosynthesis in fungal host strains has acquired considerable interest in relation to the production of secreted heme-containing peroxidases. Class II peroxidase enzymes have been suggested as eco-friendly replacements of polluting chemical processes in industry. These peroxidases are naturally

  20. Functional conservation and divergence of Miscanthus lutarioriparius GT43 gene family in xylan biosynthesis.

    Science.gov (United States)

    Wang, Xiaoyu; Tang, Qi; Zhao, Xun; Jia, Chunlin; Yang, Xuanwen; He, Guo; Wu, Aimin; Kong, Yingzhen; Hu, Ruibo; Zhou, Gongke

    2016-04-26

    Xylan is the most abundant un-cellulosic polysaccharides of plant cell walls. Much progress in xylan biosynthesis has been gained in the model plant species Arabidopsis. Two homologous pairs Irregular Xylem 9 (IRX9)/9L and IRX14/14L from glycosyltransferase (GT) family 43 have been proved to play crucial roles in xylan backbone biosynthesis. However, xylan biosynthesis in grass such as Miscanthus remains poorly understood. We characterized seven GT43 members in M. lutarioriparius, a promising bioenergy crop. Quantitative real-time RT-PCR (qRT-PCR) analysis revealed that the expression of MlGT43 genes was ubiquitously detected in the tissues examined. In-situ hybridization demonstrated that MlGT43A-B and MlGT43F-G were specifically expressed in sclerenchyma, while MlGT43C-E were expressed in both sclerenchyma and parenchyma. All seven MlGT43 proteins were localized to Golgi apparatus. Overexpression of MlGT43A-E but not MlGT43F and MlGT43G in Arabidopsis irx9 fully or partially rescued the mutant defects, including morphological changes, collapsed xylem and increased xylan contents, whereas overexpression of MlGT43F and MlGT43G but not MlGT43A-E complemented the defects of irx14, indicating that MlGT43A-E are functional orthologues of IRX9, while MlGT43F and MlGT43G are functional orthologues of IRX14. However, overexpression of all seven MlGT43 genes could not rescue the mucilage defects of irx14 seeds. Furthermore, transient transactivation analyses of MlGT43A-E reporters demonstrated that MlGT43A and MlGT43B but not MlGT43C-E were differentially activated by MlSND1, MlMYB46 or MlVND7. The results demonstrated that all seven MlGT43s are functionally conserved in xylan biosynthesis during secondary cell wall formation but diversify in seed coat mucilage xylan biosynthesis. The results obtained provide deeper insight into xylan biosynthesis in grass, which lay the foundation for genetic modification of grass cell wall components and structure to better suit for next

  1. Genetic structure of Potentilla acaulis (Rosaceae) populations ...

    African Journals Online (AJOL)

    Jane

    2011-07-18

    Jul 18, 2011 ... Ecol. 10: 1811-1819. Bohonak AJ (1999). Dispersal, gene flow, and population structure. Q. Rev. Biol., 74: 21-45. Chen S, Xia T, Chen S, Zhou Y (2005). RAPD Profiling in Detecting. Genetic Variation in Endemic Coelonema (Brassicaceae) of Qinghai-. Tibet Plateau of China. Biochem. Genet., 43: 189-201.

  2. Unedoside derivatives in Nuxia and their biosynthesis

    DEFF Research Database (Denmark)

    Jensen, Søren Rosendal; Ravnkilde, Lene; Schripsema, Jan

    1998-01-01

    An investigation of two species of Nuxia showed that this genus is characterized by the presence of the eight-carbon iridoid glucoside unedoside and/or its derivatives. From N. floribunda unedoside, nuxioside (6-O-alpha-L-rhamnopyranosyl-unedoside) and 2 "-acetyl-3 "-cinnamoyl-nuxioside were...... isolated, while from N. oppositifolia 2 "-acetyl-3 "-benzoyl-nuxioside was obtained. Both plants contained verbascoside. The biosynthesis of unedoside in N. floribunda was investigated and deoxyloganic acid was found to be a precursor, similar to wh;lt was found for the eight-carbon iridoids in Thunbergia...

  3. Biosynthesis of Gold Nanoparticles Using Pseudomonas Aeruginosa

    International Nuclear Information System (INIS)

    Abd El-Aziz, M.; Badr, Y.; Mahmoud, M. A.

    2007-01-01

    Pseudomonas aeruginosa were used for extracellular biosynthesis of gold nanoparticles (Au NPs). Consequently, Au NPs were formed due to reduction of gold ion by bacterial cell supernatant of P. aeruginos ATCC 90271, P. aeruginos (2) and P. aeruginos (1). The UV-Vis. and fluorescence spectra of the bacterial as well as chemical prepared Au NPs were recorded. Transmission electron microscopy (TEM) micrograph showed the formation of well-dispersed gold nanoparticles in the range of 15-30 nm. The process of reduction being extracellular and may lead to the development of an easy bioprocess for synthesis of Au NPs

  4. Biosynthesis of silver nanoparticles using Saccharomyces cerevisiae.

    Science.gov (United States)

    Korbekandi, Hassan; Mohseni, Soudabeh; Mardani Jouneghani, Rasoul; Pourhossein, Meraj; Iravani, Siavash

    2016-01-01

    The objectives of this study were the biosynthesis of silver nanoparticles (NPs) by biotransformations using Saccharomyces cerevisiae and analysis of the sizes and shapes of the NPs produced. Dried and freshly cultured S. cerevisiae were used as the biocatalyst. Dried yeast synthesized few NPs, but freshly cultured yeast produced a large amount of them. Silver NPs were spherical, 2-20 nm in diameter, and the NPs with the size of 5.4 nm were the most frequent ones. NPs were seen inside the cells, within the cell membrane, attached to the cell membrane during the exocytosis, and outside of the cells.

  5. Metabolic engineering of ketocarotenoid biosynthesis in higher plants.

    Science.gov (United States)

    Zhu, Changfu; Naqvi, Shaista; Capell, Teresa; Christou, Paul

    2009-03-15

    Ketocarotenoids such as astaxanthin and canthaxanthin have important applications in the nutraceutical, cosmetic, food and feed industries. Astaxanthin is derived from beta-carotene by 3-hydroxylation and 4-ketolation at both ionone end groups. These reactions are catalyzed by beta-carotene hydroxylase and beta-carotene ketolase, respectively. The hydroxylation reaction is widespread in higher plants, but ketolation is restricted to a few bacteria, fungi, and some unicellular green algae. The recent cloning and characterization of beta-carotene ketolase genes in conjunction with the development of effective co-transformation strategies permitting facile co-integration of multiple transgenes in target plants provided essential resources and tools to produce ketocarotenoids in planta by genetic engineering. In this review, we discuss ketocarotenoid biosynthesis in general, and characteristics and functional properties of beta-carotene ketolases in particular. We also describe examples of ketocarotenoid engineering in plants and we conclude by discussing strategies to efficiently convert beta-carotene to astaxanthin in transgenic plants.

  6. Biosynthesis and pathway engineering of antifungal polyene macrolides in actinomycetes.

    Science.gov (United States)

    Kong, Dekun; Lee, Mi-Jin; Lin, Shuangjun; Kim, Eung-Soo

    2013-06-01

    Polyene macrolides are a large family of natural products typically produced by soil actinomycetes. Polyene macrolides are usually biosynthesized by modular and large type I polyketide synthases (PKSs), followed by several steps of sequential post-PKS modifications such as region-specific oxidations and glycosylations. Although known as powerful antibiotics containing potent antifungal activities (along with additional activities against parasites, enveloped viruses and prion diseases), their high toxicity toward mammalian cells and poor distribution in tissues have led to the continuous identification and structural modification of polyene macrolides to expand their general uses. Advances in in-depth investigations of the biosynthetic mechanism of polyene macrolides and the genetic manipulations of the polyene biosynthetic pathways provide great opportunities to generate new analogues. Recently, a novel class of polyene antibiotics was discovered (a disaccharide-containing NPP) that displays better pharmacological properties such as improved water-solubility and reduced hemolysis. In this review, we summarize the recent advances in the biosynthesis, pathway engineering, and regulation of polyene antibiotics in actinomycetes.

  7. ADP1 Affects Plant Architecture by Regulating Local Auxin Biosynthesis

    Science.gov (United States)

    Li, Shibai; Qin, Genji; Novák, Ondřej; Pěnčík, Aleš; Ljung, Karin; Aoyama, Takashi; Liu, Jingjing; Murphy, Angus; Gu, Hongya; Tsuge, Tomohiko; Qu, Li-Jia

    2014-01-01

    Plant architecture is one of the key factors that affect plant survival and productivity. Plant body structure is established through the iterative initiation and outgrowth of lateral organs, which are derived from the shoot apical meristem and root apical meristem, after embryogenesis. Here we report that ADP1, a putative MATE (multidrug and toxic compound extrusion) transporter, plays an essential role in regulating lateral organ outgrowth, and thus in maintaining normal architecture of Arabidopsis. Elevated expression levels of ADP1 resulted in accelerated plant growth rate, and increased the numbers of axillary branches and flowers. Our molecular and genetic evidence demonstrated that the phenotypes of plants over-expressing ADP1 were caused by reduction of local auxin levels in the meristematic regions. We further discovered that this reduction was probably due to decreased levels of auxin biosynthesis in the local meristematic regions based on the measured reduction in IAA levels and the gene expression data. Simultaneous inactivation of ADP1 and its three closest homologs led to growth retardation, relative reduction of lateral organ number and slightly elevated auxin level. Our results indicated that ADP1-mediated regulation of the local auxin level in meristematic regions is an essential determinant for plant architecture maintenance by restraining the outgrowth of lateral organs. PMID:24391508

  8. Thermodynamic analysis of biochemical systems

    International Nuclear Information System (INIS)

    Yuan, Y.; Fan, L.T.; Shieh, J.H.

    1989-01-01

    Introduction of the concepts of the availability (or exergy), datum level materials, and the dead state has been regarded as some of the most significant recent developments in classical thermodynamics. Not only the available energy balance but also the material and energy balances of a biological system may be established in reference to the datum level materials in the dead state or environment. In this paper these concepts are illustrated with two examples of fermentation and are shown to be useful in identifying sources of thermodynamic inefficiency, thereby leading naturally to the rational definition of thermodynamic efficiency of a biochemical process

  9. Biochemical structure of Calendula officinalis.

    Science.gov (United States)

    Korakhashvili, A; Kacharava, T; Kiknavelidze, N

    2007-01-01

    Calendula officinalis is a well known medicinal herb. It is common knowledge that its medicinal properties are conditioned on biologically active complex substances of Carotin (Provitamin A), Stearin, Triterpiniod, Plavonoid, Kumarin, macro and micro compound elements. Because of constant need in raw material of Calendula officinalis, features of its ontogenetic development agro-biological qualities in various eco regions of Georgia were investigated. The data of biologically active compounds, biochemical structure and the maintenance both in flowers and in others parts of plant is presented; the pharmacological activity and importance in medicine was reviewed.

  10. Enzyme and biochemical producing fungi

    DEFF Research Database (Denmark)

    Lübeck, Peter Stephensen; Lübeck, Mette; Nilsson, Lena

    2010-01-01

    We are developing a biorefinery concept for biological production of chemicals, drugs, feed and fuels using plant biomass as raw material in well-defined cell-factories. Among the important goals is the discovery of new biocatalysts for production of enzymes, biochemicals and fuels and already our...... screening of a large collection of fungal strains isolated from natural habitats have resulted in identification of strains with high production of hydrolytic enzymes and excretion of organic acids. Our research focuses on creating a fungal platform based on synthetic biology for developing new cell...

  11. Biochemical Abnormalities in Batten's Syndrome

    DEFF Research Database (Denmark)

    Clausen, Jytte Lene; Nielsen, Gunnar Gissel; Jensen, Gunde Egeskov

    1978-01-01

    The present data indicate that a group of ten patients with Batten's syndrome showed reduced activity of erythrocyte glutathione (GSH) peroxidase (Px) (glutathione: H2O2 oxidoreductase, EC 1.1.1.9.) using H2O2 as peroxide donor. Assay of erythrocyte GSHPx using H2O2, cumene hydroperoxide and t......-butyl hydroperoxide as donors also makes it possible biochemically to divide Batten's syndrome into two types: (1) one type with decreased values when H2O2 and cumene hydroperoxide are used, and (2) one type with increased values when t-butyl hydroperoxide is used. Furthermore an increased content of palmitic, oleic...

  12. Genus-wide physicochemical evidence of extracellular crystalline silver nanoparticles biosynthesis by Morganella spp.

    Directory of Open Access Journals (Sweden)

    Rasesh Y Parikh

    Full Text Available This study was performed to determine whether extracellular silver nanoparticles (AgNPs production is a genus-wide phenotype associated with all the members of genus Morganella, or only Morganella morganii RP-42 isolate is able to synthesize extracellular Ag nanoparticles. To undertake this study, all the available Morganella isolates were exposed to Ag+ ions, and the obtained nanoproducts were thoroughly analyzed using physico-chemical characterization tools such as transmission electron microscopy (TEM, UV-visible spectrophotometry (UV-vis, and X-ray diffraction (XRD analysis. It was identified that extracellular biosynthesis of crystalline silver nanoparticles is a unique biochemical character of all the members of genus Morganella, which was found independent of environmental changes. Significantly, the inability of other closely related members of the family Enterobacteriaceae towards AgNPs synthesis strongly suggests that AgNPs synthesis in the presence of Ag+ ions is a phenotypic character that is uniquely associated with genus Morganella.

  13. Non-ribosomal Peptide Synthases from Pseudomonas aeruginosa Play a Role in Cyclodipeptide Biosynthesis, Quorum-Sensing Regulation, and Root Development in a Plant Host.

    Science.gov (United States)

    González, Omar; Ortíz-Castro, Randy; Díaz-Pérez, César; Díaz-Pérez, Alma L; Magaña-Dueñas, Viridiana; López-Bucio, José; Campos-García, Jesús

    2017-04-01

    Diverse molecules mediate cross-kingdom communication between bacteria and their eukaryotic partners and determine pathogenic or symbiotic relationships. N-acyl-L-homoserine lactone-dependent quorum-sensing signaling represses the biosynthesis of bacterial cyclodipeptides (CDPs) that act as auxin signal mimics in the host plant Arabidopsis thaliana. In this work, we performed bioinformatics, biochemical, and plant growth analyses to identify non-ribosomal peptide synthase (NRPS) proteins of Pseudomonas aeruginosa, which are involved in CDP synthesis. A reverse genetics strategy allowed the identification of the genes encoding putative multi-modular-NRPS (MM-NRPS). Mutations in these genes affected the synthesis of the CDPs cyclo(L-Pro-L-Val), cyclo(L-Pro-L-Leu), and cyclo(L-Pro-L-Tyr), while showing wild-type-like levels of virulence factors, such as violacein, elastase, and pyocyanin. When analyzing the bioactivity of purified, naturally produced CDPs, it was found that cyclo(L-Pro-L-Tyr) and cyclo(L-Pro-L-Val) were capable of antagonizing quorum-sensing-LasR (QS-LasR)-dependent signaling in a contrasting manner in the cell-free supernatants of the selected NRPS mutants, which showed QS induction. Using a bacteria-plant interaction system, we further show that the pvdJ, ambB, and pchE P. aeruginosa mutants failed to repress primary root growth, but improved root branching in A. thaliana seedlings. These results indicated that the CDP production in P. aeruginosa depended on the functional MM-NRPS, which influences quorum-sensing of bacteria and plays a role in root architecture remodeling.

  14. Citrus fruit flavor and aroma biosynthesis: isolation, functional characterization, and developmental regulation of Cstps1, a key gene in the production of the sesquiterpene aroma compound valencene.

    Science.gov (United States)

    Sharon-Asa, Liat; Shalit, Moshe; Frydman, Ahuva; Bar, Einat; Holland, Doron; Or, Etti; Lavi, Uri; Lewinsohn, Efraim; Eyal, Yoram

    2003-12-01

    Citrus fruits possess unique aromas rarely found in other fruit species. While fruit flavor is composed of complex combinations of soluble and volatile compounds, several low-abundance sesquiterpenes, such as valencene, nootkatone, alpha-sinensal, and beta-sinensal, stand out in citrus as important flavor and aroma compounds. The profile of terpenoid volatiles in various citrus species and their importance as aroma compounds have been studied in detail, but much is still lacking in our understanding of the physiological, biochemical, and genetic regulation of their production. Here, we report on the isolation, functional expression, and developmental regulation of Cstps1, a sesquiterpene synthase-encoding gene, involved in citrus aroma formation. The recombinant enzyme encoded by Cstps1 was shown to convert farnesyl diphosphate to a single sesquiterpene product identified as valencene by gas chromatography-mass spectrometry (GC-MS). Phylogenetic analysis of plant terpene synthase genes localized Cstps1 to the group of angiosperm sesquiterpene synthases. Within this group, Cstps1 belongs to a subgroup of citrus sesquiterpene synthases. Cstps1 was found to be developmentally regulated: transcript was found to accumulate only towards fruit maturation, corresponding well with the timing of valencene accumulation in fruit. Although citrus fruits are non-climacteric, valencene accumulation and Cstps1 expression were found to be responsive to ethylene, providing further evidence for the role of ethylene in the final stages of citrus fruit ripening. Isolation of the gene encoding valencene synthase provides a tool for an in-depth study of the regulation of aroma compound biosynthesis in citrus and for metabolic engineering for fruit flavor characteristics.

  15. A comparative modeling and molecular docking study on Mycobacterium tuberculosis targets involved in peptidoglycan biosynthesis.

    Science.gov (United States)

    Fakhar, Zeynab; Naiker, Suhashni; Alves, Claudio N; Govender, Thavendran; Maguire, Glenn E M; Lameira, Jeronimo; Lamichhane, Gyanu; Kruger, Hendrik G; Honarparvar, Bahareh

    2016-11-01

    An alarming rise of multidrug-resistant Mycobacterium tuberculosis strains and the continuous high global morbidity of tuberculosis have reinvigorated the need to identify novel targets to combat the disease. The enzymes that catalyze the biosynthesis of peptidoglycan in M. tuberculosis are essential and noteworthy therapeutic targets. In this study, the biochemical function and homology modeling of MurI, MurG, MraY, DapE, DapA, Alr, and Ddl enzymes of the CDC1551 M. tuberculosis strain involved in the biosynthesis of peptidoglycan cell wall are reported. Generation of the 3D structures was achieved with Modeller 9.13. To assess the structural quality of the obtained homology modeled targets, the models were validated using PROCHECK, PDBsum, QMEAN, and ERRAT scores. Molecular dynamics simulations were performed to calculate root mean square deviation (RMSD) and radius of gyration (Rg) of MurI and MurG target proteins and their corresponding templates. For further model validation, RMSD and Rg for selected targets/templates were investigated to compare the close proximity of their dynamic behavior in terms of protein stability and average distances. To identify the potential binding mode required for molecular docking, binding site information of all modeled targets was obtained using two prediction algorithms. A docking study was performed for MurI to determine the potential mode of interaction between the inhibitor and the active site residues. This study presents the first accounts of the 3D structural information for the selected M. tuberculosis targets involved in peptidoglycan biosynthesis.

  16. Dissection of the transcriptional program regulating secondary wall biosynthesis during wood formation in poplar.

    Science.gov (United States)

    Zhong, Ruiqin; McCarthy, Ryan L; Lee, Chanhui; Ye, Zheng-Hua

    2011-11-01

    Wood biomass is mainly made of secondary cell walls; hence, elucidation of the molecular mechanisms underlying the transcriptional regulation of secondary wall biosynthesis during wood formation will be instrumental to design strategies for genetic improvement of wood biomass. Here, we provide direct evidence demonstrating that the poplar (Populus trichocarpa) wood-associated NAC domain transcription factors (PtrWNDs) are master switches activating a suite of downstream transcription factors, and together, they are involved in the coordinated regulation of secondary wall biosynthesis during wood formation. We show that transgenic poplar plants with dominant repression of PtrWNDs functions exhibit a drastic reduction in secondary wall thickening in woody cells, and those with PtrWND overexpression result in ectopic deposition of secondary walls. Analysis of PtrWND2B overexpressors revealed up-regulation of the expression of a number of wood-associated transcription factors, the promoters of which were also activated by PtrWND6B and the Eucalyptus EgWND1. Transactivation analysis and electrophoretic mobility shift assay demonstrated that PtrWNDs and EgWND1 activated gene expression through direct binding to the secondary wall NAC-binding elements, which are present in the promoters of several wood-associated transcription factors and a number of genes involved in secondary wall biosynthesis and modification. The WND-regulated transcription factors PtrNAC150, PtrNAC156, PtrNAC157, PtrMYB18, PtrMYB74, PtrMYB75, PtrMYB121, PtrMYB128, PtrZF1, and PtrGATA8 were able to activate the promoter activities of the biosynthetic genes for all three major wood components. Our study has uncovered that the WND master switches together with a battery of their downstream transcription factors form a transcriptional network controlling secondary wall biosynthesis during wood formation.

  17. Cloning and characterization of a potato StAN11 gene involved in anthocyanin biosynthesis regulation.

    Science.gov (United States)

    Li, Wang; Wang, Bing; Wang, Man; Chen, Min; Yin, Jing-Ming; Kaleri, Ghullam Murtaza; Zhang, Rui-Jie; Zuo, Tie-Niu; You, Xiong; Yang, Qing

    2014-04-01

    Anthocyanins are a class of products of plant secondary metabolism and are responsible for tubers color in potato. The biosynthesis of anthocyanins is a complex biological process, in which multiple genes are involved including structural genes and regulatory genes. In this study, StAN11, a WD40-repeat gene, was cloned from potato cultivar Chieftain (Solanum tuberosum L.). StAN11 (HQ599506) contained no intron and its open reading frame (ORF) was 1,029 bp long, encoding a putative protein of 342 amino acids. In order to verify its role in anthocyanin biosynthesis, StAN11 was inserted behind the CaMV-35S promoter of pCMBIA1304 and the recombination vector was introduced into the potato cultivar Désirée plants by Agrobacterium-mediated transformation. The color of transgenic tuber skin was significantly deepened, compared to the wild-type control, which was highly consistent with the accumulation of anthocyanin and expression of StAN11 in transgenic lines tuber skin. Further analysis on the expression of Flavonone-3-hydroxylase (F3H), Dihydroflavonol reductase (DFR), Anthocyanidin synthase (ANS), and Flavonoid 3-O-glucosyl transferase (3GT) in transgenic plants revealed that only DFR was upregulated. This result suggested that StAN11 regulated anthocyanin biosynthesis in potato by controlling DFR expression and accumulation of anthocyanin could be increased through overexpression of StAN11 in the tubers with the genetic background of anthocyanin biosynthesis. © 2013 Institute of Botany, Chinese Academy of Sciences.

  18. Characterization of Human and Yeast Mitochondrial Glycine Carriers with Implications for Heme Biosynthesis and Anemia.

    Science.gov (United States)

    Lunetti, Paola; Damiano, Fabrizio; De Benedetto, Giuseppe; Siculella, Luisa; Pennetta, Antonio; Muto, Luigina; Paradies, Eleonora; Marobbio, Carlo Marya Thomas; Dolce, Vincenza; Capobianco, Loredana

    2016-09-16

    Heme is an essential molecule in many biological processes, such as transport and storage of oxygen and electron transfer as well as a structural component of hemoproteins. Defects of heme biosynthesis in developing erythroblasts have profound medical implications, as represented by sideroblastic anemia. The synthesis of heme requires the uptake of glycine into the mitochondrial matrix where glycine is condensed with succinyl coenzyme A to yield δ-aminolevulinic acid. Herein we describe the biochemical and molecular characterization of yeast Hem25p and human SLC25A38, providing evidence that they are mitochondrial carriers for glycine. In particular, the hem25Δ mutant manifests a defect in the biosynthesis of δ-aminolevulinic acid and displays reduced levels of downstream heme and mitochondrial cytochromes. The observed defects are rescued by complementation with yeast HEM25 or human SLC25A38 genes. Our results identify new proteins in the heme biosynthetic pathway and demonstrate that Hem25p and its human orthologue SLC25A38 are the main mitochondrial glycine transporters required for heme synthesis, providing definitive evidence of their previously proposed glycine transport function. Furthermore, our work may suggest new therapeutic approaches for the treatment of congenital sideroblastic anemia. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  19. One-carbon metabolism and nucleotide biosynthesis as attractive targets for anticancer therapy.

    Science.gov (United States)

    Shuvalov, Oleg; Petukhov, Alexey; Daks, Alexandra; Fedorova, Olga; Vasileva, Elena; Barlev, Nickolai A

    2017-04-04

    Cancer-related metabolism has recently emerged as one of the "hallmarks of cancer". It has several important features, including altered metabolism of glucose and glutamine. Importantly, altered cancer metabolism connects different biochemical pathways into the one fine-tuned metabolic network, which stimulates high proliferation rates and plasticity to malignant cells. Among the keystones of cancer metabolism are one-carbon metabolism and nucleotide biosynthesis, which provide building blocks to anabolic reactions. Accordingly, the importance of these metabolic pathways for anticancer therapy has well been documented by more than fifty years of clinical use of specific metabolic inhibitors - methotrexate and nucleotides analogs. In this review we discuss one-carbon metabolism and nucleotide biosynthesis as common and specific features of many, if not all, tumors. The key enzymes involved in these pathways also represent promising anti-cancer therapeutic targets. We review different aspects of these metabolic pathways including their biochemistry, compartmentalization and expression of the key enzymes and their regulation at different levels. We also discuss the effects of known inhibitors of these pathways as well as the recent data on other enzymes of the same pathways as perspective pharmacological targets.

  20. Low-Molecular-Weight Metabolites from Diatoms: Structures, Biological Roles and Biosynthesis.

    Science.gov (United States)

    Stonik, Valentin; Stonik, Inna

    2015-06-09

    Diatoms are abundant and important biological components of the marine environment that biosynthesize diverse natural products. These microalgae are rich in various lipids, carotenoids, sterols and isoprenoids, some of them containing toxins and other metabolites. Several groups of diatom natural products have attracted great interest due to their potential practical application as energy sources (biofuel), valuable food constituents, and prospective materials for nanotechnology. In addition, hydrocarbons, which are used in climate reconstruction, polyamines which participate in biomineralization, new apoptotic agents against tumor cells, attractants and deterrents that regulate the biochemical communications between marine species in seawaters have also been isolated from diatoms. However, chemical studies on these microalgae are complicated by difficulties, connected with obtaining their biomass, and the influence of nutrients and contaminators in their environment as well as by seasonal and climatic factors on the biosynthesis of the corresponding natural products. Overall, the number of chemically studied diatoms is lower than that of other algae, but further studies, particularly those connected with improvements in the isolation and structure elucidation technique as well as the genomics of diatoms, promise both to increase the number of studied species with isolated biologically active natural products and to provide a clearer perception of their biosynthesis.

  1. Genomic analysis of the human gut microbiome suggests novel enzymes involved in quinone biosynthesis

    Directory of Open Access Journals (Sweden)

    Dmitry A Ravcheev

    2016-02-01

    Full Text Available Ubiquinone and menaquinone are membrane lipid-soluble carriers of electrons that are essential for cellular respiration. Eukaryotic cells can synthesize ubiquinone but not menaquinone, whereas prokaryotes can synthesize both quinones. So far, most of the human gut microbiome (HGM studies have been based on metagenomic analysis. Here, we applied an analysis of individual HGM genomes to the identification of ubiquinone and menaquinone biosynthetic pathways. In our opinion, the shift from metagenomics to analysis of individual genomes is a pivotal milestone in investigation of bacterial communities, including the HGM. The key results of this study are as follows. (i The distribution of the canonical pathways in the HGM genomes was consistent with previous reports and with the distribution of the quinone-dependent reductases for electron acceptors. (ii The comparative genomics analysis identified four alternative forms of the previously known enzymes for quinone biosynthesis. (iii Genes for the previously unknown part of the futalosine pathway were identified, and the corresponding biochemical reactions were proposed. We discuss the remaining gaps in the menaquinone and ubiquinone pathways in some of the microbes, which indicate the existence of further alternate genes or routes. Together, these findings provide further insight into the biosynthesis of quinones in bacteria and the physiology of the HGM.

  2. Bistability in biochemical signaling models.

    Science.gov (United States)

    Sobie, Eric A

    2011-09-20

    This Teaching Resource provides lecture notes, slides, and a student assignment for a two-part lecture on the principles underlying bistability in biochemical signaling networks, which are illustrated with examples from the literature. The lectures cover analog, or graded, versus digital, all-or-none, responses in cells, with examples from different types of biological processes requiring each. Rate-balance plots are introduced as a method for determining whether generic one-variable systems exhibit one or several stable steady states. Bifurcation diagrams are presented as a more general method for detecting the presence of bistability in biochemical signaling networks. The examples include an artificial toggle switch, the lac operon in bacteria, and the mitogen-activated protein kinase cascade in both Xenopus oocytes and mammalian cells. The second part of the lecture links the concepts of bistability more closely to the mathematical tools provided by dynamical systems analysis. The examples from the first part of the lecture are analyzed with phase-plane techniques and bifurcation analysis, using the scientific programming language MATLAB. Using these programs as a template, the assignment requires the students to implement a model from the literature and analyze the stability of this model's steady states.

  3. Biochemical abnormalities in Pearson syndrome.

    Science.gov (United States)

    Crippa, Beatrice Letizia; Leon, Eyby; Calhoun, Amy; Lowichik, Amy; Pasquali, Marzia; Longo, Nicola

    2015-03-01

    Pearson marrow-pancreas syndrome is a multisystem mitochondrial disorder characterized by bone marrow failure and pancreatic insufficiency. Children who survive the severe bone marrow dysfunction in childhood develop Kearns-Sayre syndrome later in life. Here we report on four new cases with this condition and define their biochemical abnormalities. Three out of four patients presented with failure to thrive, with most of them having normal development and head size. All patients had evidence of bone marrow involvement that spontaneously improved in three out of four patients. Unique findings in our patients were acute pancreatitis (one out of four), renal Fanconi syndrome (present in all patients, but symptomatic only in one), and an unusual organic aciduria with 3-hydroxyisobutyric aciduria in one patient. Biochemical analysis indicated low levels of plasma citrulline and arginine, despite low-normal ammonia levels. Regression analysis indicated a significant correlation between each intermediate of the urea cycle and the next, except between ornithine and citrulline. This suggested that the reaction catalyzed by ornithine transcarbamylase (that converts ornithine to citrulline) might not be very efficient in patients with Pearson syndrome. In view of low-normal ammonia levels, we hypothesize that ammonia and carbamylphosphate could be diverted from the urea cycle to the synthesis of nucleotides in patients with Pearson syndrome and possibly other mitochondrial disorders. © 2015 Wiley Periodicals, Inc.

  4. Isoprenoid Biosynthesis Inhibitors Targeting Bacterial Cell Growth.

    Science.gov (United States)

    Desai, Janish; Wang, Yang; Wang, Ke; Malwal, Satish R; Oldfield, Eric

    2016-10-06

    We synthesized potential inhibitors of farnesyl diphosphate synthase (FPPS), undecaprenyl diphosphate synthase (UPPS), or undecaprenyl diphosphate phosphatase (UPPP), and tested them in bacterial cell growth and enzyme inhibition assays. The most active compounds were found to be bisphosphonates with electron-withdrawing aryl-alkyl side chains which inhibited the growth of Gram-negative bacteria (Acinetobacter baumannii, Klebsiella pneumoniae, Escherichia coli, and Pseudomonas aeruginosa) at ∼1-4 μg mL -1 levels. They were found to be potent inhibitors of FPPS; cell growth was partially "rescued" by the addition of farnesol or overexpression of FPPS, and there was synergistic activity with known isoprenoid biosynthesis pathway inhibitors. Lipophilic hydroxyalkyl phosphonic acids inhibited UPPS and UPPP at micromolar levels; they were active (∼2-6 μg mL -1 ) against Gram-positive but not Gram-negative organisms, and again exhibited synergistic activity with cell wall biosynthesis inhibitors, but only indifferent effects with other inhibitors. The results are of interest because they describe novel inhibitors of FPPS, UPPS, and UPPP with cell growth inhibitory activities as low as ∼1-2 μg mL -1 . © 2016 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  5. BIOSYNTHESIS AND PROPERTIES OF ANTIBIOTIC BATUMIN

    Directory of Open Access Journals (Sweden)

    V. V. Klochko

    2014-12-01

    Full Text Available Biosynthesis of antistaphylococcal antibiotic batumin under periodic conditions of Pseudomonas batumici growth has been studied. Antibiotic synthesis in fermenter occurred across the culture growth and achieved its maximal value after 50–55 hours. The active oxygen utilization by the producing strain was observed during 20–55 hours of fermentation with maximum after 40–45 hours. Antibiotic yield was 175–180 mg/l and depended on intensity of aeration. contrast to «freshly isolated» antibiotic after fermentation the long-term kept batumin has shown two identical by molecular mass peaks according to the chromato-mass spectrometric analysis. Taking into account of batumin molecule structure the conclusion has been made that the most probable isomerization type is keto-enolic tautomerism. At the same time batumin is diastereoisomer of kalimantacin A which has the same chemical structure. The optic rotation angle is [α]d25 = +56.3° for kalimantacin and [α]d25 = –13.5° for batumin. The simultaneous P. batumici growth and antibiotic biosynthesis and the ability of this molecule to optical isomerisation and keto-enolic forms formation allow us to suppose that batumin plays a certain role in metabolism of the producing strain.

  6. Essences in Metabolic Engineering of Lignan Biosynthesis

    Directory of Open Access Journals (Sweden)

    Honoo Satake

    2015-05-01

    Full Text Available Lignans are structurally and functionally diverse phytochemicals biosynthesized in diverse plant species and have received wide attentions as leading compounds of novel drugs for tumor treatment and healthy diets to reduce of the risks of lifestyle-related non-communicable diseases. However, the lineage-specific distribution and the low-amount of production in natural plants, some of which are endangered species, hinder the efficient and stable production of beneficial lignans. Accordingly, the development of new procedures for lignan production is of keen interest. Recent marked advances in the molecular and functional characterization of lignan biosynthetic enzymes and endogenous and exogenous factors for lignan biosynthesis have suggested new methods for the metabolic engineering of lignan biosynthesis cascades leading to the efficient, sustainable, and stable lignan production in plants, including plant cell/organ cultures. Optimization of light conditions, utilization of a wide range of elicitor treatments, and construction of transiently gene-transfected or transgenic lignan-biosynthesizing plants are mainly being attempted. This review will present the basic and latest knowledge regarding metabolic engineering of lignans based on their biosynthetic pathways and biological activities, and the perspectives in lignan production via metabolic engineering.

  7. Fatty acid biosynthesis in pea root plastids

    International Nuclear Information System (INIS)

    Stahl, R.J.; Sparace, S.A.

    1989-01-01

    Fatty acid biosynthesis from [1- 14 C]acetate was optimized in plastids isolated from primary root tips of 7-day-old germinating pea seeds. Fatty acid synthesis was maximum at approximately 80 nmoles/hr/mg protein in the presence of 200 μM acetate, 0.5 mM each of NADH, NADPH and CoA, 6 mM each of ATP and MgCl 2 , 1 mM each of the MnCl 2 and glycerol-3-phosphate, 15 mM KHCO 3 , and 0.1M Bis-tris-propane, pH 8.0 incubated at 35C. At the standard incubation temperature of 25C, fatty acid synthesis was linear from up to 6 hours with 80 to 100 μg/mL plastid protein. ATP and CoA were absolute requirements, whereas KHCO 3 , divalent cations and reduced nucleotides all improved activity by 80 to 85%. Mg 2+ and NADH were the preferred cation and nucleotide, respectively. Dithiothreitol and detergents were generally inhibitory. The radioactive products of fatty acid biosynthesis were approximately 33% 16:0, 10% 18:0 and 56% 18:1 and generally did not vary with increasing concentrations of each cofactor

  8. Molecular Regulation of Antibiotic Biosynthesis in Streptomyces

    Science.gov (United States)

    Liu, Gang; Chandra, Govind; Niu, Guoqing

    2013-01-01

    SUMMARY Streptomycetes are the most abundant source of antibiotics. Typically, each species produces several antibiotics, with the profile being species specific. Streptomyces coelicolor, the model species, produces at least five different antibiotics. We review the regulation of antibiotic biosynthesis in S. coelicolor and other, nonmodel streptomycetes in the light of recent studies. The biosynthesis of each antibiotic is specified by a large gene cluster, usually including regulatory genes (cluster-situated regulators [CSRs]). These are the main point of connection with a plethora of generally conserved regulatory systems that monitor the organism's physiology, developmental state, population density, and environment to determine the onset and level of production of each antibiotic. Some CSRs may also be sensitive to the levels of different kinds of ligands, including products of the pathway itself, products of other antibiotic pathways in the same organism, and specialized regulatory small molecules such as gamma-butyrolactones. These interactions can result in self-reinforcing feed-forward circuitry and complex cross talk between pathways. The physiological signals and regulatory mechanisms may be of practical importance for the activation of the many cryptic secondary metabolic gene cluster pathways revealed by recent sequencing of numerous Streptomyces genomes. PMID:23471619

  9. Rare cause of post-squalene disorder of cholesterol biosynthesis ...

    African Journals Online (AJOL)

    Errors of cholesterol biosynthesis represent a heterogeneous group of metabolic disorders. The aim of the authors of this article is to present a case of a patient with typical symptoms of a rare post-squalene disorder of cholesterol biosynthesis, its diagnostics and progress in neonatal period. The differential diagnosis of a ...

  10. Rapid biosynthesis of cadmium sulfide (CdS) nanoparticles using ...

    African Journals Online (AJOL)

    Rapid biosynthesis of cadmium sulfide (CdS) nanoparticles using culture supernatants of Escherichia coli ATCC 8739, Bacillus subtilis ATCC 6633 and Lactobacillus ... The process of extracellular and fast biosynthesis may help in the development of an easy and eco-friendly route for the synthesis of CdS nanoparticles.

  11. Ant trail pheromone biosynthesis is triggered by a neuropeptide hormone.

    Directory of Open Access Journals (Sweden)

    Man-Yeon Choi

    Full Text Available Our understanding of insect chemical communication including pheromone identification, synthesis, and their role in behavior has advanced tremendously over the last half-century. However, endocrine regulation of pheromone biosynthesis has progressed slowly due to the complexity of direct and/or indirect hormonal activation of the biosynthetic cascades resulting in insect pheromones. Over 20 years ago, a neurohormone, pheromone biosynthesis activating neuropeptide (PBAN was identified that stimulated sex pheromone biosynthesis in a lepidopteran moth. Since then, the physiological role, target site, and signal transduction of PBAN has become well understood for sex pheromone biosynthesis in moths. Despite that PBAN-like peptides (∼200 have been identified from various insect Orders, their role in pheromone regulation had not expanded to the other insect groups except for Lepidoptera. Here, we report that trail pheromone biosynthesis in the Dufour's gland (DG of the fire ant, Solenopsis invicta, is regulated by PBAN. RNAi knock down of PBAN gene (in subesophageal ganglia or PBAN receptor gene (in DG expression inhibited trail pheromone biosynthesis. Reduced trail pheromone was documented analytically and through a behavioral bioassay. Extension of PBAN's role in pheromone biosynthesis to a new target insect, mode of action, and behavioral function will renew research efforts on the involvement of PBAN in pheromone biosynthesis in Insecta.

  12. Establishing a toolkit for precursor-directed polyketide biosynthesis: exploring substrate promiscuities of acid-CoA ligases.

    Science.gov (United States)

    Go, Maybelle Kho; Chow, Jeng Yeong; Cheung, Vivian Wing Ngar; Lim, Yan Ping; Yew, Wen Shan

    2012-06-05

    Polyketides are chemically diverse and medicinally important biochemicals that are biosynthesized from acyl-CoA precursors by polyketide synthases. One of the limitations to combinatorial biosynthesis of polyketides has been the lack of a toolkit that describes the means of delivering novel acyl-CoA precursors necessary for polyketide biosynthesis. Using five acid-CoA ligases obtained from various plants and microorganisms, we biosynthesized an initial library of 79 acyl-CoA thioesters by screening each of the acid-CoA ligases against a library of 123 carboxylic acids. The library of acyl-CoA thioesters includes derivatives of cinnamyl-CoA, 3-phenylpropanoyl-CoA, benzoyl-CoA, phenylacetyl-CoA, malonyl-CoA, saturated and unsaturated aliphatic CoA thioesters, and bicyclic aromatic CoA thioesters. In our search for the biosynthetic routes of novel acyl-CoA precursors, we discovered two previously unreported malonyl-CoA derivatives (3-thiophenemalonyl-CoA and phenylmalonyl-CoA) that cannot be produced by canonical malonyl-CoA synthetases. This report highlights the utility and importance of determining substrate promiscuities beyond conventional substrate pools and describes novel enzymatic routes for the establishment of precursor-directed combinatorial polyketide biosynthesis.

  13. Starch Biosynthesis in Guard Cells But Not in Mesophyll Cells Is Involved in CO2-Induced Stomatal Closing1[OPEN

    Science.gov (United States)

    Stephan, Aaron B.; Schroeder, Julian I.

    2016-01-01

    Starch metabolism is involved in stomatal movement regulation. However, it remains unknown whether starch-deficient mutants affect CO2-induced stomatal closing and whether starch biosynthesis in guard cells and/or mesophyll cells is rate limiting for high CO2-induced stomatal closing. Stomatal responses to [CO2] shifts and CO2 assimilation rates were compared in Arabidopsis (Arabidopsis thaliana) mutants that were either starch deficient in all plant tissues (ADP-Glc-pyrophosphorylase [ADGase]) or retain starch accumulation in guard cells but are starch deficient in mesophyll cells (plastidial phosphoglucose isomerase [pPGI]). ADGase mutants exhibited impaired CO2-induced stomatal closure, but pPGI mutants did not, showing that starch biosynthesis in guard cells but not mesophyll functions in CO2-induced stomatal closing. Nevertheless, starch-deficient ADGase mutant alleles exhibited partial CO2 responses, pointing toward a starch biosynthesis-independent component of the response that is likely mediated by anion channels. Furthermore, whole-leaf CO2 assimilation rates of both ADGase and pPGI mutants were lower upon shifts to high [CO2], but only ADGase mutants caused impairments in CO2-induced stomatal closing. These genetic analyses determine the roles of starch biosynthesis for high CO2-induced stomatal closing. PMID:27208296

  14. Assessment of genetic diversity of Lycoris longituba (Amaryllidaceae ...

    Indian Academy of Sciences (India)

    To take right measures to protect wild resources of L. longituba, it is important to study the range of genetic variation, genetic structure, diversification trend and other factors affecting genetic structure of the populations of this important and sparsely distributed species. Presently, many biochemical and DNA-based markers.

  15. Cloning and Biochemical Characterization of HIV Integrase

    Science.gov (United States)

    1989-12-15

    biochemical assays for the interaction of the HIV Intebration protein, IN, with a specific DNA target, the viral LTRs. The gene encoding IN has been subcloned ...FILE COPY CO CONTRACT NO.: DAMD7-88-C-8126 r TITLE: CLONING AND BIOCHEMICAL CHARACTERIZATION OF HIV INTEGRASE I- SPRINCIPAL INVESTIGATOR: ELLEN...ELEMENT No. NO. 392- NO. IACCESSION NO. 1 . TITLE (indude Security Classification)630A I315D49C08 (U) Cloning and Biochemical Characterization of the HIV

  16. Genetic characterization of a brangus-ibage cattle population: biochemical polymorphisms and reproductive efficiency Caracterização genética de uma população de bovinos brangus-ibagé: polimorfismos bioquímicos e eficiência reprodutiva

    Directory of Open Access Journals (Sweden)

    Luiz Ernani Henkes

    2000-10-01

    Full Text Available Biochemical techniques were used to investigate the genetic variability in a Brangus-Ibage population by determining allele frequencies of 18 blood protein systems: Hemogloin beta-Chain (Hb, Albumin (Alb, Amylase (Am, Transferrin (Tf, Carbonic Anhydrase (CA, Ceruloplasmin (Cp, Malic Enzyme (ME, Diaphorase I and II (Dia I and Dia II, Slow Alpha 2 Macroglobulin (Ap, Acid Phosphatase (ACP, Esterase B and D (EstB and EstD, Phosphogluconate Dehydrogenase (PGD, Glucose-6-Phosphate Dehydrogenase (G-6-PD, Glucose-Phosphate-Isomerase (GPI, Superoxide Dismutase (SOD and Glyoxalase I (GLO. The percentage of polymorphic loci were estimated at 0.27, the mean number of alleles was 1.33 and the mean heterozygosity was 0.07. There was a good agreement between expected and observed heterozygosity values. The population was in agreement with Hardy-Weinberg expectations in all systems. Reproductive records allowed to estimate three parameters of reproductive efficiency: mean age at first calving (1152.15 ± 166.60 days, mean calving interval (539.23 ± 124.10 days and mean weight at first calving (391.02 ± 37.59kg. No relationship was found between reproductive efficiency and genetic systems.Técnicas bioquímicas foram utilizadas para determinar a variabilidade genética numa população de bovinos da raça Brangus-Ibagé com relação a 18 sistemas protéicos sangüíneos: Hemoglobina - Cadeia beta (Hb, Albumina (Alb, Amilase (Am, Transferrina (Tf, Anidrase Carbônica (CA, Ceruloplasmina (Cp, Enzima Málica (ME, Diaforase I and II (Dia I and Dia II, Macroglobulina alfa2 lenta (Ap, Fosfatase Ácida (ACP, Esterase B and D (EstB and EstD, Fosfogliconato Desidrogenase (PGD, Glicose-6-Fosfato Desidrogenase (G-6-PD, Glicose-Fosfato-Isomerase (GPI, Superóxido Dismutase (SOD e Glioxalase I (GLO. O percentual de locos polimórficos foi estimado em 0,27, o número médio de alelos foi 1,33 e a heterozigosidade média foi de 0,07. Houve boa concordância entre a

  17. Engineering plastid fatty acid biosynthesis to improve food quality and biofuel production in higher plants.

    Science.gov (United States)

    Rogalski, Marcelo; Carrer, Helaine

    2011-06-01

    The ability to manipulate plant fatty acid biosynthesis by using new biotechnological approaches has allowed the production of transgenic plants with unusual fatty acid profile and increased oil content. This review focuses on the production of very long chain polyunsaturated fatty acids (VLCPUFAs) and the increase in oil content in plants using molecular biology tools. Evidences suggest that regular consumption of food rich in VLCPUFAs has multiple positive health benefits. Alternative sources of these nutritional fatty acids are found in cold-water fishes. However, fish stocks are in severe decline because of decades of overfishing, and also fish oils can be contaminated by the accumulation of toxic compounds. Recently, there is also an increase in oilseed use for the production of biofuels. This tendency is partly associated with the rapidly rising costs of petroleum, increased concern about the environmental impact of fossil oil and the attractive need to develop renewable sources of fuel. In contrast to this scenario, oil derived from crop plants is normally contaminant free and less environmentally aggressive. Genetic engineering of the plastid genome (plastome) offers a number of attractive advantages, including high-level foreign protein expression, marker-gene excision and transgene containment because of maternal inheritance of plastid genome in most crops. Here, we describe the possibility to improve fatty acid biosynthesis in plastids, production of new fatty acids and increase their content in plants by genetic engineering of plastid fatty acid biosynthesis via plastid transformation. © 2011 The Authors. Plant Biotechnology Journal © 2011 Society for Experimental Biology, Association of Applied Biologists and Blackwell Publishing Ltd.

  18. Biochemical diagnosis of mitochondrial disorders

    NARCIS (Netherlands)

    Rodenburg, R.J.T.

    2011-01-01

    Establishing a diagnosis in patients with a suspected mitochondrial disorder is often a challenge. Both knowledge of the clinical spectrum of mitochondrial disorders and the number of identified disease-causing molecular genetic defects are continuously expanding. The diagnostic examination of

  19. Biochemical adaptation of phytopathogenic fungi, Sclerotium rolfsii ...

    African Journals Online (AJOL)

    Biochemical adaptation of phytopathogenic fungi, Sclerotium rolfsii, in response to temperature stress. Natthiya Buensanteai, Kanjana Thumanu, Khanistha Kooboran, Dusit Athinuwat, Sutruedee Prathuangwong ...

  20. Biochemical Markers in Neurocritical Care

    Directory of Open Access Journals (Sweden)

    Omidvar Rezae

    2016-07-01

    Full Text Available During the past two decades, a variety of serum or cerebrospinal fluid (CSF biochemical markers in daily clinical practice have been recommended to diagnose and monitor diverse diseases or pathologic situations. It will be essential to develop a panel of biomarkers, to be suitable for evaluation of treatment efficacy, representing distinct phases of injury and recovery and consider the temporal profile of those. Among the possible and different biochemical markers, S100b appeared to fulfill many of optimized criteria of an ideal marker. S100b, a cytosolic low molecular weight dimeric calciumbinding protein from chromosome 21, synthesized in glial cells throughout the CNS, an homodimeric diffusible, belongs to a family of closely related protein, predominantly expressed by astrocytes and Schwann cells and a classic immunohistochemical marker for these cells, is implicated in brain development and neurophysiology. Of the 3 isoforms of S-100, the BB subunit (S100B is present in high concentrations in central and peripheral glial and Schwann cells, Langerhans and anterior pituitary cells, fat, muscle, and bone marrow tissues. The biomarker has shown to be a sensitive marker of clinical and subclinical cerebral damage, such as stroke, traumatic brain injury, and spinal cord injury. Increasing evidence suggests that the biomarker plays a double function as an intracellular regulator and an extracellular signal of the CNS. S100b is found in the cytoplasm in a soluble form and also is associated with intracellular membranes, centrosomes, microtubules, and type III intermediate filaments. Their genomic organization now is known, and many of their target proteins have been identified, although the mechanisms of regulating S100b secretion are not completely understood and appear to be related to many factors, such as the proinflammatory cytokines, tumor necrosis factor alpha (TNF-a, interleukin (IL-1b, and metabolic stress. 

  1. Linear analysis near a steady-state of biochemical networks: control analysis, correlation metrics and circuit theory

    Directory of Open Access Journals (Sweden)

    Qian Hong

    2008-05-01

    Full Text Available Abstract Background: Several approaches, including metabolic control analysis (MCA, flux balance analysis (FBA, correlation metric construction (CMC, and biochemical circuit theory (BCT, have been developed for the quantitative analysis of complex biochemical networks. Here, we present a comprehensive theory of linear analysis for nonequilibrium steady-state (NESS biochemical reaction networks that unites these disparate approaches in a common mathematical framework and thermodynamic basis. Results: In this theory a number of relationships between key matrices are introduced: the matrix A obtained in the standard, linear-dynamic-stability analysis of the steady-state can be decomposed as A = SRT where R and S are directly related to the elasticity-coefficient matrix for the fluxes and chemical potentials in MCA, respectively; the control-coefficients for the fluxes and chemical potentials can be written in terms of RT BS and ST BS respectively where matrix B is the inverse of A; the matrix S is precisely the stoichiometric matrix in FBA; and the matrix eAt plays a central role in CMC. Conclusion: One key finding that emerges from this analysis is that the well-known summation theorems in MCA take different forms depending on whether metabolic steady-state is maintained by flux injection or concentration clamping. We demonstrate that if rate-limiting steps exist in a biochemical pathway, they are the steps with smallest biochemical conductances and largest flux control-coefficients. We hypothesize that biochemical networks for cellular signaling have a different strategy for minimizing energy waste and being efficient than do biochemical networks for biosynthesis. We also discuss the intimate relationship between MCA and biochemical systems analysis (BSA.

  2. Role of MbtH-like Proteins in the Adenylation of Tyrosine during Aminocoumarin and Vancomycin Biosynthesis*

    Science.gov (United States)

    Boll, Björn; Taubitz, Tatjana; Heide, Lutz

    2011-01-01

    MbtH-like proteins consist of ∼70 amino acids and are encoded in the biosynthetic gene clusters of non-ribosomally formed peptides and other secondary metabolites derived from amino acids. Recently, several MbtH-like proteins have been shown to be required for the adenylation of amino acid in non-ribosomal peptide synthesis. We now investigated the role of MbtH-like proteins in the biosynthesis of the aminocoumarin antibiotics novobiocin, clorobiocin, and simocyclinone D8 and of the glycopeptide antibiotic vancomycin. The tyrosine-adenylating enzymes CloH, SimH, and Pcza361.18, involved in the biosynthesis of clorobiocin, simocyclinone D8, and vancomycin, respectively, required the presence of MbtH-like proteins in a 1:1 molar ratio, forming heterotetrameric complexes. In contrast, NovH, involved in novobiocin biosynthesis, showed activity in the absence of MbtH-like proteins. Comparison of the active centers of CloH and NovH showed only one amino acid to be different, i.e. Leu-383 versus Met-383. Mutation of this amino acid in CloH (L383M) indeed led to MbtH-independent adenylating activity. All investigated tyrosine-adenylating enzymes exhibited remarkable promiscuity for MbtH-like proteins from different pathways and organisms. YbdZ, the MbtH-like protein from the expression host Escherichia coli, was found to bind to adenylating enzymes during expression and to influence their biochemical properties markedly. Therefore, the use of ybdZ-deficient expression hosts is important in biochemical studies of adenylating enzymes. PMID:21890635

  3. Stress-induced neutral lipid biosynthesis in microalgae - Molecular, cellular and physiological insights.

    Science.gov (United States)

    Zienkiewicz, Krzysztof; Du, Zhi-Yan; Ma, Wei; Vollheyde, Katharina; Benning, Christoph

    2016-09-01

    Photosynthetic microalgae have promise as biofuel feedstock. Under certain conditions, they produce substantial amounts of neutral lipids, mainly in the form of triacylglycerols (TAGs), which can be converted to fuels. Much of our current knowledge on the genetic and molecular basis of algal neutral lipid metabolism derives mainly from studies of plants, i.e. seed tissues, and to a lesser extent from direct studies of algal lipid metabolism. Thus, the knowledge of TAG synthesis and the cellular trafficking of TAG precursors in algal cells is to a large extent based on genome predictions, and most aspects of TAG metabolism have yet to be experimentally verified. The biofuel prospects of microalgae have raised the interest in mechanistic studies of algal TAG biosynthesis in recent years and resulted in an increasing number of publications on lipid metabolism in microalgae. In this review we summarize the current findings on genetic, molecular and physiological studies of TAG accumulation in microalgae. Special emphasis is on the functional analysis of key genes involved in TAG synthesis, molecular mechanisms of regulation of TAG biosynthesis, as well as on possible mechanisms of lipid droplet formation in microalgal cells. This article is part of a Special Issue entitled: Plant Lipid Biology edited by Kent D. Chapman and Ivo Feussner. Copyright © 2016 Elsevier B.V. All rights reserved.

  4. Nature's combinatorial biosynthesis and recently engineered production of nucleoside antibiotics in Streptomyces.

    Science.gov (United States)

    Chen, Shawn; Kinney, William A; Van Lanen, Steven

    2017-04-01

    Modified nucleosides produced by Streptomyces and related actinomycetes are widely used in agriculture and medicine as antibacterial, antifungal, anticancer and antiviral agents. These specialized small-molecule metabolites are biosynthesized by complex enzymatic machineries encoded within gene clusters in the genome. The past decade has witnessed a burst of reports defining the key metabolic processes involved in the biosynthesis of several distinct families of nucleoside antibiotics. Furthermore, genome sequencing of various Streptomyces species has dramatically increased over recent years. Potential biosynthetic gene clusters for novel nucleoside antibiotics are now apparent by analysis of these genomes. Here we revisit strategies for production improvement of nucleoside antibiotics that have defined mechanisms of action, and are in clinical or agricultural use. We summarize the progress for genetically manipulating biosynthetic pathways for structural diversification of nucleoside antibiotics. Microorganism-based biosynthetic examples are provided and organized under genetic principles and metabolic engineering guidelines. We show perspectives on the future of combinatorial biosynthesis, and present a working model for discovery of novel nucleoside natural products in Streptomyces.

  5. Polyamine biosynthesis during germination of yeast ascospores.

    Science.gov (United States)

    Brawley, J V; Ferro, A J

    1979-01-01

    The role of the diamine putrescine during germination and outgrowth of ascospores of Saccharomyces cerevisiae was examined. Ornithine decarboxylase activity increased and declined rapidly during germination and outgrowth; peak activity was attained after the cells had proceeded through the G1 interval of the cell cycle, whereas minimal activity was present at the completion of the first cell division. alpha-Methylornithine inhibited both ornithine decarboxylase activity and the in vivo accumulation of putrescine. In the presence of alpha-methylornithireak dormancy and proceed through one cell division. Subsequent cellular growth, however, was retarded but not completely inhibited. The supplementation of Methylglyoxal bis(guanylhydrazone) to sporulation medium greatly inhibited this sexual process. These data suggest that the synthesis of putrescine is not required for the breaking of spore dormancy, but that polyamine biosynthesis may be essential for meiosis and sporulation. PMID:387744

  6. Biosynthesis and function of plant lipids

    International Nuclear Information System (INIS)

    Thomson, W.W.; Mudd, J.B.; Gibbs, M.

    1983-01-01

    The Sixth Annual Symposium in Botany and Plant Physiology was held January 13-15, 1983, at the University of California, Riverside. This volume comprises the papers that were presented. Subjects discussed at the symposium covered a wide range in the field of plant lipids. Biosynthesis of lipids occupied an important fraction of the presentations at the symposium. Subjects included detailed studies of the enzymes of fatty acid synthesis, several discussions of the incorporation of fatty acids into glycerolipids and the further modification of the fatty acids, and the synthesis of glycerolipids and desaturation of fatty acids in both maturing oilseeds and chloroplasts. The physicochemical studies of glycerolipids and sterols in artificial membranes have led to distinct conclusions about their behaviour which must be relevant in the biological membrane. Results on the functional consequences of modifying the galactolipid composition in the chloroplast were an encouraging sign of progress in the attempts to relate membrane lipid composition to physiological function

  7. Biosurfactant Mediated Biosynthesis of Selected Metallic Nanoparticles

    Science.gov (United States)

    Płaza, Grażyna A.; Chojniak, Joanna; Banat, Ibrahim M.

    2014-01-01

    Developing a reliable experimental protocol for the synthesis of nanomaterials is one of the challenging topics in current nanotechnology particularly in the context of the recent drive to promote green technologies in their synthesis. The increasing need to develop clean, nontoxic and environmentally safe production processes for nanoparticles to reduce environmental impact, minimize waste and increase energy efficiency has become essential in this field. Consequently, recent studies on the use of microorganisms in the synthesis of selected nanoparticles are gaining increased interest as they represent an exciting area of research with considerable development potential. Microorganisms are known to be capable of synthesizing inorganic molecules that are deposited either intra- or extracellularly. This review presents a brief overview of current research on the use of biosurfactants in the biosynthesis of selected metallic nanoparticles and their potential importance. PMID:25110864

  8. Terpenoids and Their Biosynthesis in Cyanobacteria

    Directory of Open Access Journals (Sweden)

    Bagmi Pattanaik

    2015-01-01

    Full Text Available Terpenoids, or isoprenoids, are a family of compounds with great structural diversity which are essential for all living organisms. In cyanobacteria, they are synthesized from the methylerythritol-phosphate (MEP pathway, using glyceraldehyde 3-phosphate and pyruvate produced by photosynthesis as substrates. The products of the MEP pathway are the isomeric five-carbon compounds isopentenyl diphosphate and dimethylallyl diphosphate, which in turn form the basic building blocks for formation of all terpenoids. Many terpenoid compounds have useful properties and are of interest in the fields of pharmaceuticals and nutrition, and even potentially as future biofuels. The MEP pathway, its function and regulation, and the subsequent formation of terpenoids have not been fully elucidated in cyanobacteria, despite its relevance for biotechnological applications. In this review, we summarize the present knowledge about cyanobacterial terpenoid biosynthesis, both regarding the native metabolism and regarding metabolic engineering of cyanobacteria for heterologous production of non-native terpenoids.

  9. Terpenoids and Their Biosynthesis in Cyanobacteria

    Science.gov (United States)

    Pattanaik, Bagmi; Lindberg, Pia

    2015-01-01

    Terpenoids, or isoprenoids, are a family of compounds with great structural diversity which are essential for all living organisms. In cyanobacteria, they are synthesized from the methylerythritol-phosphate (MEP) pathway, using glyceraldehyde 3-phosphate and pyruvate produced by photosynthesis as substrates. The products of the MEP pathway are the isomeric five-carbon compounds isopentenyl diphosphate and dimethylallyl diphosphate, which in turn form the basic building blocks for formation of all terpenoids. Many terpenoid compounds have useful properties and are of interest in the fields of pharmaceuticals and nutrition, and even potentially as future biofuels. The MEP pathway, its function and regulation, and the subsequent formation of terpenoids have not been fully elucidated in cyanobacteria, despite its relevance for biotechnological applications. In this review, we summarize the present knowledge about cyanobacterial terpenoid biosynthesis, both regarding the native metabolism and regarding metabolic engineering of cyanobacteria for heterologous production of non-native terpenoids. PMID:25615610

  10. A Molecular Description of Cellulose Biosynthesis

    Science.gov (United States)

    McNamara, Joshua T.; Morgan, Jacob L.W.; Zimmer, Jochen

    2016-01-01

    Cellulose is the most abundant biopolymer on Earth, and certain organisms from bacteria to plants and animals synthesize cellulose as an extracellular polymer for various biological functions. Humans have used cellulose for millennia as a material and an energy source, and the advent of a lignocellulosic fuel industry will elevate it to the primary carbon source for the burgeoning renewable energy sector. Despite the biological and societal importance of cellulose, the molecular mechanism by which it is synthesized is now only beginning to emerge. On the basis of recent advances in structural and molecular biology on bacterial cellulose synthases, we review emerging concepts of how the enzymes polymerize glucose molecules, how the nascent polymer is transported across the plasma membrane, and how bacterial cellulose biosynthesis is regulated during biofilm formation. Additionally, we review evolutionary commonalities and differences between cellulose synthases that modulate the nature of the cellulose product formed. PMID:26034894

  11. Biosurfactant Mediated Biosynthesis of Selected Metallic Nanoparticles

    Directory of Open Access Journals (Sweden)

    Grażyna A. Płaza

    2014-08-01

    Full Text Available Developing a reliable experimental protocol for the synthesis of nanomaterials is one of the challenging topics in current nanotechnology particularly in the context of the recent drive to promote green technologies in their synthesis. The increasing need to develop clean, nontoxic and environmentally safe production processes for nanoparticles to reduce environmental impact, minimize waste and increase energy efficiency has become essential in this field. Consequently, recent studies on the use of microorganisms in the synthesis of selected nanoparticles are gaining increased interest as they represent an exciting area of research with considerable development potential. Microorganisms are known to be capable of synthesizing inorganic molecules that are deposited either intra- or extracellularly. This review presents a brief overview of current research on the use of biosurfactants in the biosynthesis of selected metallic nanoparticles and their potential importance.

  12. Phylogenetic and evolutionary patterns in microbial carotenoid biosynthesis are revealed by comparative genomics.

    Directory of Open Access Journals (Sweden)

    Jonathan L Klassen

    Full Text Available BACKGROUND: Carotenoids are multifunctional, taxonomically widespread and biotechnologically important pigments. Their biosynthesis serves as a model system for understanding the evolution of secondary metabolism. Microbial carotenoid diversity and evolution has hitherto been analyzed primarily from structural and biosynthetic perspectives, with the few phylogenetic analyses of microbial carotenoid biosynthetic proteins using either used limited datasets or lacking methodological rigor. Given the recent accumulation of microbial genome sequences, a reappraisal of microbial carotenoid biosynthetic diversity and evolution from the perspective of comparative genomics is warranted to validate and complement models of microbial carotenoid diversity and evolution based upon structural and biosynthetic data. METHODOLOGY/PRINCIPAL FINDINGS: Comparative genomics were used to identify and analyze in silico microbial carotenoid biosynthetic pathways. Four major phylogenetic lineages of carotenoid biosynthesis are suggested composed of: (i Proteobacteria; (ii Firmicutes; (iii Chlorobi, Cyanobacteria and photosynthetic eukaryotes; and (iv Archaea, Bacteroidetes and two separate sub-lineages of Actinobacteria. Using this phylogenetic framework, specific evolutionary mechanisms are proposed for carotenoid desaturase CrtI-family enzymes and carotenoid cyclases. Several phylogenetic lineage-specific evolutionary mechanisms are also suggested, including: (i horizontal gene transfer; (ii gene acquisition followed by differential gene loss; (iii co-evolution with other biochemical structures such as proteorhodopsins; and (iv positive selection. CONCLUSIONS/SIGNIFICANCE: Comparative genomics analyses of microbial carotenoid biosynthetic proteins indicate a much greater taxonomic diversity then that identified based on structural and biosynthetic data, and divides microbial carotenoid biosynthesis into several, well-supported phylogenetic lineages not evident

  13. Biochemical survey for children of A-bomb survivors

    International Nuclear Information System (INIS)

    Sato, Chiyoko

    1992-01-01

    The Radiation Effects Research Foundation has conducted biochemical survey in children of A-bomb survivors, with the purpose of elucidating whether or not the rate of genetic mutation in genital cells is increased. This paper describes the previous surveys done at protein levels. Two kinds of indicators have been used: (1) 'rare mutation type' reflecting base substitution mutation, base deletion, and insertion; (2) 'mutation type' reflecting the decrease of red cell enzyme activity. According to the DS86 dosimetry system, the children population of A-bomb survivors were examined by dividing into the exposed group (n=11,364) of their parents exposed to 0.01 Sv or more and the control group (n=12,297) of those exposed to less than 0.01 Sv. 'Rare mutation type' was detected using electrophoresis in a total of 1,233 children in both groups. Of these children, 2 in the exposed group and 4 in the control group had a new 'mutation', i.e., mutation that was considered to have occurred in genital cells of their parents. Survey for genetic foci has revealed mutation in 2 children in the exposed group and 4 children in the control group, with the rate of mutation being 0.37 x 10 -5 /genetic foci/generation and 0.68 x 10 -5 /genetic foci/generation, respectively. Mutation type reflecting the decrease in red cell enzyme activity was seen in 26 in the exposed group and 21 in the control group. A total of 41 children were found to have been inherited from their parents. In the survey for genetic foci, only one had mutation in the exposed group, with the rate of mutation being 1.7 x 10 -5 /genetic foci/generation. These findings have revealed no evidence of significant difference in the rate of mutation between the exposed and control groups. Finally, the future genetic surveys at molecular levels are briefly discussed. (N.K.)

  14. The logic, experimental steps, and potential of heterologous natural product biosynthesis featuring the complex antibiotic erythromycin A produced through E. coli.

    Science.gov (United States)

    Jiang, Ming; Zhang, Haoran; Pfeifer, Blaine A

    2013-01-13

    The heterologous production of complex natural products is an approach designed to address current limitations and future possibilities. It is particularly useful for those compounds which possess therapeutic value but cannot be sufficiently produced or would benefit from an improved form of production. The experimental procedures involved can be subdivided into three components: 1) genetic transfer; 2) heterologous reconstitution; and 3) product analysis. Each experimental component is under continual optimization to meet the challenges and anticipate the opportunities associated with this emerging approach. Heterologous biosynthesis begins with the identification of a genetic sequence responsible for a valuable natural product. Transferring this sequence to a heterologous host is complicated by the biosynthetic pathway complexity responsible for product formation. The antibiotic erythromycin A is a good example. Twenty genes (totaling >50 kb) are required for eventual biosynthesis. In addition, three of these genes encode megasynthases, multi-domain enzymes each ~300 kDa in size. This genetic material must be designed and transferred to E. coli for reconstituted biosynthesis. The use of PCR isolation, operon construction, multi-cystronic plasmids, and electro-transformation will be described in transferring the erythromycin A genetic cluster to E. coli. Once transferred, the E. coli cell must support eventual biosynthesis. This process is also challenging given the substantial differences between E. coli and most original hosts responsible for complex natural product formation. The cell must provide necessary substrates to support biosynthesis and coordinately express the transferred genetic cluster to produce active enzymes. In the case of erythromycin A, the E. coli cell had to be engineered to provide the two precursors (propionyl-CoA and (2S)-methylmalonyl-CoA) required for biosynthesis. In addition, gene sequence modifications, plasmid copy number

  15. The Missing Link in Leguminous Pterocarpan Biosynthesis is a Dirigent Domain-Containing Protein with Isoflavanol Dehydratase Activity.

    Science.gov (United States)

    Uchida, Kai; Akashi, Tomoyoshi; Aoki, Toshio

    2017-02-01

    Pterocarpan forms the basic structure of leguminous phytoalexins, and most of the isoflavonoid pathway genes encoding the enzymes responsible for its biosynthesis have been identified. However, the last step of pterocarpan biosynthesis is a ring closure reaction, and the enzyme that catalyzes this step, 2'-hydroxyisoflavanol 4,2'-dehydratase or pterocarpan synthase (PTS), remains as an unidentified 'missing link'. This last ring formation is assumed to be the key step in determining the stereochemistry of pterocarpans, which plays a role in their antimicrobial activity. In this study, a cDNA clone encoding PTS from Glycyrrhiza echinata (GePTS1) was identified through functional expression fractionation screening of a cDNA library, which requires no sequence information, and orthologs from soybean (GmPTS1) and Lotus japonicus (LjPTS1) were also identified. These proteins were heterologously expressed in Escherichia coli and biochemically characterized. Surprisingly, the proteins were found to include amino acid motifs characteristic of dirigent proteins, some of which control stereospecific phenoxy radical coupling in lignan biosynthesis. The stereospecificity of substrates and products was examined using four substrate stereoisomers with hydroxy and methoxy derivatives at C-4'. The results showed that the 4R configuration was essential for the PTS reaction, and (-)- and (+)-pterocarpans were produced depending on the stereochemistry at C-3. In suspension-cultured soybean cells, levels of the GmPTS1 transcript increased temporarily prior to the peak in phytoalexin accumulation, strongly supporting the possible involvement of PTS in pterocarpan biosynthesis. © The Author 2017. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved.

  16. Cloning and characterization of genes involved in nostoxanthin biosynthesis of Sphingomonas elodea ATCC 31461.

    Directory of Open Access Journals (Sweden)

    Liang Zhu

    Full Text Available Most Sphingomonas species synthesize the yellow carotenoid nostoxanthin. However, the carotenoid biosynthetic pathway of these species remains unclear. In this study, we cloned and characterized a carotenoid biosynthesis gene cluster containing four carotenogenic genes (crtG, crtY, crtI and crtB and a β-carotene hydroxylase gene (crtZ located outside the cluster, from the gellan-gum producing bacterium Sphingomonas elodea ATCC 31461. Each of these genes was inactivated, and the biochemical function of each gene was confirmed based on chromatographic and spectroscopic analysis of the intermediates accumulated in the knockout mutants. Moreover, the crtG gene encoding the 2,2'-β-hydroxylase and the crtZ gene encoding the β-carotene hydroxylase, both responsible for hydroxylation of β-carotene, were confirmed by complementation studies using Escherichia coli producing different carotenoids. Expression of crtG in zeaxanthin and β-carotene accumulating E. coli cells resulted in the formation of nostoxanthin and 2,2'-dihydroxy-β-carotene, respectively. Based on these results, a biochemical pathway for synthesis of nostoxanthin in S. elodea ATCC 31461 is proposed.

  17. Cloning and characterization of genes involved in nostoxanthin biosynthesis of Sphingomonas elodea ATCC 31461.

    Science.gov (United States)

    Zhu, Liang; Wu, Xuechang; Li, Ou; Qian, Chaodong; Gao, Haichun

    2012-01-01

    Most Sphingomonas species synthesize the yellow carotenoid nostoxanthin. However, the carotenoid biosynthetic pathway of these species remains unclear. In this study, we cloned and characterized a carotenoid biosynthesis gene cluster containing four carotenogenic genes (crtG, crtY, crtI and crtB) and a β-carotene hydroxylase gene (crtZ) located outside the cluster, from the gellan-gum producing bacterium Sphingomonas elodea ATCC 31461. Each of these genes was inactivated, and the biochemical function of each gene was confirmed based on chromatographic and spectroscopic analysis of the intermediates accumulated in the knockout mutants. Moreover, the crtG gene encoding the 2,2'-β-hydroxylase and the crtZ gene encoding the β-carotene hydroxylase, both responsible for hydroxylation of β-carotene, were confirmed by complementation studies using Escherichia coli producing different carotenoids. Expression of crtG in zeaxanthin and β-carotene accumulating E. coli cells resulted in the formation of nostoxanthin and 2,2'-dihydroxy-β-carotene, respectively. Based on these results, a biochemical pathway for synthesis of nostoxanthin in S. elodea ATCC 31461 is proposed.

  18. Analysis of genetic diversity within and among four rabbit genotypes ...

    African Journals Online (AJOL)

    The results show a variation in biochemical activity levels since, esterase profiles showed higher percentage of polymorphism (67.21%) than peroxidase (34.78%) and SDS-protein profiles (39.11 %). Moreover, the mean of the genetic similarity within the genotypes based on overall biochemical markers were 0.81 (APRI), ...

  19. Deep sequencing of Lotus corniculatus L. reveals key enzymes and potential transcription factors related to the flavonoid biosynthesis pathway.

    Science.gov (United States)

    Wang, Ying; Hua, Wenping; Wang, Jian; Hannoufa, Abdelali; Xu, Ziqin; Wang, Zhezhi

    2013-04-01

    Lotus corniculatus L. is used worldwide as a forage crop due to its abundance of secondary metabolites and its ability to grow in severe environments. Although the entire genome of L. corniculatus var. japonicus R. is being sequenced, the differences in morphology and production of secondary metabolites between these two related species have led us to investigate this variability at the genetic level, in particular the differences in flavonoid biosynthesis. Our goal is to use the resulting information to develop more valuable forage crops and medicinal materials. Here, we conducted Illumina/Solexa sequencing to profile the transcriptome of L. corniculatus. We produced 26,492,952 short reads that corresponded to 2.38 gigabytes of total nucleotides. These reads were then assembled into 45,698 unigenes, of which a large number associated with secondary metabolism were annotated. In addition, we identified 2,998 unigenes based on homology with L. japonicus transcription factors (TFs) and grouped them into 55 families. Meanwhile, a comparison of four tag-based digital gene expression libraries, built from the flowers, pods, leaves, and roots, revealed distinct patterns of spatial expression of candidate unigenes in flavonoid biosynthesis. Based on these results, we identified many key enzymes from L. corniculatus which were different from reference genes of L. japonicus, and five TFs that are potential enhancers in flavonoid biosynthesis. Our results provide initial genetics resources that will be valuable in efforts to manipulate the flavonoid metabolic pathway in plants.

  20. Mutagenesis of NosM Leader Peptide Reveals Important Elements in Nosiheptide Biosynthesis

    Science.gov (United States)

    Jin, Liang; Wu, Xuri; Xue, Yanjiu; Jin, Yue; Wang, Shuzhen

    2016-01-01

    ABSTRACT Nosiheptide, a typical member of the ribosomally synthesized and posttranslationally modified peptides (RiPPs), exhibits potent activity against multidrug-resistant Gram-positive bacterial pathogens. The precursor peptide of nosiheptide (NosM) is comprised of a leader peptide with 37 amino acids and a core peptide containing 13 amino acids. To pinpoint elements in the leader peptide that are essential for nosiheptide biosynthesis, a collection of mutants with unique sequence features, including N- and C-terminal motifs, peptide length, and specific sites in the leader peptide, was generated by mutagenesis in vivo. The effects of various mutants on nosiheptide biosynthesis were evaluated. In addition to the necessity of a conserved motif LEIS box, native length and the N-terminal 12 amino acid residues were indispensable, and single-site substitutions of these 12 amino acid residues resulted in changes ranging from a greater-than-5-fold decrease to a 2-fold increase of nosiheptide production, depending on the sites and substituted residues. Moreover, although the C-terminal motif is not conservative, significant effects of this portion on nosiheptide production were also evident. Taken together, the present results further highlight the importance of the leader peptide in nosiheptide biosynthesis, and provide new insights into the diversity and specificity of leader peptides in the biosynthesis of various RiPPs. IMPORTANCE As a representative thiopeptide, nosiheptide exhibits excellent antibacterial activity. Although the biosynthetic gene cluster and several modification steps have been revealed, the presence and roles of the leader peptide within the precursor peptide of the nosiheptide gene cluster remain elusive. Thus, identification of specific elements in the leader peptide can significantly facilitate the genetic manipulation of the gene cluster for increasing nosiheptide production or generating diverse analogues. Given the complexity of the